Low oxygen tension stimulates collagen synthesis and COL1A1 transcription through the action of TGF-β1

Recent findings point to low oxygen tension (hypoxia) as an important mechanism for the expression of several eukaryotic genes. We have previously shown that hypoxia (2% O-2), when compared to standard oxygen tension (20% O-2), upregulates the mRNA levels of the human alpha1 (1) (COL1A1) procollagen gene and transforming growth factor-betal TGF-beta1) in human dermal fibroblasts. In this report, we determined the effect of hypoxia on collagen synthesis and transcription. Exposure of human dermal fibroblasts to hypoxia for 24-72 h led to a threefold, dose-dependent increase in collagenous protein (P<0.0001; r = 0.9794) and to enhanced type I procollagen deposition, as shown by direct immunofluorescence. Transient transfections with a series of luciferase- and CAT-promoter constructs of the human COL1A1 gene (spanning from - 2.5 kb to + 113 bp) showed that hypoxia increases the transcriptional activity of constructs having 5' endpoints between - 804 bp and - 107 bp, with loss of stimulation at - 84 bp. Maximal increase in promoter activity in hypoxia was observed between - 190 and - 174 bp of the proximal promoter, once a cKrox repressor site (-199 to -224 bp) was deleted. Upregulation of COL1A1 mRNA levels in hypoxia was blocked by a TGF-beta1 anti-sense oligonucleotide, and failed to occur in fibroblasts from TGF-beta1 knock-out mice. Co-transfection and overexpression with a Smad7 construct abrogated the increase in COL1A1 promoter activity observed in hypoxia. Upregulated transcriptional activity of the TGF-beta1 promoter in hypoxia was found to be maximal between - 453 and - 175 bp from the transcriptional start site. Since hypoxia is a critical feature of the early phases of wound repair, we conclude that it may act as a potent physiologic stimulus for collagen synthesis. TGF-beta1 appears to be a critical component of this response.