左归饮对D-半乳糖致亚急性衰老模型大鼠肝肾功能及尿液代谢组学的影响
发布时间：2018-07-22 17:33
【摘要】：目的:以补益肾阴名方左归饮为研究对象,从中医临床功效出发,采用现代药理学研究方法,探讨左归饮对衰老模型大鼠肝肾器官功能的影响,并对其作用机制进行初步研究,为左归饮延缓机体衰老的理论提供更多实验依据,同时也为左归饮临床合理用药提供科学指导。方法:将实验大鼠分正常对照组、模型组、给药组(维生素E组、六味地黄丸组、左归饮高、低剂量组),采用D-半乳糖颈部皮下注射方法复制亚急性衰老大鼠模型,造模同时,各给药组灌胃给予相应药物进行干预,连续给药8周后,进行以下实验研究:(1)采用行为学实验方法(水迷宫和跳台实验)对衰老模型大鼠进行评价,考察左归饮对衰老模型大鼠学习记忆能力的影响;(2)通过对衰老模型大鼠肝肾功能指标及其组织中自由基进行检测,考察左归饮对衰老模型大鼠肝肾功能及自由基代谢水平的影响;(3)采用病理学实验方法对衰老模型大鼠肝肾组织病理进行研究,考察左归饮对衰老模型大鼠肝肾组织细胞的形态结构和病理变化的影响;(4)采用免疫组化方法对肝肾组织中细胞凋亡相关蛋白Bax、Bcl-2、Caspase-3进行研究,考察左归饮对衰老模型大鼠肝肾组织细胞凋亡相关蛋白表达水平的影响;(5)采用代谢组学研究方法,通过LC-MS技术对衰老模型大鼠尿液进行分析,比较左归饮对衰老模型大鼠尿液中内源性生物标记物的调节作用,考察左归饮对衰老模型大鼠尿液代谢组学的影响。结果:(1)在大鼠行为学评价中,左归饮能缩短衰老模型大鼠在水迷宫、跳台实验的潜伏时间,增加水迷宫实验穿越隐藏平台次数,减少跳台实验错误次数;(2)在生化指标测试中,除ALT外,左归饮能够降低衰老模型大鼠血清中AST、ALP、CREA、BUN指标水平,提高衰老模型大鼠肝肾组织中SOD和T-AOC活性,降低MDA含量;(3)肝肾组织形态的病理学实验结果表明,左归饮在一定程度上能够减轻衰老模型大鼠肝肾组织细胞的病理损伤,保持组织细胞形态结构的完整性;(4)肝肾组织免疫组化实验结果表明,左归饮能够增强衰老模型大鼠肝肾组织细胞中Bcl-2的阳性表达,降低肝肾组织细胞中Bax、Caspase-3的阳性表达;(5)大鼠尿液代谢组学分析结果共鉴定出29个生物标记物,找出9个相关代谢通路,结果表明,左归饮能够使衰老模型大鼠尿液代谢轮廓趋于正常,对23个生物标记物有回调作用,且对7个相关代谢通路产生影响。结论:左归饮能够通过提高衰老模型大鼠学习记忆能力,改善衰老模型大鼠肝肾功能,增强对衰老模型大鼠体内自由基的代谢及清除能力,保护肝肾组织细胞免于损伤,发挥延缓衰老作用。其作用可能通过影响肝肾组织细胞中相关凋亡蛋白的表达,调整衰老模型大鼠机体的异常代谢等机制起到延缓机体衰老作用。
[Abstract]:Objective: to study the effect of Zuo Gui Yin, a famous prescription of tonifying kidney yin, on the function of liver and kidney organs in aging model rats. It provides more experimental basis for the theory of Zuo Gui Yin to delay the aging of organism, and also provides scientific guidance for the rational use of Zuo Gui Yin in clinic. Methods: the experimental rats were divided into normal control group, model group, administration group (vitamin E group, Liuwei Dihuang pill group, Zuo Guiyin high dose group, low dose group). Subacute aging rat model was induced by subcutaneous injection of D-galactose. After 8 weeks of continuous administration, the following experimental studies were carried out: (1) the aging model rats were evaluated by behavioral experiment (water maze and jumper test). To investigate the effect of Zuoguiyin on learning and memory ability of aging model rats. (2) to detect the indexes of liver and kidney function and free radicals in tissues of aging model rats. To investigate the effects of ZuoGuiyin on liver and kidney function and free radical metabolism in aging model rats. (3) to study the pathological changes of liver and kidney tissues in aging model rats by pathological experiment. To investigate the effect of Zuoguiyin on the morphology and pathological changes of liver and kidney cells in aging model rats. (4) to study the apoptosis related protein Baxia Bcl-2Caspase-3 in liver and kidney by immunohistochemical method. To investigate the effect of Zuoguiyin on the expression of apoptosis-related protein in liver and kidney tissues of aging model rats. (5) to analyze the urine of aging model rats by LC-MS with the method of metabolomics. To compare the effects of Zuoguiyin on endogenous biomarkers in urine of aging model rats, and to investigate the effect of Zuogui Yin on urine metabolism of aging model rats. Results: (1) in the behavioral evaluation of rats, Zuo Gui Yin can shorten the latency time in water maze and platform jumping experiment, increase the number of water maze experiment traversing hidden platform, reduce the number of errors in platform jumping experiment; (2) in biochemical index test, In addition to alt, Zuoguiyin could decrease the serum level of ASTPPA-CREAEABUN, increase the activities of SOD and T-AOC and decrease the content of MDA in the liver and kidney of aging model rats. (3) the pathological results of liver and kidney morphology showed that Zuo Gui Yin can alleviate the pathological injury of liver and kidney cells in aging model rats to some extent and maintain the integrity of histocyte morphology. (4) Immunohistochemical experiment of liver and kidney tissue showed that, ZuoGuiyin could enhance the expression of Bcl-2 and decrease the positive expression of Baxfen Caspase-3 in the liver and kidney of aging model rats. (5) 29 biomarkers were identified by the results of urine metabolism in rats. Nine related metabolic pathways were identified. The results showed that Zuoguiyin could make the metabolic profile of urine of aging model rats normal, have a callback effect on 23 biomarkers, and have an effect on 7 related metabolic pathways. Conclusion: ZuoGuiyin can improve the learning and memory ability of aging model rats, improve the liver and kidney function of aging model rats, enhance the metabolism and scavenging ability of free radicals in aging model rats, and protect liver and kidney tissue cells from injury. Play a role in delaying senescence. Its effect may delay senescence by affecting the expression of apoptosis protein in liver and kidney tissues and regulating the abnormal metabolism of aging model rats.
【学位授予单位】：黑龙江中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5;R-332
本文编号：2138225
[Abstract]:Objective: to study the effect of Zuo Gui Yin, a famous prescription of tonifying kidney yin, on the function of liver and kidney organs in aging model rats. It provides more experimental basis for the theory of Zuo Gui Yin to delay the aging of organism, and also provides scientific guidance for the rational use of Zuo Gui Yin in clinic. Methods: the experimental rats were divided into normal control group, model group, administration group (vitamin E group, Liuwei Dihuang pill group, Zuo Guiyin high dose group, low dose group). Subacute aging rat model was induced by subcutaneous injection of D-galactose. After 8 weeks of continuous administration, the following experimental studies were carried out: (1) the aging model rats were evaluated by behavioral experiment (water maze and jumper test). To investigate the effect of Zuoguiyin on learning and memory ability of aging model rats. (2) to detect the indexes of liver and kidney function and free radicals in tissues of aging model rats. To investigate the effects of ZuoGuiyin on liver and kidney function and free radical metabolism in aging model rats. (3) to study the pathological changes of liver and kidney tissues in aging model rats by pathological experiment. To investigate the effect of Zuoguiyin on the morphology and pathological changes of liver and kidney cells in aging model rats. (4) to study the apoptosis related protein Baxia Bcl-2Caspase-3 in liver and kidney by immunohistochemical method. To investigate the effect of Zuoguiyin on the expression of apoptosis-related protein in liver and kidney tissues of aging model rats. (5) to analyze the urine of aging model rats by LC-MS with the method of metabolomics. To compare the effects of Zuoguiyin on endogenous biomarkers in urine of aging model rats, and to investigate the effect of Zuogui Yin on urine metabolism of aging model rats. Results: (1) in the behavioral evaluation of rats, Zuo Gui Yin can shorten the latency time in water maze and platform jumping experiment, increase the number of water maze experiment traversing hidden platform, reduce the number of errors in platform jumping experiment; (2) in biochemical index test, In addition to alt, Zuoguiyin could decrease the serum level of ASTPPA-CREAEABUN, increase the activities of SOD and T-AOC and decrease the content of MDA in the liver and kidney of aging model rats. (3) the pathological results of liver and kidney morphology showed that Zuo Gui Yin can alleviate the pathological injury of liver and kidney cells in aging model rats to some extent and maintain the integrity of histocyte morphology. (4) Immunohistochemical experiment of liver and kidney tissue showed that, ZuoGuiyin could enhance the expression of Bcl-2 and decrease the positive expression of Baxfen Caspase-3 in the liver and kidney of aging model rats. (5) 29 biomarkers were identified by the results of urine metabolism in rats. Nine related metabolic pathways were identified. The results showed that Zuoguiyin could make the metabolic profile of urine of aging model rats normal, have a callback effect on 23 biomarkers, and have an effect on 7 related metabolic pathways. Conclusion: ZuoGuiyin can improve the learning and memory ability of aging model rats, improve the liver and kidney function of aging model rats, enhance the metabolism and scavenging ability of free radicals in aging model rats, and protect liver and kidney tissue cells from injury. Play a role in delaying senescence. Its effect may delay senescence by affecting the expression of apoptosis protein in liver and kidney tissues and regulating the abnormal metabolism of aging model rats.
【学位授予单位】：黑龙江中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R285.5;R-332
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