Guo XiaoHong, Zhang HaiYan, Zhang QianQian, Li XiuQing, Liu LiXin
aDepartment of Gastroenterology and Hepatology bExperimental Center of Science and Research, The First Clinical Hospital of Shanxi Medical University cKey Laboratory of Cell Physiology, Provincial Department of the Ministry of Education, Shanxi Medical University, Taiyuan, Shanxi Province, China.
Eur J Gastroenterol Hepatol. 2016 Jul;28(7):762-72. doi: 10.1097/MEG.0000000000000631.
Our previous studies characterized insulin-like growth factor-binding protein-related protein 1 (IGFBPrP1) as a molecule that promotes hepatic fibrogenesis, but its mechanism has not been fully elucidated. Here, we have investigated the effect of IGFBPrP1 on gene expression in the hepatic fibrosis-related pathway.
Sprague-Dawley rats received injections of an adenovirus carrying IGFBPrP1 or EGFP cDNA into their tail veins. In hepatic preparations, hepatic stellate cell activation was determined by α-smooth muscle actin expression and hepatic fibrosis by Sirius red staining and hydroxyproline content analysis. IGFBPrP1-inducible genes of the hepatic fibrosis-related pathway were assessed by PCR array. Expression of IGFBPrP1 and transforming growth factor β1 (TGFβ1) and array results were evaluated by quantitative real-time PCR and western blotting.
IGFBPrP1-overexpressing rats showed an increase in α-smooth muscle actin expression and collagen and hydroxyproline content in the liver. The PCR array results indicated that some genes were upregulated and some were downregulated in Ad-IGFBPrP1-infected rats. Among these, Egr1, MAP2K2 (MEK2) and MAPK3 (ERK1) expression increased, whereas PTEN and Hhip mRNA expression decreased. Egr1 protein levels increased and peaked 2 weeks after infection and then decreased gradually. PTEN protein decreased gradually in Ad-IGFBPrP1-infected rats with a concurrent increase in the degree of hepatic fibrosis. TGFβ1 levels increased during hepatic fibrosis development in liver tissues.
Egr1, PTEN, Hhip, MAP2K2 (MEK2) and MAPK3 (ERK1) were identified as candidate genes of the IGFBPrP1-induced hepatic fibrosis-related pathway. IGFBPrP1 promoted hepatic fibrosis mainly by enhancing the TGFβ1 expression that it triggered.
我们之前的研究将胰岛素样生长因子结合蛋白相关蛋白1（IGFBPrP1）鉴定为一种促进肝纤维化的分子，但其机制尚未完全阐明。在此，我们研究了IGFBPrP1对肝纤维化相关途径中基因表达的影响。
将携带IGFBPrP1或EGFP cDNA的腺病毒经尾静脉注射到Sprague-Dawley大鼠体内。在肝脏标本中，通过α-平滑肌肌动蛋白表达确定肝星状细胞活化情况，通过天狼星红染色和羟脯氨酸含量分析确定肝纤维化情况。通过PCR阵列评估肝纤维化相关途径中IGFBPrP1诱导的基因。通过定量实时PCR和蛋白质印迹法评估IGFBPrP1、转化生长因子β1（TGFβ1）的表达及阵列结果。
过表达IGFBPrP1的大鼠肝脏中α-平滑肌肌动蛋白表达、胶原蛋白和羟脯氨酸含量增加。PCR阵列结果表明，在感染Ad-IGFBPrP1的大鼠中，一些基因上调，一些基因下调。其中，Egr1、MAP2K2（MEK2）和MAPK3（ERK1）表达增加，而PTEN和Hhip mRNA表达降低。Egr1蛋白水平在感染后2周升高并达到峰值，随后逐渐下降。在感染Ad-IGFBPrP1的大鼠中，PTEN蛋白逐渐减少，同时肝纤维化程度增加。肝组织中TGFβ1水平在肝纤维化发展过程中升高。
Egr1、PTEN、Hhip、MAP2K2（MEK2）和MAPK3（ERK1）被确定为IGFBPrP1诱导的肝纤维化相关途径的候选基因。IGFBPrP1主要通过增强其触发的TGFβ1表达促进肝纤维化。
