The Experimental Study Of The Proliferation And Osteogenic Differention Effects Of Zuogui Pill On Rat Mesenchymal Stem Cells

Objective:To investigate the influences of Zuogui Pill on the proliferation and differentiation of mesenchymal stem cells cultured in vitro.Methods:1.Whole marrow method was used to separate and culture rat mesenchymal stem cells,and then they were identified.2.The medicated serum concentration of Zuogui Pill(high,middle,low)was prepared,and MTT method was used to detect the effect of medicated serum on proliferation,it also was used to comprehend dose-effect relationship.3.The optimal medicated serum concentration of Zuogui Pill was used to induce the differentiation of mesenchymal stem cells into osteoblasts.Five groups were divided:the high concentration medicated serum of Zuogui Pill,the middle concentration medicated serum of Zuogui Pill,the low concentration medicated serum of Zuogui Pill,classic group and blank control group.4.Von-Kossa staining was used to study the effect of medicated serum of Zuogui Pill on osteoplastic induction.5.â… collagen was detected by immunohistochemistry and and gray analysis was done,which was used to demonstrate the effects of medicated serum of Zuogui Pill on osteoplastic induction.Results:1.Primary cultured MSCs were adhered to plastic surface within 24 hours and reached confluence within 12-15 days.To be conditioned induced,MSCs were suceessfully differentiated into osteoblast.2.Flow cytometry(FCM)analysis indicated that the third generation MSCs of the G₀/G₁ stage were 96.7%,G₂stage were1.2%,S stage were 2.1%and MSCs were positive for CD44,CD105,CD29and negative for CD34,CD45,CD19,which was consistent with characteristics of MSCs; 3.Compare with blank control group,the different concentration medicated serum of Zuogui Pill promoted the proliferation and showed a certain quantity-efficiency relationship(P＜0.05);4.The black mineraliztion node were positive in MSCs after osteogenic differentiation and Von-Kossa staining;5.Compare with blank control group,the different concentration medicated serum of Zuogui Pill improved the average gray expression of I ollagen(P＜0.05) except for the low concentration medicated serum of Zuogui Pill after 7 days' induction(P＞0.05).Conclusion:The different concentration medicated serum of Zuogui Pill can enhance the proliferation of MSCs,showed a certain quantity-efficiency relationship.Each concentration medicated serum could accelerate the proliferation and induce the differentiation to osteoblast of MSCs,so Zuogui Pill was used to prevent and cure osteoporosis.The mechanism of Zuogui Pill on preventing and curing osteoporosis may be related to accelerating the proliferation as well as inducing the osteodifferentiation by enhancing the ability of osteogenic differentiation.