Effects of in vitro cultured adipose-derived stem cells of rats on the tympanic membrane fibroblasts

Abstract

To explore the effects of rats adipose-derived stem cell (ASC) on the proliferation and migration of fibroblasts from the tympanic membrane, and to discuss the possibility and significance of therapy with ASC for tympanic membrane (TM) healing and regeneration. Wistar rats were sacrificed, and then the isolation, culture and identification of both ASC and the TM fibroblasts were performed respectively. To verify the effect of ASC on fibroblasts proliferation, transwell co-culture system was used. To examine the effect of ASC on fibroblasts migration, cell migration assay with transwell was also applied. All the data were analyzed under a confocal laser scan microscopy system. Immunofluorescence of cell surface markers indicated that rats ASC were positive for both of CD44 and CD29, but negative for CD34. The rat TM fibroblasts were positive for vimentin. The fibroblasts co-cultured with ASC proliferated faster than the fibroblasts of control group, and the difference of the cell counting number between the two groups was significant (t = 6.75, P = 0.003). Compared with the control group, the fibroblasts cultured with ASC conditioned culture medium migrated significantly faster, and the space between the fibroblasts and the polycarbonate membrane pore was significantly shortened at different time point (1, 2.5 and 4 h, P < 0.05). The cell number of the fibroblasts that had migrated through the polycarbonate membrane had been significantly increased 5 h after migration (P < 0.05). ASC might promote TM fibroblasts proliferation and migration by paracrine activation, and it will facilitate the regeneration of TM fibrous layer.