Abstract
The effects of various concentrations of serum, ascorbic acid, and sodium diphenylhydantoin (DPH) in culture medium on cell proliferation and collagen fibrogenesis were morphologically investigated in rat skin fibroblasts.
The cell proliferative rate appeared to be dependent upon the serum concentration both in αMEM and DM-160 medium. But, the rate in αMEM medium was much higher than that in DM-160 medium. Fibroblasts in αMEM medium produced extracellular fibers stained positively by silver impregnation as early as 1 day after planting and accumulated collagen fibers in the intercellular matrix continuously throughout their logarithmic growth. This fibrogenesis was not affected by the serum concentration in the culture medium. The degree of fibrogenesis in DM-160 medium was less up to 3 weeks, and the amount of collagen in the cell layer increased thereafter.
Fibroblasts grown in DM-160 medium containing less than 1μg/ml ascorbic acid showed evidence of ascorbic acid deficiency when compared with cells grown in αMEM medium containing 50μg/ml ascorbic acid. Ultrastructural study demonstrated that ascorbic acid deficiency prevented the deposition of typical 64 nm-banded collagen fibrils; instead, finer, unbanded fibrils and microfibrils were deposited extracellularly. These results indicate that the fibroblasts require ascorbic acid to produce collagen fibers.
Addition of DPH to cultured fibroblasts resulted in stimulation of their proliferation and enhancement of collagen fibrogenesis.
