【作者】 张晓忠；

【导师】 邹伟；

【作者基本信息】 黑龙江中医药大学 ， 中西医结合临床， 2011， 博士

【摘要】 目的：研究“百会”透“曲鬓”针法对脑出血大鼠脑组织MMP-2、NGB、CytC和VCAM-1蛋白及MMP-2、NGBmRNA基因表达的影响,探讨其对脑组织的神经保护作用及可能机制,为临床应用提供实验依据。方法：选取健康雄性SD大鼠,先随机分为三个组：假手术组、模型组和针刺组,各组又分为术后6h、1d、2d、3d、7d五个时间点,每个时相点10只大鼠。自体血脑内注射制备脑出血动物模型,然后针刺动物模型百会穴透曲鬓穴,贯穿模型动物顶额颞三区,采用多个穴位一针透刺疗法进行治疗。采用Berderson神经体征评分法观察神经行为学变化；干湿比重法进行脑组织含水量测定；光镜观察脑组织病理形态学变化；采用免疫组化法检测脑组织中MMP-2、NGB、CytC和VCAM-1蛋白表达；采用原位杂交法检测脑组织MMP-2、NGBmRNA基因表达。结果：1、光镜下脑组织病理改变结果针刺组较模型组改变显著。2、针刺组各时间点大鼠神经功能缺损体征有明显恢复,与模型组比较,1d,2d,3d,7d评分差异明显,有统计学意义,P<0.05或P<0.01。3.针刺组各时点脑水含量均小于模型组相应时间点,与模型组比较,其中1d,2d,3d,7d有统计学意义,P<0.05或P<0.01。4.模型组各时间点CytC蛋白表达随时间增长而升高,1d时达高峰,随后逐渐下降,7d时仍高于假手术组,与假手术比较有统计学意义,P<0.01。针刺治疗后,各时间点大鼠脑组织中CytC蛋白表达明显降低,与同时间点模型组比较,有统计学意义,P<0.01。5.模型组各时间点VCAM-1蛋白表达随时间增长而增强,2d时达高峰,随后逐渐下降,7d时仍高于假手术组,与假手术比较有统计学意义,P<0.01。针刺治疗后,各时间点大鼠脑组织中VCAM-1蛋白表达明显降低,与同时间点模型组比较,有统计学意义,P<0.01。6.模型组各时间点MMP-2蛋白和MMP-2 mRNA基因表达随时间增长而增强,1d时达高峰,随后逐渐下降,7d时仍高于假手术组,与假手术比较有统计学意义,P<0.01。针刺治疗后,各时间点大鼠脑组织中MMP-2蛋白和MMP-2mRNA基因表达明显降低,与同时间点模型组比较,有统计学意义,P<0.01。7.模型组各时间点NGB蛋白和NGBmRNA基因表达随时间增长而增强,1d时达高峰,随后逐渐下降,7d时仍高于假手术组,与假手术比较有统计学意义, P<0.01。针刺治疗后,各时间点大鼠脑组织中NGB蛋白和NGBmRNA基因表达明显升高,与同时间点模型组比较,有统计学意义,P<0.01。结论：1.“百会”透“曲鬓”针法能够显著降低脑组织含水量,减轻脑组织水肿,改善脑组织病理学损伤状态,从而改善了大鼠脑出血不同时间的神经功能损伤症状。2.“百会”透“曲鬓”针法能够抑制脑出血大鼠脑组织CytC蛋白表达,减少神经细胞的凋亡,从而起到神经元保护作用。3.“百会”透“曲鬓”针法能够抑制脑出血大鼠脑组织VCAM-1蛋白表达,保证脑血管内皮细胞的完整性,减轻炎症级联反应所引起的脑水肿。4.“百会”透“曲鬓”针法能够抑制脑出血大鼠脑组织MMP-2蛋白和MMP-2 mRNA基因表达,从而减轻脑出血对基底膜及细胞外基质的降解。5.“百会”透“曲鬓”针法能够提高脑出血大鼠脑组织NGB蛋白和NGBmRNA基因表达,改善脑组织缺血、缺氧状态,从而对神经细胞起到保护作用。更多还原

【Abstract】 Objective:Observe the expression of MMP-2、NGB、CytC and VCAM-1 protein inintracerebral hemorrhage in rat brain through "Baihui" Acupuncture "QuBin" to explore the brain tissue Neuroprotective effects and possible mechanisms to provide experimental basis for clinical application.Methods:SD male rats were first randomly divided into three groups:sham operation groups, model groups and the acupuncture groups, each group was divided after 6h, 1d,2d,3d, 7d, every point included 10 rats. Preparation of autologous blood injection ICH model through the use of acupuncture Baihui QuBin hole in the head through the top three frontal and temporal area,using a needle through the multi-point method of treatment.Using Berderson to estimate neurological signs of nerve behavior change; wet and dry weight method for determination of brain tissue water content; observing pathological changes in brain tissue; MMP-2, NGB, CytC and VCAM-1 protein expression were detected by immunohistochemistry in brain tissue; by in situ hybridization detection of brain tissue MMP-2, NGBmRNA gene expression.Results:1.Under the light microscope, in the acupuncture group,the results of brain tissue pathological changes compared with model group changed significantly.2.The acupuncture group at each time point signs of neurological deficit in rats significantly restored, compared with the model group, 1d,2d,3d,7d had significant differences, with statistical significance, P<0.05 or P<0.01.3. Acupuncture group at each time point brain water content was less than model group at corresponding time points, compared with model group, 1d,2d,3d,7d was statistically significant, P<0.05 or P<0.01.4. Model group at different time points CytC protein expression increased over time, 1d reached the peak, then decreased gradually,7d, remains higher than the sham group with sham operation was statistically significant, P<0.01. Acupuncture treatment, at each time point CytC rat brain protein expression was significantly reduced, and compared the model group at the same time, was statistically significant, P<0.01.5. Model group at different time points the expression of VCAM-1 protein enhanced the growth over time,2d reached the peak, then decreased gradually,7d, remains higher than the sham group with sham operation was statistically significant, P<0.01. Acupuncture treatment, at each time point in rat brain expression of VCAM-1 protein was significantly reduced, and the same time the model group, was statistically significant, P<0.01.6. Model group at each time point MMP-2 protein and MMP-2 mRNA expression increased, 1d reached the peak, then decreased gradually,7d, remains higher than the sham group, compared with the sham operation was statistically significant, P<0.01. Acupuncture treatment, at each time point in rat brain tissue MMP-2 protein and gene expression of MMP-2mRNA significantly lower, with the same time the model group, was statistically significant, P<0.01.7. Model group at different time points and NGBmRNA NGB protein expression increased, Id reached the peak, then decreased gradually,7d, remains higher than the sham group with sham operation was statistically significant, P<0.01. Acupuncture treatment, at each time point in the NGB protein in rat brain gene expression and NGBmRNA significantly increased, at the same time the model group, was statistically significant, P<0.01. Conclusion:1. "Baihui" through "QuBin" acupuncture can significantly reduce brain water content, reduce the brain edema and improve the state of brain tissue pathological damage, thereby improving cerebral hemorrhage in different time symptoms of neurological damage.2."Baihui" through "QuBin" Acupuncture can inhibit CytC during intracerebral hemorrhage in rat brain, reduce neuronal apoptosis.3."Baihui" through "QuBin" Acupuncture can inhibit intracerebral hemorrhage in rat brain expression of VCAM-1, to ensure the integrity of brain vascular endothelial cells, reducing the inflammatory cascade caused by cerebral edema.4."Baihui" through "QuBin" Acupuncture can inhibit intracerebral hemorrhage in rat brain MMP-2 protein and MMP-2 mRNA gene expression, thereby reducing cerebral hemorrhage degradation on the basement membrane and extracellular matrix5."Baihui" through "QuBin" Acupuncture can improve the rat brain hemorrhage NGBmRNA NGB protein and gene expression, to improve cerebral ischemia, hypoxia, and thus play a protective role in nerve cells.更多还原