ABSTRACT

Crocin is the main monomer of saffron, which is a momentous component of traditional Chinese medicine Lang Qing A Ta. Here, we tried to probe into the role of crocin in liver fibrosis. Hematoxylin-eosin staining and Sirius Red staining were used to observe the pathological changes of liver tissues. After hepatic stellate cells (HSCs) were isolated from liver tissues, lnc-LFAR1, MTF-1, GDNF, and α-SMA expressions were detected by qRT-PCR and western blot. Immunohistochemistry and immunofluorescence were used to detect α-SMA expression. Chromatin immunoprecipitation was used to analyze the binding of MTF-1 to the GDNF promoter. Moreover, the dual-luciferase reporter gene, RNA pull-down, and RNA immunoprecipitation were used to clarify the interaction between MTF-1 and GDNF, lnc-LFAR1 and MTF-1. The degree of liver fibrosis was more severe in the mice from the liver fibrosis model, while the liver fibrosis was alleviated by the injection of crocin. lnc-LFAR1, GDNF, and α-SMA were up-regulated, and MTF-1 was down-regulated in liver fibrosis tissues and cells, while these trends were reversed after the injection of crocin. Besides, lnc-LFAR1 negatively regulated MTF-1 expression, and positively regulated GDNF and α-SMA expressions, and MTF-1 was enriched in the promoter region of GDNF. Furthermore, the cellular direct interactions between MTF-1 and GDNF, lnc-LFAR1 and MTF-1 were verified. In vivo experiments confirmed the relief of crocin on liver fibrosis. Our research expounded that crocin restrained the activation of HSCs through the lnc-LFAR1/MTF-1/GDNF axis, thereby ameliorating liver fibrosis.

摘要

藏红花是藏红花的主要单体，是中药朗青阿塔的重要成分。在这里，我们试图探讨藏红花素在肝纤维化中的作用。苏木精-伊红染色和天狼星红染色观察肝组织病理变化。从肝组织中分离肝星状细胞（HSC）后，通过qRT-PCR和蛋白质印迹检测lnc-LFAR1、MTF-1、GDNF和α-SMA的表达。免疫组织化学和免疫荧光用于检测α-SMA的表达。染色质免疫沉淀用于分析 MTF-1 与 GDNF 启动子的结合。此外，双荧光素酶报告基因、RNA 下拉和 RNA 免疫沉淀用于阐明 MTF-1 和 GDNF、lnc-LFAR1 和 MTF-1 之间的相互作用。肝纤维化模型小鼠肝纤维化程度更重，而注射藏红花素后肝纤维化程度有所缓解。lnc-LFAR1、GDNF和α-SMA在肝纤维化组织和细胞中上调，MTF-1下调，而这些趋势在注射藏红花素后逆转。此外，lnc-LFAR1负调控MTF-1的表达，正调控GDNF和α-SMA的表达，且MTF-1在GDNF的启动子区富集。此外，验证了 MTF-1 和 GDNF、lnc-LFAR1 和 MTF-1 之间的细胞直接相互作用。而这些趋势在注射藏红花素后发生了逆转。此外，lnc-LFAR1负调控MTF-1的表达，正调控GDNF和α-SMA的表达，且MTF-1在GDNF的启动子区富集。此外，验证了 MTF-1 和 GDNF、lnc-LFAR1 和 MTF-1 之间的细胞直接相互作用。而这些趋势在注射藏红花素后发生了逆转。此外，lnc-LFAR1负调控MTF-1的表达，正调控GDNF和α-SMA的表达，且MTF-1在GDNF的启动子区富集。此外，验证了 MTF-1 和 GDNF、lnc-LFAR1 和 MTF-1 之间的细胞直接相互作用。体内实验证实了藏红花素对肝纤维化的缓解作用。我们的研究阐述了藏红花素通过lnc-LFAR1/MTF-1/GDNF轴抑制HSCs的活化，从而改善肝纤维化。