基于肝窦内皮细胞微尺度生物力学性质研究芪术颗粒抗肝纤维化机制

Qizhu granule is TCM famous doctor’s antifibrotic experience side，also Guang An Men Hospital preparation.This drug has been used in clinical for over 30 years, the efficacy against liver fibrosis is exactly, preliminary studies have been carried in different ways,such as the molecular pathways of immune regulation,cytokine expression.We also foud that the major active components of the pharmaceutical composition of Qizhu granule have significant effects on the microscale mechanical properties and the distribution of fenestrae of endothelial cells of sinusoidal, also the expansion of sinusoida,that is liver capillaries.From the perspective of biomechanopharmacology,according to interactions among blood, blood flowing and blood vessels ,we put forward the following scientific hypothesis:"The regulation of Mechanical microenvironment internal sinusoidal is an important mechanism of Qizhu granule anti fibrosis".In this study, atomic force microscopy, scanning electron microscopy, fluorescence microscopy and RT-PCR technology will be used to first observed Qizhu granules and effective active ingredients of benefiting qi Astragalus, activating blood circulation Zedoary on behalf at the nanometer level of the impact on mechanical properties, microscopic morphology, distribution of fenestrae, NO synthesis and gene expression of sinusoidal endothelial cells to verify the hypothesis of objectivity.It opens up a new avenues for anti-hepatic fibrosis mechanism of Chinese medicine,designed in the name of pass on clinical experience of famous TCM and provides a scientific for antifibrotic,also to provide new ideas and new methods for the treatment of liver fibrosis.

芪术颗粒是名老中医抗肝纤维化的经验传承方,是广安门医院院内制剂，临床运用30余年，抗肝纤维化疗效确切，前期已从分子通路、免疫调节、细胞因子表达等角度进行了机制研究。课题组同时发现芪术颗粒主要组成药物活性组分对肝脏微血管—肝窦的内皮细胞微尺度力学性质、窗孔分布及肝窦扩张均有明显作用。本课题从生物力药理学角度，根据血液/血流/血管相互作用的理论，提出科学假说：“对肝窦力学微环境的调整是芪术颗粒抗肝纤维化作用的重要机制之一”。利用原子力显微镜、环境扫描电镜、荧光显微镜及生物信息学技术等，首次从纳米层次观察芪术颗粒及其益气、活血代表药黄芪、莪术有效活性组分对肝窦内皮细胞力学性质、微观形貌、窗孔分布、内分泌及基因表达等的影响，验证假说的客观性。为中医药抗肝纤维化机制研究开辟了新途径，旨在为传承、挖掘名老中医经验并揭示其内涵提供科学依据，并为抗肝纤维化的临床和研究提供新思路和新方法。

芪术颗粒是广安门医院院内制剂，名老中医抗肝纤维化的经验传承方，临床研究证明其抗肝纤维化疗效确切，前期已从分子通路、免疫调节、细胞因子表达等角度进行了机制研究，同时课题组发现芪术颗粒主要组成药物活性组分对肝脏微血管—肝窦的内皮细胞微尺度力学性质、窗孔分布及肝窦扩张均有明显作用。本课题从生物力药理学角度，根据血液/血流/血管相互作用的理论，主要利用原子力显微镜、环境扫描电镜、荧光显微镜及生物信息学技术等，从纳米层次观察芪术颗粒及其益气、活血代表药黄芪、莪术有效活性组分对肝窦内皮细胞的杨氏模量力学性质、表面窗孔微观形貌、NO释放及相关基因表达等的影响。结果显示：（1）芪术颗粒及其有效成分黄芪多糖、莪术油能够明显改善损伤血清引起的肝窦内皮细胞杨氏模量下降，提高细胞刚度，改善细胞力学性质；（2）电子扫描显微镜结果显示，中药组对于维持肝窦内皮细胞表面窗孔形貌具有重要作用，有效保持细胞表面窗孔数量；（3）NO荧光检测结果显示：中药组明显提高肝窦内皮细胞的NO释放，并且提高eNOs蛋白表达；（4）对各组转录组学差异基因鉴定结果显示：各组之间基因表达具有明显差异，差异基因主要富集在代谢、炎症、细胞外基质相关、血流动力及黏性等相关通路，为后续进一步的机制研究提示了研究方向。本研究为中医药抗肝纤维化的机制研究开辟了新途径，为传承、挖掘名老中医经验并揭示其内涵提供了科学依据，并为抗肝纤维化的临床和研究提供新思路和新方法。