脾气虚证大鼠肠神经-ICC-平滑肌网络损伤和针刺足三里、天枢穴修复机制

The syndrome of dificiency of spleen-Qi is a commen dificiency syndrome in Chinese traditional medicine which is the basis of other spleen syndrome. The results of clinical study in integrated Chinese traditional and western medicine showed that there were serious gastrointestinal motility dysfunction in the patients with syndrome of dificiency of spleen-Qi, and acupuncture on ZU SAN LI(S36) and TIAN SHU (S25) can correct the gastrointestinal motility dysfunction and effectively treat the syndrome of dificiency of spleen-Qi. But the detail mechanism was not distinct. The results of recent study in modern medicine proved that the signal transduction in the network of enteral nervous system(ENS)-interstitial cell of Cajal(ICC)-smooth muscle cell(SMC) was closely correlated with the gastrointestinal motility function..In our study， the immuno-fluoro-histochemical method, confocal microscopy, immuno-electric microscopy, mFISH, TUNEL, BrdU，and the analysis of gene chips would be used; the morphology and function in the network of ENS- ICC- SMC would be observed to study the changes in the signal transduction in the network of ENS- ICC- SMC in Wistar rats with the syndrome of dificiency of spleen-Qi; and to observe the therapeutic mechanism of acupuncture on ZU SAN LI(S36) and TIAN SHU (S25). The innate character of pathophysiologic changes in the syndrome of dificiency of spleen-Qi would be explored, the scientific connotation of "theory of spleen-stomach" in Chinese traditional medicine would be repleshed, and the theory basis for development, popularization and utilization in acupuncture on ZU SAN LI(S36) and TIAN SHU (S25) would be established in our study.

脾气虚是祖国医学最常见的虚证之一，也是脾脏其它证候的基础。中西医结合临床观察结果显示脾气虚证患者存在明显胃肠运动功能障碍，针刺足三里和天枢穴可纠正胃肠运动功能障碍，有效治疗脾气虚证，但详细机理还不清楚。最近研究证明胃肠神经-Cajal间质细胞（ICC）-平滑肌网络信号转导通路与胃肠运动密切相关。本研究应用免疫荧光组化多重染色、共聚焦显微镜观测、免疫电镜超微结构检测和TUNEL、BrdU、 mFISH、基因芯片等方法；在胃肠运动神经、ICC和平滑肌细胞形态、功能，肠神经-ICC间信号转导和ICC-平滑肌细胞间缝隙连接等多个层面；在组织、细胞和基因水平，从形态、功能和细胞间通讯角度，观察脾气虚证大鼠肠神经-ICC-平滑肌网络信号转导通路变化和针刺阳明胃经足三里、天枢穴干预机制。进一步探讨脾气虚证病理生理改变本质，丰富脾胃学说科学内涵，为针刺足三里、天枢穴治疗适应症进一步开发和推广应用奠定基础。

中西医结合临床观察结果显示脾气虚证患者存在明显胃肠运动功能障碍，针刺足三里和天枢穴可纠正胃肠运动功能障碍，有效治疗脾气虚证，但详细机理还不清楚。. 最近现代医学研究证明Cajal间质细胞（ICC）是胃肠慢波的起搏细胞，参与肠神经系统（ENS）信号向平滑肌细胞（SMC）的传导。肠神经、ICC、平滑肌细胞，三者相互连接，共同构成ENS - ICC-平滑肌网络，做为胃肠运动的基本功能单位，参与到胃肠动力发生和调控中。. 本研究应用免疫荧光组化多重染色标记、共聚焦显微镜观测、免疫电镜超威结构检测和基因芯片等方法；观察了脾气虚证大鼠ENS -ICC-平滑肌网络信号通路变化和针刺阳明胃经足三里、天枢穴干预机制。. 结果显示：①针刺足三里穴和天枢穴能够提高脾气虚证大鼠摄食量和体 重，改善脾气虚证大鼠肠道高敏症；②针刺足三里穴和天枢穴能够改善脾气虚证大鼠胃肠道ICC损伤和改善脾气虚证大鼠免疫功能；③针刺足三里穴及天枢穴治疗能够显着减轻脾气虚证小肠充血水肿，改善小肠功能。④脾气虚时，小肠PGP 9.5、ACh、SP、VIP和NO神经纤维数量明显减少，肠神经网络结构破坏严重；针刺足三里穴及天枢穴能够减轻脾气虚证肠神经的损坏，维持肠神经网络结构的完整。⑤针刺足三里穴和天枢穴可减轻脾气虚时ICC形态结构和ICC-SMC间连接的损伤。⑥在基因表达水平，足三里针刺组与自然恢复组具有显着差别。提示针刺足三里穴治疗对脾气虚证的改善是通过多通路进行的，主要包括脂类代谢，多糖代谢，类固醇激素的代谢，细胞的增殖和分化，矿物质的代谢，胆汁的代谢等。. 本研究在ENS、ICC和平滑肌细胞功能和形态、肠神经—ICC—平滑肌网络等多个层面；在组织、细胞和基因水平，从形态、功能和细胞间通讯角度，观察脾气虚证大鼠肠神经-ICC-平滑肌网络信号转导通路变化和针刺足 三里等穴干预机制。结果提示，针刺足三里穴和天枢穴可通过作用在机体多 靶点上协调胃肠运动，有效治疗脾气虚证。为针刺足阳明胃经足三里等穴治 疗脾气虚证提供理论依据。