lncRNA-HELF通过调控肝星状细胞和巨噬细胞活化及交互作用促进肝纤维化的机制研究

Long non-coding RNAs (lncRNAs) are involved in numerous biological functions and pathological processes. However, little is known that lncRNAs regulate the progression of liver fibrosis. We have performed microarray assay and identified a novel lncRNA ENSMUST00000147617, named Highly Expressed in Liver Fibrosis (lnc-HELF), which is remarkably up-regulated in mouse fibrotic livers and has homology in human and mouse. We have found that lnc-HELF was mainly localized in the nucleus of hepatic stellate cells (HSCs) and hepatic macrophages (HMs). Moreover, we have observed that the level of lnc-hHELF was increased in liver tissue and serum from patients with advanced hepatic fibrosis. In addition, knockdown of lnc-HELF attenuates liver fibrosis in vivo. Therefore, we hypothesize that lnc-HELF promotes liver fibrosis via regulating the activation of HSCs and HMs. Further investigations will focus on the signaling through which lnc-HELF activates both HSCs and HMs and subsequently induces liver fibrosis on the cellular level and with the use of animal models; illustrating lnc-HELF-modulated interaction between HSCs and HMs; investigating the mechanism of lnc-HELF over-expression; investigating the level of lnc-hHELF in liver and serum with enlarged samples, thus, identifying lnc-HELF as a biomarker for liver fibrosis and a potential target for treatment clinically.

lncRNAs与多种疾病的发生相关，但在肝纤维化中的研究较少。我们前期应用基因芯片技术从小鼠纤维化肝组织中筛选出一个显著过表达且具有物种同源性的lncRNA-HELF。确定了小鼠HELF和人同源转录本hHELF的序列；并明确HELF主要存在于肝星状细胞（HSCs）和肝巨噬细胞（HMs）内，且主要定位在细胞核；我们发现hHELF在肝纤维化和肝硬化患者的肝组织和血清标本中过表达；敲低肝组织内HELF抑制小鼠肝纤维化。因此，我们提出HELF通过调控HSCs和HMs的活化及交互作用进而促进肝纤维化发生的假说。拟进一步在细胞水平和动物水平揭示HELF激活HSCs和HMs诱发肝纤维化的信号通路及分子机制；明确HELF调控HSCs与HMs相互作用的途径；揭示HELF过表达的机理；并扩大样本在患者肝组织和血清中检测HELF的表达，确定HELF成为肝纤维化早期诊断标志物及治疗靶标。

肝纤维化是慢性肝病共同的病理过程，是由肝脏受到损伤后造成的反复刺激和愈合反应引起的，严重的可发展为肝硬化、肝癌。越来越多的研究表明长链非编码RNA（lncRNA）参与细胞的多种生理病理过程。本课题组通过基因芯片技术从小鼠的纤维化肝组织中筛选出一个显著过表达、且具有物种同源性的lncRNA：ENSMUST00000147617，将其命名为lncRNA-HELF（Highly Expressed in Liver Fibrosis）。研究结果表明lnc-Helf在人和小鼠的纤维化肝组织中、激活的肝星状细胞（HSCs）中过表达。体内试验证实通过鼠尾静脉注射腺相关病毒（AAV8）为载体的lnc-Helf-shRNA敲低lnc-Helf可显著减轻四氯化碳（CCl4）和胆管结扎（BDL）诱导的小鼠肝纤维化及炎症。细胞水平研究表明，lnc-Helf一方面促进肝巨噬细胞（HMs）M1型活化及增殖；另一方面通过促进HSCs的增殖和激活进而诱导肝纤维化进展。机制研究表明lnc-Helf通过与PTBP1结合，促进PTBP1与PIK3R5 mRNA结合并维持其mRNA的稳定性，进而诱导下游AKT磷酸化，促进肝纤维化发生，以上研究结果证实lnc-HELF可成为肝纤维化早期诊断标志物及治疗靶标。