Abstract: Objective To observe the vascular network formation by co-culturing rat bone marrow mesenchymal stem cells(BMSCs)and endothelial cells(ECs)under conditions of two-dimensional co-culturing with/without serum,which would provide a minimum basal co-culture condition for the further study of the interactions between the cells.M ethods Rat BMSCs were isolated by direct adherence of the whole bonemarrow,and were identified by cellmorphology,cell surface antigens and three-lineage differentiation.Low-glucose DMEM with/without 10%FBSwas used during the co-culture of rat BMSCs and ECs,followed by a consecutive observation of the vascular network formation.CM-Dil solely labeled rat BMSCs or ECs were then co-cultured to verify the cell type within the network.Finally,immunofluorescent staining of EC marker CD31 and smoothmuscle cellmarker SM22αand Calponin was used to identify the cell differentiation during the co-culture. Resu lts Whole marrow direct adherence was used to isolate rat BMSCs successfully.With medium containing 10%FBS, co-cultured cells assembled vascular network in 8 days,which was formed together by rat BMSCs and ECs.In serum-free medium,however,self-assembled vascular network was formed together by co-cultured cells within 24 hours.In both settings,CD31⁺cells and SM22αor Calponin weremostly seen in the network structure.Conclusion Rat BMSCs and ECscan be co-cultured on two-dimensional plateswith/without serum-containingmedium for the induction of the self-assembled vascular network.

Key words: Bonemarrow mesenchymal stem cell, Endothelial cell, Co-culture, Vascular network