Role of Hydrogen Sulfide in the Survival of Fibroblasts and Fibroblast-mediated Contraction of Collagen Gel

ABSTRACT

PURPOSE: To investigate the role of hydrogen sulfide in the survival and collagen gel contraction of cultured human Tenon's capsule fibroblasts (HTCFs). METHODS: Primarily cultured HTCFs were exposed to 0, 100, 200, or 300 µM hydrogen sulfide (sodium hydrogen sulfide, NaHS) for 2 days. Cellular survival was assessed by MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) assay. Degree of apoptosis was assessed with flow cytometry using annexin-V/propidium iodide double staining. To evaluate the effect of NaHS on cellular transdifferentiation, HTCFs were stimulated with 5 ng/mL TGF-β1 and the level of expression of α-smooth muscle actin (SMA) mRNA was assessed using reverse-transcription polymerase chain reaction. The cells were embedded in collagen gel, and the amount of gel contraction was measured. RESULTS: NaHS at 300 µM reduced HTCF survival (p = 0.013); NaHS at both 200 and 300 µM increased apoptosis in a dose-dependent manner (p = 0.013 and p = 0.016). TGF-β1 increased the expression of α-SMA mRNA (p = 0.041); co-treatment with 100 µM NaHS decreased TGF-β1-induced α-SMA mRNA expression (p = 0.039) and inhibited collagen gel contraction. CONCLUSIONS: NaHS at high concentration reduced cellular survival and increased HTCF apoptosis. NaHS decreased TGF-β 1-induced increases in α-SMA mRNA expression and collagen gel contraction. Thus, hydrogen sulfide may suppress scar formation by inhibiting HTCF transdifferentiation and contraction of collagen gels.