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  • 1.
    Alenkvist, Ida
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Dyachok, Oleg
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Tian, Geng
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Li, Jia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mehrabanfar, Saba
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jin, Yang
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience.
    Birnir, Bryndis
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Neuroscience, Physiology.
    Tengholm, Anders
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Absence of Shb impairs insulin secretion by elevated FAK activity in pancreatic islets2014In: Journal of Endocrinology, ISSN 0022-0795, E-ISSN 1479-6805, Vol. 223, no 3, p. 267-275Article in journal (Refereed)
    Abstract [en]

    The Src homology-2 domain containing protein B (SHB) has previously been shown to function as a pleiotropic adapter protein, conveying signals from receptor tyrosine kinases to intracellular signaling intermediates. The overexpression of Shb in β-cells promotes β-cell proliferation by increased insulin receptor substrate (IRS) and focal adhesion kinase (FAK) activity, whereas Shb deficiency causes moderate glucose intolerance and impaired first-peak insulin secretion. Using an array of techniques, including live-cell imaging, patch-clamping, immunoblotting, and semi-quantitative PCR, we presently investigated the causes of the abnormal insulin secretory characteristics in Shb-knockout mice. Shb-knockout islets displayed an abnormal signaling signature with increased activities of FAK, IRS, and AKT. β-catenin protein expression was elevated and it showed increased nuclear localization. However, there were no major alterations in the gene expression of various proteins involved in the β-cell secretory machinery. Nor was Shb deficiency associated with changes in glucose-induced ATP generation or cytoplasmic Ca(2) (+) handling. In contrast, the glucose-induced rise in cAMP, known to be important for the insulin secretory response, was delayed in the Shb-knockout compared with WT control. Inhibition of FAK increased the submembrane cAMP concentration, implicating FAK activity in the regulation of insulin exocytosis. In conclusion, Shb deficiency causes a chronic increase in β-cell FAK activity that perturbs the normal insulin secretory characteristics of β-cells, suggesting multi-faceted effects of FAK on insulin secretion depending on the mechanism of FAK activation.

  • 2.
    Andersson, Arne
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Eriksson, Ulf J.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jansson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Sandler, Stellan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Nils
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Claes Hellerström: a friendly islet explorer2007In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 50, no 2, p. 4 p following 496-Article in journal (Refereed)
  • 3.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lindholm, Cecilia K.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kriz, Vitezslav
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The FRK/RAK-SHB signaling cascade: a versatile signal-transduction pathway that regulates cell survival, differentiation and profileration2003In: Current molecular medicine, ISSN 1566-5240, E-ISSN 1875-5666, Vol. 3, no 4, p. 313-324Article in journal (Refereed)
    Abstract [en]

    Recent experiments have unravelled novel signal transduction pathways that involve the SRC homology 2 (SH2) domain adapter protein SHB. SHB is ubiquitously expressed and contains proline rich motifs, a phosphotyrosine binding (PTB) domain, tyrosine phosphorylation sites and an SH2 domain and serves a role in generating signaling complexes in response to tyrosine kinase activation. SHB mediates certain responses in platelet-derived growth factor (PDGF) receptor-, fibroblast growth factor (FGF) receptor-, neural growth factor (NGF) receptor TRKA-, T cell receptor-, interleukin-2 (IL-2) receptor- and focal adhesion kinase- (FAK) signaling. Upstream of SHB in some cells lies the SRC-like FYN-Related Kinase FRK / RAK (also named BSK / IYK or GTK). FRK / RAK and SHB exert similar effects when overexpressed in rat phaeochromocytoma (PC12) and β-cells, where they both induce PC12 cell differentiation and β-cell proliferation. Furthermore, β-cell apoptosis is augmented by these proteins under conditions that cause β-cell degeneration. The FRK / RAK-SHB responses involve FAK and insulin receptor substrates (IRS) -1 and -2.

    Besides regulating apoptosis, proliferation and differentiation, SHB is also a component of the T cell receptor (TCR) signaling response. In Jurkat T cells, SHB links several signaling components with the TCR and is thus required for IL-2 production. In endothelial cells, SHB both promotes apoptosis under conditions that are anti-angiogenic, but is also required for proper mitogenicity, spreading and tubular morphogenesis. In embryonic stem cells, dominant-negative SHB (R522K) prevents early cavitation of embryoid bodies and reduces differentiation to cells expressing albumin, amylase, insulin and glucagon, suggesting a role of SHB in development.

    In summary, SHB is a versatile signal transduction molecule that produces diverse biological responses in different cell types under various conditions. SHB operates downstream of GTK in cells that express this kinase.

  • 4.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Reedquist, K A
    Bos, J L
    Welsh, M
    GTK, a Src-related tyrosine kinase, induces nerve growth factor-independent neurite outgrowth in PC12 cells through activation of the Rap1 pathway: Relationship to Shb tyrosine phosphorylation and elevated levels of focal adhesion kinase2000In: J Biol Chem, Vol. 275, p. 29153-29161Article in journal (Refereed)
  • 5.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, M
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Increased cytokine-induced cytotoxicity of pancreatic islet cells from transgenic mice expressing the Src-like tyrosine kinase GTK2001In: Mol Med., Vol. 7, p. 301-310Article in journal (Refereed)
  • 6.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    GTK Tyrosine Kinase-induced Alteration of IRS-protein Signalling in Insulin Producing Cells2002In: Molecular medicine (Cambridge, Mass. Print), ISSN 1076-1551, E-ISSN 1528-3658, Vol. 8, no 11, p. 705-713Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Insulin receptor substrate proteins (IRS) mediate various effects of insulin, including regulation of glucose homeostasis, cell growth and survival. To understand the underlying mechanisms explaining the effects of the Src-related tyrosine kinase GTK on beta-cell proliferation and survival, insulin-signalling pathways involving IRS-1 and IRS-2 were studied in islet cells and RINm5F cells overexpressing wild-type and two different mutants of the SRC-related tyrosine kinase GTK. MATERIALS AND METHODS: Islets isolated from transgenic mice and RINm5F cells overexpressing wild-type and mutant GTK were analysed for IRS-1, IRS-2, SHB, AKT and ERK phosphorylation/activity by Western blot analysis. RESULTS: RINm5F cells expressing the kinase active mutant Y504F-GTK and islet cells from GTK(Y504F) -transgenic mice exhibited reduced insulin-induced tyrosine phosphorylation of IRS-1 and IRS-2. In RINm5F cells, the diminished IRS-phosphorylation was accompanied by a reduced insulin-stimulated activation of phosphatidylinositol 3-kinase (PI3K), AKT and Extracellular Signal-Regulated Kinase, partly due to an increased basal activity. In addition, increased tyrosine phosphorylation of the SHB SH2 domain-adaptor protein and its association with IRS-2, IRS-1 and focal adhesion kinase was observed in these cells. RINm5F cells overexpressing wild-type GTK also exhibited reduced activation of IRS-2, PI3K and AKT, whereas cells expressing a GTK mutant with lower kinase activity (GTK(Y394F)) exhibited insignificantly altered responses to insulin compared to the mock transfected cells. Moreover, GTK was shown to associate with and phosphorylate SHB in transiently transfected COS-7 cells, indicating that SHB is a specific substrate for GTK. CONCLUSIONS: The results suggest that GTK signals via SHB to modulate insulin-stimulated pathways in beta cells and this may explain previous results showing an increased beta-cell mass in GTK-transgenic mice.

  • 7.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Role of the Bsk/lyk non-receptor tyrosine kinase for the control of growth and hormone production in RINm5F cells2000In: Growth Factors, ISSN 0897-7194, E-ISSN 1029-2292, Vol. 17, p. 233-247Article in journal (Refereed)
    Abstract [en]

    Bsk/Iyk, a murine non-receptor-tyrosine kinase which is expressed in fetal and adult islet of Langerhans was previously found to decrease NIH3T3 cell proliferation when expressed as a Y497/504F-mutant. We presently wanted to determine the effects of Bsk/Iyk on the proliferation of insulin producing cells. Cells expressing Bsk/IykY497/504F and Bsk/IykY504F display a decreased proliferation rate and express higher levels of the cell cycle inhibitor p27/Kip1 compared to control cells. These mutants also conferred diminished cell viability in response to INF-gamma and IL-1beta and contain higher levels of glucagon mRNA. Wild-type Bsk/Iyk is mainly localized at the plasma membrane whereas mutant Bsk/Iyk can enter the nucleus. In vitro kinase reactions using an exogenous substrate indicate a complicated mode of regulation of kinase activity by Y497 and Y504 with the latter being homologous to Y527 in pp60c-Src. These findings suggest that Bsk/Iyk might play a role in inhibiting cell proliferation, transducing cytokine-induced cytotoxicity and regulating hormone production of endocrine pancreatic cells.

  • 8.
    Annerén, Cecilia
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jansson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Glucose intolerance and reduced islet blood flow in transgenic mice expressing the FRK tyrosine kinase under the control of the rat insulin promoter2007In: American Journal of Physiology. Endocrinology and Metabolism, ISSN 0193-1849, E-ISSN 1522-1555, Vol. 292, no 4, p. E1183-E1190Article in journal (Refereed)
    Abstract [en]

    The FRK tyrosine kinase has previously been shown to transduce β-cell cytotoxic signals in response to cytokines and streptozotocin and to promote β-cell proliferation and an increased β-cell mass. We therefore aimed to further evaluate the effects of overexpression of FRK tyrosine kinase in β-cells. A transgenic mouse expressing kinase-active FRK under control of the insulin promoter (RIP-FRK) was studied with regard to islet endocrine function and vascular morphology. Mild glucose intolerance develops in RIP-FRK male mice of at least 4 mo of age. This effect is accompanied by reduced glucose-stimulated insulin secretion in vivo and reduced second-phase insulin secretion in response to glucose and arginine upon pancreas perfusion. Islets isolated from the FRK transgenic mice display a glucose-induced insulin secretory response in vitro similar to that of control islets. However, islet blood flow per islet volume is decreased in the FRK transgenic mice. These mice also exhibit a reduced islet capillary lumen diameter as shown by electron microscopy. Total body weight and pancreas weight are not significantly affected, but the β-cell mass is increased. The data suggest that long-term expression of active FRK in β-cells causes an in vivo insulin-secretory defect, which may be the consequence of islet vascular abnormalities that yield a decreased islet blood flow.

  • 9.
    Barbu, Andreea R.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Bodin, Birgitta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jansson, Leif
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Nils
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    A perfusion protocol for highly efficient transduction of intact pancreatic islets of Langerhans2006In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 49, no 10, p. 2388-2391Article in journal (Refereed)
    Abstract [en]

    Successful gene transfer to pancreatic islets might be a powerful tool for dissecting the biological pathways involved in the functional impairment and destruction of beta cells in type 1 diabetes. In the long run, such an approach may also prove useful for promoting islet graft survival after transplantation in diabetic patients. However, efficient genetic modification of primary insulin-producing cells is limited by the specific compact structure of the pancreatic islet. We present here a whole-pancreas perfusion-based transduction procedure for genetic modification of intact pancreatic islets.

    We used flow cytometry analysis and confocal microscopy to evaluate the efficiency of in vitro and perfusion-based transduction protocols that use adenoviral and lentiviral vectors expressing green fluorescent protein. Islet cell viability was assessed by fluorescence microscopy and beta cell function was determined via glucose-stimulated insulin secretion.

    In intact rat and human pancreatic islets, adenoviral and lentiviral vectors mediated gene transfer to about 30% of cells, but they did not reach the inner cellular mass within the islet core. Using the whole-pancreas perfusion protocol, we demonstrate that at least in rodent models the centrally located insulin-producing cells can be transduced with high efficiency, while preserving the structural integrity of the islet. Moreover, islet cell viability and function are not impaired by this procedure.

    These results support the view that perfusion-based transduction protocols may significantly improve the yield of successfully engineered primary insulin-producing cells for diabetes research.

  • 10.
    Calounova, Gabriela
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Livera, Gabriel
    INSERM/CEA/paris Diderot-Paris 7.
    Zhang, X-Q
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Liu, Kui
    Umeå Universitet.
    Gosden, Roger G
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Src homology 2 domain-containing adapter protein B (SHB) regulates mouse oocyte maturation2010In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 5, no 6, p. e11155-Article in journal (Refereed)
    Abstract [en]

    SHB (Src homology 2 domain-containing adapter protein B) is involved in receptor tyrosine kinase signaling. Mice deficient in the Shb gene have been found to exhibit a transmission ratio distortion with respect to inheritance of the Shb null allele among offspring and this phenomenon was linked to female gamete production. Consequently, we postulated that Shb plays a role for oocyte biology and thus decided to investigate oocyte formation, meiotic maturation, and early embryo development in relation to absence of the Shb gene. Oogenesis was apparently accelerated judging from the stages of oocyte development on fetal day 18.5 and one week postnatally in Shb -/- mice; but in adulthood ovarian follicle maturation was impaired in these mice. Completion of meiosis I (first polar body extrusion) was less synchronized, with a fraction of oocytes showing premature polar body extrusion in the absence of Shb. In vitro fertilization of mature oocytes isolated from Shb +/+, +/- and -/- mice revealed impaired early embryo development in the -/- embryos. Moreover, the absence of Shb enhanced ERK (extracellular-signal regulated kinase) and RSK (ribosomal S6 kinase) signaling in oocytes and these effects were paralleled by an increased ribosomal protein S6 phosphorylation and activation. It is concluded that SHB regulates normal oocyte and follicle development and that perturbation of SHB signaling causes defective meiosis I and early embryo development.

  • 11.
    Christoffersson, Gustav
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology, Integrative Physiology.
    Zang, Guangxiang
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Zhuang, Zhen W.
    Vågesjö, Evelina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology, Integrative Physiology.
    Simons, Michael
    Phillipson, Mia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology, Integrative Physiology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Vascular adaptation to a dysfunctional endothelium as a consequence of Shb deficiency2012In: Angiogenesis, ISSN 0969-6970, E-ISSN 1573-7209, Vol. 15, no 3, p. 469-480Article in journal (Refereed)
    Abstract [en]

    Vascular endothelial growth factor (VEGF)-A regulates angiogenesis, vascular morphology and permeability by signaling through its receptor VEGFR-2. The Shb adapter protein has previously been found to relay certain VEGFR-2 dependent signals and consequently vascular physiology and structure was assessed in Shb knockout mice. X-ray computed tomography of vessels larger than 24 mm diameter (micro-CT) after contrast injection revealed an increased frequency of 48-96 µm arterioles in the hindlimb calf muscle in Shb knockout mice. Intravital microscopy of the cremaster muscle demonstrated a less regular vasculature with fewer branch points and increased vessel tortuosity, changes that led to an increased blood flow velocity. Reduced in vivo angiogenesis was observed in Shb knockout MatrigelTM plugs. Unlike the wild-type situation, VEGF-A did not provoke a dissociation of VE-cadherin from adherens junctions in Shb knockout venules. The reduced angiogenesis and altered properties of junctions had consequences for two patho-physiological responses to arterial occlusion: vascular permeability was reduced in the Shb knockout cremaster muscle after ligation of one supplying artery and heat-induced blood flow determined by Laser-Doppler measurements was decreased in the hindlimb after ligation of the femoral artery. Consequently, the Shb knockout mouse exhibited structural and functional (angiogenesis and vascular permeability) vascular abnormalities that have implications for understanding the function of VEGF-A under physiological conditions.

  • 12.
    Claesson-Welsh, Lena
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    VEGFA and tumour angiogenesis2013In: Journal of Internal Medicine, ISSN 0954-6820, E-ISSN 1365-2796, Vol. 273, no 2, p. 114-127Article, review/survey (Refereed)
    Abstract [en]

    In this review we summarize the current understanding of signal transduction downstream of vascular endothelial growth factor A (VEGFA) and its receptor VEGFR2, and the relationship between these signal transduction pathways and the hallmark responses of VEGFA, angiogenesis and vascular permeability. These physiological responses involve a number of effectors, including extracellular signal-regulated kinases (ERKs), Src, phosphoinositide 3 kinase (PI3K)/Akt, focal adhesion kinase (FAK), Rho family GTPases, endothelial NO and p38 mitogen-activated protein kinase (MAPK). Several of these factors are involved in the regulation of both angiogenesis and vascular permeability. Tumour angiogenesis primarily relies on VEGFA-driven responses, which to a large extent result in a dysfunctional vasculature. The reason for this remains unclear, although it appears that certain aspects of the VEGFA-stimulated angiogenic milieu (high level of microvascular density and permeability) promote tumour expansion. The high degree of redundancy and complexity of VEGFA-driven tumour angiogenesis may explain why tumours commonly develop resistance to anti-angiogenic therapy targeting VEGFA signal transduction.

  • 13.
    Claesson-Welsh, Lena
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Ito, Nobuyuki
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Anand-Apte, Bela
    Soker, Shay
    Setter, Bruce
    O´Reilly, Michael
    Folkman, Judah
    Angiostatin induces endothelial cell apoptosis and activation of focal adhesion kinase independently of the integrin-binding motif RGD1998In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 95, no 10, p. 5579-5583Article in journal (Refereed)
    Abstract [en]

    Angiostatin, a fragment of plasminogen, has been identified and characterized as an endogenous inhibitor of neovascularization. We show that angiostatin treatment of endothelial cells in the absence of growth factors results in an increased apoptotic index whereas the proliferation index is unchanged. Angiostatin also inhibits migration and tube formation of endothelial cells. Angiostatin treatment has no effect on growth factor-induced signal transduction but leads to an RGD-independent induction of the kinase activity of focal adhesion kinase, suggesting that the biological effects of angiostatin relate to subversion of adhesion plaque formation in endothelial cells.

  • 14.
    Cross, Michael J
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Lu, Lingge
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Magnusson, Peetra
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Nyqvist, Daniel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Holmqvist, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Claesson-Welsh, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    The Shb Adaptor Protein Binds to Tyrosine 766 in the FGFR-1 and Regulatesthe Ras/MEK/MAPK Pathway via FRS2 Phosphorylation in Endothelial Cells2002In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 13, no 8, p. 2881-2893Article in journal (Refereed)
    Abstract [en]

    Stimulation of fibroblast growth factor receptor-1 (FGFR-1) is known to result in phosphorylation of tyrosine 766 and the recruitment and subsequent activation of phospholipase C-γ (PLC-γ). To assess the role of tyrosine 766 in endothelial cell function, we generated endothelial cells expressing a chimeric receptor, composed of the extracellular domain of the PDGF receptor-α and the intracellular domain of FGFR-1. Mutation of tyrosine 766 to phenylalanine prevented PLC-γ activation and resulted in a reduced phosphorylation of FRS2 and reduced activation of the Ras/MEK/MAPK pathway relative to the wild-type chimeric receptor. However, FGFR-1–mediated MAPK activation was not dependent on PKC activation or intracellular calcium, both downstream mediators of PLC-γ activation. We report that the adaptor protein Shb is also able to bind tyrosine 766 in the FGFR-1, via its SH2 domain, resulting in its subsequent phosphorylation. Overexpression of an SH2 domain mutant Shb caused a dramatic reduction in FGFR-1–mediated FRS2 phosphorylation with concomitant perturbment of the Ras/MEK/MAPK pathway. Expression of the chimeric receptor mutant and the Shb SH2 domain mutant resulted in a similar reduction in FGFR-1–mediated mitogenicity. We conclude, that Shb binds to tyrosine 766 in the FGFR-1 and regulates FGF-mediated mitogenicity via FRS2 phosphorylation and the subsequent activation of the Ras/MEK/MAPK pathway.

  • 15.
    Davoodpour, Padideh
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Landström, Maréne
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , Ludwig Institute for Cancer Research.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Reduced tumor growth in vivo and increased c-Abl activity in PC3 prostate cancer cells overexpressing the Shb adapter protein2007In: BMC Cancer, ISSN 1471-2407, E-ISSN 1471-2407, Vol. 7, p. 161-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Induction of apoptosis is one strategy for treatment of prostate cancer. The Shb adapter protein has been found to regulate apoptosis in various cell types and consequently human prostate cancer 3 (PC3) cells were transfected to obtain cells overexpressing Shb in order to increase our understanding of the mechanisms regulating PC3 cell apoptosis. METHODS: Human prostate cancer cells (PC3) were transfected with control vector or a vector containing the Shb cDNA. Clones overexpressing Shb were studied with respect to apoptosis (Dapi, M30) and c-Abl activation (Western blot for pY-245-Abl). The cells were exposed to the anti-tumor agent 2-methoxyestradiol (2-ME) and the p38 MAPK and c-Abl inhibitors SB203580 and STI-571, respectively, after which cell death was determined. In vivo tumor growth and tumor cell proliferation (Ki-67 staining) or apoptosis (active caspase 3 staining) were also determined in nude mice. RESULTS: PC3 cells overexpressing Shb exhibited increased rates of apoptosis in the presence of the anti-tumor agent 2-ME. The Shb cells displayed increased activity of the pro-apoptotic kinase c-Abl. Pre-treatment with p38 MAPK (SB203580) or c-Abl (STI-571) inhibitors completely blocked 2-ME-induced apoptosis, implicating these two pathways in the response. The PC3-Shb cells displayed reduced tumor growth in vivo, an effect occurring as a consequence of increased apoptosis and reduced DNA synthesis. CONCLUSION: It is concluded that Shb promotes 2-ME-induced PC3 cell apoptosis by increased pro-apoptotic signaling via the c-Abl pathway and that this causes reduced tumor growth in vivo.

  • 16.
    Dixelius, Johan
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Cancer and Vascular Biology.
    Larsson, Helena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Cancer and Vascular Biology.
    Sasaki, Takako
    Holmqvist, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lu, Lingge
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Engström, Åke
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Timpl, Rupert
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Claesson-Welsh, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology, Cancer and Vascular Biology.
    Endostatin-induced tyrosine kinase signaling through the Shb adaptor protein regulates endothelial cell apoptosis2000In: Blood, ISSN 0006-4971, E-ISSN 1528-0020, Vol. 95, no 11, p. 3403-3411Article in journal (Refereed)
    Abstract [en]

    Endostatin, which corresponds to the C-terminal fragment of collagen XVIII, is a potent inhibitor of angiogenesis. Fibroblast growth factor-2 (FGF-2)-induced angiogenesis in the chicken chorioallantoic membrane was inhibited by endostatin, but not by an endostatin mutant R158/270A, lacking heparin-binding ability. Endostatin was internalized by endothelial cells, but not by mouse fibroblasts. Treatment of murine brain endothelial (IBE) cells with endostatin reduced the proportion of cells in S phase, whereas growth-arrested IBE cells in collagen gels treated with endostatin displayed enhanced tubular morphogenesis. IBE cells overexpressing Shb, an adaptor protein implicated in angiostatin-induced apoptosis, displayed elevated apoptosis and decreased tubular morphogenesis in collagen gels in response to endostatin when added together with FGF-2. Induction of apoptosis was dependent on the heparin-binding ability of endostatin and the expression of Shb with a functional Src homology 2 (SH2)-domain. Endostatin treatment for 10 minutes or 24 hours induced tyrosine phosphorylation of Shb and formation of multiprotein complexes. An Shb SH2 domain fusion protein precipitated a 125-kd phosphotyrosyl protein in endostatin-treated cells. The 125-kd component either contained intrinsic tyrosine kinase activity or occurred in complex with a tyrosine kinase. In conclusion, our data show that endostatin induces tyrosine kinase activity and enhanced apoptosis in FGF-treated endothelial cells.

  • 17.
    Funa, Nina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Reddy, Kalpana
    Bhandarkar, Sulochana
    Kurenova, Elena
    Yang, Lily
    Cance, William
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Arbiser, Jack
    Shb gene knockdown increases the susceptibility of SVR endothelial tumor cells to apoptotic stimuli in vitro and in vivo2008In: Journal of Investigative Dermatology, ISSN 0022-202X, E-ISSN 1523-1747, Vol. 128, no 3, p. 710-716Article in journal (Refereed)
    Abstract [en]

    The Shb adapter protein is an Src homology 2-domain containing signaling intermediate operating downstream of several tyrosine kinase receptors, including vascular endothelial growth factor receptor-2. Shb is multifunctional and apoptosis is one response that Shb regulates. Inhibition of angiogenesis can be used in cancer therapy, and one way to achieve this is by inducing endothelial cell apoptosis. The angiosarcoma cell line SVR is of endothelial origin and can be used as a tool for studying in vivo inhibition of angiogenesis, and we thus employed an Shb-knockdown strategy using an inducible lentiviral system to reduce Shb levels in SVR cells and to study their responses. Shb knockdown increases the susceptibility of SVR cells to the apoptotic agents, cisplatin and staurosporine. Simultaneously, Shb knockdown causes reduced focal adhesion kinase (FAK) activation, monitored as phosphorylation of the regulatory residues tyrosines 576/577. No detectable effects on Akt or extracellular signal-regulated kinase activity were noted. The altered FAK activity coincided with an elongated cell phenotype that was particularly noticeable in the presence of staurosporine. In order to relate the effects of Shb knockdown to in vivo tumorigenicity, cells were exposed to the angiogenesis inhibitor honokiol, and again the cells with reduced Shb content exhibited increased apoptosis. Tumor growth in vivo was strongly reduced in the Shb-knockdown cells upon honokiol treatment. It is concluded that Shb regulates apoptosis and cell shape in tumor endothelial cells via FAK, and that Shb is a potential target for inhibition of angiogenesis.

  • 18.
    Funa, Nina S
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kriz, Vitezslav
    Zang, Guangxiang
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Calounova, Gabriela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Åkerblom, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mares, Jaroslav
    Larsson, Erik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Sun, Ying
    Betsholtz, Christer
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Dysfunctional microvasculature as a consequence of shb gene inactivation causes impaired tumor growth2009In: Cancer Research, ISSN 0008-5472, E-ISSN 1538-7445, Vol. 69, no 5, p. 2141-2148Article in journal (Refereed)
    Abstract [en]

    Shb (Src homology 2 protein B) is an adapter protein downstream of the vascular endothelial growth factor receptor receptor-2 (VEGFR-2). Previous experiments have suggested a role for Shb in endothelial cell function. Recently, the Shb gene was inactivated and Shb null mice were obtained on a mixed genetic background, but not on C57Bl6 mice. The present study was performed to address endothelial function in the Shb knockout mouse and its relevance for tumor angiogenesis. Tumor growth was retarded in Shb mutant mice, and this correlated with decreased angiogenesis both in tumors and in Matrigel plugs. Shb null mice display an abnormal endothelial ultrastructure in liver sinusoids and heart capillaries with cytoplasmic extensions projecting toward the lumen. Shb null heart VE-cadherin staining was less distinct than that of control heart, exhibiting in the former case a wavy and punctuate pattern. Experiments on isolated endothelial cells suggest that these changes could partly reflect cytoskeletal abnormalities. Vascular permeability was increased in Shb null mice in heart, kidney, and skin, whereas VEGF-stimulated vascular permeability was reduced in Shb null mice. It is concluded that Shb plays an important role in maintaining a functional vasculature in adult mice, and that interference with Shb signaling may provide novel means to regulate tumor angiogenesis.

  • 19.
    Funa, Nina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Saldeen, Johan
    Åkerblom, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Interdependent fibroblast growth factor and activin A signaling promotes the expression of endodermal genes in differentiating mouse embryonic stem cells expressing Src Homology 2-domain inactive Shb2008In: Differentiation, ISSN 0301-4681, E-ISSN 1432-0436, Vol. 76, no 5, p. 443-453Article in journal (Refereed)
    Abstract [en]

    The mechanisms controlling endodermal development during stem cell differentiation have been only partly elucidated, although previous studies have suggested the participation of fibroblast growth factor (FGF) and activin A in these processes. Shb is a Src homology 2 (SH2) domain-containing adapter protein that has been implicated in FGF receptor 1 (FGFR1) signaling. To study the putative crosstalk between activin A and Shb-dependent FGF signaling in the differentiation of endoderm from embryonic stem (ES) cells, embryoid bodies (EBs) derived from mouse ES cells overexpressing wild-type Shb or Shb with a mutated SH2 domain (R522K-Shb) were cultured in the presence of activin A. We show that expression of R522K-Shb results in up-regulation of FGFR1 and FGF2 in EBs. Addition of activin A to the cultures enhances the expression of endodermal genes primarily in EBs expressing mutant Shb. Inhibition of FGF signaling by the addition of the FGFR1 inhibitor SU5402 completely counteracts the synergistic effects of R522K-Shb and activin A. In conclusion, the present results suggest that expression of R522K-Shb enhances certain signaling pathways downstream of FGF and that an interplay between FGF and activin A participates in ES cell differentiation to endoderm.

  • 20.
    Gao, Xiang
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Sallstrom, Johan
    Ma, Zufu
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Persson, Erik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    SHB deficient mice display an increased GFR and augmented renal arteriolar contractions to both Adenosine and Ang II2013In: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 27, p. 909.14-Article in journal (Other academic)
  • 21.
    Gao, Xiang
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology, Integrative Physiology.
    Sallstrom, Johan
    Ma, Zufu
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Persson, Erik A.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Role of Src homology 2 domain-containing protein B in the regulation of GFR and renal afferent arteriole responsiveness2014In: The FASEB Journal, ISSN 0892-6638, E-ISSN 1530-6860, Vol. 28, no 1, article id 1088.12Article in journal (Other academic)
  • 22.
    Gustafsson, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Calounova, Gabriela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Hjelm, Fredrik
    Kriz, Vitezslav
    Heyman, Birgitta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Grönvik, Kjell-Olov
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Mostoslavsky, Gustavo
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Shb deficient mice display an augmented TH2 response in peripheral CD4+ T cells2011In: BMC Immunology, ISSN 1471-2172, E-ISSN 1471-2172, Vol. 12, p. 3-Article in journal (Refereed)
    Abstract [en]

    Background: Shb, a ubiquitously expressed Src homology 2 domain-containing adaptor protein has previously been implicated in the signaling of various tyrosine kinase receptors including the TCR. Shb associates with SLP76, LAT and Vav, all important components in the signaling cascade governing T cell function and develeopment. A Shb knockout mouse was recently generated and the aim of the current study was to address the importance of Shb deficiency on T cell development and function.

    Results: Shb knockout mice did not display any major changes in thymocyte development despite an aberrant TCR signaling pattern, including increased basal activation and reduced stimulation-induced phosphorylation. The loss of Shb expression did however affect peripheral CD4+ TH cells resulting in an increased proliferative response to TCR stimulation and an elevated IL-4 production level of naïve TH cells. This suggests a TH2 skewing of the Shb knockout immune system, seemingly caused by an altered TCR signaling pattern.

    Conclusion: Our results indicate that Shb appears to play an important modulating role on TCR signaling, thus regulating the peripheral CD4+ TH2 cell response.

  • 23.
    Gustafsson, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Heffner, Garrett
    Wenzel, Pamela L.
    Curran, Matthew
    Grawé, Jan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    McKinney-Freeman, Shannon L.
    Daley, Georg Q.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Src Homology 2 Protein Shb Promotes Cell Cycle Progression In Murine Hematopoietic Stem Cells By Regulation Of Focal Adhesion Kinase Activity2013In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 319, no 12, p. 1852-1864Article in journal (Refereed)
    Abstract [en]

    The widely expressed adaptor protein Shb has previously been reported to contribute to T cell function due to its association with the T cell receptor and furthermore, several of Shb's known interaction partners are established regulators of blood cell development and function. In addition, Shb deficient embryonic stem cells displayed reduced blood cell colony formation upon differentiation in vitro. The aim of the current study was therefore to explore hematopoietic stem and progenitor cell function in the Shb knockout mouse. Shbdeficient bone marrow contained reduced relative numbers of long-term hematopoietic stem cells (LT-HSCs) that exhibited lower proliferation rates. Despite this, Shb knockout LT-HSCs responded promptly by entering the cell cycle in response to genotoxic stress by 5-fluorouracil treatment. In competitive LT-HSC transplantations, Shb null cells initially engrafted as well as the wild-type cells but provided less myeloid expansion over time. Moreover, Shb knockout bone marrow cells exhibited elevated basal activities of focal adhesion kinase/Rac1/p21-activated kinase signaling and reduced responsiveness to Stem Cell Factor stimulation. Consequently, treatment with a focal adhesion kinase inhibitor increased Shb knockout LT-HSC proliferation. The altered signaling characteristics thus provide a plausible mechanistic explanation for the changes in LT-HSC proliferation since these signaling intermediates have all been shown to participate in LT-HSC cell cycle control. In summary, the loss of Shb dependent signaling in bone marrow cells, resulting in elevated focal adhesion kinase activity and reduced proliferative responses in LT-HSCs under steady state hematopoiesis, confers a disadvantage to the maintenance of LT-HSCs over time.

  • 24.
    Gustafsson, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jamalpour, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Trinh, Camilla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kharas, Michael G
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Src homology-2 protein Shb modulates focal adhesion kinase signaling in a BCR-ABL myeloproliferative disorder causing accelerated progression of disease2014In: Journal of hematology & oncology, ISSN 1756-8722, Vol. 7, no 1, p. 45-Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: The Src homology-2 domain protein B (Shb) is an adapter protein operating downstream of several tyrosine kinase receptors and consequently Shb regulates various cellular responses. Absence of Shb was recently shown to reduce hematopoietic stem cell proliferation through activation of focal adhesion kinase (FAK) and thus we sought to investigate Shb's role in the progression of leukemia.

    METHODS: Wild type and Shb knockout bone marrow cells were transformed with a retroviral BCR-ABL construct and subsequently transplanted to wild type or Shb knockout recipients. Disease latency, bone marrow and peripheral blood cell characteristics, cytokine expression, signaling characteristics and colony formation were determined by flow cytometry, qPCR, western blotting and methylcellulose colony forming assays.

    RESULTS: It was observed that Shb knockout BCR-ABL-transformed bone marrow cells produced a disease with death occurring at earlier time points compared with corresponding wild type controls due to elevated proliferation of transformed bone marrow cells. Moreover, significantly elevated interleukin-6 and granulocyte colony-stimulation factor mRNA levels were observed in Shb knockout c-Kit + leukemic bone marrow cells providing a plausible explanation for the concurrent peripheral blood neutrophilia. Shb knockout leukemic bone marrow cells also showed increased ability to form colonies in methylcellulose devoid of cytokines that was dependent on the concomitantly observed increased activity of FAK. Transplanting BCR-ABL-transformed Shb knockout bone marrow cells to Shb knockout recipients revealed decreased disease latency without neutrophilia, thus implicating the importance of niche-derived cues for the increase of blood granulocytes.

    CONCLUSIONS: Absence of Shb accelerates disease progression by exerting dual roles in BCR-ABL-induced leukemia: increased cell expansion due to elevated FAK activity and neutrophilia in peripheral blood, the latter dependent on the genetic background of the leukemic niche.

  • 25.
    Gustafsson, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jamalpour, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Absence of the Shb adaptor protein accelerates leukemia progression by control of cytokine responses2013In: Experimental Hematology, ISSN 0301-472X, E-ISSN 1873-2399, Vol. 41, no 8, p. S50-S50Article in journal (Other academic)
  • 26.
    Gustafsson, Karin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Willebrand, Elsa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Absence of the adaptor protein Shb potentiates the TH2 response in a mouse model of atopic dermatitis2014In: Immunology, ISSN 0019-2805, E-ISSN 1365-2567, Vol. 143, no 1, p. 33-41Article in journal (Refereed)
    Abstract [en]

    Aberrant regulation of T helper (Th) cell maturation is associated with a number of autoimmune conditions, including allergic disorders and rheumatoid arthritis. The Src homology domain protein B (Shb) adaptor protein was recently implicated as a regulator of Th cell differentiation. Shb is an integral component of the T-cell receptor (TCR) signalling complex and in the absence of Shb the TCR is less responsive to stimulation, resulting in the preferential development of Th2 responses under conditions of in vitro stimulation. In the present study, we extend those observations to an in vivo situation using a murine model of atopic dermatitis. Shb knockout mice develop more pronounced symptoms of atopic dermatitis with increased localized oedema, epidermal hyperplasia and IgE production. Dermal infiltration of mast cells, eosinophils, CD4(+) Th cells and F4/80(+) macrophages was also significantly increased in Shb-deficient mice. This correlated with elevated transcription of the hallmark Th2 cytokines interleukin-4 and interleukin-5. The loss of Shb therefore alters TCR signalling ability, thereby favouring the development of Th2-driven inflammation and exacerbating symptoms of allergy.

  • 27.
    He, Qi
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Li, Xiujuan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Cyrus Tang Hematology Center, Soochow University, Suzhou, China.
    Singh, Kailash
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Luo, Zhengkang
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Meija-Cordova, Mariela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jamalpour, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lindahl, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kriz, Vitezslav
    Vuolteenaho, Reetta
    Ulvmar, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Cdh5-CreERT2 transgene causes conditional Shb gene deletion in hematopoietic cells with consequences for immune cell responses to tumors2019In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 9, article id 7548Article in journal (Refereed)
    Abstract [en]

    The tamoxifen-responsive conditional Cdh5-CreERT2 is commonly used for endothelial cell specific conditional deletion of loxP-flanked gene sequences. To address the role of endothelial cell Shb gene for B16F10 melanoma immune responses, tamoxifen-injected Cdh5-CreERT2/WT and Cdh5-CreERT2/Shbflox/flox mice received subcutaneous tumor cell injections. We observed a decrease of tumor myeloid cell Shb mRNA in the tamoxifen treated Cdh5-CreERT2/Shbflox/flox mice, which was not present when the mice had undergone a preceding bone marrow transplantation using wild type bone marrow. Differences in CD4+/FoxP3+ Tregs were similarly abolished by a preceding bone marrow transplantation. In ROSA26-mTmG mice, Cdh5-CreERT2 caused detectable floxing in certain bone marrow populations and in spleen cells. Floxing in bone marrow could be detected two months after tamoxifen treatment. In the spleen, however, floxing was undetectable two months after tamoxifen treatment, suggesting that Cdh5-CreERT2 is operating in a non-renewable population of hematopoietic cells in this organ. These data suggest that conditional gene deletion in hematopoietic cells is a potential confounder in experiments attempting to assess the role of endothelial specific effects. A cautious approach to achieve an endothelial-specific phenotype would be to adopt a strategy that includes a preceding bone marrow transplantation.

  • 28.
    Holmqvist, Kristina
    et al.
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
    Cross, Michael J
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Genetics and Pathology.
    Rolny, Charlotte
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
    Hägerkvist, Robert
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
    Rahimi, Nader
    Matsumoto, Taro
    Claesson-Welsh, Lena
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Genetics and Pathology.
    Welsh, Michael
    Uppsala University, Medicinska vetenskapsområdet, Faculty of Medicine, Department of Medical Cell Biology.
    The adaptor protein Shb binds to tyrosine 1175 in vascular endothelial growth factor (VEGF) receptor-2 and regulates VEGF-dependent cellular migration.2004In: J Biological chemistry, Vol. 279, no 21, p. 22267-22275Article in journal (Refereed)
  • 29.
    Holmqvist, Kristina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Cross, Michael
    Riley, Debbie
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Shb adaptor protein causes Src-dependent cell spreading and activation of focal adhesion kinase in murine brain endothelial cells2003In: Cell Signal, Vol. 15, no 2, p. 171-179Article in journal (Refereed)
  • 30.
    Holmqvist, Kristina
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lu, Lingge
    Center for Matrix Biology, Department of Medicine, Dana 513, Beth Israel Deaconess Medical Center and Harvard Medical School, 330 Brookline Ave, Boston, 02215 Massachusetts, USA.
    A role of the protein Cbl in FGF-2-induced angiogenesis in murine brain endothelial cells2005In: Cellular Signalling, ISSN 0898-6568, E-ISSN 1873-3913, Vol. 17, no 11, p. 1433-1438Article in journal (Refereed)
    Abstract [en]

    The Cbl protein functions both as a multivalent adaptor and a negative regulator of receptor tyrosine kinases (RTKs), the latter by directing polyubiquitination of RTKs. To study the function of Cbl in endothelial cell signalling and angiogenesis, wild-type Cbl and tyrosine kinase binding (TKB) domain mutated Cbl (G306E) were overexpressed in murine immortalised brain endothelial (IBE) cells. Wild-type Cbl cells exhibited enhanced proliferation in low serum compared with the control and G306E Cbl cells. Furthermore, up-regulated phosphorylation of fibroblast growth factor receptor 1 (FGFR-1) and Akt were observed in wild-type Cbl cells upon FGF-2 stimulation. A Cbl TKB domain mutant, G306E, disrupted the phosphorylation of the FGFR-1 but not that of FRS2. In the tubular morphogenesis assay, cells expressing wild-type Cbl initially formed tubular structures. These showed decreased stability and converted into cell aggregates, possibly due to a failure to cease proliferating. Our data support the idea that the wild-type Cbl cells exhibit enhanced proliferation, and thus lose their ability to differentiate appropriately. The present study reveals a role of the Cbl protein in FGF-2 dependent signalling in endothelial cells by its destabilisation of tubular structures.

  • 31.
    Hooshmand-Rad, Roya
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Lu, Lingge
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Heldin, Carl-Henrik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwig Institute for Cancer Research.
    Claesson-Welsh, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Platelet-Derived Growth Factor-Mediated Signaling through the Shb Adaptor Protein: Effects on Cytoskeletal Organization2000In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 257, no 2, p. 245-254Article in journal (Refereed)
    Abstract [en]

    The Src homology (SH) 2 domain adaptor protein Shb has previously been shown to interact with the platelet-derived growth factor (PDGF)-β receptor. In this study we show an association between Shb and the PDGF-α receptor which is mediated by the SH2 domain of Shb and involves tyrosine residue 720 in the kinase insert domain of the receptor. To assess the role of Shb in PDGF-mediated signaling, we have overexpressed wild-type Shb or Shb carrying a mutation (R522K) which renders the SH2 domain inactive, in Patch mouse (PhB) fibroblasts expressing both PDGF receptors (PhB/Rα). Overexpression of wild-type Shb, but not the R522K Shb mutant, affected PDGF-mediated reorganization of the cytoskeleton by decreasing membrane ruffle formation and stimulating the generation of filopodia relative the parental control cells. In addition, the PDGF-induced receptor-associated phosphatidylinositol 3′-kinase activity and phosphorylation of Akt was similar in both PhB/Rα/Shb and PhB/Rα/ShbR522K cells compared with the parental control, whereas the activation of Rac in response to PDGF-BB was diminished only in the PhB/Rα/Shb cells. We conclude that Shb plays a role in PDGF-dependent regulation of certain cytoskeletal changes by modulating the ability of PDGF to activate Rac.

  • 32.
    Hägerkvist, Robert
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mokhtari, Dariush
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lindholm, Cecilia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Farnebo, Filip
    Mostoslavsky, Gustavo
    Mulligan, Richard C.
    Welsh, Nils
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Consequences of Shb and c-Abl interactions for cell death in response to various stress stimuli2007In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 313, no 2, p. 284-291Article in journal (Refereed)
    Abstract [en]

    The adaptor protein Shb has previously been shown to regulate apoptosis in response to cytokines and inhibitors of angiogenesis although the mechanisms governing these effects have remained obscure. We currently demonstrate interactions between Shb and c-Abl and that Shb regulates c-Abl kinase activity. The data suggest that c-Abl binds to tyrosine phosphorylated Shb via a concerted effort involving both the c-Abl SH3 and SH2 domains. The biological significance of the Shb/c-Abl interaction was presently tested in overexpression experiments and was found to promote hydrogen peroxide-induced cell death. We also show by Shb knockdown experiments that Shb regulates c-Abl activity and modulates cell death in response to the genotoxic agent cisplatin and the endoplasmic reticulum stress-inducer tunicamycin. The findings are in agreement with the notion of Shb playing a pivotal role in modulating c-Abl pro-apoptotic signaling in response to various stress stimuli.

  • 33.
    Jamalpour, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Li, Xiujuan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Cavelier, Lucia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology.
    Gustafsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mostoslavsky, Gustavo
    Center for Regenerative Medicine (CReM), Department of Medicine, School of Medicine, Boston University..
    Höglund, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Tumor SHB gene expression affects disease characteristics in human acute myeloid leukemia2017In: Tumor Biology, ISSN 1010-4283, E-ISSN 1423-0380, Vol. 39, no 10Article in journal (Refereed)
    Abstract [en]

    Objective: The mouse Shb gene coding for the Src Homology 2-domain containing adapter protein B has recently been placed in context of BCRABL1-induced myeloid leukemia in mice and the current study was performed in order to relate SHB to human acute myeloid leukemia (AML). Methods: Publicly available AML databases were mined for SHB gene expression and patient survival. SHB gene expression was determined in the Uppsala cohort of AML patients by qPCR. Cell proliferation was determined after SHB gene knockdown in leukemic cell lines. Results: Despite a low frequency of SHB gene mutations, many tumors overexpressed SHB mRNA compared with normal myeloid blood cells. AML patients with tumors expressing low SHB mRNA displayed longer survival times. A subgroup of AML exhibiting a favorable prognosis, acute promyelocytic leukemia (APL) with a PMLRARA translocation, expressed less SHB mRNA than AML tumors in general. When examining genes co-expressed with SHB in AML tumors, four other genes (PAX5, HDAC7, BCORL1, TET1) related to leukemia were identified. A network consisting of these genes plus SHB was identified that relates to certain phenotypic characteristics, such as immune cell, vascular and apoptotic features. SHB knockdown in the APL PMLRARA cell line NB4 and the monocyte/macrophage cell line MM6 adversely affected proliferation, linking SHB gene expression to tumor cell expansion and consequently to patient survival. Conclusions: It is concluded that tumor SHB gene expression relates to AML survival and its subgroup APL. Moreover, this gene is included in a network of genes that plays a role for an AML phenotype exhibiting certain immune cell, vascular and apoptotic characteristics.

  • 34.
    Jamalpour, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Li, Xiujuan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Cavelier, Lucia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Medicinsk genetik och genomik.
    Gustafsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mostoslavsky, Gustavo
    Center for Regenerative Medicine (CReM), Department of Medicine, School of Medicine, Boston University, Boston, MA, USA.
    Höglund, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Haematology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Tumor SHB gene expression affects disease characteristics in human acute myeloid leukemia2017In: Tumor Biology, ISSN 1010-4283, E-ISSN 1423-0380, Vol. 39, no 10, article id 1010428317720643Article in journal (Refereed)
    Abstract [en]

    The mouse Shb gene coding for the Src Homology 2-domain containing adapter protein B has recently been placed in context of BCRABL1-induced myeloid leukemia in mice and the current study was performed in order to relate SHB to human acute myeloid leukemia (AML). Publicly available AML databases were mined for SHB gene expression and patient survival. SHB gene expression was determined in the Uppsala cohort of AML patients by qPCR. Cell proliferation was determined after SHB gene knockdown in leukemic cell lines. Despite a low frequency of SHB gene mutations, many tumors overexpressed SHB mRNA compared with normal myeloid blood cells. AML patients with tumors expressing low SHB mRNA displayed longer survival times. A subgroup of AML exhibiting a favorable prognosis, acute promyelocytic leukemia (APL) with a PMLRARA translocation, expressed less SHB mRNA than AML tumors in general. When examining genes co-expressed with SHB in AML tumors, four other genes (PAX5, HDAC7, BCORL1, TET1) related to leukemia were identified. A network consisting of these genes plus SHB was identified that relates to certain phenotypic characteristics, such as immune cell, vascular and apoptotic features. SHB knockdown in the APL PMLRARA cell line NB4 and the monocyte/macrophage cell line MM6 adversely affected proliferation, linking SHB gene expression to tumor cell expansion and consequently to patient survival. It is concluded that tumor SHB gene expression relates to AML survival and its subgroup APL. Moreover, this gene is included in a network of genes that plays a role for an AML phenotype exhibiting certain immune cell, vascular and apoptotic characteristics.

  • 35.
    Jamalpour, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Li, Xiujuan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Gustafsson, Karin
    Harvard Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA, USA.;Center for Regenerative Medicine and the Cancer Center, Massachusetts General Hospital, MA, USA.
    Tyner, Jeffrey
    Cell, Developmental & Cancer Biology, Oregon Health & Science University, Portland, OR, USA.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Disparate effects of Shb-gene deficiency on disease characteristics in murine models of myeloid, B-cell and T-cell leukemia2018In: Tumor Biology, ISSN 1010-4283, E-ISSN 1423-0380, Vol. 40, no 4, p. 1-13Article in journal (Refereed)
    Abstract [en]

    The Src homology-2 domain protein B is an adaptor protein operating downstream of tyrosine kinases. The Shb gene knockout has been found to accelerate p210 Breakpoint cluster region-cAbl oncogene 1 tyrosine kinase-induced leukemia. In human myeloid leukemia were tumors with high Src homology-2 domain protein B mRNA content, tumors were, however, associated with decreased latency and myeloid leukemia exhibiting immune cell characteristics. Thus, the aim of this study was to investigate the effects of Shb knockout on the development of leukemia in three additional models, that is, colony stimulating factor 3 receptor-T618I–induced neutrophilic leukemia, p190 Breakpoint cluster region-cAbl oncogene 1 tyrosine kinase-induced B-cell leukemia, and G12D-Kras-induced T-cell leukemia/thymic lymphoma. Wild-type or Shb knockout bone marrow cells expressing the oncogenes were transplanted to bone marrow–deficient recipients. Organs from moribund mice were collected and further analyzed. Shb knockout increased the development of CSF3RT618I-induced leukemia and increased the white blood cell count at the time of death. In the p190 Breakpoint cluster region-cAbl oncogene 1 tyrosine kinase B-cell model, Shb knockout reduced white blood cell counts without affecting latency, whereas in the G12D-Kras T-cell model, thymus size was increased without major effects on latency, suggesting that Shb knockout accelerates the development thymic lymphoma. Cytokine secretion plays a role in the progression of leukemia, and consequently Shb knockout bone marrows exhibited lower expression of granulocyte colony stimulating factor and interleukin 6 in the neutrophilic model and interleukin 7 and chemokine C-X-C motif ligand 12 (C-X-C motif chemokine 12) in the B-cell model. It is concluded that in experimental mouse models, the absence of the Shb gene exacerbates the disease in myeloid leukemia, whereas it alters the disease characteristics without affecting latency in B- and T-cell leukemia. The results suggest a role of Shb in modulating the disease characteristics depending on the oncogenic insult operating on hematopoietic cells. These findings help explain the outcome of human disease in relation to Src homology-2 domain protein B mRNA content.

  • 36.
    Karlsson, Torbjörn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kullander, Klas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The Src homology 2 domain protein shb transmits basic fibroblast growth factor - and nerve growth factor - dependent differentiation signals in PC12 cells11998In: Cell growth & differentiation, ISSN 1044-9523, Vol. 9, no 9, p. 757-766Article in journal (Refereed)
    Abstract [en]

    To assess a possible role for the Src homology 2 (SH2) domain adaptor protein Shb in PC12 cell signal transduction and differentiation, we have investigated the expression of Shb in PC12 cells and found that the differentiation factors nerve growth factor (NGF) and basic fibroblast growth factor (bFGF), as well as the PC12 cell mitogen epidermal growth factor, increased Shb protein expression and Shb mRNA steady-state levels. Two PC12 cell clones stably overexpressing the Shb cDNA exhibited enhanced NGF- or bFGF-induced differentiation, assessed as neurite outgrowth. This effect showed no direct correlation to mitogen-activated protein kinase activation, although the mitogen-activated protein kinase/kinase inhibitor PD-98059 caused a partial inhibition of neurite outgrowth. Furthermore, it was found that the Shb-overexpressing cells extended neurites in response to epidermal growth factor. The effects of Shb overexpression on neurite outgrowth required a functional SH2 domain because PC12 cells expressing Shb with an inactivated SH2 domain did not differentiate more readily in response to NGF. Tyrosine phosphorylation of the p13 Suc1-associated neurotrophic factor-induced tyrosine-phosphorylated target protein in response to bFGF was strongly inhibited by Shb overexpression, without correlating with the corresponding rate of neurite outgrowth. NGF-induced tyrosine phosphorylation of phospholipase Cgamma, TrkA, and Shc was unaltered in the Shb-overexpressing cells, whereas that of Shb was greatly enhanced in these cells compared with control PC12-neo cells. In addition, an NGF-activated Mr 140,000 phosphotyrosine protein was found to be associated with Shb in immunoprecipitation experiments. The data in this study suggest that Shb is involved in transmission of NGF- and bFGF-dependent differentiation signals in PC12 cells.

  • 37.
    Karlsson, Torbjörn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Songyang, Z
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Landgren, E
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lavergne, C
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Di Fiore, P P
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Anafi, M
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Pawson, T
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Cantley, L C
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Claesson-Welsh, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Molecular interactions of the Src homology 2 domain protein Shb with phosphotyrosine residues, tyrosine kinase receptors and Src homology 3 domain proteins1995In: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 10, no 8, p. 1475-1483Article in journal (Refereed)
    Abstract [en]

    The molecular interactions of the Src homology 2 (SH2) domain and the N-terminal proline-rich sequence motifs (pro-1 to pro-5) of the SH2 protein Shb with other components were presently characterised. Using a degenerate phosphopeptide library the preferred binding site for the Shb SH2 domain was determined to pTyr-Thr/Val/Ile-X-Leu at positions +1 to +3 relative the phosphotyrosine residue. Experiments with competing peptides and platelet-derived growth factor (PDGF) beta-receptor mutants with Y to F substitutions in autophosphorylation sites revealed multiple binding sites for the Shb SH2 domain in the receptor. The Shb SH2 domain also binds to in vitro phosphorylated fibroblast growth factor receptor-1 (FGFR-1) mainly through position Y776. The receptor experiments suggest that other residues besides the +1 to +3 positions may also be of significance for Shb binding. The pro-4/pro-5 motif of Shb binds in vitro particularly well to the Src, p85 alpha PI3-kinase and Eps8 SH3 domains expressed as GST fusion proteins. However, the GST-SH3 domain fusion proteins tested bind in vitro to peptides corresponding to the pro-1 to pro-5 motifs of Shb with low affinity and selectivity, suggesting that sequences outside the core proline motif may also be important for Shb-SH3 domain interactions. In vivo association between Shb-SH3 domain proteins v-Src and Eps8 was detected by coimmunoprecipitation. PDGF treatment did not affect the association between Eps8 and Shb. The data suggest that Shb is an adaptor protein linking SH3 domain proteins to tyrosine kinases or other tyrosine phosphorylated proteins.

  • 38.
    Karlsson, Torbjörn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Apoptosis of NIH3T3 cells overexpressing the Src homology 2 domain protein Shb1996In: Oncogene, ISSN 0950-9232, E-ISSN 1476-5594, Vol. 13, no 5, p. 955-961Article in journal (Refereed)
    Abstract [en]

    To understand the role of the Src homology 2 (SH2) domain protein Shb in the signal transduction of tyrosine kinase receptor, NIH3T3 cells were transfected with a DNA construct expressing the Shb cDNA (NIHSHB cells). The NIHSHB cells expressed elevated levels of proteins with the estimated molecular weights of 77, 66 and 55 kDa as determined by immunoblotting. In contrast to the control cells, the NIHSHB cells failed to increase in cell number in the presence of 1% serum. This effect was largely due to apoptosis, since staining of pyknotic nuclei was observed using the terminal transferase labeling method. The NIHSHB cells displayed similar levels of c-myc mRNA and decreased contents of the p53 protein after culture in 1% serum compared with control cells. The addition of platelet-derived growth factor (PDGF-BB) restored the growth of the NIHSHB cells, whereas insulin-like growth factor-1 (IGF-1) failed to affect the proliferation of Shb overexpressing cells in 1% serum. We conclude that Shb overexpression is associated with cell degeneration under certain conditions, and that Shb could transduce apoptotic signals from tyrosine kinase receptors.

  • 39.
    Karlsson, Torbjörn
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Modulation of Src homology 3 proteins by the proline-rich adaptor protein Shb1997In: Experimental Cell Research, ISSN 0014-4827, E-ISSN 1090-2422, Vol. 231, no 2, p. 269-275Article in journal (Refereed)
    Abstract [en]

    In the present study we have investigated a possible role for the proline-rich SH2 domain protein Shb as a regulator of expression or activity of certain SH3 domain proteins and MAP kinase. The expression of the Shb binding proteins Eps8, Src, and p85 PI3-kinase, PI3-kinase activity, and MAP kinase activation were assessed in wild-type NIH3T3 cells and in NIH3T3 cells overexpressing the Shb cDNA. In addition, the expression of the SH3 domain STAT1 proteins was assessed in wild-type and Shb overexpressing cells. The Eps8 protein content and Eps8 mRNA steady-state levels were downregulated, whereas the protein contents of Src and p85 PI3-kinase were unaffected by Shb overexpression. There was, however, an increased basal PI3-kinase activity in Shb transfected cells after a 3-h serum starvation. Increased steady-state levels of STAT1 mRNA were accompanied by an increased STAT1 protein content in Shb overexpressing cells. Shb overexpression was not associated with an altered activation of p44 or p42 MAP kinases in response to PDGF stimulation. The data presented in this study suggest novel functions for the adaptor protein Shb regulating the expression of certain signal-transducing SH3 domain proteins and modulating PI3-kinase activity.

  • 40.
    Kriz, Vitezslav
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Anneren, Cecilia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lai, Cheng
    Karlsson, Jacob
    Mares, Jaroslav
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The SHB protein is required for efficient multilineage differentiation of mouse embryonic stem cells2003In: Exp Cell Res, Vol. 286, no (1), p. 40-56Article in journal (Refereed)
  • 41.
    Kriz, Vitezslav
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mares, Jaroslav
    Wentzel, Parri
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Funa, Nina S.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Calounova, Gabriela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Zhang, X-Q
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Forsberg-Nilsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Forsberg, Maud
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Shb null allele is inherited with a transmission ratio distortion and causes reduced viability in utero2007In: Developmental Dynamics, ISSN 1058-8388, E-ISSN 1097-0177, Vol. 236, no 9, p. 2485-2492Article in journal (Refereed)
    Abstract [en]

    SHB is an Src homology 2 domain-containing adapter protein that has been found to be involved in numerous cellular responses. We have generated an Shb knockout mouse. No Shb-/- pups or embryos were obtained on the C57Bl6 background, indicating an early defect as a consequence of Shb- gene inactivation on this genetic background. Breeding heterozygotes for Shb gene inactivation (Shb+/-) on a mixed genetic background (FVB/C57Bl6/129Sv) reveals a distorted transmission ratio of the null allele with reduced numbers of Shb+/+ and Shb-/- animals, but increased number of Shb+/- animals. The Shb- allele is associated with various forms of malformations, explaining the relative reduction in the number of Shb-/- offspring. Shb-/- animals that were born were viable, fertile, and showed no obvious defects. However, Shb+/- female mice ovulated preferentially Shb- oocytes explaining the reduced frequency of Shb+/+ mice. Our study suggests a role of SHB during reproduction and development.

  • 42.
    Kriz, Vitezslav
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Ågren, Nina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lindholm, Cecilia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lenell, Samuel
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Saldeen, Johan
    Mares, Jaroslav
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The SHB adapter protein is required for normal maturation of mesoderm during in vitro differentiation of embryonic stem cells2006In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 281, no 45, p. 34484-34491Article in journal (Refereed)
    Abstract [en]

    Definitive mesoderm arises from a bipotent mesendodermal population, and to study processes controlling its development at this stage, embryonic stem (ES) cells can be employed. SHB ((S) under bar rc (h) under bar omology 2 protein in beta-cells) is an adapter protein previously found to be involved in ES cell differentiation to mesoderm. To further study the role of SHB in this context, we have established ES cell lines deficient for one (SHB+/-) or both SHB alleles (SHB-/-). Differentiating embryoid bodies (EBs) derived from these ES cell lines were used for gene expression analysis. Alternatively, EBs were stained for the blood vessel marker CD31. For hematopoietic differentiation, EBs were differentiated in methylcellulose. SHB-/- EBs exhibited delayed down-regulation of the early mesodermal marker Brachyury. Later mesodermal markers relatively specific for the hematopoietic, vascular, and cardiac lineages were expressed at lower levels on day 6 or 8 of differentiation in EBs lacking SHB. The expression of vascular endothelial growth factor receptor-2 and fibroblast growth factor receptor-1 was also reduced in SHB-/- EBs. SHB-/- EBs demonstrated impaired blood vessel formation after vascular endothelial growth factor stimulation. In addition, the SHB-/- ES cells formed fewer blood cell colonies than SHB+/+ ES cells. It is concluded that SHB is required for appropriate hematopoietic and vascular differentiation and that delayed down-regulation of Brachyury expression may play a role in this context.

  • 43.
    Lavergne, Corinne
    et al.
    INSERM CJF 93-13, Hopital Robert Debré and INSERM U55, Hopital Saint-Antoine, Paris, France.
    Mares, Jaroslav
    Institute of Biology and Medical Genetics, Charles University, Prague, Czech Republic.
    Karlsson, Torbjörn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Breant, Bernadette
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Control of SHB gene expression by protein phosphorylation1996In: Cellular Signalling, ISSN 0898-6568, E-ISSN 1873-3913, Vol. 8, no 1, p. 55-58Article in journal (Refereed)
    Abstract [en]

    To increase our understanding of the role of the Src homology 2 (SH2) domain-containing protein Shb in the mitogenic signal transduction, Shb mRNA contents were determined in the fibroblast-like NIH3T3 cells and the insulin producing βTC-1 cells under various conditions. In NIH3T3 cells, the serine/ threonine phosphatase inhibitor okadaic acid and the tyrosine kinase inhibitor genistein increased Shb mRNA contents, the protein kinase C activating phorbol ester 12-O-tetradecanoyl 13-acetate (TPA) decreased the Shb mRNA content, whereas the tyrosine kinase inhibitor tyrphostin 25 and the mitogen platelet-derived growth factor (PDGF-BB) had no effect. In βTC-1 cells, okadaic acid and genistein increased the Shb mRNA content, whereas tyrphostin 25 and serum were without effect. Okadaic acid and genistein decreased the rates of βTC-1 cell DNA synthesis. It is concluded that expression of the SHB gene is under a complex mode of regulation involving at least three different protein kinases. As a consequence of this, it is likely that SHB gene expression is significantly modulated by conditions of specific activation of certain pathways, whereas its expression appears little influenced by serum and a mitogen.

  • 44.
    Li, Xiujuan
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology. Soochow University, Suzhou, China.
    Singh, Kailash
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Luo, Zhengkang
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Mejia Cordova, Mariela
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Jamalpour, Maria
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Lindahl, Björn
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Zhang, Ganlin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sandler, Stellan
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Pro-tumoral immune cell alterations in wild type and Shb-deficient mice in response to 4T1 breast carcinomas2018In: OncoTarget, ISSN 1949-2553, E-ISSN 1949-2553, Vol. 9, no 27, p. 18720-18733Article in journal (Refereed)
    Abstract [en]

    To assess mechanisms responsible for breast carcinoma metastasis, 4T1 breast carcinomas were grown orthotopically in wild type or Shb knockout mice. Tumor growth, metastasis, vascular characteristics and immune cell properties were analyzed. Absence of Shb did not affect tumor growth although it increased lung metastasis. Shb knockout mouse tumors showed decreased redness and less developed vascular plexa located at the periphery of the tumors. No difference in overall tumor vascular density, leakage or pericyte coverage was noted between the genotypes although the average vessel size was smaller in the knockout. Tumors induced an increase of CD11b+ cells in spleen, lymph node, thymus, bone marrow and blood. Numbers of Shb knockout CD11b/CD8+ cells were decreased in lymph nodes and bone marrow of tumor bearing mice. Mice with tumors had reduced numbers of CD4+ lymphocytes in blood/lymphoid organs, whereas in most of these locations the proportion of CD4+ cells co-expressing FoxP3 was increased, suggesting a relative increase in Treg cells. This finding was reinforced by increased blood interleukin-35 (IL-35) in wild type tumor bearing mice. Shb knockout blood showed in addition an increased proportion of IL-35 expressing Treg cells, supporting the notion that absence of Shb further promotes tumor evasion from immune cell recognition. This could explain the increased number of lung metastases observed under these conditions. In conclusion, 4T1 tumors alter immune cell responses that promote tumor expansion, metastasis and escape from T cell recognition in an Shb dependent manner. 

  • 45.
    Lindholm, Cecilia K
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Frantz, Daniel J
    Shoelson, Steven E
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Shf, a Shb-like Adapter protein, Is Involved in PDGF-a-receptor Regulation of Apoptosis2000In: Biochemical and Biophysical Research Communications - BBRC, ISSN 0006-291X, E-ISSN 1090-2104, Vol. 278, no 3, p. 537-543Article in journal (Refereed)
    Abstract [en]

    Recent work has implicated the importance of adapter proteins in signal transduction. To identify homologues of the previously identified adapter protein Shb, database searches were performed. A Shb-like protein was found which we have named Shf. Shf contains an SH2 domain and four putative tyrosine phosphorylation sites and is mainly expressed in skeletal muscle, brain, liver, prostate, testis, ovary, small intestine, and colon. The SH2 domain of Shf bound to the PDGF-alpha-receptor at tyrosine-720, but not to the PDGF-beta-receptor in PAE cells. Pervanadate induced tyrosine phosphorylation of Shf in NIH3T3 fibroblasts overexpressing this protein, whereas PDGF-AA alone had no detectable effect. NIH3T3 cells overexpressing Shf displayed significantly lower rates of apoptosis than control cells in the presence of PDGF-AA. Our findings suggest a role for the novel adapter Shf in PDGF-receptor signaling and regulation of apoptosis.

  • 46.
    Lindholm, Cecilia K
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Gylfe, Erik
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Zhang, Weigu
    Samelson, Lawrence E
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Requirement of the Src homology 2 domain protein Shb for T cell receptor-dependent activation of the Interleukin-2 gene nuclear factor for activation of T cells element in Jurkat T cells1999In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 274, no 39, p. 28050-28057Article in journal (Refereed)
    Abstract [en]

    Stimulation of the T cell antigen receptor (TCR) induces tyrosine phosphorylation of numerous intracellular proteins. We have recently investigated the role of the adaptor protein Shb in the early events of T cell signaling and observed that Shb associates with Grb2, linker for activation of T cells (LAT) and the TCR zeta-chain in Jurkat cells. We now report that Shb also associates with phospholipase C-gamma1 (PLC-gamma1) in these cells. Overexpression of Src homology 2 domain defective Shb caused diminished phosphorylation of LAT and consequently the activation of mitogen-activated protein kinases was decreased upon TCR stimulation. In addition, the Shb mutant also blocked phosphorylation of PLC-gamma1 and the increase in cytoplasmic Ca(2+) following TCR stimulation. Nuclear factor for activation of T cells is a major target for Ras and calcium signaling pathways in T cells following TCR stimulation, and the overexpression of the mutant Shb prevented TCR-dependent activation of the nuclear factor for activation of T cells. Consequently, endogenous interleukin-2 production was decreased under these conditions. The results indicate a role for Shb as a link between the TCR and downstream signaling events involving LAT and PLC-gamma1 and resulting in the activation of transcription of the interleukin-2 gene.

  • 47.
    Lindholm, Cecilia K
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Henriksson, Maria L
    Hallberg, Bengt
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Shb links SLP-76 and Vav with the CD3 complex in Jurkat T cells2002In: European Journal of Biochemistry, ISSN 0014-2956, E-ISSN 1432-1033, Vol. 269, no 13, p. 3279-3288Article in journal (Refereed)
    Abstract [en]

    This study addresses the interactions between the adaptor protein Shb and components involved in T cell signalling, including SLP-76, Gads, Vav and ZAP70. We show that both SLP-76 and ZAP70 co-immunoprecipitate with Shb in Jurkat T cells and that Shb and Vav co-immunoprecipitate when cotransfected in COS cells. We also demonstrate, utilizing fusion protein constructs, that SLP-76, Gads and Vav associate independently of each other to different domains or regions, of Shb. Overexpression of an SH2 domain-defective Shb causes diminished phosphorylation of SLP-76 and Vav and consequently decreased activation of c-Jun kinase upon T cell receptor (TCR) stimulation. Shb was also found to localize to glycolipid-enriched membrane microdomains (GEMs), also called lipid rafts, after TCR stimulation. Our results indicate that upon TCR stimulation, Shb is targeted to these lipid rafts where Shb aids in recruiting the SLP-76–Gads–Vav complex to the T cell receptor ζ-chain and ZAP70.

  • 48.
    Lu, Lingge
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Anneren, Cecilia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Reedquist, Kris A.
    Bos, Johannes L.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    NGF-dependent neurite outgrowth in PC12 cells overexpressing the Src homology 2-domain protein Shb requires activation of the Rap1 pathway2000In: Exp. Cell Res., Vol. 259, no 2, p. 370-377Article in journal (Refereed)
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    Holmqvist, Kristina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Cross, Michael
    Welsh, Michael
    Role of the Src Homology 2 domain-containing protein Shb in murine brain endothelial cell proliferation and differentiation2002In: Cell Growth Differ, Vol. 13, no 3, p. 141-148Article in journal (Refereed)
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    Massena, Sara
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    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Christoffersson, Gustaf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Vågesjö, Evelina
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Gustafsson, Karin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Kutschera, Simone
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology.
    Welsh, Michael
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    Claesson-Welsh, Lena
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Cancer and Vascular Biology.
    Phillipson, Mia
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Cell Biology.
    The mechanisms of VEGF-A-induced recruitment of pro-angiogenic neutrophils2013In: European Journal of Clinical Investigation, ISSN 0014-2972, E-ISSN 1365-2362, Vol. 43, no SI, p. 27-27Article in journal (Other academic)
12 1 - 50 of 77
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