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目的研究清心养神方药对应激大鼠心肌细胞凋亡的影响。方法采用高效液相检测血清儿茶酚胺;采用TUNEL、DNA Landder、流式细胞仪等方法,检测心肌细胞凋亡。整体实验设清心养神方药组、应激空白对照组、心得安组和朱砂安神丸组,离体实验设清心养神方药组、应激空白对照组和心得安组。结果血浆去甲肾上腺素(NE)在应激4周时升高(P<0.05),肾上腺素、多巴胺在应激1、4周时升高(P<0.05);清心养神组、心得安组和朱砂安神丸组均使束缚应激大鼠升高的血浆儿茶酚胺水平降低。而2周时,清心养神组和朱砂安神丸组NE、多巴胺水平较应激组升高。束缚应激1、2、4周后,TUNEL染色结果:清心养神组、心得安组和朱砂安神丸组平均每视野心肌细胞凋亡个数,均较应激组明显减少(P<0.05)。流式细胞仪检测结果:去甲肾上腺素诱导心肌细胞在12、24、48h时无论早期凋亡还是晚期凋亡,均明显高于正常心肌细胞组;在24、48h时清心养神组、心得安组细胞凋亡率降低,与应激组比较有统计学意义(P<0.05);并且清心养神方药对早期凋亡率的降低较心得安组显著(P<0.05)。DNA Landder检测整体实验中各组未见到凋亡特征性条带。结论清心养神方药对应激大鼠血浆儿茶酚胺水平具有双相调节作用,且具有抗应激诱发的心肌细胞凋亡作用。
Objective To study the effect of Qingxin Nourishing Prescription on cardiomyocyte apoptosis in stress rats. Methods Serum catecholamines were detected by HPLC. TUNEL, DNA Lander, flow cytometry and other methods were used to detect cardiomyocyte apoptosis. The whole experiment was set up Qingxin Yangshen prescription group, stress control group, propranolol group and Zhusha Anshen pill group. In vitro experiments were set up Qingxin Yangshen prescription group, stress control group and propranolol group. Results Plasma norepinephrine (NE) increased at 4 weeks of stress (P<0.05), epinephrine, and dopamine increased at 1 and 4 weeks of stress (P<0.05); Qingxin Shenxin group, propranolol group, and Zhusha Anshen Pill Both groups reduced the elevated plasma catecholamine levels in restraint-stressed rats. At 2 weeks, the levels of NE and dopamine in the Qingxin Yangshen group and Zhusha Anshen Pill group were higher than those in the stress group. After 1, 2, and 4 weeks of restraint stress, TUNEL staining results showed that the number of myocardial cell apoptosis in each field of the Qingxin Shenshen group, the propranolol group, and the Zhusha Anshen Pill group was significantly lower than that of the stress group (P<0.05). Flow cytometry results: Norepinephrine-induced cardiomyocytes were significantly higher than normal myocardium group at 12, 24, and 48h, regardless of early apoptosis or late apoptosis. At 24 and 48 hours, the heart and mind rejuvenation group, propranolol The apoptosis rate of the cells in the group was significantly lower than that in the stressed group (P<0.05), and the decrease of the early apoptotic rate in the Qing Xin Yang Shen Recipe was significant (P<0.05). Apoptotic characteristic bands were not seen in the DNA Landder assay in the whole experiment. Conclusion Qingxin Nourishing Decoction has biphasic regulation on plasma catecholamine levels in stress rats, and has an anti-stress-induced cardiomyocyte apoptosis.