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[摘要]
目的 比较瓜蒌薤白半夏汤(GXB)不同提取部位对缺血缺氧心肌细胞的保护作用,初步探寻该经典方剂干预胸痹、心痛的物质基础。方法 称取瓜蒌240 g、薤白90 g、半夏120 g,50%乙醇回流提取,分别用石油醚、氯仿、醋酸乙酯、水饱和正丁醇、水5种不同极性溶剂萃取,得到相应提取部位,及药渣水煎液部位。培养大鼠原代心肌细胞,制备缺血缺氧模型,给予系列浓度(100、10、1、0.1、0.01 mg/mL)的GXB 6种提取部位及瓜蒌皮注射液(阳性对照,100、50、25、5、1 mg/mL)。MTT法检测细胞存活率,倒置显微镜观察细胞形态变化,生化试剂盒检测培养液中肌酸激酶(CK)、肌酸激酶同工酶(CK-MB)活性。结果 与对照组比较,模型组细胞凋亡、坏死明显,细胞存活率显著降低,细胞培养液中CK、CK-MB的活性显著升高(P < 0.05);质量浓度均为0.1 mg/mL的GXB水饱和正丁醇部位、水溶性部位及药渣水煎部位可以显著逆转缺血缺氧心肌细胞的生长状态、存活率以及培养液中CK、CK-MB活性(P < 0.05);石油醚、氯仿及醋酸乙酯部位处理的细胞,与模型组无明显差异。结论 GXB的水饱和正丁醇、水溶性及药渣水煎部位对缺血缺氧损伤的心肌细胞具有显著的保护作用。
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[Abstract]
objective To compare the protective effect of each part of Gualou-xiebai-banxia Decoction on cardiomyocytes induced by ischemia-hypoxia, and initially explore the material basis of this classical prescriptions intervention on thoracic obstruction and heartache. Methods Weighting 240 g trichosanthes kirilowii, 90 g allium macrostemon and 120 g pinellia ternata for preparation. After 50% ethanol reflux extraction, five different polar solvents including petroleum ether, chloroform, ethyl acetate, n-butanol, and water were used for extraction to obtain their extracts from different sites and decocting part of drug residue. Primary rat myocardial cells were cultured to establish ischemic anoxia model, and a series of concentrations (100, 10, 1, 0.1 and 0.01 mg/mL) of six GXB extracts and gualoupi injection (positive control, 100, 50, 25, 5, 1 mg/mL) were added for treatment. The survival rate of cells was detected by MTT and the morphological changes were observed under inverted microscope, biochemical reagent kit was used to test creatine kinase (CK) and reatine kinase isoenzyme (CK-MB) activity in the liquid culture. Results Compared with control group, the cells in the model group showed apoptosis and necrosis, cell viability significantly decreased, and the activity of CK and CK-MB in cell culture medium increased significantly (P < 0.05). The treatment of water saturated n-butanol site, water soluble site and decocting site of drug residue of 0.1 mg/mL can significantly reverse the growth state, survival rate and myocardial enzyme activity of ischemic and anoxic cardiomyocytes (P < 0.05). However, the cells treated with petroleum ether, chloroform and ethyl acetate had no significant difference with the hypoxia ischemia group. Conclusion The water saturated n-butanol site, water soluble site and decocting site of drug residue of GXB have significant protective effects on myocardial cells injured by ischemia and hypoxia.
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[基金项目]
国家自然科学基金项目(81173164、81573852);山东省教育厅高等学校科研项目(J10LF54)