Objective   To establish a co-culture system between mesenchymal stem cells (MSCs) and smooth muscle cells (SMCs) in vitro

and study the influence of SMCs on the differentiation of bone marrow MSCs (BMSCs) into SMCs in the co-culture system.    Methods   Density gradient centrifugation was used to separate and culture BMSCs

and collagenase digestion was employed to separate and culture bladder smooth muscle cells(BSMCs). Then the 2 types of cells were identified respectively. Then the cells were divided into 4 groups

BMSCs culture group (positive control)

BSMCs cells culture group (negative control)

BMSCs and BSMCs co-culture groups with contact or non-contact. The expression of α-smooth muscle actin (α-SMA) and calponin were detected by immunofluorescence staining. Western blot analysis was employed to determine protein expression of calponin.    Results   In contact co-culture group

the positive rate of CD90 abd CD44 were 99.78% and 60.27% respectively in BMSCs

while only 0.21% for CD45. Western blotting showed that BMSCs expressed little calponin

but those contacting with BSMCs had an increasingly stronger expression of calponin with the elapse of contact culture time

while there was no significant change in its expression in BMSCs with contact. Immunofluorescence staining indicated that part of BMSCs were calponin positive at the 8th day after culture

but no positive cell was observed in non-contact culture group and BMSCs culture group.    Conclusion   The contact between BSMCs and BMSCs can promote the differentiation of BMSCs into SMCs.