Comparison Of Chondrogenic Potential Between Mesenchymal Stem Cells From Rabbits' Bone Marrow And Adipose Tissue

Objective:To compare the chondrogenic potential of rabbit bone marrow-derived mesenchymal stem cells (BMSCs) and adipose tissue-derived stem cells (ADSCs) in vitro.Methods:ADSCs were isolated from the subcutaneous adipose tissue which was from 6-month-old New Zealand white rabbits'abdomen by mechanical digestion and enzyme digestion, and then cultured and amplified in vitro. BMSCs were obtained from femurs of healthy 6-month-old New Zealand white rabbits, which isolated by whole bone marrow culture, and then purified in vitro. The morphology and proliferation of both ADSCs and BMSCs were observed using inverted phase contrast microscope in the whole time. The generation time of the primary,2nd and 3rd passage cells was compared between ADSCs and BMSCs.Both ADSCs and BMSCs at passage 3 were used for drawing growth curve and observing doubling time,and then the growth curve and doubling time of ADSCs and BMSCs were compared separately.The 3nd passage of ADSCs and BMSCs were used for induction of chondrogenesis. The experiment was divided into four groups:ADSCs cultured under condition of chondrogenic media(A1), ADSCs cultured in normal DMEM media (A2), BMSCs cultured under condition of chondrogenic media (B1), BMSCs cultured in normal DMEM media (B2). A1 and B1 were experimental groups,while A2 and B2 were control groups.The morphology and proliferation of all groups were observed using inverted phase contrast microscope.After cultured for 14 days, the experimental groups were detected by collagen type II immunohistochemistry staining and toluidine blue staining of proteoglycan,and the control groups were detected by collagen type II immunohistochemistry staining.The chondrogenic potential of A1 and B1 were compared by the method of gray value of immunocytochemical staining on collagen II.Results:Primary cells of BMSCs showed aggregative growth,while ADSCs were diffuse. But since the 2nd passage, BMSCs showed diffuse growth just like ADSCs. Both BMSCs and ADSCs exhibited a fibroblast-like phenotype (flat, long-spindle) in monolayer cultures, without any obvious variation in cell morphology. The generation time of primary ADSCs was 6 days,which was shorter than that of primary BMSCs(10 days). Also, the generation time of 2nd and 3rd passage of ADSCs (3 days) was shorter than that of BMSCs (5days).Both BMSCs and ADSCs proliferated fast and stably, but the proliferation speed of ADSCs at passages 3 was obviously faster than BMSCs according to the growth curve. The doubling time of ADSCs was 26h,which was less than BMSCs (36h). When cells were cultured under condition of chondrogenic media, the proliferation of the experimental groups'cells became slow,and the morphology of which changed from long-spindle to round gradually. The morphology and proliferation of the control groups were just like before.After cultured for 14 days, A2 and B2 showed negtive results in collagen typeâ…¡immunohistochemistry staining.Both A1 and B1 presented positive,while detected by collagen typeâ…¡immunohistochemistry staining,and the same results were detected by toluidine blue staining of proteoglycan. But the expression of collagen type II in B1 was more than in Al according to the results of the gray value of immunocytochemical staining on collagen type II (p<0.05).Conclusion:The morphology of BMSCs and ADSCs had no obvious variation. The proliferative ability of ADSCs was better than BMSCs. Both of them could express chondrogenic phenotype while they were cultured under condition of chondrogenic media,but BMSCs seemed to be more potential.