uu.seUppsala University Publications
Change search
Refine search result
123 1 - 50 of 127
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf
Rows per page
  • 5
  • 10
  • 20
  • 50
  • 100
  • 250
Sort
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
  • Standard (Relevance)
  • Author A-Ö
  • Author Ö-A
  • Title A-Ö
  • Title Ö-A
  • Publication type A-Ö
  • Publication type Ö-A
  • Issued (Oldest first)
  • Issued (Newest first)
  • Created (Oldest first)
  • Created (Newest first)
  • Last updated (Oldest first)
  • Last updated (Newest first)
  • Disputation date (earliest first)
  • Disputation date (latest first)
Select
The maximal number of hits you can export is 250. When you want to export more records please use the 'Create feeds' function.
  • 1. Agudo, Marta
    et al.
    Perez-Marin, Maria Cruz
    Sobrado-Calvo, Paloma
    Lönngren, Ulrika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Salinas-Navarro, Manuel
    Canovas, Isabel
    Nadal-Nicolas, Francisco Manuel
    Miralles-Imperial, Jaime
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Vidal-Sanz, Manuel
    Immediate Upregulation of Proteins Belonging to Different Branches of the Apoptotic Cascade in the Retina after Optic Nerve Transection and Optic Nerve Crush2009In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 50, no 1, p. 424-431Article in journal (Refereed)
    Abstract [en]

    Purpose:

    To further investigate the molecular signals underlying optic nerve (ON) injury we have analyzed in adult control, ON transected and ON crushed retinas, the expression pattern and time-course regulation of the following proteins, all of which are linked to apoptosis through different pathways: Stat 1, Caspase 11 (inflammation and death), Cathepsins C and B (lysosomal death pathway), Calpain 1 (endoplasmic reticulum stress), Calreticulin (apoptosis marker), Jun (early response) and Ahr (cell cycle arrest).

    Methods:

    Adult female rats were subjected to either intraorbital optic nerve transection (IONT) or intraorbital optic nerve crush (IONC). Protein from naive and ON injured adult rat retinas was extracted at increasing time-points post-lesion and western blotting experiments carried out. For immnuhistofluorescence analyses, retinal ganglion cells (RGCs) were retrogradelly identified with fluorogold applied to the superior colliculi one week before injury.

    Results:

    Western blotting analyses revealed up-regulation of all the analyzed proteins as soon as 12 hours post-lesion (hpl) peaking at 48hpl, in agreement with our previous RNA studies1. Furthermore, immunohistofluorescence to radial sections show that all of them, but Stat1, are expressed by the primarily injured neurons, the RGCs, as seen by colocalization with FG.

    Conclusions:

    All analyzed proteins were up-regulated in the retina after IONT or IONC as soon as 12hpl, indicating that ON injury regulates several branches of the apoptotic cascade and suggesting that commitment to death might be an earlier event than previously anticipated.

  • 2. Agudo, Marta
    et al.
    Pérez-Marín, Maria Cruz
    Lönngren, Ulrika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Sobrado, Paloma
    Conesa, Ana
    Cánovas, Isabel
    Salinas-Navarro, Manuel
    Miralles-Imperial, Jaime
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Vidal-Sanz, Manuel
    Time course profiling of the retinal transcriptome after optic nerve transection and optic nerve crush2008In: Molecular Vision, ISSN 1090-0535, E-ISSN 1090-0535, Vol. 14, p. 1050-1063Article in journal (Refereed)
    Abstract [en]

    PURPOSE:

    A time-course analysis of gene regulation in the adult rat retina after intraorbital nerve crush (IONC) and intraorbital nerve transection (IONT).

    METHODS:

    RNA was extracted from adult rat retinas undergoing either IONT or IONC at increasing times post-lesion. Affymetrix RAE230.2 arrays were hybridized and analyzed. Statistically regulated genes were annotated and functionally clustered. Arrays were validated by means of quantative reverse transcription polymerase chain reaction (qRT-PCR) on ten regulated genes at two times post-lesion. Western blotting and immunohistofluorescence for four pro-apoptotic proteins were performed on naïve and injured retinas. Finally, custom signaling maps for IONT- and IONC-induced death response were generated (MetaCore, Genego Inc.).

    RESULTS:

    Here we show that over time, 3,219 sequences were regulated after IONT and 1,996 after IONC. Out of the total of regulated sequences, 1,078 were commonly regulated by both injuries. Interestingly, while IONT mainly triggers a gene upregulation-sustained over time, IONC causes a transitory downregulation. Functional clustering identified the regulation of high interest biologic processes, most importantly cell death wherein apoptosis was the most significant cluster. Ten death-related genes upregulated by both injuries were used for array validation by means of qRT-PCR. In addition, western blotting and immunohistofluorescence of total and active Caspase 3 (Casp3), tumor necrosis factor receptor type 1 associated death domain (TRADD), tumor necrosis factor receptor superfamily member 1a (TNFR1a), and c-fos were performed to confirm their protein regulation and expression pattern in naïve and injured retinas. These analyses demonstrated that for these genes, protein regulation followed transcriptional regulation and that these pro-apoptotic proteins were expressed by retinal ganglion cells (RGCs). MetaCore-based death-signaling maps show that several apoptotic cascades were regulated in the retina following optic nerve injury and highlight the similarities and differences between IONT and IONC in cell death profiling.

    CONCLUSIONS:

    This comprehensive time course retinal transcriptome study comparing IONT and IONC lesions provides a unique valuable tool to understand the molecular mechanisms underlying optic nerve injury and to design neuroprotective protocols.

  • 3.
    Althini, Sanna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bengtsson, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Usoskin, Dmitry
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Söderström, Stine
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kylberg, Annika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lindqvist, Eva
    Chuva de Sousa Lopes, Susana
    Olson, Lars
    Lindeberg, Jonas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Normal Nigrostriatal Innervation but Dopamine Dysfunction in Mice Carrying Hypomorphic Tyrosine Hydroxylase Alleles2003In: Journal of Neuroscience Research, ISSN 0360-4012, E-ISSN 1097-4547, Vol. 72, no 4, p. 444-453Article in journal (Refereed)
    Abstract [en]

    We investigated the use of the mouse tyrosine hydroxylase (TH) gene to drive knock-in constructs in catecholaminergic neurons. Two targeting constructs representing truncated forms of either of the BMP receptors ALK-2 or BMPR-II preceded by an internal ribosome entry site (IRES) were introduced into the 3' untranslated region of TH. An frt-flanked neomycin-resistance (neo(r)) cassette was placed in the 3' end of the targeting constructs. Mice homozygous for the knock-in alleles showed various degrees of hypokinetic behavior, depending mainly on whether the neo(r) cassette was removed. In situ hybridization and immunohistochemistry showed that TH mRNA and protein were variously down-regulated in these mouse strains. Reduced levels of dopamine and noradrenalin were found in several brain areas. However, number and morphology of neurons in substantia nigra and their projections to striatum appeared normal in the neo(r)-positive TH hypomorphic mice as examined by markers for L-aromatic amino acid decarboxylase and the dopamine transporter. Elimination of the neo(r) cassette from the knock-in alleles partially restored TH and dopamine levels. The present neo(r)-positive TH hypomorphic mice show that nigrostriatal innervation develops independently of TH and should find use as a model for conditions of reduced catecholamine synthesis, as seen in, for example, L-dihydroxyphenylalanine-responsive dystonia/infantile parkinsonism.

  • 4.
    Althini, Susanna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Usoskin, Dmitry
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kylberg, Annika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kaplan, Paul L.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Blocked MAP kinase activity selectively enhances neurotrophic growth responses2004In: Molecular and Cellular Neuroscience, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 25, no 2, p. 345-354Article in journal (Refereed)
    Abstract [en]

    Bone morphogenetic proteins (BMPs) 4 and 6 as well as MEK inhibitors PD98059 and U0126 potentiate neurotrophin 3 (NT3)- and neurturin (NTN)-induced neurite outgrowth and survival of peripheral neurons from the E9 chicken embryo. Preexposure to BMP4 or PD98059 was sufficient to prime the potentiation of subsequently added NT3. Phosphorylation of Erk2, induced by NT3, was reduced by MEK inhibition but unaffected by BMP signaling. Real-time PCR showed that neither BMP stimulation nor MEK inhibition increased Trk receptor expression and that the BMP-induced genes Smad6 and Id1 were not upregulated by PD98059. In contrast, both MEK inhibition and BMP signaling suppressed transcription of the serum-response element (SRE)-driven Egr1 gene. A reporter assay using NGF-stimulated PC12 cells demonstrated that MEK/Erk/Elk-driven transcriptional activity was inhibited by Smad1/5 and by PD98059. Thus, suppression of SRE-controlled transcription represents a likely convergence point for pathways regulating neurotrophic responses.

  • 5.
    Althini, Susanna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Usoskin, Dmitry
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kylberg, Annika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    ten Dijke, Peter
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bone morphogenetic protein signalling in NGF-stimulated PC12 cells2003In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 307, no 3, p. 632-639Article in journal (Refereed)
    Abstract [en]

    Bone morphogenetic proteins (BMPs) are shown to potentiate NGF-induced neuronal differentiation in PC12 phaeochromocytoma cells grown on collagen under low-serum conditions. Whereas, cell bodies remained rounded in control medium or with only BMPs present, addition of BMP4 or BMP6 robustly increased the neuritogenic effect of NGF within 2 days. NGF-increased phosphorylation of p44(Erk1) and p42(Erk2) between 2 and 24h was unaffected by addition of BMP6. PC12 cells transfected with the SBE(4x)-luc reporter showed that BMP4 significantly increased receptor-activated Smad activity. Expression of constitutively active BMP receptor ALK2 activating Smad1 and Smad5 resulted in a strong increase in the SBE(4x)-luc reporter response. Adding the inhibitory Smad7 drastically reduced this signal. In contrast to wild-type (wt) Smad5, a Smad5 variant lacking five Erk phosphorylation sites in the linker region (designated Smad5/5SA) showed a strong background transcriptional activity. A fusion construct (Gal4-Smad5/5SA) was also highly transcriptionally active. Addition of the MEK inhibitor U0126 to PC12 cells expressing Gal4-Smad5/wt did not increase background transcriptional activity. However, upon activation by constitutively active ALK2 both Gal4-Smad5/wt and Gal4-Smad5/5SA strongly stimulated transcription. The data show that serine residues of the linker region of Smad5 reduce spontaneous transcriptional activity and that NGF-activated Erk does not antagonise BMP signalling at this site. Hence, NGF and BMP signals are likely to interact further downstream at the transcriptional level in neuronal differentiation of the PC12 cells.

  • 6. Apostolova, Galina
    et al.
    Dorn, Roland
    Ka, Sojeong
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lundeberg, Joakim
    Liser, Keren
    Hakim, Vicky
    Brodski, Claude
    Michaelidis, Theologos M.
    Dechant, Georg
    Neurotransmitter phenotype-specific expression changes in developing sympathetic neurons2007In: Molecular and Cellular Neuroscience, ISSN 1044-7431, E-ISSN 1095-9327, Vol. 35, no 3, p. 397-408Article in journal (Refereed)
    Abstract [en]

    During late developmental phases individual sympathetic neurons undergo a switch from noradrenergic to cholinergic neurotransmission. This phenomenon of plasticity depends on target-derived signals in vivo and is triggered by neurotrophic factors in neuronal cultures. To analyze genome-wide expression differences between the two transmitter phenotypes we employed DNA microarrays. RNA expression profiles were obtained from chick paravertebral sympathetic ganglia, treated with neurotrophin 3, glial cell line-derived neurotrophic factor or ciliary neurotrophic factor, all of which stimulate cholinergic differentiation. Results were compared with the effect of nerve growth factor, which functions as a pro-noradrenergic stimulus. The gene set common to all three comparisons defined the noradrenergic and cholinergic synexpression groups. Several functional categories, such as signal transduction, G-protein-coupled signaling, cation transport, neurogenesis and synaptic transmission, were enriched in these groups. Experiments based on the prediction that some of the identified genes play a role in the neurotransmitter switch identified bone morphogenetic protein signaling as an inhibitor of cholinergic differentiation.

  • 7.
    Bengtsson, Henrik
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bone Morphogenetic Protein Receptors in the Nervous System: Neurotrophic Functions with Emphasis on Catecholaminergic Neurons2001Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Members of the transforming growth factor-β (TGF-β) superfamily exhibit a range of effects on a host of different cell types. They signal through heteromeric complexes of serine/threonine kinase receptors of type I and type II. Gene targeted mutations of both factors and receptors have revealed that many of them are involved in early embryonic development. This thesis examines the receptors for this superfamily in the nervous system, especially bone morphogenetic protein receptor type II (BMPR-II). It was cloned from chicken nervous tissue, and its and other receptors’ expression in peripheral ganglia, spinal cord and brain of chicken, rat and mouse were examined. BMPR-II, ActR-II and ActR-IA were abundantly expressed throughout development in the nervous system, however with temporal regulation. One ligand of BMPR-II, BMP-7, was found to act synergistically with NT-3 and GDNF on subsets of peripheral neurons to promote survival and neurite outgrowth. A knock-in mouse was generated, encoding a truncated form of BMPR-II coupled to the endogenous tyrosine hydroxylase (TH) gene with an internal ribosome entry site (IRES). For ES-cell selection, a neomycin resistance gene was incorporated into the construct. Homozygous mice carrying the knock-in allele exhibited a small, hypokinetic phenotype. Levels of dopamine, noradrenaline and serotonin were measured, and the catecholamines were found to be lowered, dopamine as much as 97% in the caudate nucleus. The low catecholamine levels may not be an effect of the truncated BMPR-II, but rather a consequence of the knock-in construct reducing TH transcriptional rate. The TH hypomorphic mouse strain generated could find use as a model for catecholamine impaired systems, as seen in Parkinson’s disease.

  • 8.
    Bengtsson, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Epifantseva, Irina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Åbrink, Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kylberg, Annika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Usoskin, Dmitry
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Generation and characterization of a Gdf1 conditional null allele2008In: Genesis, ISSN 1526-954X, E-ISSN 1526-968X, Vol. 46, no 7, p. 368-372Article in journal (Refereed)
    Abstract [en]

    Growth differentiation factor-1 (GDF1), a TGF-beta superfamily member, participates in early embryo patterning. Later functions are implied by the Gdf1 expression in the peripheral and central nervous system. Such roles of the gene have been difficult to study, because Gdf1 null mice die during late embryogenesis. Here, we report the production of a mouse carrying a conditional Gdf1 allele, with exon 2 flanked by loxP sites. Crossing these mice with CaMKIIalpha-Cre mice resulted in Gdf1 ablation in the forebrain postnatally. Such mice displayed no behavioral changes or altered expression levels in a set of hippocampal genes examined. However, excision of the floxed Gdf1 exon caused increased expression of the remaining part of the bicistronic Uog1-Gdf1 transcript in the hippocampus. This indicates that the transcript level is regulated by a negative feedback-loop, sensing presence of either the protein or the mRNA region encoded by Gdf1 exon 2.

  • 9.
    Blixt, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Keeping up with retinal photoreceptors and horizontal cells: Labelling and mapping of cells in the normal and diseased embryonic chicken retina2017Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    The childhood eye cancer retinoblastoma originates from the retina and its development is initiated while the foetus is in the uterus. Retinoblastoma has a reported incidence of 1 in 15-18 000 live births, and approximately 90% of all patients are diagnosed before the age of 5. The occurrence of retinoblastoma is usually detected by the parents and the most frequent symptoms are leukocoria (white pupillary reflex), strabismus (squinting) or if the child complains of visual problems. Retinoblastoma is diagnosed by examination under anaesthesia and documentation by RetCam. It is treated with various cytostatic agents, or by laser. If the treatment is unsuccessful, or there is a risk that the tumour cells will spread and form metastases, the eye is removed.

    Previous studies have indicated that the cell type from which the tumour arises, the cell-of-origin, may be the cone photoreceptors and/or their immediate interneuron, the horizontal cells. Determining the cell-of-origin for retinoblastoma is an important goal, however, understanding the molecular mechanisms that distinguish the photoreceptors and the horizontal cells from the other retinal cells may prove just as important for understanding this disease.

    The aim of my project has been to develop, optimise and validate methods to label, map and target expression to photoreceptors and horizontal cells in the chicken embryonic retina. We have successfully established several methods that test the expression pattern of conserved, regulatory DNA sequences, and have performed short- and long-term expression of various genes that have been reported to be involved in cell cycle regulation and cell fate determination. One of my most important findings was that a region from the RXRγ gene allowed us to specifically target the photoreceptors and horizontal cells. Our previous knowledge, together with the newly established tools, puts us an important step closer towards understanding the development and behaviour of the retinal photoreceptors and horizontal cells, however, further studies are of course needed.

    List of papers
    1. Whole Retinal Explants from Chicken Embryos for Electroporation and Chemical Reagent Treatments
    Open this publication in new window or tab >>Whole Retinal Explants from Chicken Embryos for Electroporation and Chemical Reagent Treatments
    2015 (English)In: Journal of Visualized Experiments, ISSN 1940-087X, E-ISSN 1940-087X, no 103, article id e53202Article in journal (Refereed) Published
    Abstract [en]

    The retina is a good model for the developing central nervous system. The large size of the eye and most importantly the accessibility for experimental manipulations in ovo/in vivo makes the chicken embryonic retina a versatile and very efficient experimental model. Although the chicken retina is easy to target in ovo by intraocular injections or electroporation, the effective and exact concentration of the reagents within the retina may be difficult to fully control. This may be due to variations of the exact injection site, leakage from the eye or uneven diffusion of the substances. Furthermore, the frequency of malformations and mortality after invasive manipulations such as electroporation is rather high. This protocol describes an ex ovo technique for culturing whole retinal explants from chicken embryos and provides a method for controlled exposure of the retina to reagents. The protocol describes how to dissect, experimentally manipulate, and culture whole retinal explants from chicken embryos. The explants can be cultured for approximately 24 hr and be subjected to different manipulations such as electroporation. The major advantages are that the experiment is not dependent on the survival of the embryo and that the concentration of the introduced reagent can be varied and controlled in order to determine and optimize the effective concentration. Furthermore, the technique is rapid, cheap and together with its high experimental success rate, it ensures reproducible results. It should be emphasized that it serves as an excellent complement to experiments performed in ovo.

    Keywords
    Developmental Biology, Issue 103, Culture, explant, eye, inhibitors, in ovo, in vitro, plasmid
    National Category
    Neurosciences
    Identifiers
    urn:nbn:se:uu:diva-268713 (URN)10.3791/53202 (DOI)000364222300048 ()
    Funder
    Swedish Childhood Cancer Foundation, PR2013-0104Swedish Research Council, 12187-18-3
    Available from: 2015-12-09 Created: 2015-12-09 Last updated: 2018-01-10
    2. The p53 co-activator Zac1 neither induces cell cycle arrest nor apoptosis in chicken Lim1 horizontal progenitor cells
    Open this publication in new window or tab >>The p53 co-activator Zac1 neither induces cell cycle arrest nor apoptosis in chicken Lim1 horizontal progenitor cells
    2015 (English)In: Cell Death Discovery, ISSN 2058-7716, Vol. 1, article id 15023Article in journal (Refereed) Published
    Abstract [en]

    Chicken horizontal progenitor cells are able to enter their final mitosis even in the presence of DNA damage despite having a functional p53-p21 system. This suggests that they are resistant to DNA damage and that the regulation of the final cell cycle of horizontal progenitor cells is independent of the p53-p21 system. The activity of p53 is regulated by positive and negative modulators, including the zinc finger containing transcription factor Zac1 (zinc finger protein that regulates apoptosis and cell cycle arrest). Zac1 interacts with and enhances the activity of p53, thereby inducing cell cycle arrest and apoptosis. In this work, we use a gain-of-function assay in which mouse Zac1 (mZac1) is overexpressed in chicken retinal progenitor cells to study the effect on the final cell cycle of horizontal progenitor cells. The results showed that overexpression of mZac1 induced expression of p21 in a p53-dependent way and arrested the cell cycle as well as triggered apoptosis in chicken non-horizontal retinal progenitor cells. The negative regulation of the cell cycle by mZac1 is consistent with its proposed role as a tumour-suppressor gene. However, the horizontal cells were not affected by mZac1 overexpression. They progressed into S- and late G2/M-phase despite overexpression of mZac1. The inability of mZac1 to arrest the cell cycle in horizontal progenitor cells support the notion that the horizontal cells are less sensitive to events that triggers the p53 system during their terminal and neurogenic cell cycle, compared with other retinal cells. These properties are associated with a cell that has a propensity to become neoplastic and thus with a cell that may develop retinoblastoma.

    Place, publisher, year, edition, pages
    Nature Publishing Group, 2015
    National Category
    Neurology Developmental Biology
    Research subject
    Neurology
    Identifiers
    urn:nbn:se:uu:diva-279034 (URN)10.1038/cddiscovery.2015.23 (DOI)
    Available from: 2016-02-28 Created: 2016-02-28 Last updated: 2017-02-17Bibliographically approved
    3. The zinc finger gene Nolz1 is controlled by retinoic acid and regulates the formation of chicken retinal progenitors and Lim3 expressing bipolar cells
    Open this publication in new window or tab >>The zinc finger gene Nolz1 is controlled by retinoic acid and regulates the formation of chicken retinal progenitors and Lim3 expressing bipolar cells
    (English)Article in journal (Refereed) Submitted
    National Category
    Neurosciences
    Identifiers
    urn:nbn:se:uu:diva-315608 (URN)
    Available from: 2017-02-16 Created: 2017-02-16 Last updated: 2018-01-13
    4. A regulatory sequence from the retinoid X receptor gamma gene directs expression to horizontal cells and photoreceptors in the embryonic chicken retina
    Open this publication in new window or tab >>A regulatory sequence from the retinoid X receptor gamma gene directs expression to horizontal cells and photoreceptors in the embryonic chicken retina
    2016 (English)In: Molecular Vision, ISSN 1090-0535, E-ISSN 1090-0535, Vol. 22, p. 1405-1420Article in journal (Refereed) Published
    Abstract [en]

    PURPOSE: Combining techniques of episomal vector gene-specific Cre expression and genomic integration using the piggyBac transposon system enables studies of gene expression-specific cell lineage tracing in the chicken retina. In this work, we aimed to target the retinal horizontal cell progenitors. METHODS: A 208 bp gene regulatory sequence from the chicken retinoid X receptor gammagene (RXRgamma208) was used to drive Cre expression. RXRgamma is expressed in progenitors and photoreceptors during development. The vector was combined with a piggyBac "donor" vector containing a floxed STOP sequence followed by enhanced green fluorescent protein (EGFP), as well as a piggyBac helper vector for efficient integration into the host cell genome. The vectors were introduced into the embryonic chicken retina with in ovo electroporation. Tissue electroporation targets specific developmental time points and in specific structures. RESULTS: Cells that drove Cre expression from the regulatory RXRgamma208 sequence excised the floxed STOP-sequence and expressed GFP. The approach generated a stable lineage with robust expression of GFP in retinal cells that have activated transcription from the RXRgamma208 sequence. Furthermore, GFP was expressed in cells that express horizontal or photoreceptor markers when electroporation was performed between developmental stages 22 and 28. Electroporation of a stage 12 optic cup gave multiple cell types in accordance with RXRgamma gene expression in the early retina. CONCLUSIONS: In this study, we describe an easy, cost-effective, and time-efficient method for testing regulatory sequences in general. More specifically, our results open up the possibility for further studies of the RXRgamma-gene regulatory network governing the formation of photoreceptor and horizontal cells. In addition, the method presents approaches to target the expression of effector genes, such as regulators of cell fate or cell cycle progression, to these cells and their progenitor.

    National Category
    Developmental Biology
    Identifiers
    urn:nbn:se:uu:diva-313804 (URN)000393909400001 ()28003731 (PubMedID)
    Funder
    Swedish Research Council, MH521.2013.3346Swedish Childhood Cancer Foundation
    Available from: 2017-01-24 Created: 2017-01-24 Last updated: 2017-11-29Bibliographically approved
    5. Overexpression of MYCN in the developing chicken retina increases the generation of photoreceptor cells
    Open this publication in new window or tab >>Overexpression of MYCN in the developing chicken retina increases the generation of photoreceptor cells
    Show others...
    (English)Manuscript (preprint) (Other academic)
    National Category
    Neurosciences
    Identifiers
    urn:nbn:se:uu:diva-315611 (URN)
    Available from: 2017-02-16 Created: 2017-02-16 Last updated: 2018-01-13
  • 10.
    Blixt, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    A regulatory sequence from the retinoid X receptor gamma gene directs expression to horizontal cells and photoreceptors in the embryonic chicken retina2016In: Molecular Vision, ISSN 1090-0535, E-ISSN 1090-0535, Vol. 22, p. 1405-1420Article in journal (Refereed)
    Abstract [en]

    PURPOSE: Combining techniques of episomal vector gene-specific Cre expression and genomic integration using the piggyBac transposon system enables studies of gene expression-specific cell lineage tracing in the chicken retina. In this work, we aimed to target the retinal horizontal cell progenitors. METHODS: A 208 bp gene regulatory sequence from the chicken retinoid X receptor gammagene (RXRgamma208) was used to drive Cre expression. RXRgamma is expressed in progenitors and photoreceptors during development. The vector was combined with a piggyBac "donor" vector containing a floxed STOP sequence followed by enhanced green fluorescent protein (EGFP), as well as a piggyBac helper vector for efficient integration into the host cell genome. The vectors were introduced into the embryonic chicken retina with in ovo electroporation. Tissue electroporation targets specific developmental time points and in specific structures. RESULTS: Cells that drove Cre expression from the regulatory RXRgamma208 sequence excised the floxed STOP-sequence and expressed GFP. The approach generated a stable lineage with robust expression of GFP in retinal cells that have activated transcription from the RXRgamma208 sequence. Furthermore, GFP was expressed in cells that express horizontal or photoreceptor markers when electroporation was performed between developmental stages 22 and 28. Electroporation of a stage 12 optic cup gave multiple cell types in accordance with RXRgamma gene expression in the early retina. CONCLUSIONS: In this study, we describe an easy, cost-effective, and time-efficient method for testing regulatory sequences in general. More specifically, our results open up the possibility for further studies of the RXRgamma-gene regulatory network governing the formation of photoreceptor and horizontal cells. In addition, the method presents approaches to target the expression of effector genes, such as regulators of cell fate or cell cycle progression, to these cells and their progenitor.

  • 11.
    Blixt, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lineage tracing of horizontal and photoreceptor cells in the embryonic chicken retina2016In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 57, no 12Article in journal (Refereed)
  • 12.
    Blixt, Maria K. E.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Konjusha, Dardan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ring, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Zinc finger gene nolz1 regulates the formation of retinal progenitor cells and suppresses the Lim3/Lhx3 phenotype of retinal bipolar cells in chicken retina2018In: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 247, no 4, p. 630-641Article in journal (Refereed)
    Abstract [en]

    Background: The zinc-finger transcription factor Nolz1 regulates spinal cord neuron development by interacting with the transcription factors Isl1, Lim1, and Lim3, which are also important for photoreceptors, horizontal and bipolar cells during retinal development. We, therefore, studied Nolz1 during retinal development.

    Results: Nolz1 expression was seen in two waves during development: one early (peak at embryonic day 3-4.5) in retinal progenitors and one late (embryonic day 8) in newly differentiated cells in the inner nuclear layer. Overexpression and knockdown showed that Nolz1 decreases proliferation and stimulates cell cycle withdrawal in retinal progenitors with effects on the generation of retinal ganglion cells, photoreceptors, and horizontal cells without triggering apoptosis. Overexpression of Nolz1 gave more p27 positive cells. Sustained overexpression of Nolz1 in the retina gave fewer Lim3/Lhx3 bipolar cells.

    Conclusions: We conclude that Nolz1 has multiple functions during development and suggest a mechanism in which Nolz1 initially regulates the proliferation state of the retinal progenitor cells and then acts as a repressor that suppresses the Lim3/Lhx3 bipolar cell phenotype at the time of bipolar cell differentiation.

  • 13.
    Blixt, Martin K. E.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Shirazi Fard, Shahrzad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    All-Ericsson, C
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Adding another piece to the retinoblastoma puzzle2015In: Cell Death and Disease, ISSN 2041-4889, E-ISSN 2041-4889, Vol. 6, article id e1957Article in journal (Refereed)
  • 14.
    Boije, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Generation of Retinal Neurons: Focus on the Proliferation and Differentiation of the Horizontal Cells and their Subtypes2011Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    We have used the chicken retina as a model for investigating cell cycle regulation and cell fate commitment during central nervous system development. This thesis focuses on the characterization of and commitment to the horizontal cell fate in the retina. Horizontal cells are interneurons that provide intraretinal signal processing prior to information relay to the brain. We have identified molecular markers that selectively distinguish the three subtypes of horizontal cells, previously described in the chicken retina based on morphology. Subtype specific birth-dating revealed that horizontal cell subtypes are generated consecutively by biased progenitors that are sensitive to the inhibitory effects of follistatin. Follistatin stimulates proliferation in progenitors by repressing the differentiation signal of activin. Initially, injection of follistatin led to a decrease in committed horizontal cells but as the inhibitory effect dissipated it resulted in an increased number of horizontal cells. During development committed horizontal cell progenitors migrate to the vitreal side of the retina where they become arrested in G2-phase for approximately two days. When the arrest is overcome the horizontal cell progenitors undergo ectopic mitosis followed by migration to their designated layer. The G2-phase arrest is not triggered or maintained by any of the classic G2-arrest pathways such as DNA damage or stress. Nevertheless, we show that the cyclin B1-Cdk1 complex has a central role in maintaining this G2-phase arrest. Two transcription factors, FoxN4 and Ptf1a, are required for the generation of horizontal cells. We show that these factors are also sufficient to promote horizontal cell fate. Overexpression of FoxN4 and Ptf1a resulted in an overproduction of horizontal- and amacrine cells at the expense of ganglion- and photoreceptor cells. We identified Atoh7, a transcription factor required for the generation of ganglion cells, as a Ptf1a transcriptional target for downregulation. Our data support a common horizontal/amacrine lineage separated from the ganglion/photoreceptor lineage by the action of Ptf1a. In conclusion, these data describe several novel characteristics of horizontal cells enhancing our understanding of neural development and cell fate commitment.

    List of papers
    1. Axon-bearing and axon-less horizontal cell subtypes are generated consecutively during chick retinal development from progenitors that are sensitive to follistatin
    Open this publication in new window or tab >>Axon-bearing and axon-less horizontal cell subtypes are generated consecutively during chick retinal development from progenitors that are sensitive to follistatin
    Show others...
    2008 (English)In: BMC Developmental Biology, ISSN 1471-213X, E-ISSN 1471-213X, Vol. 8, p. 46-Article in journal (Refereed) Published
    Abstract [en]

    BACKGROUND:

    Horizontal cells are retinal interneurons that modulate the output from photoreceptors. A rich literature on the morphological classification and functional properties of HCs in different animals exists, however, the understanding of the events underlying their development is still limited. In most vertebrates including chicken, two main horizontal cell (HC) subtypes are identified based on the presence or absence of an axon.

    RESULTS:

    In this work we have molecularly characterized three HC subtypes based on Lim1, Isl1, GABA and TrkA, a classification that is consistent with three chick HC subtypes previously defined by morphology. The axon-bearing and axon-less HC subpopulations molecularly defined by Lim1 and Isl1, are born consecutively on embryonic day (E) 3-4 and E4-5, respectively, and exhibit temporally distinguishable periods of migration. Their relative numbers are not adjusted by apoptosis. A sharp decrease of high endogenous levels of the activin-inhibitor follistatin at E3 coincides with the appearance of the Lim1 positive cells. Extending the follistatin exposure of the HC retinal progenitor cells by injection of follistatin at E3 increased the number of both Lim1- and Isl1 positive HCs when analysed at E9.

    CONCLUSION:

    The results imply that the axon-bearing and axon-less HC subgroups are defined early and are generated consecutively from a retinal progenitor cell population that is sensitive to the inhibitory action of follistatin. The results are consistent with a model wherein added follistatin causes HC-generating progenitors to proliferate beyond the normal period of HC generation, thus producing extra HCs of both types that migrate to the HC layer.

    National Category
    Biological Sciences
    Identifiers
    urn:nbn:se:uu:diva-102172 (URN)10.1186/1471-213X-8-46 (DOI)000255933400001 ()18439241 (PubMedID)
    Available from: 2009-05-05 Created: 2009-05-05 Last updated: 2017-12-13Bibliographically approved
    2. Horizontal cell progenitors arrest in G2-phase and undergo terminal mitosis on the vitreal side of the chick retina
    Open this publication in new window or tab >>Horizontal cell progenitors arrest in G2-phase and undergo terminal mitosis on the vitreal side of the chick retina
    2009 (English)In: Developmental Biology, ISSN 0012-1606, E-ISSN 1095-564X, Vol. 330, no 1, p. 105-113Article in journal (Refereed) Published
    Abstract [en]

    We have addressed the question when horizontal cells in the chick retina are generated and undergo their terminal mitosis. Horizontal cell progenitors replicate their DNA early and migrate bi-directionally to the horizontal cell layer. It was hypothesized that the cells undergo mitosis directly after replication and migrate as post-mitotic transition cells before differentiating to horizontal cells. However, our results show that cells expressing markers for the axon-bearing and the axon-less subtypes of horizontal cells undergo terminal mitosis while residing on the vitreal side of the retina. By combining horizontal cell transcription factors Lim1, Isl1 and Prox1 labeling with phospho-histone H3, a marker for mitosis, we demonstrate that all or a clear majority of vitreal mitoses are undertaken by the horizontal cell committed progenitors. The pattern of cells that incorporated the thymidine analogue EdU implied that the progenitors replicated their genome while migrating towards the vitreal side. Upon arrival to the vitreal retina they become arrested for about two days prior to mitosis. Hence, cells expressing horizontal cell markers are arrested in G2-phase on the vitreal side of the retina. These results support the existence of committed progenitors that give rise to horizontal cells and that those cells become arrested in G2-phase before undergoing terminal mitosis on the vitreal side of the retina followed by migration to the horizontal cell layer. The results also indicate that the regulation of the transition from G2-phase to mitosis is important for the development of these committed progenitor cells.

    Place, publisher, year, edition, pages
    Elsevier, 2009
    Keywords
    Cell cycle, Differentiation, Development, EdU, Prox1, Lim1, Stem cell
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-120381 (URN)10.1016/j.ydbio.2009.03.013 (DOI)000266300800010 ()19324032 (PubMedID)
    Available from: 2010-03-11 Created: 2010-03-11 Last updated: 2017-12-12Bibliographically approved
    3. The terminal mitosis of chicken retinal horizontal cells is preceded by a G2-Phase arrest that relies on the cyclin B1-Cdk1 complex but is independent of DNA damage.
    Open this publication in new window or tab >>The terminal mitosis of chicken retinal horizontal cells is preceded by a G2-Phase arrest that relies on the cyclin B1-Cdk1 complex but is independent of DNA damage.
    (English)Article in journal (Other academic) Submitted
    Identifiers
    urn:nbn:se:uu:diva-150879 (URN)
    Available from: 2011-04-11 Created: 2011-04-07 Last updated: 2011-07-01
    4. Temporal and spatial expression of transcription factors FoxN4, Ptf1a, Prox1, Isl1 and Lim1 mRNA in the developing chick retina
    Open this publication in new window or tab >>Temporal and spatial expression of transcription factors FoxN4, Ptf1a, Prox1, Isl1 and Lim1 mRNA in the developing chick retina
    2008 (English)In: Gene Expression Patterns, ISSN 1567-133X, E-ISSN 1872-7298, Vol. 8, no 2, p. 117-123Article in journal (Refereed) Published
    Abstract [en]

    Transcription factors are pivotal in regulating cell fate and development. We analyzed five transcription factors - FoxN4, Ptf1a, Prox1, Isl1 and Lim1 - with putative functions in the formation of early-generated retinal interneurons. A full-length chicken FoxN4 cDNA was characterized and in situ as well as RT-PCR showed that FoxN4 expression commenced already in the stage 12-14 optic vesicles. Ptf1a, Prox1, Isl1 and Lim1 expression appeared later by stage 20-24, concomitant with the first post-mitotic ganglion-, amacrine- and horizontal cells. The FoxN4 and Ptf1a expression was transient with peak levels by stage 32-35. Expression disappeared as the retinal progenitor cells differentiated. Prox1, Isl1 and Lim1 expression remained in several differentiated cells including the horizontal cells. The order of expression supports a scheme where Ptf1a and Prox1 is downstream of FoxN4 and that FoxN4 and Ptf1a have transient roles during fate specification while Prox1, Isl1 and Lim1 have roles that are important for the generation of the neuronal subtypes.

    Keywords
    In situ hybridization, Optic vesicle, Retina, Quantitative RT-PCR, RACE
    National Category
    Medical and Health Sciences
    Identifiers
    urn:nbn:se:uu:diva-87576 (URN)10.1016/j.modgep.2007.09.004 (DOI)000252714300009 ()18006384 (PubMedID)
    Available from: 2009-01-05 Created: 2008-12-30 Last updated: 2017-12-14Bibliographically approved
    5. Ptf1a/Rbpj complex inhibits ganglion cell fate and drives the specification of all horizontal cell subtypes in the chick retina
    Open this publication in new window or tab >>Ptf1a/Rbpj complex inhibits ganglion cell fate and drives the specification of all horizontal cell subtypes in the chick retina
    Show others...
    2011 (English)In: Developmental Biology, ISSN 0012-1606, E-ISSN 1095-564X, Vol. 358, no 2, p. 296-308Article in journal (Refereed) Published
    Abstract [en]

    During development, progenitor cells of the retina give rise to six principal classes of neurons and the Müller glial cells found within the adult retina. The pancreas transcription factor 1 subunit a (Ptf1a) encodes a basic-helix–loop–helix transcription factor necessary for the specification of horizontal cells and the majority of amacrine cell subtypes in the mouse retina. The Ptf1a-regulated genes and the regulation of Ptf1a activity by transcription cofactors during retinogenesis have been poorly investigated. Using a retrovirus-mediated gene transfer approach, we reported that Ptf1a was sufficient to promote the fates of amacrine and horizontal cells from retinal progenitors and inhibit retinal ganglion cell and photoreceptor differentiation in the chick retina. Both GABAergic H1 and non-GABAergic H3 horizontal cells were induced following the forced expression of Ptf1a. We describe Ptf1a as a strong, negative regulator of Atoh7 expression. Furthermore, the Rbpj-interacting domains of Ptf1a protein were required for its effects on cell fate specification. Together, these data provide a novel insight into the molecular basis of Ptf1a activity on early cell specification in the chick retina.

    Keywords
    Cell specification, Horizontal cell, Atoh7/Ath5, Retina; Chick
    National Category
    Medical and Health Sciences Basic Medicine
    Research subject
    Neuroscience
    Identifiers
    urn:nbn:se:uu:diva-150878 (URN)10.1016/j.ydbio.2011.07.033 (DOI)000295603900003 ()21839069 (PubMedID)
    Available from: 2011-04-09 Created: 2011-04-07 Last updated: 2018-01-12Bibliographically approved
    6. FoxN4 is sufficient for commitment to the retinal horizontal cell fate and is able to instigate differentiation programs in neural progenitors
    Open this publication in new window or tab >>FoxN4 is sufficient for commitment to the retinal horizontal cell fate and is able to instigate differentiation programs in neural progenitors
    (English)Manuscript (preprint) (Other academic)
    Identifiers
    urn:nbn:se:uu:diva-150880 (URN)
    Available from: 2011-04-11 Created: 2011-04-07 Last updated: 2011-07-01
  • 15.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Edqvist, Per-Henrik D.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Horizontal cell progenitors arrest in G2-phase and undergo terminal mitosis on the vitreal side of the chick retina2009In: Developmental Biology, ISSN 0012-1606, E-ISSN 1095-564X, Vol. 330, no 1, p. 105-113Article in journal (Refereed)
    Abstract [en]

    We have addressed the question when horizontal cells in the chick retina are generated and undergo their terminal mitosis. Horizontal cell progenitors replicate their DNA early and migrate bi-directionally to the horizontal cell layer. It was hypothesized that the cells undergo mitosis directly after replication and migrate as post-mitotic transition cells before differentiating to horizontal cells. However, our results show that cells expressing markers for the axon-bearing and the axon-less subtypes of horizontal cells undergo terminal mitosis while residing on the vitreal side of the retina. By combining horizontal cell transcription factors Lim1, Isl1 and Prox1 labeling with phospho-histone H3, a marker for mitosis, we demonstrate that all or a clear majority of vitreal mitoses are undertaken by the horizontal cell committed progenitors. The pattern of cells that incorporated the thymidine analogue EdU implied that the progenitors replicated their genome while migrating towards the vitreal side. Upon arrival to the vitreal retina they become arrested for about two days prior to mitosis. Hence, cells expressing horizontal cell markers are arrested in G2-phase on the vitreal side of the retina. These results support the existence of committed progenitors that give rise to horizontal cells and that those cells become arrested in G2-phase before undergoing terminal mitosis on the vitreal side of the retina followed by migration to the horizontal cell layer. The results also indicate that the regulation of the transition from G2-phase to mitosis is important for the development of these committed progenitor cells.

  • 16.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Edqvist, Per-Henrik D
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Temporal and spatial expression of transcription factors FoxN4, Ptf1a, Prox1, Isl1 and Lim1 mRNA in the developing chick retina2008In: Gene Expression Patterns, ISSN 1567-133X, E-ISSN 1872-7298, Vol. 8, no 2, p. 117-123Article in journal (Refereed)
    Abstract [en]

    Transcription factors are pivotal in regulating cell fate and development. We analyzed five transcription factors - FoxN4, Ptf1a, Prox1, Isl1 and Lim1 - with putative functions in the formation of early-generated retinal interneurons. A full-length chicken FoxN4 cDNA was characterized and in situ as well as RT-PCR showed that FoxN4 expression commenced already in the stage 12-14 optic vesicles. Ptf1a, Prox1, Isl1 and Lim1 expression appeared later by stage 20-24, concomitant with the first post-mitotic ganglion-, amacrine- and horizontal cells. The FoxN4 and Ptf1a expression was transient with peak levels by stage 32-35. Expression disappeared as the retinal progenitor cells differentiated. Prox1, Isl1 and Lim1 expression remained in several differentiated cells including the horizontal cells. The order of expression supports a scheme where Ptf1a and Prox1 is downstream of FoxN4 and that FoxN4 and Ptf1a have transient roles during fate specification while Prox1, Isl1 and Lim1 have roles that are important for the generation of the neuronal subtypes.

  • 17.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Harun-Or-Rashid, Mohammad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lee, Yu-Jen
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiology.
    Imsland, Freyja
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Bruneau, Nicolas
    Vieaud, Agathe
    Gourichon, David
    Tixier-Boichard, Michèle
    Bed’hom, Bertrand
    Andersson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Sonic Hedgehog-Signalling Patterns the Developing Chicken Comb as Revealed by Exploration of the Pea-comb Mutation2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 12, p. e50890-Article in journal (Refereed)
    Abstract [en]

    The genetic basis and mechanisms behind the morphological variation observed throughout the animal kingdom is stillrelatively unknown. In the present work we have focused on the establishment of the chicken comb-morphology byexploring the Pea-comb mutant. The wild-type single-comb is reduced in size and distorted in the Pea-comb mutant. Peacombis formed by a lateral expansion of the central comb anlage into three ridges and is caused by a mutation in SOX5,which induces ectopic expression of the SOX5 transcription factor in mesenchyme under the developing comb. Analysis ofdifferential gene expression identified decreased Sonic hedgehog (SHH) receptor expression in Pea-comb mesenchyme. Byexperimentally blocking SHH with cyclopamine, the wild-type single-comb was transformed into a Pea-comb-likephenotype. The results show that the patterning of the chicken comb is under the control of SHH and suggest that ectopicSOX5 expression in the Pea-comb change the response of mesenchyme to SHH signalling with altered combmorphogenesis as a result. A role for the mesenchyme during comb morphogenesis is further supported by the recentfinding that another comb-mutant (Rose-comb), is caused by ectopic expression of a transcription factor in combmesenchyme. The present study does not only give knowledge about how the chicken comb is formed, it also adds to ourunderstanding how mutations or genetic polymorphisms may contribute to inherited variations in the human face.

  • 18.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Ring, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Fard, Shahrzad Shirazi
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Grundberg, Ida
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Nilsson, Mats
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular tools. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Hallbook, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Alternative Splicing of the Chromodomain Protein Morf4l1 Pre-mRNA Has Implications on Cell Differentiation in the Developing Chicken Retina2013In: Journal of Molecular Neuroscience, ISSN 0895-8696, E-ISSN 1559-1166, Vol. 51, no 2, p. 615-628Article in journal (Refereed)
    Abstract [en]

    The proliferation, cell cycle exit and differentiation of progenitor cells are controlled by several different factors. The chromodomain protein mortality factor 4-like 1 (Morf4l1) has been ascribed a role in both proliferation and differentiation. Little attention has been given to the existence of alternative splice variants of the Morf4l1 mRNA, which encode two Morf41l isoforms: a short isoform (S-Morf4l1) with an intact chromodomain and a long isoform (L-Morf4l1) with an insertion in or in the vicinity of the chromodomain. The aim of this study was to investigate if this alternative splicing has a function during development. We analysed the temporal and spatial distribution of the two mRNAs and over-expressed both isoforms in the developing retina. The results showed that the S-Morf4l1 mRNA is developmentally regulated. Over-expression of S-Morf4l1 using a retrovirus vector produced a clear phenotype with an increase of early-born neurons: retinal ganglion cells, horizontal cells and cone photoreceptor cells. Over-expression of L-Morf4l1 did not produce any distinguishable phenotype. The over-expression of S-Morf4l1 but not L-Morf4l1 also increased apoptosis in the infected regions. Our results suggest that the two Morf4l1 isoforms have different functions during retinogenesis and that Morf4l1 functions are fine-tuned by developmentally regulated alternative splicing. The data also suggest that Morf4l1 contributes to the regulation of cell genesis in the retina.

  • 19.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Ring, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lopez-Gallardo, Meritxell
    Prada, Carmen
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Pax2 Is Expressed in a Subpopulation of Muller Cells in the Central Chick Retina2010In: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 239, no 6, p. 1858-1866Article in journal (Refereed)
    Abstract [en]

    Muller cells in the chick retina are generally thought to be a homogeneous population. We show that the transcription factor Pax2 is expressed by Muller cells in the central chick retina and its expression was first observed at stage 32 (embryonic day [E] 7.5). Birth-dating indicated that the majority of Pax2-positive Muller cells are generated between stage 29 and 33 (E5.5-E8). At stage 42 (E16), several Muller cell markers, such as Sox2 and 2M6, had reached the peripheral retina, while the Pax2 labeling extended approximately half-way. A similar pattern was maintained in the 6-month-old chicken. Neither the Pax2-positive nor the Pax2-negative Muller cells could be specifically associated to proliferative responses in the retina induced by growth factors or N-methyl-D-aspartate. Pax2 was not detected in Muller cells in mouse, rat, guinea-pig, rabbit, or pig retinas; but the zebrafish retina displayed a similar pattern of central Pax2-expressing Muller cells.

  • 20.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Shirazi Fard, S.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ring, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Forkheadbox N4 (FoxN4) triggers context-dependent differentiation in the developing chick retina and neural tube2013In: Differentiation, ISSN 0301-4681, E-ISSN 1432-0436, Vol. 85, no 1-2, p. 11-19Article in journal (Refereed)
    Abstract [en]

    FoxN4, a forkhead box transcription factor, is expressed in the chicken eye field and in retinal progenitor cells (RPCs) throughout development. FoxN4 labelling overlapped with that of Pax6 and Sox2, two crucial transcription factors for RPCs. Later, during neurogenesis in the retina, some cells were intensely and transiently labelled for FoxN4. These cells co-labelled for Lim1, a transcription factor expressed in early-born horizontal cells. The result suggests that high levels of FoxN4 combined with expression of Lim1 define a population of RPCs committed to the horizontal cell fate prior to their last apical mitosis. As these prospective horizontal cells develop, their FoxN4 expression is down-regulated. Previous results suggested that FoxN4 is important for the generation of horizontal and amacrine cells but that it is not sufficient for the generation of horizontal cells (Li et al., 2004). We found that over-expression of FoxN4 in embryonic day 3 chicken retina could activate horizontal cell markers Prox1 and Lim1, and that it generated numerous and ectopically located horizontal cells of both main subtypes. However, genes expressed in photoreceptors, amacrine and ganglion cells were also activated, indicating that FoxN4 triggered the expression of several differentiation factors. This effect was not exclusive for the retina but was also seen when FoxN4 was over-expressed in the mesencephalic neural tube. Combining the results from over-expression and wild-type expression data we suggest a model where a low level of FoxN4 is maintained in RPCs and that increased levels during a restricted period trigger neurogenesis and commitment of RPCs to the horizontal cell fate.

  • 21.
    Boije, Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Shirazi Fard, Shahrzad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ring, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Forkhead box N4 (FoxN4) triggers context-dependent differentiation in the developing chick retina and neural tubeIn: Differentiation, ISSN 0301-4681, E-ISSN 1432-0436Article in journal (Refereed)
  • 22. Chavarría, Teresa
    et al.
    Valenciano, Ana I.
    Mayordomo, Raquel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Egea, Joaquim
    Comella, Joan X.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    de Pablo, Flora
    de la Rosa, Enrique J.
    Differential, age-dependent MEK-ERK and PI3K-Akt activation by insulin acting as a survival factor during embryonic retinal development2007In: Developmental Neurobiology, ISSN 1932-8451, E-ISSN 1932-846X, Vol. 67, no 13, p. 1777-1788Article in journal (Refereed)
    Abstract [en]

    Programmed cell death is a genuine developmental process of the nervous system, affecting not only projecting neurons but also proliferative neuroepithelial cells and young neuroblasts. The embryonic chick retina has been employed to correlate in vivo and in vitro studies on cell death regulation. We characterize here the role of two major signaling pathways, PI3K-Akt and MEK-ERK, in controlled retinal organotypic cultures from embryonic day 5 (E5) and E9, when cell death preferentially affects proliferating neuroepithelial cells and ganglion cell neurons, respectively. The relative density of programmed cell death in vivo was much higher in the proliferative and early neurogenic stages of retinal development (E3-E5) than during neuronal maturation and synaptogenesis (E8-E19). In organotypic cultures from E5 and E9 retinas, insulin, as the only growth factor added, was able to completely prevent cell death induced by growth factor deprivation. Insulin activated both the PI3K-Akt and the MEK-ERK pathways. Insulin survival effect, however, was differentially blocked at the two stages. At E5, the effect was blocked by MEK inhibitors, whereas at E9 it was blocked by PI3K inhibitors. The cells which were found to be dependent on insulin activation of the MEK-ERK pathway at E5 were mostly proliferative neuroepithelial cells. These observations support a remarkable specificity in the regulation of early neural cell death.

  • 23.
    Clausen, Fredrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Hånell, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Israelsson, Charlotte
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hedin, Johanna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Mir, Anis K.
    Gram, Hermann
    Marklund, Niklas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Neutralization of interleukin-1 beta reduces cerebral edema and tissue loss and improves late cognitive outcome following traumatic brain injury in mice2011In: European Journal of Neuroscience, ISSN 0953-816X, E-ISSN 1460-9568, Vol. 34, no 1, p. 110-123Article in journal (Refereed)
    Abstract [en]

    Increasing evidence suggests that interleukin-1 beta (IL-1 beta) is a key mediator of the inflammatory response following traumatic brain injury (TBI). Recently, we showed that intracerebroventricular administration of an IL-1 beta-neutralizing antibody was neuroprotective following TBI in mice. In the present study, an anti-IL-1 beta antibody or control antibody was administered intraperitoneally following controlled cortical injury (CCI) TBI or sham injury in 105 mice and we extended our histological, immunological and behavioral analysis. First, we demonstrated that the treatment antibody reached target brain regions of brain-injured animals in high concentrations (> 11 nm) remaining up to 8 days post-TBI. At 48 h post-injury, the anti-IL-1b treatment attenuated the TBI-induced hemispheric edema (P < 0.05) but not the memory deficits evaluated using the Morris water maze (MWM). Neutralization of IL-1 beta did not influence the TBI-induced increases (P < 0.05) in the gene expression of the Ccl3 and Ccr2 chemokines, IL-6 or Gfap. Up to 20 days post-injury, neutralization of IL-1 beta was associated with improved visuospatial learning in the MWM, reduced loss of hemispheric tissue and attenuation of the microglial activation caused by TBI (P < 0.05). Motor function using the rotarod and cylinder tests was not affected by the anti-IL-1 beta treatment. Our results suggest an important negative role for IL-1 beta in TBI. The improved histological and behavioral outcome following anti-IL-1 beta treatment also implies that further exploration of IL-1 beta-neutralizing compounds as a treatment option for TBI patients is warranted.

  • 24.
    Dorshorst, Ben
    et al.
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Harun-Or-Rashid, Mohammad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bagherpoor, Alireza Jian
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Rubin, Carl-Johan
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Ashwell, Chris
    Gourichon, David
    Tixier-Boichard, Michèle
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Andersson, Leif
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    A Genomic Duplication is Associated with Ectopic Eomesodermin Expression in the Embryonic Chicken Comb and Two Duplex-comb Phenotypes2015In: PLOS Genetics, ISSN 1553-7390, E-ISSN 1553-7404, Vol. 11, no 3, article id e1004947Article in journal (Refereed)
    Abstract [en]

    Duplex-comb (D) is one of three major loci affecting comb morphology in the domestic chicken. Here we show that the two Duplex-comb alleles, V-shaped (D*V) and Buttercup (D*C), are both associated with a 20 Kb tandem duplication containing several conserved putative regulatory elements located 200 Kb upstream of the eomesodermin gene (EOMES). EOMES is a T-box transcription factor that is involved in mesoderm specification during gastrulation. In D*V and D*C chicken embryos we find that EOMES is ectopically expressed in the ectoderm of the comb-developing region as compared to wild-type embryos. The confinement of the ectopic expression of EOMES to the ectoderm is in stark contrast to the causal mechanisms underlying the two other major comb loci in the chicken (Rose-comb and Pea-comb) in which the transcription factors MNR2 and SOX5 are ectopically expressed strictly in the mesenchyme. Interestingly, the causal mutations of all three major comb loci in the chicken are now known to be composed of large-scale structural genomic variants that each result in ectopic expression of transcription factors. The Duplex-comb locus also illustrates the evolution of alleles in domestic animals, which means that alleles evolve by the accumulation of two or more consecutive mutations affecting the phenotype. We do not yet know whether the V-shaped or Buttercup allele correspond to the second mutation that occurred on the haplotype of the original duplication event.

  • 25.
    Dorshorst, Ben
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Molin, Anna-Maja
    Rubin, Carl-Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Johansson, Anna M.
    Stromstedt, Lina
    Pham, Manh-Hung
    Chen, Chih-Feng
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ashwell, Chris
    Andersson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    A Complex Genomic Rearrangement Involving the Endothelin 3 Locus Causes Dermal Hyperpigmentation in the Chicken2011In: PLoS Genetics, ISSN 1553-7390, Vol. 7, no 12, p. e1002412-Article in journal (Refereed)
    Abstract [en]

    Dermal hyperpigmentation or Fibromelanosis (FM) is one of the few examples of skin pigmentation phenotypes in the chicken, where most other pigmentation variants influence feather color and patterning. The Silkie chicken is the most widespread and well-studied breed displaying this phenotype. The presence of the dominant FM allele results in extensive pigmentation of the dermal layer of skin and the majority of internal connective tissue. Here we identify the causal mutation of FM as an inverted duplication and junction of two genomic regions separated by more than 400 kb in wild-type individuals. One of these duplicated regions contains endothelin 3 (EDN3), a gene with a known role in promoting melanoblast proliferation. We show that EDN3 expression is increased in the developing Silkie embryo during the time in which melanoblasts are migrating, and elevated levels of expression are maintained in the adult skin tissue. We have examined four different chicken breeds from both Asia and Europe displaying dermal hyperpigmentation and conclude that the same structural variant underlies this phenotype in all chicken breeds. This complex genomic rearrangement causing a specific monogenic trait in the chicken illustrates how novel mutations with major phenotypic effects have been reused during breed formation in domestic animals.

  • 26.
    Edqvist, Per-Henrik D
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lek, Madelen
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Boije, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lindbäck, Sarah M
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Axon-bearing and axon-less horizontal cell subtypes are generated consecutively during chick retinal development from progenitors that are sensitive to follistatin2008In: BMC Developmental Biology, ISSN 1471-213X, E-ISSN 1471-213X, Vol. 8, p. 46-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND:

    Horizontal cells are retinal interneurons that modulate the output from photoreceptors. A rich literature on the morphological classification and functional properties of HCs in different animals exists, however, the understanding of the events underlying their development is still limited. In most vertebrates including chicken, two main horizontal cell (HC) subtypes are identified based on the presence or absence of an axon.

    RESULTS:

    In this work we have molecularly characterized three HC subtypes based on Lim1, Isl1, GABA and TrkA, a classification that is consistent with three chick HC subtypes previously defined by morphology. The axon-bearing and axon-less HC subpopulations molecularly defined by Lim1 and Isl1, are born consecutively on embryonic day (E) 3-4 and E4-5, respectively, and exhibit temporally distinguishable periods of migration. Their relative numbers are not adjusted by apoptosis. A sharp decrease of high endogenous levels of the activin-inhibitor follistatin at E3 coincides with the appearance of the Lim1 positive cells. Extending the follistatin exposure of the HC retinal progenitor cells by injection of follistatin at E3 increased the number of both Lim1- and Isl1 positive HCs when analysed at E9.

    CONCLUSION:

    The results imply that the axon-bearing and axon-less HC subgroups are defined early and are generated consecutively from a retinal progenitor cell population that is sensitive to the inhibitory action of follistatin. The results are consistent with a model wherein added follistatin causes HC-generating progenitors to proliferate beyond the normal period of HC generation, thus producing extra HCs of both types that migrate to the HC layer.

  • 27.
    Edqvist, Per-Henrik D
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Molecular and Morphological Pathology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Niklasson, Mia
    Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology.
    Vidal-Sanz, Manuel
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Forsberg-Nilsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology. Uppsala University, Science for Life Laboratory, SciLifeLab.
    Platelet-derived growth factor over-expression in retinal progenitors results in abnormal retinal vessel formation2012In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 7, no 8, p. e42488-Article in journal (Refereed)
    Abstract [en]

    Platelet-derived growth factor (PDGF) plays an important role in development of the central nervous system, including the retina. Excessive PDGF signaling is associated with proliferative retinal disorders. We reported previously that transgenic mice in which PDGF-B was over-expressed under control of the nestin enhancer, nes/tk-PdgfB-lacZ, exhibited enhanced apoptosis in the developing corpus striatum. These animals display enlarged lateral ventricles after birth as well as behavioral aberrations as adults. Here, we report that in contrast to the relatively mild central nervous system phenotype, development of the retina is severely disturbed in nes/tk-PdgfB-lacZ mice.

    In transgenic retinas all nuclear layers were disorganized and photoreceptor segments failed to develop properly. Since astrocyte precursor cells did not populate the retina, retinal vascular progenitors could not form a network of vessels. With time, randomly distributed vessels resembling capillaries formed, but there were no large trunk vessels and the intraocular pressure was reduced. In addition, we observed a delayed regression of the hyaloid vasculature. The prolonged presence of this structure may contribute to the other abnormalities observed in the retina, including the defective lamination.

  • 28.
    Edqvist, Per-Henrik
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Myers, Sarah
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Early identification of retinal subtypes in the developing, pre-laminated chick retina using the transcription factors Prox1, Lim1, Ap2α, Pax6, Isl1, Isl2, Lim3 and Chx102006In: European journal of histochemistry, ISSN 1121-760X, E-ISSN 2038-8306, Vol. 50, no 2, p. 147-154Article in journal (Refereed)
    Abstract [en]

    In this study, antibodies toward the transcription factors Prox1, Lim1, Ap2a, Pax6, Isl1, Isl2, Lim3 and Chx10 were used to identify and distinguish between developing cell types in the pre-laminated chick retina. The spatio-temporal expression patterns were analysed from embryonic day 3 (E3) to E9, thus covering a time-span from the onset of retinal cell-fate determination to when retinal laminas can be distinguished. Most transcription factors were found at early stages of development, enabling us to trace various precursor cell populations throughout the lamination process. With time, each transcription factor expression became restricted to distinct laminas or sub-laminas of the maturing retina. These early emerging patterns were compared and found to be consistent with those of the hatched chick retina, where the outer nuclear layer label for Lim3, Isl1 and Isl2. In the inner nuclear layer, horizontal cells labeled for Prox1, Lim1, Isl1, Ap2a and Pax6, bipolar cell labeled for Lim3, Isl1 and Chx10 and amacrine cells labeled for Ap2a, Isl1 and Pax6. The ganglion cell layer labeled for Isl1, Pax6 and Isl2. The immunolabeling patterns of Lim3 and Isl2 have not previously been described in detail.

  • 29.
    Ekegren, Titti
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Grundström, Eva
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Lindholm, Dan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Aquilonius, Sten-Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Upregulation of Bax protein and increased DNA degradation in ALS spinal cord motor neurons1999In: Acta Neurologica Scandinavica, ISSN 0001-6314, E-ISSN 1600-0404, Vol. 100, no 5, p. 317-321Article in journal (Refereed)
    Abstract [en]

    Objectives

    To investigate if degeneration of motor neurons in amyotrophic lateral sclerosis (ALS) is related to altered levels of the apoptosis regulating proteins Bcl-2 and Bax. In addition, immunoreactivity of the cysteine protease ICH-1L and detection of motor neurons with DNA fragmentation, indicative of apoptosis, was also studied.

    Material and methods

    The immunoreactivity of Bcl-2, Bax and ICH-1L were compared in ALS and control spinal cord motor neurons by immunohistochemical analysis and motor neurons with DNA fragmentation were identified by the TUNEL-method.

    Results

    The results demonstrate an increased expression of Bax in the ALS material as compared to controls but no change in Bcl-2 and ICH-1L expressions. Moreover, a larger proportion of motor neurons stained positive for TUNEL in ALS spinal cords.

    Conclusion

    Present study suggest an upregulation of the cell death promoting protein Bax and increased DNA degradation, indicative of apoptosis, in spinal motor neurons of ALS patients.

  • 30.
    Fard, Shahrzad Shirazi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    All-Ericsson, Charlotta
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    The heterogenic final cell cycle of chicken retinal Lim1 horizontal cells is not regulated by the DNA damage response pathway2014In: Cell Cycle, ISSN 1538-4101, E-ISSN 1551-4005, Vol. 13, no 3, p. 408-417Article in journal (Refereed)
    Abstract [en]

    Cells with aberrations in chromosomal ploidy are normally removed by apoptosis. However, aneuploid neurons have been shown to remain functional and active both in the cortex and in the retina. Lim1 horizontal progenitor cells in the chicken retina have a heterogenic final cell cycle, producing some cells that enter S-phase without proceeding into M-phase. The cells become heteroploid but do not undergo developmental cell death. This prompted us to investigate if the final cell cycle of these cells is under the regulation of an active DNA damage response. Our results show that the DNA damage response pathway, including gamma-H2AX and Rad51 foci, is not triggered during any phase of the different final cell cycles of horizontal progenitor cells. However, chemically inducing DNA adducts or double-strand breaks in Lim1 horizontal progenitor cells activated the DNA damage response pathway, showing that the cells are capable of a functional response to DNA damage. Moreover, manipulation of the DNA damage response pathway during the final cell cycle using inhibitors of ATM/ATR, Chk1/2, and p38MAPK, neither induced apoptosis nor mitosis in the Lim1 horizontal progenitor cells. We conclude that the DNA damage response pathway is functional in the Lim1 horizontal progenitor cells, but that it is not directly involved in the regulation of the final cell cycle that gives rise to the heteroploid horizontal cell population.

  • 31.
    Fard, Shahrzad Shirazi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Blixt, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    The p53 co-activator Zac1 neither induces cell cycle arrest nor apoptosis in chicken Lim1 horizontal progenitor cells2015In: Cell Death Discovery, ISSN 2058-7716, Vol. 1, article id 15023Article in journal (Refereed)
    Abstract [en]

    Chicken horizontal progenitor cells are able to enter their final mitosis even in the presence of DNA damage despite having a functional p53-p21 system. This suggests that they are resistant to DNA damage and that the regulation of the final cell cycle of horizontal progenitor cells is independent of the p53-p21 system. The activity of p53 is regulated by positive and negative modulators, including the zinc finger containing transcription factor Zac1 (zinc finger protein that regulates apoptosis and cell cycle arrest). Zac1 interacts with and enhances the activity of p53, thereby inducing cell cycle arrest and apoptosis. In this work, we use a gain-of-function assay in which mouse Zac1 (mZac1) is overexpressed in chicken retinal progenitor cells to study the effect on the final cell cycle of horizontal progenitor cells. The results showed that overexpression of mZac1 induced expression of p21 in a p53-dependent way and arrested the cell cycle as well as triggered apoptosis in chicken non-horizontal retinal progenitor cells. The negative regulation of the cell cycle by mZac1 is consistent with its proposed role as a tumour-suppressor gene. However, the horizontal cells were not affected by mZac1 overexpression. They progressed into S- and late G2/M-phase despite overexpression of mZac1. The inability of mZac1 to arrest the cell cycle in horizontal progenitor cells support the notion that the horizontal cells are less sensitive to events that triggers the p53 system during their terminal and neurogenic cell cycle, compared with other retinal cells. These properties are associated with a cell that has a propensity to become neoplastic and thus with a cell that may develop retinoblastoma.

  • 32.
    Fard, Shahrzad Shirazi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Blixt, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Whole Retinal Explants from Chicken Embryos for Electroporation and Chemical Reagent Treatments2015In: Journal of Visualized Experiments, ISSN 1940-087X, E-ISSN 1940-087X, no 103, article id e53202Article in journal (Refereed)
    Abstract [en]

    The retina is a good model for the developing central nervous system. The large size of the eye and most importantly the accessibility for experimental manipulations in ovo/in vivo makes the chicken embryonic retina a versatile and very efficient experimental model. Although the chicken retina is easy to target in ovo by intraocular injections or electroporation, the effective and exact concentration of the reagents within the retina may be difficult to fully control. This may be due to variations of the exact injection site, leakage from the eye or uneven diffusion of the substances. Furthermore, the frequency of malformations and mortality after invasive manipulations such as electroporation is rather high. This protocol describes an ex ovo technique for culturing whole retinal explants from chicken embryos and provides a method for controlled exposure of the retina to reagents. The protocol describes how to dissect, experimentally manipulate, and culture whole retinal explants from chicken embryos. The explants can be cultured for approximately 24 hr and be subjected to different manipulations such as electroporation. The major advantages are that the experiment is not dependent on the survival of the embryo and that the concentration of the introduced reagent can be varied and controlled in order to determine and optimize the effective concentration. Furthermore, the technique is rapid, cheap and together with its high experimental success rate, it ensures reproducible results. It should be emphasized that it serves as an excellent complement to experiments performed in ovo.

  • 33.
    Fard, Shahrzad Shirazi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Jarrin, Miguel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Boije, Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Fillon, Valerie
    All-Eriksson, Charlotta
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Heterogenic Final Cell Cycle by Chicken Retinal Lim1 Horizontal Progenitor Cells Leads to Heteroploid Cells with a Remaining Replicated Genome2013In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 8, no 3, p. e59133-Article in journal (Refereed)
    Abstract [en]

    Retinal progenitor cells undergo apical mitoses during the process of interkinetic nuclear migration and newly generated post-mitotic neurons migrate to their prospective retinal layer. Whereas this is valid for most types of retinal neurons, chicken horizontal cells are generated by delayed non-apical mitoses from dedicated progenitors. The regulation of such final cell cycle is not well understood and we have studied how Lim1 expressing horizontal progenitor cells (HPCs) exit the cell cycle. We have used markers for S-and G2/M-phase in combination with markers for cell cycle regulators Rb1, cyclin B1, cdc25C and p27Kip1 to characterise the final cell cycle of HPCs. The results show that Lim1+ HPCs are heterogenic with regards to when and during what phase they leave the final cell cycle. Not all horizontal cells were generated by a non-apical (basal) mitosis; instead, the HPCs exhibited three different behaviours during the final cell cycle. Thirty-five percent of the Lim1+ horizontal cells was estimated to be generated by non-apical mitoses. The other horizontal cells were either generated by an interkinetic nuclear migration with an apical mitosis or by a cell cycle with an S-phase that was not followed by any mitosis. Such cells remain with replicated DNA and may be regarded as somatic heteroploids. The observed heterogeneity of the final cell cycle was also seen in the expression of Rb1, cyclin B1, cdc25C and p27Kip1. Phosphorylated Rb1-Ser608 was restricted to the Lim1+ cells that entered S-phase while cyclin B1 and cdc25C were exclusively expressed in HPCs having a basal mitosis. Only HPCs that leave the cell cycle after an apical mitosis expressed p27Kip1. We speculate that the cell cycle heterogeneity with formation of heteroploid cells may present a cellular context that contributes to the suggested propensity of these cells to generate cancer when the retinoblastoma gene is mutated.

  • 34.
    Galindo-Romero, Caridad
    et al.
    Uppsala University.
    Harun-Or-Rashid, Mohammad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience. Uppsala University.
    Jiménez-López, Manuel
    Vidal-Sanz, Manuel
    Agudo-Barriuso, Marta
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Neuroprotection by α2-Adrenergic Receptor Stimulation after Excitotoxic Injury of Chicken Retinal Ganglion Cells: A Population StudyManuscript (preprint) (Other academic)
    Abstract [en]

    Retinal ganglion cells (RGCs) loss is one of the most common causes of blindness in worldwide. In this work we studied the RGC population in normal and excitotoxically injured chicken retina after pretreatment with the α2-adrenergic receptor (α2-ADR) agonist brimonidine. The main objective of this work was to study the effects of brimonidine on injured chicken RGCs. A lesion was inflicted by intraocular injection of N-methyl-D-aspartate (NMDA) at embryonic day 18 and the total population of retinal ganglion cells was studied using an automated cell counting of cells positive for the retinal ganglion cell marker Brn3a in flat-mounted retinas. Surviving Brn3a positive RGCs and their distribution in the retina were analyzed 7 and 14 days post lesion. In addition, the total population of retinal ganglion cells was analyzed in a series of normal embryonic day 8 to post-hatch day 11 retinas. The result showed the distribution of total population of RGCs both in embryonic and post-nantal chicken retina. The pretreatment with brimonidine in excitotoxic retina significantly reduced RGC death as seen both 7 and 14 days post lesion. The excitotoxic lesion was more severe in the dorsal quadrants of the retina than in the ventral ones. The regional difference was also seen in the effect of brimonidine. Thus, we conclude that α2-ADR signaling protects RGCs against the excitotoxic injury in the chicken retina.

  • 35.
    Grundström, Eva
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Askmark, Håkan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Lindeberg, Jonas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Nygren, Ingela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Aquilonius, Sten-Magnus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurology.
    Increased expression of glial cell line-derived neurotrophic factor mRNA in muscle biopsies from patients with amyotrophic lateral sclerosis1999In: Journal of the Neurological Sciences, ISSN 0022-510X, E-ISSN 1878-5883, Vol. 162, no 2, p. 169-173Article in journal (Refereed)
    Abstract [en]

    The expression of glial cell line-derived neurotrophic factor (GDNF) mRNA and brain-derived neurotrophic factor (BDNF) mRNA were studied in muscle biopsies from five patients with amyotrophic lateral sclerosis (ALS), six patients with other neuromuscular diseases and eight healthy control persons. All five patients with ALS had higher GDNF mRNA expressions in their biopsies than the healthy control group (almost a three fold increase). Among the other patients only one, who had a rapidly progressing toxic polyneuropathy, showed a GDNF mRNA expression above those of the controls. The BDNF mRNA expressions in the biopsies from the ALS patients were in the same range as those from the healthy controls, although the mean value of the ALS patients was higher. The only biopsy that showed a markedly higher BDNF mRNA expression was taken from one patient with progressive muscular atrophy. These results suggest that increased GDNF mRNA expression in muscle is an unspecific response to ongoing denervation and that this response is maintained in ALS, at least temporarily. If increased GDNF mRNA in muscle proves to be a constant finding in ALS the rationale for the use of GDNF as a therapeutic agent in ALS must be questioned.

  • 36.
    Haitina, Tatjana
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Olsson, Fredrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Stephansson, Olga
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Alsiö, Johan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Roman, Erika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Schiöth, Helgi B.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Fredriksson, Robert
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Functional Pharmacology.
    Expression profile of the entire family of Adhesion G protein-coupled receptors in mouse and rat2008In: BMC neuroscience (Online), ISSN 1471-2202, E-ISSN 1471-2202, Vol. 9, p. 43-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND

    The Adhesion G protein-coupled receptors (GPCRs) are membrane-bound receptors with long N termini. This family has 33 members in humans. Several Adhesion GPCRs are known to have important physiological functions in CNS development and immune system response mediated by large cell surface ligands. However, the majority of Adhesion GPCRs are still poorly studied orphans with unknown functions.

    RESULTS

    In this study we performed the extensive tissue localization analysis of the entire Adhesion GPCR family in rat and mouse. By applying the quantitative real-time PCR technique we have produced comparable expression profile for each of the members in the Adhesion family. The results are compared with literature data and data from the Allen Brain Atlas project. Our results suggest that the majority of the Adhesion GPCRs are either expressed in the CNS or ubiquitously. In addition the Adhesion GPCRs from the same phylogenetic group have either predominant CNS or peripheral expression, although each of their expression profile is unique.

    CONCLUSION

    Our findings indicate that many of Adhesion GPCRs are expressed, and most probably, have function in CNS. The related Adhesion GPCRs are well conserved in their structure and interestingly have considerable overlap in their expression profiles, suggesting similarities among the physiological roles for members within many of the phylogenetically related clusters.

  • 37.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Evolution of the vertebrate neurotrophin and Trk receptor gene families1999In: Current Opinion in Neurobiology, ISSN 0959-4388, E-ISSN 1873-6882, Vol. 9, no 5, p. 616-621Article in journal (Refereed)
    Abstract [en]

    Studies of neurotrophins and Trk receptors in jawless fish have shed light on the course of events underlying the formation of these gene families. They evolved early in vertebrate history during major gene duplication events, before the appearance of cartilaginous fish. The existence of multiple genes has permitted the diversification of neurotrophin and Trk receptor expression, and thereby enabling the acquisition of specific functions in selective neuronal populations.

  • 38.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Evolution of the vertebrate neurotrophin and Trk receptor gene families1999In: Current Opinion in Neurobiology, ISSN 0959-4388, E-ISSN 1873-6882, Vol. 9, no 5, p. 616-621Article in journal (Refereed)
    Abstract [en]

    Studies of neurotrophins and Trk receptors in jawless fish have shed light on the course of events underlying the formation of these gene families. They evolved early in vertebrate history during major gene duplication events, before the appearance of cartilaginous fish. The existence of multiple genes has permitted the diversification of neurotrophin and Trk receptor expression, and thereby enabling the acquisition of specific functions in selective neuronal populations.

  • 39.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bäckström, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Carri, Nestor G
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Expression of neurotrophins and trk receptors in the avian retina1996In: Journal of Comparative Neurology, ISSN 0021-9967, E-ISSN 1096-9861, Vol. 364, no 4, p. 664-676Article in journal (Refereed)
    Abstract [en]

    Using the RNase protection assay, we have found that nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) are expressed in the avian retina during development. The expression peaks around embryonic days 12-15, with decreasing levels at later stages of development. Abundant levels of NGF and BDNF but low levels of NT-3 mRNA were found in the adult retina. We also found that light/darkness regulated the levels of NGF and BDNF mRNAs but not the levels of NT-3 mRNA in the 5-day-old chicken retina. It was demonstrated that NGF and BDNF mRNA levels were up-regulated by light exposure. The cellular localization of mRNA expression for the neurotrophins and neurotrophin receptors TrkA, TrkB, and TrkC in the retina was studied using in situ hybridization. The patterns of NGF and trkA mRNA expression were very similar and were localized to the external part of the inner nuclear layer on the border with the outer plexiform layer and corresponded to the localization of horizontal cells. NT-3 labeling was also found over the external part of the inner nuclear layer, whereas trkC mRNA was found over all layers in the retina. BDNF labeling was found over all layers in the retina, whereas TrkB labeling was intense over cells in the ganglion cell layer, which is in agreement with the response of ganglion cells to BDNF stimulation. Functional neurotrophin receptors were suggested by the response of retinal explants to neurotrophin stimulation. These data indicate that the neurotrophins play local roles in the retina that involve interactions between specific neuronal populations, which were identified by the localization of the Trk receptor expression. The data also suggest that NGF and BDNF expression is regulated by normal neuron usage in the retina.

  • 40.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Bäckström, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Kylberg, Annika
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Williams, Reg
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Ebendal, Ted
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Neurotrophins and their receptors in chicken neuronal development1995In: International Journal of Developmental Biology, ISSN 0214-6282, E-ISSN 1696-3547, Vol. 39, no 5, p. 855-868Article in journal (Refereed)
    Abstract [en]

    A review on current studies of chicken neurotrophins and their receptors is given. Chicken NGF, BDNF and NT-3 have been cloned and sequences have been used to synthesize oligonucleotides for specific localization of expression during development. Also, chicken TrkA, TrkB and TrkC have been cloned, sequenced and studied by in situ hybridization. Recombinant NT-3 was applied to chicken ganglia at different developmental stages to examine acquirement of responsiveness to NT-3 compared to NGF. Phylogenetic analyses of the chicken neurotrophins and Trk receptors were carried out based on parsimony. Finally, some data on apoptosis in chicken embryo sympathetic ganglia are presented.

  • 41.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Catsicas, M
    Staple, J K
    Grenningloh, G
    Catsicas, S
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Gene functional analysis in nervous system2000In: Methods in Enzymology, ISSN 0076-6879, E-ISSN 1557-7988, Vol. 314, p. 148-167Article in journal (Refereed)
  • 42.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Catsicas, M
    Staple, JK
    Grenningloh, G
    Catsicas, S
    Gene functional analysis in the nervous system2000In: Methods in Enzymology, ISSN 0076-6879, E-ISSN 1557-7988, Vol. 314, p. 148-167Article in journal (Refereed)
  • 43.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Fritzsch, B
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Distribution of BDNF and trkB mRNA in the otic region of 3.5 and 4.5 day chick embryos as revealed with a combination of in situ hybridization and tract tracing1997In: International Journal of Developmental Biology, ISSN 0214-6282, E-ISSN 1696-3547, Vol. 41, no 5, p. 725-732Article in journal (Refereed)
    Abstract [en]

    We have used a recently developed technique which combines fluorescent tract tracing and in situ hybridization to study co-localization of neurotrophin mRNA and neurotrophin receptor mRNA expression simultaneously with the pattern of innervation in the developing chick ear. Efferent and afferent fibersfrom the VII/VIIIth cranial nerves were retrogradely and anterogradely filled with Dextran amines conjugated to Texas red and the brain stem was incubated for 2 hours in tissue culture medium. The tissue was subsequently fixed, sectioned frozen, mounted and subjected to in situ hybridization analysis using probes for brain-derived neurotrophic factor (BDNF) and its tyrosine kinase receptor, trkB. The results show that afferent and efferent fibers to the ear innervate areas of the developing otocyst which express BDNF mRNA. We also found that neurons in the stato-acoustic ganglion express high levels of trkB mRNA whereas the subset of facial motor neurons that is efferent to the ear only had no or very low levels of trkB mRNA. From our results we conclude that chicken otic efferent fibers preferentially project to areas with BDNF mRNA expression. The very low levels of trkB mRNA in these motor neurons compared to afferent neurons innervating the same region suggest that other factors, perhaps co-expressed with BDNF, may support efferents. A possible involvement of afferents in guiding efferents to specific areas of the ear is suggested.

  • 44.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lundin, Lars-Gustav
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Lampetra fluviatilis neurotrophin homolog, descendant of a neurotrophin ancestor, discloses the early molecular evolution of neurotrophins in the vertebrate subphylum1998In: Journal of Neuroscience, ISSN 0270-6474, E-ISSN 1529-2401, Vol. 18, no 21, p. 8700-8711Article in journal (Refereed)
    Abstract [en]

    We have isolated a neurotrophin from the lamprey that permitted us to perform a phylogenetic analysis of the neurotrophin gene family that dates back more than 460 million years to the early vertebrate ancestors. The results show that the neurotrophin gene family was originally formed by two subsequent duplications. The duplication that formed nerve growth factor, neurotrophin-3, brain-derived neurotrophic factor, and neurotrophin-4/5 occurred after the split of lampreys but before the split of cartilaginous fish from the main vertebrate lineage. Compilation of chromosomal gene maps around the neurotrophins shows that they are located in paralogous regions, suggesting that the genes were formed at major duplication events possibly by complete genome doubling. Analysis of two isolated Trk receptor sequences shows similar results as for the lamprey neurotrophin. Multiple neurotrophin and Trk genes, including neurotrophin-6 and -7, have been found in bony fish, and we suggest that the extra genes were formed by an additional duplication in the bony fish lineage. Analysis of lamprey Trk mRNA expression in the adult brain shows that the genes are expressed in all regions analyzed so far. Together, the results suggest that the duplications of ancestral neurotrophin and Trk genes at an early vertebrate stage have permitted evolution to bring about differential neurotrophin and Trk expression, thereby allowing the formation of specific functions in selective neuronal populations.

  • 45.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Sahlén, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Catsicas, S
    Characterization and evaluation of antisense oligonucleotides: inhibition of NGF synthesis in transfected COS cells1997In: Antisense and Nucleic Acid Drug Development, ISSN 1087-2906, E-ISSN 2168-6599, Vol. 7, no 2, p. 89-100Article in journal (Refereed)
    Abstract [en]

    We present a system for the assessment of the inhibiting capacity of antisense oligonucleotides. The aim of this study was to identify an oligonucleotide that can inhibit chicken nerve growth factor (NGF) synthesis. Five antisense chicken NGF phosphorothioate oligonucleotides, AS1–5, were designed and were tested for their capacity to inhibit NGF expression in COS cells. COS cells that transiently expressed chicken NGF were treated with the oligonucleotides, and NGF expression was analyzed using a bioassay and Western blotting for NGF protein. Two oligonucleotides, AS 1 and AS 5, were more capable than the others of inhibiting expression compared with nonsense oligonucleotide, and they targeted the translational initiation and stop sites. The chicken NGF is expressed at a high level from an adenovirus major late promoter, and AS 1 was capable of inhibiting more than 80% of the NGF expression as determined using the bioassay and Western blotting. Expression of another member of the NGF gene family, neurotrophin-4, was not affected by treatment of the antisense oligonucleotides. A 10-fold lower concentration of the AS 1 oligonucleotide could be used to inhibit NGF synthesis if the cellular uptake was facilitated using lipofectin compared with addition of oligonucleotide directly to the culture medium. The amount of oligonucleotide taken up by the cells was similar in the lipofectin-treated cells as in the cells treated by a 10-fold higher concentration of medium-supplemented nucleotide. This system based on COS cells can facilitate evaluation of the capacity of inhibiting antisense oligonucleotides, particularly targeting those genes in which endogenous products are present in low levels and are difficult to analyze.

  • 46.
    Hallböök, Finn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Wilson, Karen
    Thorndyke, Mike
    Olinski, Robert Piotr
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Formation and evolution of the chordate neurotrophin and Trk receptor genes2006In: Brain, behavior, and evolution, ISSN 0006-8977, E-ISSN 1421-9743, Vol. 68, no 3, p. 133-144Article in journal (Refereed)
    Abstract [en]

    Neurotrophins are structurally related neurotrophic polypeptide factors that regulate neuronal differentiation and are essential for neuronal survival, neurite growth and plasticity. It has until very recently been thought that the neurotrophin system appeared with the vertebrate species, but identification of a cephalochordate neurotrophin receptor (Trk), and more recently neurotrophin sequences in several genomes of deuterostome invertebrates, show that the system already existed at the stem of the deuterostome group. Comparative genomics supports the hypothesis that two whole genome duplications produced many of the vertebrate gene families, among those the neurotrophin and Trk families. It remains to be proven to what extent the whole genome duplications have driven macroevolutionary change, but it appears certain that the formation of the multi-gene copy neurotrophin and Trk receptor families at the stem of vertebrates has provided a foundation from which the various functions and pleiotropic effects produced by each of the four extant neurotrophins have evolved.

  • 47.
    Hanell, Anders
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Clausen, Fredrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Djupsjö, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Vallstedt, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Patra, Kalicharan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Israelsson, Charlotte
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Larhammar, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Björk, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Paixao, Sonia
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Genetics.
    Marklund, Niklas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Neurosurgery.
    Functional and Histological Outcome after Focal Traumatic Brain Injury Is Not Improved in Conditional EphA4 Knockout Mice2012In: Journal of Neurotrauma, ISSN 0897-7151, E-ISSN 1557-9042, Vol. 29, no 17, p. 2660-2671Article in journal (Refereed)
    Abstract [en]

    We investigated the role of the axon guidance molecule EphA4 following traumatic brain injury (TBI) in mice. Neutralization of EphA4 improved motor function and axonal regeneration following experimental spinal cord injury (SCI). We hypothesized that genetic absence of EphA4 could improve functional and histological outcome following TBI. Using qRT-PCR in wild-type (WT) mice, we evaluated the EphA4 mRNA levels following controlled cortical impact (CCI) TBI or sham injury and found it to be downregulated in the hippocampus (p < 0.05) but not the cortex ipsilateral to the injury at 24 h post-injury. Next, we evaluated the behavioral and histological outcome following CCI using WT mice and Emx1-Cre-driven conditional knockout (cKO) mice. In cKO mice, EphA4 was completely absent in the hippocampus and markedly reduced in the cortical regions from embryonic day 16, which was confirmed using Western blot analysis. EphA4 cKO mice had similar learning and memory abilities at 3 weeks post-TBI compared to WT controls, although brain-injured animals performed worse than sham-injured controls (p < 0.05). EphA4 cKO mice performed similarly to WT mice in the rotarod and cylinder tests of motor function up to 29 days post-injury. TBI increased cortical and hippocampal astrocytosis (GFAP immunohistochemistry, p < 0.05) and hippocampal sprouting (Timm stain, p < 0.05) and induced a marked loss of hemispheric tissue (p < 0.05). EphA4 cKO did not alter the histological outcome. Although our results may argue against a beneficial role for EphA4 in the recovery process following TBI, further studies including post-injury pharmacological neutralization of EphA4 are needed to define the role for EphA4 following TBI.

  • 48.
    Harun-Or-Rashid, Mohammad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience. Uppsala University.
    Modulation of the Progenitor Cell and Homeostatic Capacities of Müller Glia Cells in Retina: Focus on α2-Adrenergic and Endothelin Receptor Signaling Systems2016Doctoral thesis, comprehensive summary (Other academic)
    Abstract [en]

    Müller cells are major glial cells in the retina and have a broad range of functions that are vital for the retinal neurons. During retinal injury gliotic response either leads to Müller cell dedifferentiation and formation of a retinal progenitor or to maintenance of mature Müller cell functions. The overall aim of this thesis was to investigate the intra- and extracellular signaling of Müller cells, to understand how Müller cells communicate during an injury and how their properties can be regulated after injury. Focus has been on the α2-adrenergic receptor (α2-ADR) and endothelin receptor (EDNR)-induced modulation of Müller cell-properties after injury.

    The results show that α2-ADR stimulation by brimonidine (BMD) triggers Src-kinase mediated ligand-dependent and ligand-independent transactivation of epidermal growth factor receptor (EGFR) in both chicken and human Müller cells. The effects of this transactivation in injured retina attenuate injury-induced activation and dedifferentiation of Müller cells by attenuating injury-induced ERK signaling. The attenuation was concomitant with a synergistic up-regulation of negative ERK- and RTK-feedback regulators during injury. The data suggest that adrenergic stress-signals modulate glial responses during retinal injury and that α2-ADR pharmacology can be used to modulate glial injury-response. We studied the effects of this attenuation of Müller cell dedifferentiation on injured retina from the perspective of neuroprotection. We analyzed retinal ganglion cell (RGC) survival after α2-ADR stimulation of excitotoxically injured chicken retina and our results show that α2-ADR stimulation protects RGCs against the excitotoxic injury. We propose that α2-ADR-induced protection of RGCs in injured retina is due to enhancing the attenuation of the glial injury response and to sustaining mature glial functions. Moreover, we studied endothelin-induced intracellular signaling in Müller cells and our results show that stimulation of EDNRB transactivates EGFR in Müller cells in a similar way as seen after α2-ADR stimulation. These results outline a mechanism of how injury-induced endothelins may modulate the gliotic responses of Müller cells.

    The results obtained in this thesis are pivotal and provide new insights into glial functions, thereby uncovering possibilities to target Müller cells by designing neuroprotective treatments of retinal degenerative diseases or acute retinal injury.

    List of papers
    1. Transactivation of EGF Receptors in Chicken Muller Cells by α2A-Adrenergic Receptors Stimulated by Brimonidine
    Open this publication in new window or tab >>Transactivation of EGF Receptors in Chicken Muller Cells by α2A-Adrenergic Receptors Stimulated by Brimonidine
    2014 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 55, no 6, p. 3385-3394Article in journal (Refereed) Published
    Abstract [en]

    Purpose: α2-Adrenergic receptor agonists are used in glaucoma treatment and have been shown to have some neuroprotective effects. We performed this study to test the hypothesis that epidermal growth factor receptors on chicken Müller cells are transactivated by α2-adrenergic receptors and we focused on the extracellular signal-activated kinases 1/2 (ERK) pathway. Methods: Embryonic chicken retina and cultures of primary Müller cells were stimulated by α2-adrenergic receptor agonist brimonidine. Immunostaining, qRT-PCR and western blot techniques in combination with Src-, epidermal growth factor receptor kinase-, and matrix metalloproteinase inhibitors were used for analysis of the cellular responses. Results: Our results showed that Müller cells express α2A-adrenergic receptors in vivo and in vitro and that brimonidine triggered a robust and transient phosphorylation of ERK1/2. This ERK-response was Src-kinase dependent, associated with tyrosine phosphorylation of epidermal growth factor receptors (phospho-Y1068, Y1173) and was mediated by matrix metalloproteinase-activity on the Müller cells. Conclusions: Müller cells express the α2A-adrenergic receptor and brimonidine triggers both Src-kinase- and matrix metalloproteinase-mediated autocrine ligand-dependent activation of epidermal growth factor receptors on Müller cell. This response is consistent with transactivation of epidermal growth factor receptors by stimulation of α2-adrenergic receptors.

    National Category
    Ophthalmology
    Identifiers
    urn:nbn:se:uu:diva-224134 (URN)10.1167/iovs.13-13823 (DOI)000339485800003 ()24781942 (PubMedID)
    Available from: 2014-05-05 Created: 2014-05-05 Last updated: 2017-12-05Bibliographically approved
    2. Alpha2-Adrenergic Agonist Brimonidine Stimulates ERK1/2 and AKT Signaling Via Transactivation of EGF Receptors in MIO-M1 Human Müller Cells
    Open this publication in new window or tab >>Alpha2-Adrenergic Agonist Brimonidine Stimulates ERK1/2 and AKT Signaling Via Transactivation of EGF Receptors in MIO-M1 Human Müller Cells
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Alpha2-adrenergic receptor (α2-ADR) agonist brimonidine is clinically used in glaucoma and ocular hypertension treatment and has been shown to protect the retina from adverse effects. The purpose of this study to test the hypothesis that brimonidine stimulates extracellular signal-regulated kinases (ERK) 1/2 and AKT signaling via transactivation of epidermal growth factor receptors in MIO-M1 human Müller cells. MIO-M1 cells were treated with brimonidine in combination with Src-kinase, epidermal growth factor receptor kinase, and matrix metalloproteinase inhibitors and analyzed by immunocytochemistry, quantitative PCR and western blot techniques. Our results show that human MIO-M1 cells express α2A-ADR and stimulation of that by brimonidine caused a robust increase of ERK1/2 and AKT (Thr-308) phosphorylation in MIO-M1 cells. The P-ERK1/2 and P-AKT (Thr-308) signaling were mediated by Src-kinase, associated with phosphorylation of tyrosine residue of epidermal growth factor receptor (P-EGFR Y1173) and activation of matrix metalloproteinase. These effects could be blocked by Src-kinase inhibitors (PP1, PP2), EGFR-kinase inhibitor (AG1478) and matrix metalloproteinases inhibitor (GM6001). These results conclude that human MIO-M1 cells express α2A-ADR and brimonidine triggers Srckinase mediated both ligand-dependent and ligand-independent EGFR transactivation. Transactivation leads to activation of ERK1/2 and AKT signaling in MIO-M1 human Müller cells.

    Keywords
    AKT pathway, Alpha 2-adrenergic receptors, Brimonidine, EGF receptor, ERK1/2, Matrix metalloproteinases, MIO-M1 human Müller cell, and Src-kinase
    National Category
    Neurosciences
    Research subject
    Medical Science
    Identifiers
    urn:nbn:se:uu:diva-281576 (URN)
    Available from: 2016-03-24 Created: 2016-03-24 Last updated: 2018-01-10
    3. Alpha2-Adrenergic-Agonist Brimonidine Stimulates Negative Feedback and Attenuates Injury-Induced Phospho-ERK and Dedifferentiation of Chicken Müller Cells
    Open this publication in new window or tab >>Alpha2-Adrenergic-Agonist Brimonidine Stimulates Negative Feedback and Attenuates Injury-Induced Phospho-ERK and Dedifferentiation of Chicken Müller Cells
    2015 (English)In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 56, no 10, p. 5933-5945Article in journal (Refereed) Published
    Abstract [en]

    Purpose:

    Retinal injury induces Müller cell dedifferentiation by activating extracellular signal-regulated kinase (ERK) signaling. Stimulation of α2-adrenergic receptors protects against injury but also activates ERK in Müller cells. The purpose of this work was to study the effect of α2-adrenergic signaling on injury-induced ERK and Müller cell dedifferentiation. We tested the hypothesis that α2-stimulation triggers negative feedback regulation of the injury-induced ERK pathway that attenuates Müller cell dedifferentiation.

    Methods:

    Chicken retina injured by N-methyl-D-aspartate and cultured primary Müller cells were stimulated by the α2-adrenergic agonist brimonidine. Immunostaining, quantitative RT-PCR, and Western blot techniques in combination with receptor blockers were used for analysis of the cellular responses.

    Results:

    Alpha2-adrenergic receptor stimulation attenuated injury-induced ERK activation and dedifferentiation of Müller cells as seen by decreased phospho-ERK, expression of transitin, and retinal progenitor cell genes. The attenuation was concomitant with a synergistic upregulation of several negative ERK-signal feedback regulators including ERK-phosphatases, Raf1-, and growth factor receptor–binding proteins. The results were also seen in cultures of primary Müller cells.

    Conclusions:

    Alpha2-adrenergic signaling on Müller cells elicits an intracellular attenuation of the injury response that comprises negative ERK-signaling feedback leading to attenuated Müller cell dedifferentiation. The implications of this study are that adrenergic stress signals may directly modulate glial function in retina and that α2-adrenergic receptor pharmacology may be used to control glial injury response.

    National Category
    Neurosciences
    Identifiers
    urn:nbn:se:uu:diva-262700 (URN)10.1167/iovs.15-16816 (DOI)000368426300032 ()
    Funder
    Swedish Research Council, M 12187
    Available from: 2015-09-18 Created: 2015-09-18 Last updated: 2018-01-11Bibliographically approved
    4. Neuroprotection by α2-Adrenergic Receptor Stimulation after Excitotoxic Injury of Chicken Retinal Ganglion Cells: A Population Study
    Open this publication in new window or tab >>Neuroprotection by α2-Adrenergic Receptor Stimulation after Excitotoxic Injury of Chicken Retinal Ganglion Cells: A Population Study
    Show others...
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Retinal ganglion cells (RGCs) loss is one of the most common causes of blindness in worldwide. In this work we studied the RGC population in normal and excitotoxically injured chicken retina after pretreatment with the α2-adrenergic receptor (α2-ADR) agonist brimonidine. The main objective of this work was to study the effects of brimonidine on injured chicken RGCs. A lesion was inflicted by intraocular injection of N-methyl-D-aspartate (NMDA) at embryonic day 18 and the total population of retinal ganglion cells was studied using an automated cell counting of cells positive for the retinal ganglion cell marker Brn3a in flat-mounted retinas. Surviving Brn3a positive RGCs and their distribution in the retina were analyzed 7 and 14 days post lesion. In addition, the total population of retinal ganglion cells was analyzed in a series of normal embryonic day 8 to post-hatch day 11 retinas. The result showed the distribution of total population of RGCs both in embryonic and post-nantal chicken retina. The pretreatment with brimonidine in excitotoxic retina significantly reduced RGC death as seen both 7 and 14 days post lesion. The excitotoxic lesion was more severe in the dorsal quadrants of the retina than in the ventral ones. The regional difference was also seen in the effect of brimonidine. Thus, we conclude that α2-ADR signaling protects RGCs against the excitotoxic injury in the chicken retina.

    Keywords
    Alpha2-adrenergic receptor agonist, brimonidine, Brn3a, excitotoxin, flat-mount retina, NMDA, retinal ganglion cells, topographical distribution, development
    National Category
    Neurosciences
    Research subject
    Medical Science
    Identifiers
    urn:nbn:se:uu:diva-281583 (URN)
    Available from: 2016-03-24 Created: 2016-03-24 Last updated: 2018-01-10
    5. The Endothelin B Receptor Transactivates Epidermal Growth Factor Receptors in primary chicken Müller cells and in MIO-M1 Human Müller Cells
    Open this publication in new window or tab >>The Endothelin B Receptor Transactivates Epidermal Growth Factor Receptors in primary chicken Müller cells and in MIO-M1 Human Müller Cells
    (English)Manuscript (preprint) (Other academic)
    Abstract [en]

    Injury to the nervous system elicits signals that trigger a variety of cellular responses. Injury to the retina triggers Müller cells, the major glia cell of the retina, to dedifferentiate, proliferate, attain retinal progenitor properties and in some species generate new neurons. The epidermal growth factor receptor (EGFR) system and extracellular signal-regulated kinase (ERK) signalling are key regulators of these processes in Müller cells. The complexity of the extracellular signals that modulate and control the process are not fully understood. In this work we studied whether endothelin receptor signalling can activate EGFR and ERK signalling in Müller cells. Endothelin expression is robustly up-regulated at retinal injury and endothelin receptors have been shown to transactivate EGFRs in other cell-types. We treated chicken Müller cells in vivo, cultured primary chicken Müller cells and the human Müller cell line MIO-M1 with receptor agonists and enzyme blockers, and analyzed endothelin receptor mediated transactivation of EGFRs by using western blot analysis, quantitative reverse transcriptase PCR and immunocytochemistry. The results showed that both chicken and human Müller cells express endothelin receptor B. Stimulation by using the endothelin receptor B agonist IRL1620 caused Src-kinase mediated ligand-dependent and ligand-independent EGFR transactivation. The effects could be blocked by Src-kinase inhibitors (PP1, PP2), EGFR inhibitor (AG1478) and by inhibitors to extracellular matrix metalloproteinases (GM6001). Our data outline a mechanism how injury-induced endothelins may modulate the Müller cell responses by transactivation of EGFRs. The data give support to a view in which endothelins, among several other functions, serve as an injury-signal that regulate the gliotic response of Müller cells.

    Keywords
    AG1478, endothelin receptor B agonist IRL1620, ERK1/2, gliosis, matrix-metalloproteinases, MAPK, N-methyl-D-aspartate, excitotoxic retinal injury, Src-kinase
    National Category
    Neurosciences
    Research subject
    Medical Science
    Identifiers
    urn:nbn:se:uu:diva-281586 (URN)
    Available from: 2016-03-24 Created: 2016-03-24 Last updated: 2018-01-10
  • 49.
    Harun-Or-Rashid, Mohammad
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Diaz-DelCastillo, Marta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Galindo-Romero, Caridad
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Alpha2-Adrenergic-Agonist Brimonidine Stimulates Negative Feedback and Attenuates Injury-Induced Phospho-ERK and Dedifferentiation of Chicken Müller Cells2015In: Investigative Ophthalmology and Visual Science, ISSN 0146-0404, E-ISSN 1552-5783, Vol. 56, no 10, p. 5933-5945Article in journal (Refereed)
    Abstract [en]

    Purpose:

    Retinal injury induces Müller cell dedifferentiation by activating extracellular signal-regulated kinase (ERK) signaling. Stimulation of α2-adrenergic receptors protects against injury but also activates ERK in Müller cells. The purpose of this work was to study the effect of α2-adrenergic signaling on injury-induced ERK and Müller cell dedifferentiation. We tested the hypothesis that α2-stimulation triggers negative feedback regulation of the injury-induced ERK pathway that attenuates Müller cell dedifferentiation.

    Methods:

    Chicken retina injured by N-methyl-D-aspartate and cultured primary Müller cells were stimulated by the α2-adrenergic agonist brimonidine. Immunostaining, quantitative RT-PCR, and Western blot techniques in combination with receptor blockers were used for analysis of the cellular responses.

    Results:

    Alpha2-adrenergic receptor stimulation attenuated injury-induced ERK activation and dedifferentiation of Müller cells as seen by decreased phospho-ERK, expression of transitin, and retinal progenitor cell genes. The attenuation was concomitant with a synergistic upregulation of several negative ERK-signal feedback regulators including ERK-phosphatases, Raf1-, and growth factor receptor–binding proteins. The results were also seen in cultures of primary Müller cells.

    Conclusions:

    Alpha2-adrenergic signaling on Müller cells elicits an intracellular attenuation of the injury response that comprises negative ERK-signaling feedback leading to attenuated Müller cell dedifferentiation. The implications of this study are that adrenergic stress signals may directly modulate glial function in retina and that α2-adrenergic receptor pharmacology may be used to control glial injury response.

  • 50.
    Harun-Or-Rashid, Mohammad
    et al.
    Uppsala University.
    Hallböök, Finn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Developmental Neuroscience.
    Alpha2-Adrenergic Agonist Brimonidine Stimulates ERK1/2 and AKT Signaling Via Transactivation of EGF Receptors in MIO-M1 Human Müller CellsManuscript (preprint) (Other academic)
    Abstract [en]

    Alpha2-adrenergic receptor (α2-ADR) agonist brimonidine is clinically used in glaucoma and ocular hypertension treatment and has been shown to protect the retina from adverse effects. The purpose of this study to test the hypothesis that brimonidine stimulates extracellular signal-regulated kinases (ERK) 1/2 and AKT signaling via transactivation of epidermal growth factor receptors in MIO-M1 human Müller cells. MIO-M1 cells were treated with brimonidine in combination with Src-kinase, epidermal growth factor receptor kinase, and matrix metalloproteinase inhibitors and analyzed by immunocytochemistry, quantitative PCR and western blot techniques. Our results show that human MIO-M1 cells express α2A-ADR and stimulation of that by brimonidine caused a robust increase of ERK1/2 and AKT (Thr-308) phosphorylation in MIO-M1 cells. The P-ERK1/2 and P-AKT (Thr-308) signaling were mediated by Src-kinase, associated with phosphorylation of tyrosine residue of epidermal growth factor receptor (P-EGFR Y1173) and activation of matrix metalloproteinase. These effects could be blocked by Src-kinase inhibitors (PP1, PP2), EGFR-kinase inhibitor (AG1478) and matrix metalloproteinases inhibitor (GM6001). These results conclude that human MIO-M1 cells express α2A-ADR and brimonidine triggers Srckinase mediated both ligand-dependent and ligand-independent EGFR transactivation. Transactivation leads to activation of ERK1/2 and AKT signaling in MIO-M1 human Müller cells.

123 1 - 50 of 127
CiteExportLink to result list
Permanent link
Cite
Citation style
  • apa
  • ieee
  • modern-language-association
  • vancouver
  • Other style
More styles
Language
  • de-DE
  • en-GB
  • en-US
  • fi-FI
  • nn-NO
  • nn-NB
  • sv-SE
  • Other locale
More languages
Output format
  • html
  • text
  • asciidoc
  • rtf