Targeting proteotoxic stress responses in liver fibrosis

摘要 肝纤维化与炎症和解除调节的伤口愈合反应有关，由慢性肝炎引发 暴露于各种类型的侮辱中，包括酒精、肥胖、NASH和病毒性肝炎。新近兴起的 针对消除病原体的肝脏疗法在解决纤维化方面是有希望的。然而，为了 设计抗纤维化的治疗策略，需要对纤维化的发病机制有更好的认识。 细胞外基质积聚是肝纤维化的标志。肝星状细胞(HSCs)是 由促纤维化刺激激活，是肝脏中产生细胞外基质的主要细胞。氧化的作用 应激和炎性细胞因子在HSC激活中的作用一直受到广泛的研究。瞄准 应激途径灭活肝星状细胞是一种有吸引力的治疗策略，也是有效治疗的关键。 应激诱导的转录因子和蛋白毒性反应的主要调节因子，HSF1介导 HSP90和HSP70的诱导，应激伴侣在炎症信号转导中起重要作用，并可能在 在肝星状细胞激活和逆转中的作用。我们的初步数据显示，HSF1， 不能诱导HSP70促进HSC活化和纤维化，而且无法分解HSC 纤维化恢复模型中的活化和纤维化。胞浆HSP90AA1和内质增加 纤维化肝组织中的网状细胞(ER)、HSP90B1/Grp94/gp96可能参与了肝纤维化的发生。针对HSP90， 使用一种特定的抑制剂17-DMAG，它可以诱导HSF1激活和下游HSP70，将描绘出 HSF1-HSP70在肝星状细胞失活中的作用我们假设HSF1处于不活动状态 星状细胞激活过程中HSP90的诱导有助于肝纤维化，而HSP90的诱导 HSF1和HSPA1A/HSP70有助于纤维化的消退。具体目标是-1)拆解 应激介导的转录因子HSF1在肝纤维化调节中的作用 星状细胞特异性HSF1基因敲除对肝纤维化的影响 肝纤维化消退；评估HSF1调节的纵向表达、活性和新的靶基因 在纤维化和消退过程中。2)评估靶向HSP90AA1(胞浆)和/或 HSP90B1/Grp94/gp96(ER)通过评估HSP90B1/Grp94/gp96(ER)的诱导作用调节星状细胞的激活和纤维化 HSP90AA1和gp96在星状细胞激活中的作用；HSP90抑制剂17-DMAG对肝脏的影响 肝纤维化；测定体内星状细胞特异性抑制gp96和HSP90对肝纤维化的影响。 3)研究HSF1诱导的HSP70在星状细胞功能中的重要性： HSP70及其对HSF1的依赖，在纤维化消退过程中；调查是否过度表达 HSP70使HSC失活并解决纤维化；在17-DMAG治疗期间评估HSF1-HSP70轴 肝星状细胞活化和纤维化。我们建议的研究将揭开HSF1介导的应激通路的作用 和伴侣蛋白在星状细胞活化和纤维化中的作用。

ABSTRACT Liver fibrosis is linked to inflammation and deregulated wound healing response, triggered by chronic exposure to various types of insults including alcohol, obesity, NASH and viral hepatitis. Recently emerging liver therapies directed to eliminate the pathogenic agent suggest promise in resolving fibrosis. However, to design anti-fibrotic therapeutic strategies we need a better understanding of the pathogenesis of fibrosis. Accumulation of extracellular matrix is the hallmark of hepatic fibrosis. Hepatic stellate cells (HSCs) are activated by pro-fibrogenic stimuli and are the main ECM producing cells in the liver. The role of oxidative stress and inflammatory cytokines in HSC activation has been under extensive investigation. Targeting stress pathways to inactivate HSCs is an attractive therapeutic strategy and crucial to effective treatment. Stress induced transcription factor and master regulator of proteotoxic responses, HSF1 mediates induction of HSP90 and HSP70, stress chaperones important in inflammatory signaling and likely to play a role in HSC activation and reversion, respectively. Our preliminary data show that deficiency of HSF1, unable to induce HSP70 promotes HSC activation and fibrosis and moreover, is unable to resolve HSC activation and fibrosis in a model of fibrosis recovery. Increased cytosolic HSP90AA1 and endoplasmic reticulum (ER), HSP90B1/Grp94/gp96 in fibrotic livers may contribute to fibrogenesis. Targeting HSP90, using a specific inhibitor, 17-DMAG, which induces HSF1 activation and downstream HSP70, will delineate the role of HSF1-HSP70 in inactivation of hepatic stellate cells. We hypothesize that HSF1 inactivity and HSP90 induction during stellate cell activation contributes to liver fibrosis, whereas induction of HSF1 and HSPA1A/HSP70 contributes to resolution of fibrosis. The Specific Aims are- 1) To unravel the role of stress mediated transcription factor, HSF1 on regulation of liver fibrosis by: Examining the effect of stellate cell specific knock-out of HSF1 on fibrogenesis; Testing effect of HSF1-plasmid nanoparticles on fibrosis resolution; Evaluating longitudinal expression, activity and novel target genes regulated by HSF1 during fibrosis and resolution. 2) To assess whether targeting HSP90AA1 (cytosolic) and/or HSP90B1/Grp94/gp96(ER) regulates stellate cell activation and fibrogenesis by: Evaluating the induction of HSP90AA1 and gp96 during stellate cell activation; Investigating effect of hsp90 inhibitor, 17-DMAG on liver fibrosis; Determining effect of stellate cell-specific inhibition of gp96 and HSP90 in vivo, on hepatic fibrosis. 3) To investigate importance of HSF1 induced HSP70 in stellate cell function by: Determining induction of Hsp70 and its dependence on HSF1, during resolution of fibrosis; Investigating whether overexpression of HSP70 inactivates HSC and resolves fibrosis; Evaluating HSF1-HSP70 axis during 17-DMAG treatment on HSC activation and fibrosis. Our proposed studies will unravel the role of HSF1 mediated stress pathways and chaperones in stellate cell activation and fibrosis.