复方海蛇胶囊抗PC-12细胞凋亡研究

目的:研究复方海蛇胶囊对β-淀粉样蛋白(Aβ_1~42)诱导的PC-12细胞凋亡的影响,探讨复方海蛇胶囊应用于痴呆的治疗机制。方法:采用四氮唑蓝比色法(MTT法)确定复方海蛇胶囊对神经生长因子(NGF)诱导培养PC-12细胞的安全剂量后,加或不加Aβ,分别用MTT法、流式细胞术(FCM术)检测不同质量浓度的复方海蛇胶囊(0.01,0.1,1,5 mg.mL^-1)对PC-12细胞凋亡的影响;用蛋白免疫印迹(Western-blot)法分别检测凋亡执行酶caspase-9与caspase-3的活性。结果:复方海蛇胶囊对NGF诱导培养的PC-12细胞的安全剂量为小于10 mg.mL^-1。复方海蛇胶囊各剂量组对Aβ诱导的PC-12细胞的凋亡具有明显的抑制作用。各组数据(A)分别为:1.75±0.12,0.73±0.35,0.79±0.11,0.83±0.07,1.31±0.07和1.80±0.38,与空白组比较P<0.01,流式细胞检测中凋亡细胞的百分率降低(1.93±0.41)%,(46.17±4.08)%,(35.35±4.63)%,(28.62±3.81)%,(15.13±3.15)%,(7.84±1.76)%,与空白组比较P<0.01。另外,复方海蛇胶囊抑制了Aβ诱导PC-12细胞的caspase-9和caspase-3活性的增高。结论:复方海蛇胶囊可显著抑制Aβ诱导PC-12细胞的凋亡,其机制可能通过抑制凋亡执行酶caspase-9,caspase-3的活性实现。

Study of Fufang Haishe capsule against cell apoptosis

OBJECTIVE: To study mechanismt of Fufang Haishe capsule for dementia by observing the effect of it on PC-12 cell apoptosis, which was induced by beta-amyloid protein (Abl-42). METHOD: Nerve growth factor (NGF) was used to cultivate the PC-12 cells. Fufang Haishe capsule at different concentrations was added into the culture medium so as to identify the nontoxic concentrations with MTT. To analyze the PC-12 cell apoptosis respectively by MTT assay, Flow cytometry (FCM technique) with different concentrations of Fufang Haishe capsule (0.01, 0.1, 1, 5 mg x mL(-1)), adding Ab or not Western blot was used to detect apoptosis which was measured on the implementation of caspase-9 and caspase-3 activity. RESULT: Fufang Haishe capsule could significantly inhibit the apoptosis of PC-12 cells induced by Abeta with increased colorimetric MTT asay ( compare among the control group and concentration 0, 0.01, 0.1, 1 and 5 mg x mL(-1) group, which is the same below: 1.75 +/- 0.12, 0.73 +/- 0.35, 0.79 +/- 0.11, 0.83 +/- 0.07, 1.31 +/- 0.07, 1.80 +/- 0.38, P < 0.01) and the decreased apoptosis rate of the cells which was analysed by flow cytometry (1.93 +/- 0.41)%, (46.17 +/- 4.08)%, (35.35 +/- 4.63)%, (28.62 +/- 3.81)%, (15.13 +/- 3.15)%, (7.84 +/- 1.76)%, P < 0.01. In addition, Fufang Haishe capsule inhibited the activity of caspase-9 and caspase-3 of PC-12 cells which was induced by Abeta. CONCLUSION: Fufang Haishe capsule significantly inhibite apoptosis of PC-12 cells induced by Abeta. The mechanism might be that Fufang Haishe capsule decrease the activity of the apoptosis implementing protein,caspase-9 and caspase-3.