傣药猫须草有效组分抗肾间质纤维化作用机制研究

bstract: The research shows that the Dai medicine-Clerodendranthus Spicatus has a protective effect for the renal tissue with renal interstitial fibrosis, but its material basis and mechanism are not clear. The renal fibroblasts are the renal cells that are closely related to the generation and development of renal interstitial fibrosis, so we guess that Clerodendranthus Spicatus may inhibit their proliferation or induce their apoptosis by intervening in the expression of proteins related to the signal path of fibroblasts and then play a role in resisting renal fibrosis. In addition, the research has demonstrated that the effective ingredients having pharmacological activity in Clerodendranthus Spicatus are mainly flavonoids and phenolic acid compounds. Based on the above bases, the theme will apply Celigo Cytomter in-situ analysis technology for living cells and Merck Millipore Luminex? 200 liquid-phase suspension chip technology, screen the signal path-related proteins that affect the proliferation and apoptosis of fibroblasts in Clerodendranthus Spicatus, and determine the anti-fibrosis effective ingredients and action targets of Clerodendranthus Spicatus; and also establish a renal fibrosis model of rat, research the influence of effective ingredients in Clerodendranthus Spicatus on the in-vivo fibroblasts and the expression of signal path-related proteins through the in-vivo imaging system of the small live animal (Caliper IVIS Lumina XR), thereby further specifying the action mechanism of effective ingredients in Clerodendranthus Spicatus against renal interstitial fibrosis.

研究表明，傣药猫须草对肾间质纤维化肾组织具有保护作用，但其作用的物质基础和机制尚不明确。肾成纤维细胞是与肾间质纤维化发生、发展密切相关的肾细胞，我们推测猫须草可能通过干预成纤维细胞信号通路相关蛋白的表达而达到抑制其增殖或诱导其凋亡，从而发挥抗肾纤维化的作用。另外，已研究证实猫须草具有药理活性的有效组分主要为黄酮类和酚酸类化合物。基于以上依据，本课题拟采用Celigo Cytomter活细胞原位成像分析技术研究猫须草有效组分对离体成纤维细胞增殖、凋亡的影响；Caliper IVIS Lumina XR 小动物活体体内成像系统研究猫须草有效组分对在体成纤维细胞增殖、凋亡的影响， Merck Millipore Luminex? 200液相悬浮芯片技术研究猫须草影响成纤维细胞增殖、凋亡的信号通路蛋白表达变化，确定猫须草的抗纤维化有效组分及作用靶点，明确猫须草有效组分抗肾间质纤维化的作用机制。

本项目进行了猫须草两个有效组分抗肾间质纤维化机制的研究。利用体内大鼠5/6肾切除肾纤维化模型及原代成纤维细胞探究了猫须草黄酮及酚酸对于肾纤维化进程的抑制作用。体内研究表明猫须草黄酮与酚酸均能够改善大鼠肾功能、降低尿蛋白、降低肾小管间质纤维化并诱导肾成纤维细胞的凋亡，其治疗效果好于猫须草酚酸组；体外机制探究结果表明：猫须草黄酮与猫须草酚酸含药血清能够抑制由TGF-β诱导的成纤维细胞的增殖、细胞外基质的释放、凋亡通路的失活、Smad通路的激活，其药效明显强于猫须草酚酸组；进一步的研究表明，猫须草黄酮能够抑制Smad2/3与Smad4的结合并抑制Smad2/3的核转位，并最终抑制Smad通路的多度激活。综上，猫须草黄酮抑制肾纤维化的机制主要有以下几个方面：1）通过抑制成纤维细胞内周期蛋白的过度表达，诱导成纤维细胞的凋亡，进而抑制成纤维细胞的过度增殖；2）通过抑制成纤维细胞中Smad2/3的核转位，进而抑制Smad2/3的靶基因，抑制细胞外基质的释放，并最终抑制促纤维化因子的作用，进而抑制肾纤维化的进程。项目研究结果将对猫须草的进一步开发研究提供一定依据。.已撰写SCI论文1篇，待发表；已发表论文3篇；已录用论文2篇。已出版专著1部。培养博士研究生2名，硕士研究生1名。