健脾活骨方不同提取部位对骨髓间充质干细胞增殖和分化的影响

摘要

Objective To observe effects of different extracts of Jianpi Huogu Formula (JPHGF) on proliferation and differentiation of bone marrow mesenchymal stem cell (BMSCs). Methods Whole bone marrow adherent was used to screen, culture, and isolate BMSCs. Extracts from different parts (water, chloroform, ethyl acetate and n-butanol parts) of JPHGF were administrated for a certain time. MTS was used to detent cell proliferation;ALP staining was used to detect ALP activity;ARS staining was used to detect the formation of calcium nodules;oil red O staining was used to detect fat cell formation. Results Extracts from different parts of JPHGF could promote cell proliferation of BMSCs in different levels, followed by its strength in water, chloroform, ethyl acetate, and n-butanol parts;ALP staining results showed that the intensity of ALP expression of the order is water, acetic acid ethyl, chloroform and n-butanol parts;in promoting the formation of calcium nodules, ARS staining results showed that its intensity were water, chloroform, ethyl acetate, and n-butanol parts;oil red O staining results showed that inhibition intensity of fat cells interaction strength was formed from ethyl acetate, water, chloroform to n-butanol parts. Conclusion Extracts from different parts of JPHGF have different effects on BMSCs proliferation and differentiation. Water extraction has the strongest osteogenic differentiation and proliferation, and ethyl acetate has the best effect on the inhibition of cell formation.%目的：观察健脾活骨方不同提取部位对体外培养大鼠骨髓间充质干细胞(BMSCs)增殖及成骨、成脂分化的影响,对其健脾作用的有效部位进行初步筛选。方法采用全骨髓贴壁筛选培养法分离BMSCs,给予健脾活骨方不同提取部位(水、氯仿、正丁醇和乙酸乙酯萃取部位)作用一定时间后,MTS法检测细胞增殖,碱性磷酸酶(ALP)染色检测ALP活性,茜素红(ARS)染色检测钙结节的形成,油红O染色检测脂肪细胞的形成。结果健脾活骨方不同提取部位均能不同程度促进 BMSCs 的增殖,其强度依次为水、氯仿、正丁醇和乙酸乙酯部位；ALP染色结果显示,促进ALP表达的强度依次为水、乙酸乙酯、氯仿和正丁醇部位；ARS染色结果显示,促进钙结节的形成作用强度依次为水、氯仿、正丁醇和乙酸乙酯部位；油红O染色结果显示,抑制脂肪细胞形成的作用强度依次为乙酸乙酯、水、氯仿和正丁醇部位。结论健脾活骨方不同提取部位对BMSCs增殖和分化的影响不同,水部位促进增殖和成骨分化作用最强,乙酸乙酯抑制脂肪细胞形成作用最佳。