针刺疗法对局灶性脑缺血大鼠神经功能、大脑皮层蛋白激酶A表达及细胞凋亡率的影响

目的:观察针刺疗法对局灶性脑缺血大鼠大脑皮层蛋白激酶A(protein kinase A,PKA)表达的影响,探讨针刺疗法对局灶性脑缺血大鼠的神经保护机制。方法:选取雄性SD大鼠,随机分为假手术组、模型组、针刺组,每组30只,每组大鼠再随机等分为3d、7d、14d组,共计9个亚组。采用线栓法制备左侧大脑中动脉阻塞2h再灌注模型。针刺组大鼠行针刺"百会"水沟"及右侧"曲池"合谷"内关"足三里"三阴交"太冲"穴,每日1次,每次30min。采用神经行为评分法评价大鼠神经功能缺损情况,采用流式细胞仪测定缺血侧皮层细胞凋亡率,采用免疫组织化学法测定缺血侧大脑皮层PKA阳性细胞表达率。结果:与假手术组各时相点比较,模型组大鼠神经功能严重受限,缺血侧皮层细胞凋亡率及PKA阳性细胞表达率明显升高(均P<0.05);针刺组各时相点的神经行为缺陷评分均低于模型组(P<0.05),缺血侧皮层细胞凋亡率明显低于模型组,PKA阳性细胞表达率明显高于模型组(均P<0.05)。结论:针刺能够降低局灶性脑缺血大鼠缺血侧大脑皮层的细胞凋亡率,提高大脑皮层PKA阳性细胞表达率,促进神经修复与再生,从而降低神经行为缺损评分,改善受损神经功能。

Influence of Acupuncture Intervention on Neurologic Deficits, Cerebrocortical Cell Apoptosis and Protein Kinase A Expression in Rats with Focal Cerebral Ischemia

Objective To observe the influence of acupuncture intervention on expression of protein kinase A(PKA) in the cerebrocortex of rats with focal cerebral ischemia-reperfusion injury(CI/RI),so as to explore its underlying neuroprotection mechanism on cerebral ischemia.Methods Ninety male SD rats were randomly and evenly divided into sham-operation(sham),model and acupuncture groups which were further randomized into 3,7 and 14-days(d) subgroups(10 rats/subgroup).CI/RI model was established by occlusion of the left middle cerebral artery for 2 hours and reperfusion.Manual acupuncture stimulation was applied to "Baihui"(GV 20) and "Shuigou"(GV 26),as well as the right "Quchi"(LI 11),"Hegu"(LI 4),"Neiguan"(PC 6),"Zusanli"(ST 36),"Sanyinjiao"(SP 6) and "Taichong"(LR 3) for 30 min,once daily for 3,7 and 14 d respectively.Rats of the sham and model groups were restrained for 30 min each day.Neurological defects were assessed by ethologic scoring according to Bederson's neurologic assessment scales.Cellular apoptosis in the ischemic cortex was detected by flow cytometry and PKA expression determined by immunohistochemistry for calculating its PKA immunoreaction(IR)-positive cell rate,respectively.Results Compared with the sham group,the neurologic scores,cortical cellular apoptosis rates and PKA IR-positive cell rates of the model group were significantly increased at the time-points of day 3,7 and 14 post-treatment(P<0.05).In comparison with the model group,the neurologic scores and cortical cellular apoptosis rates of the acupuncture group at the time-points of day 3,7 and 14 post-treatment were considerably down-regulated(P<0.05),and the cortical PKA IR-positive cell rates of the acupuncture group were remarkably increased(P<0.05).In addition,along with the increase of acupuncture treatment sessions,lower apoptosis rates and more PKA IR-positive cells were found,suggesting a cumulative effect.Conclusion Acupuncture intervention can lower cellular apoptosis rate of the ischemic cerebrocortex and up-regulate cortical PKA expression level in CI/RI rats,which may be responsible for its effect in improving neurologic deficits.