高胆固醇血症兔Oddi括约肌神经-ICC-平滑肌网络的变化及芍药甘草汤的作用机制

Sphincter of Oddi(SO) dysfunction (SOD) is a common clinical disease and could affect the quality of human life Seriously. So far there is no effective treatment of Western medicine. Chinese traditional medicine considers that SOD belongs to the category of“hypochondriac pain”and“jaundice”. ShaoYaoGanCao decoction has the function of relieving spasm and pain and “sour-sweet herbs nourishing yin”(Suan Gan Hua Yin), which is used for thetreatmentof“Liver-spleen disharmony”(Gan Pi Bu He). ShaoYaoGanCao decoction can relax Sphincter of Oddi (SO) and reduce SO pressure; Studies have shown that there is the network of nervous system (NS)-interstitial cell of Cajal (ICC)-smooth muscle cells (SMC) in SO, which is closely related the function of SO and participates in the happening of SOD. But the detailed mechanisms were not clear.. In our study， the immuno-fluoro-histochemical method, confocal microscopy, immuno-electric microscopy, mFISH, TUNEL, BrdU, molecular biology method, electrophysiology method (patch clamp, etc.) and other advanced methods would be used; the distribution, morphology and function in the network of NS- ICC- SMC would be observed to study the changes in the signal transduction in the network of NS- ICC- SMC of SO in hypercholesterolemia rabbit,the SOD rabbit model; and to observe the therapeutic mechanism of ShaoYaoGanCao. The innate character of pathophysiologic changes of SOD and the therapeutic mechanism of ShaoYa GanCao decoction would be explored, thus the mechanism of ShaoYaoGanCao decoction in treating SOD could be revealed.

Oddi括约肌(SO)功能障碍（SOD）是临床常见病，严重影响人类生活质量，目前西医尚无有效治疗方法。中医认为SOD 属“胁痛”、“黄疸”范畴，芍药甘草汤能缓急止痛、 酸甘化阴，治疗肝脾不和。临床应用芍药甘草汤加减可松弛SO，治疗SOD；初步研究证明：SO存在神经-Cajal间质细胞（ICC）-平滑肌网络，与SO功能有关，参与了SOD的发生，但详细机理不清楚。本课题以高胆固醇血症家兔SOD模型SO内神经-ICC-平滑肌网络为对象，应用免疫荧光组化多重染色、共聚焦显微镜、免疫电镜超微结构检测、分子生物学和电生理(膜片钳)等手段，在神经-ICC-平滑肌分布、形态、功能，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上；在组织、细胞和分子水平，观察SOD时SO内神经-ICC-平滑肌网络及信号转导通路变化和芍药甘草汤的作用机制，进一步探讨SOD机理，揭示芍药甘草汤治疗SOD的机理。

Oddi括约肌（sphincter of Oddi,SO）是一种重要的神经肌肉复合体，具有十分复杂的生理功能，SOD( sphincter of Oddi dysfunction ,SOD)，oddi括约肌功能障碍，分为SO功能紊乱及狭窄。SOD不但是大多数胆道、胰腺炎症性及功能性疾病发病机制的主导环节，并且是这些疾患关键的继发病理改变,严重影响人类的生活质量。但目前对SOD的发病机理的了解还处于初始阶段，探讨SOD的发病机理和寻找有效的治疗方法是当前消化病领域的重要课题。目前临床以芍药甘草汤加减治疗SOD取得了很好的疗效，芍药甘草汤(简称芍甘汤) 是源于东汉张仲景《伤寒论·太阳篇》的著名方剂，是常用的缓急止痛之方，但其作用机理不明确，难以大范围推广，不利于中医药的传承、传播以及发展。肠神经系统（Enteric nervous system, ENS）、Cajal间质细胞(interstitial cell of Cajal, ICC)在胃肠道平滑肌中普遍存在，他们共同调节平滑肌的运动，维持肠道功能。ENS由肠神经胶质细胞（Enteric glial cells, EGCs）和神经元组成，EGCs通过释放各种营养因子，在胃肠道稳态中发挥重要作用，能调节神经活动以及神经元的表达，从而调节胃肠道动力功能，本项目通过大量实验研究,运用多种检测方法对蛋白基因产物9.5 (PGP9.5)、一氧化氮(NO)、SMC、ICC和增殖相关蛋白Ki67，PCNA、凋亡的相关蛋白Bcl2、cleaved caspase-3、Bax的表达以及对EGCs的标记物S100β、神经胶质细胞源性神经营养因子（glial cellderived neurotrophic factor , GDNF）、神经元的标记物PGP9.5（protein gene product 9.5）、兴奋性神经递质-P物质（P substance，SP）、抑制性神经递质-血管活性肠肽（vasoactive intestinal peptide, VIP）、NK1R、VIP2R进行检测表明，芍药甘草汤通过保护ENS-ICC-SMC网络，促进高胆固醇血症家兔Oddi功能障碍括约肌的恢复，为中医药事业发展做出了贡献,为临床上中西医结合治疗提供了重要理论依据。