Development of a chemically defined in vitro culture system to effectively stimulate the proliferation of adult human dermal fibroblasts

Despite the fact that dermal fibroblasts are a practical model for research related to cell physiology and cell therapy, an invitro culture system excluding serum, which complicates standardization and specificity and induces variability and unwanted effects, does not exist. We tried to establish a CDCS that supports effective proliferation of aHDFs. KDMEM supplemented with 5% (v/v) KSR, 12ng/ml bFGF, 5ng/ml EGF and 1g/ml hydrocortisone supported sufficient proliferation of aHDFs for 1week. However, aHDF proliferation was decreased greatly after subculture. This problem could be overcome by culturing aHDFs in CDCM in culture plates coated with 10g/ml FN. Long-term culture of aHDFs was achieved using CDCM and FN-coated culture plates for 7weeks. The optimized CDCS increased the proliferation of aHDFs significantly, without any increase in the senescence rate or alteration in morphology of aHDFs, despite long-term culture. In conclusion, we established a CDCS that improved proliferation of aHDFs while inhibiting cellular senescence. The CDCS will contribute to advances in various future research related to clinical skin regeneration.