The thrust of our application is based on two groups of findings. Our previous data demonstrate a role for profibrogenic cytokines in the occurrence of skin fibrosis in TSK mice that develop a scleroderma-like syndrome. TSK mice do not develop skin sclerosis following disruption of IL-4, IL-4R gene, or TGF-gene. The lack of a compensatory effect on collagen synthesis by TGF gein mice with a disrupted IL-4R gene suggests that IL-4 may influence the expression of TGF gene in fibroblasts. Additionally, reported data demonstrate that levels of profibrogenic cytokines are elevated in the serum of human scleroderma patients. Little is known of the molecular mechanisms by which IL-4 and IL-13 up-regulate collagen expression. Specifically, the signaling pathways that lead to over-synthesis of collagen and formation of fibrosis as well as the molecular basis of the epistatic interaction between IL-4 and TGF-genes are not known. The overall goal of this application is to address these questions.
The specific aims are as follow: 1. To characterize the role of IL-4 and IL-13 in STAT-6-dependent pathways leading to up-regulation of collagen expression in normal and TSK fibroblasts. This wilt be studied by investigating STAT-6 activation, promoter activity, collagen transcription, and collagen synthesis in fibroblasts stimulated with IL-4 and IL-13. 2. To delineate the molecular mechanisms of the IL-4 and TGF-gene interaction that regulate the expression of collagen genes. This will be studied using a set of deletion constructs containing the TGF-gene promoter followed by the CAT gene; the effects of IL-4 and IL-13 on promoter activity will be assessed. We will also study the DNA binding of STAT-6 to the TGF-gene promoter. AP-1, SP-1, NF-1, GATA3, and c-Maf transcription factors will also be examined since the DNA binding sites for these transcription factors have been identified. 3. To study the effect of the activation of MAPK modules by IL-4 and IL-13 on the expression of the collagen (I) gene in a human skin fibroblast line. The kinetics of activation of various molecules of ERK, p38, and c- Jun MAPK modules will be examined. The effect of dominant-positive and dominant-negative constructs on collagen gene expression will be studied in the presence or absence of profibrogenic cytokines.
| Bhogal, Rashpal K; Stoica, Cristina M; McGaha, Tracy L et al. (2005) Molecular aspects of regulation of collagen gene expression in fibrosis. J Clin Immunol 25:592-603 |
