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目的:研究补阳还五汤含药血清对骨髓间充质干细胞(BMSCs)增殖及细胞周期的影响。方法:以6.5,13,26 g·kg~(-1)补阳还五汤和等量蒸馏水连续灌胃SD大鼠1周,制备3种补阳还五汤含药血清和空白血清。将BMSCs分为补阳还五汤6.5 g·kg~(-1)组(用含10%6.5 g·kg~(-1)生药制备的血清培养),补阳还五汤13 g·kg~(-1)组(用含10%13 g·kg~(-1)生药制备的血清培养),补阳还五汤26 g·kg~(-1)组(用含10%26 g·kg~(-1)生药制备的血清培养),空白组(用含10%空白血清培养)。采用噻唑蓝(MTT)比色法检测各组BMSCs的增殖活性,流式细胞术检测细胞周期,蛋白免疫印迹(Western blot)法检测磷脂酰肌醇-3羟基激酶(PI3K)和磷酸化丝(苏)氨酸蛋白酶(p-Akt)(Ser473)蛋白的表达。结果:13,26 g·kg~(-1)生药制备的补阳还五汤含药血清作用BMSCs 48,72 h后,细胞增殖活性较空白组明显升高(P<0.05);3种补阳还五汤含药血清作用48 h后,BMSCs在S期的细胞比率随着药物浓度的升高而升高,与空白组比较有统计学意义(P<0.05);13,26 g·kg~(-1)生药制备的补阳还五汤含药血清作用48 h后,PI3K和p-Akt(Ser473)蛋白表达较空白组显著增强(P<0.01)。结论:13,26 g·kg~(-1)生药制备的补阳还五汤含药血清可能通过活化PI3K/Akt信号通路促使细胞进入S期,增强BMSCs增殖活性。
Objective: To study the effects of Buyang Huanwu Decoction-containing serum on proliferation and cell cycle of bone marrow-derived mesenchymal stem cells (BMSCs). Methods: SD rats were fed with Bu Yang Huan Wu decoction of 6.5, 13, 26 g · kg ~ (-1) and distilled water of equal volume for 1 week. Three kinds of drug-containing serum and blank serum of Buyang Huanwu Decoction were prepared. BMSCs were divided into Buyang Huanwu Decoction 6.5g · kg -1 (containing 10% 6.5g · kg -1 serums prepared from crude drugs), Buyanghuanwu Decoction 13g · kg ~ (-1) group (serum containing 10% 13 g · kg -1 crude drug) and Buyang Huanwu Decoction 26 g · kg -1 (containing 10% 26 g · kg -1 ~ (-1) crude drug serum preparation), blank group (containing 10% blank serum culture). The proliferative activity of BMSCs in each group was detected by MTT colorimetric assay. The cell cycle was detected by flow cytometry. The expressions of PI3K and phosphorylated phosphatidylinositol 3 - kinase (PI3K) were detected by Western blot. Threonine) cathepsin (p-Akt) (Ser473) protein expression. Results: After 48 h and 72 h, the proliferation of BMSCs was significantly increased (P <0.05) compared with the blank group (P <0.05) The cell ratio of BMSCs in S phase increased with the increase of drug concentration after 48 h of treatment with YangHuwuWu containing serum, which was statistically significant compared with the blank group (P <0.05); 13,26 g · kg The serum level of PI3K and p-Akt (Ser473) in BUHH treated with ~ (-1) crude drug for 48 h increased significantly (P <0.01). CONCLUSION: The serum of Buyang Huanwu Decoction prepared with 13,26 g · kg -1 crude drug may promote the proliferation of BMSCs by activating PI3K / Akt signaling pathway.