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An ex vivo model for functional studies of myofibroblasts
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Genetics and Pathology.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Oncology, Radiology and Clinical Immunology. Radiology. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology. (R Nyman)
2005 (English)In: Laboratory Investigation, ISSN 0023-6837, E-ISSN 1530-0307, Vol. 85, no 5, 643-54 p.Article in journal (Refereed) Published
Abstract [en]

Migration, proliferation and invasive growth of myofibroblasts are key cellular events during formation of granulation tissue in situations of wound healing, arteriosclerosis and tumor growth. To study the invasive phenotype of myofibroblasts, we established an assay where arterial tissue from chicken embryos was embedded in fibrin gels and stimulated with growth factors. Addition of serum, PDGF-BB and FGF-2, but not VEGF-A, resulted in an outgrowth of cellular sprouts with a pattern that was similar to the organization of cells invading a provisional matrix in an in vivo model of wound healing using the chicken chorioallantoic membrane. Sprouting cells were defined as myofibroblasts based on being alpha-smooth muscle actin-positive but desmin-negative. There was no contribution of endothelial cells in outgrowing sprouts. The acquired myofibroblastic phenotype was stable since sprout-derived cells resumed sprouting in a growth factor-independent manner when re-embedded as spheroids in a fibrin matrix. Invasive growth and sprouting of vascular smooth muscle cells was not limited to chicken cells since a similar response was seen when spheroids composed of purified primary human aortic smooth muscle cells were embedded in fibrin. Finally, a technique for flat visualization of the three-dimensional sprouting and a quantification method is described. This ex vivo model allows quantitative analysis of invasive growth and differentiation of vascular smooth muscle cells and fibroblasts into myofibroblasts.

Place, publisher, year, edition, pages
2005. Vol. 85, no 5, 643-54 p.
Keyword [en]
Animals, Aorta/cytology, Cell Differentiation/drug effects/*physiology, Cell Movement/drug effects/*physiology, Chick Embryo, Chorioallantoic Membrane/cytology/drug effects, Disease Models; Animal, Fibroblast Growth Factor 2/pharmacology, Fibroblasts/*cytology/drug effects, Humans, Myocytes; Smooth Muscle/*cytology/drug effects, Neovascularization; Physiologic, Phenotype, Platelet-Derived Growth Factor/pharmacology, Research Support; Non-U.S. Gov't, Spheroids; Cellular/cytology/drug effects, Wound Healing/drug effects/*physiology
National Category
Medical and Health Sciences
URN: urn:nbn:se:uu:diva-77847DOI: 10.1038/labinvest.3700255PubMedID: 15723087OAI: oai:DiVA.org:uu-77847DiVA: diva2:105760
Available from: 2006-06-29 Created: 2006-06-29 Last updated: 2011-01-11Bibliographically approved
In thesis
1. Mechanisms of Tissue Vascularization
Open this publication in new window or tab >>Mechanisms of Tissue Vascularization
2005 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Tissue neovascularization in postnatal life occurs during post-traumatic tissue healing, neoplastic growth and in the endometrium during the reproductive cycle of females. Although embryonic angiogenesis has been intensively studied, far less is known about tissue revascularization and vessel remodeling in adults due to methodological difficulties. In the current studies, we developed a novel in vivo model of neovascularization that is performed on the chicken chorioallantoic membrane (CAM). A provisional matrix placed on the CAM was vascularized in response to FGF-2. In order to distinguish new from pre-existing vessels, the matrix was separated from the CAM by a nylon grid. Techniques to visualize the three dimensional structure of vascular networks and a method for rapid and semi-automated quantification were developed. This novel model allowed us to study the effects of potential inhibitors of tissue vascularization and their effects on the pre-existing vasculature. We found that while fumagillin or inhibition of MEK and Src inhibited only neovascularization, addition of cortisol or wortmannin was toxic to pre-existing vessels.

The CAM model allowed intravital observations during extended periods of time, which together with immunohistochemical analysis revealed a novel mechanism of tissue vascularization. Tensional forces generated by myofibroblast-mediated contraction of the provisional matrix, induced and directed ingrowth of vascular tissue. During the initial stages of vascularization, the vascular network was recruited from the surrounding tissue through a non-angiogenic mechanism by elongation and enlargement of pre-existing vessels, which were moved as vascular loops with constant functional circulation. Ingrown vessels were remodeled, presumably through intussusception, fusion and pruning. The CAM model was validated by observations of neovascularization associated with healing of the injured mouse cornea.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2005. 48 p.
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 14
Pathology, vascularization, angiogenesis, granulation tissue, myofibroblast, wound healing, Patologi
National Category
Cell and Molecular Biology
urn:nbn:se:uu:diva-4819 (URN)91-554-6166-2 (ISBN)
Public defence
2005-04-08, Rudbecksalen, Rudbecklaboratoriet, Dag Hammarskjölds väg 20, Uppsala, 09:00 (English)
Available from: 2005-03-17 Created: 2005-03-17 Last updated: 2009-05-13Bibliographically approved

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