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Author(s): |
Lays Martin Sobral
Total Authors: 1
|
Document type: | Master's Dissertation |
Press: | Piracicaba, SP. |
Institution: | Universidade Estadual de Campinas (UNICAMP). Faculdade de Odontologia de Piracicaba |
Defense date: | 2007-02-23 |
Examining board members: |
Ricardo Della Coletta;
Paulo Tambasco de Oliveira;
Edgard Graner
|
Advisor: | Ricardo Della Coletta |
Abstract | |
Myofibroblasts are the main cellular type involved in extracellular matrix deposition in fibrotic diseases. Relatively little is known about the underlying mechanisms that regulate myofibroblast emergence (transdifferentiation), however the regulatory cytokine transforming growth factor-beta 1 (TGF-'beta'1) has been traditionally considered an inducer of the myofibroblastic phenotype via activation of connective tissue growth factor (CTGF)-dependent pathway. Some inflammatory cytokines, particularly interferon g (IFN'gama'), show opposite effects to TGF-'beta'1, inhibiting myofibroblast transdifferentiation. Cyclosporin A (CsA) is a widely used immunosuppressant drug that causes significant side effects such as gingival overgrowth. Normal gingival fibroblast cell lines treated with CsA express high levels of TGF-'beta'1 and collagen, which are intrinsic of myofibroblasts. In the present study we examined whether TGF-'beta'1 and IFN'gama' can modulate transdifferentiation of gingival fibroblasts into myofibroblasts, and the biologic mechanisms behind those factors in this process. Additionally, we have analyzed the presence of myofibroblasts in CsA-induced gingival overgrowth. Our results demonstrated throughout a modality of experiments that included reverse trancriptasepolymerase chain reaction (RT-PCR), western blot, immunofluorescence and flow citometry analysis, that TGF-'beta'1 induces and IFN'gama' blocks fibroblast-myofibroblast transdifferentiation in a dose- and time-dependent manner. In fibroblasts with CTGF levels know-down by specific small interference RNA, TGF-'beta'1 effect on transdifferentiation was significantly reduced, revealing that CTGF plays a crucial role in mediating TGF-'beta'1 myofibroblast transdifferentiation. IFN'gama' blocked myofibroblast transdifferentiation by stimulating Smad 7 levels, a protein that negatively regulates TGF-'beta'1 signaling. Interestingly, myofibroblasts were not found in CsA-induced gingival overgrowth, as revealed by the in vivo model of daily injections of CsA in Wistar rats and by the in vitro model of gingival fibroblast cultures treated with CsA. Although CsA treatment stimulated TGF-'beta'1 expression by NG fibroblasts, it lacked to induce CTGF levels. Our results demonstrate that TGF-'beta'1 induction of transdifferentiation of gingival fibroblasts to myofibroblasts occurs via a CTGF dependent pathway. Additionally, this study suggests that IFN'gama' may be clinically effective in the treatment of fibrotic diseases associate with myofibroblast (AU) |