Culture and identification of smooth muscle progenitor cells from rat bone marrow / 基础医学与临床

ABSTRACT

Objective To isolate and identify smooth muscle progenitor cells(SPCs) from rat bone marrow and to observe specific expression of smooth muscle progenitor cells during proliferation and differentiation in vitro.Methods MNCs were isolated by density gradient centrifugation from rat marrow and cultured in conditioned nutrient medium,identification was performed by immunofluorescent staining(?-SMA,CD14).Smooth muscle cells specific markers(?-SMA) were checked with Western blotting and Real-time PCR at different time.Results During culturing,cells adhered and became spindle shaped with outgrowth at 4 d and 7 d,and showed typical "peak" "valley" at 14 d.Both ?-SMA and CD14 were positive after 4 d.Expression of ?-SMA was not found at 1 d with Western blotting,but it gradually enhanced at 4 d and reached the peak from 10 d to 14 d,then maintaned high-level at 21 d.The results with Real-time PCR indicated that no expression of ?-SMA mRNA within non-induced cells was found,but after being induced it gradually enhanced at 4 d and got to peak at 14 d,then kept the high-level at 21 d,low-expression at 1 d was significantly different from the other ones(P