Heat shock protein 90 in alcoholic liver disease: targeting macrophage Function

摘要 肝脏免疫细胞激活是酒精相关性肝病（ALD）的核心，但人们对此知之甚少。 蛋白质稳态网络/途径在保护细胞蛋白质组免受细胞应激方面至关重要。 其中，蛋白质稳态介导剂HSP 90作为治疗靶点被广泛研究和探索 在癌症中。我们报道了HSP 90的两种亚型，HSP 90 AA 1（应激诱导的细胞质HSP 90）和HSP 90 AA 2（应激诱导的细胞质HSP 90）。 HSP 90 B1/GP 96（Endoplasmic reticulum parasites of HSP 90）在ALD中起重要作用。这些研究表明， HSP 90在ALD中的病理生理作用该项目上一个供资期间的研究表明， 在ALD中使用纳米颗粒的HSP 90抑制剂17-DMAG的肝靶向可以预防和逆转肝损伤， 巨噬细胞中GP 96/HSP 90 B1缺陷促进肝脏炎症和ALD、M-TLR 4缺陷的消退 小鼠不能免于ALD，抑制HSP 90降低NLRP 3炎性体活性，降低IL-1β 蛋白质和HSF 1部分有助于减少促炎信号，而HSF 1缺乏则有助于减少促炎信号。 没有保护ALD，这表明HSF 1对ALD的消退没有贡献。在下一个项目期间 本研究旨在探讨HSP 90 AA 1和HSP 90 B1/GP 96在酒精诱导的肝损伤中的作用机制 巨噬细胞活化导致ALD。我们认为HSP 90通过替代机制诱导炎症反应 （独立于HSP 90/TLR 4轴），例如通过表观遗传机制的巨噬细胞有氧糖酵解，或 HIF 1 α。在我们的初步数据中，我们发现在ALD的细胞核中检测到HSP 90 AA 1，并且其抑制可以 调节染色质修饰酶的表达。初步数据还显示，抑制HSP 90 AA 1 减少酒精介导的葡萄糖转运蛋白（SLC 2a 1/GLUT 1）和己糖激酶II的诱导，这两个重要的 糖酵解的介质，表明在ALD的M1巨噬细胞的代谢程序中的作用HSP 90。对 另一方面，优选在肝巨噬细胞中诱导的HSP 90 B1/GP 96通过糖酵解促进炎症 并且其抑制诱导GRP 78 + ATF 3 + Trem 2+恢复性/修复性巨噬细胞解决ALD。我们 假设酒精诱导HSP 90 AA 1通过表观遗传促进巨噬细胞有氧糖酵解 HSP 90 B1/GP 96和ER应激通过HIF 1 α促进巨噬细胞糖酵解。 ALD中HSP 90的抑制使用OXPHOS促进诱导ATF 3 + Trem 2+修复巨噬细胞 保护本发明的具体目的是：1）鉴定HSP 90 AA 1介导的表观遗传和代谢 在体内鼠ALD和体外人AH PBMC中的巨噬细胞编程。2)表征 巨噬细胞特异性HSP 90 B1/GP 96在巨噬细胞代谢中意义及对HSP 90 B1/GP 96的评价 巨噬细胞中的ATF 3和TREM 2提供对肝损伤的保护。在下一个项目中， 在此期间，我们的研究将确定新的代谢和表观遗传机制的调节热休克蛋白90和表征 修复性巨噬细胞在ALD的解决中至关重要。在小鼠ALD模型和人类中研究这些通路 本研究为进一步开发HSP 90在ALD中的临床应用奠定了基础。

ABSTRACT Liver immune cell activation is central to alcohol associated liver disease (ALD) and is poorly understood. The protein homeostasis network/pathways are crucial in safeguarding the cellular proteome from cellular stress. Amongst these, proteostasis mediator HSP90 is widely studied and extensively explored as a therapeutic target in cancer. We reported two isoforms of HSP90, HSP90AA1 (stress inducible cytoplasmic HSP90) and HSP90B1/GP96 (Endoplasmic reticulum paralog of HSP90) are important in ALD. These studies point to a pathophysiological role for HSP90 in ALD. Studies during the last funding period of this project established that liver targeting of HSP90 inhibitor 17-DMAG using nanoparticles in ALD can prevent and reverse liver injury, GP96/HSP90B1 deficiency in macrophages facilitates resolution of liver inflammation and ALD, M-TLR4 deficient mice are not protected from ALD, inhibition of HSP90 reduced NLRP3 inflammasome activity decreasing IL-1β protein and HSF1 partially contributes to reduced pro-inflammatory signaling, whereas deficiency of HSF1 did not protect from ALD, suggesting that HSF1 does not contribute to resolution of ALD. In the next project period we will investigate mechanisms mediated by HSP90AA1 and HSP90B1/GP96 in alcohol induced liver macrophage activation resulting in ALD. We propose that HSP90 induces inflammation by alternate mechanisms (independent of HSP90/TLR4 axis), for instance macrophage aerobic glycolysis via epigenetic mechanisms or HIF1α. In our preliminary data, we found that HSP90AA1 is detected in the nucleus in ALD and its inhibition can modulate expression of chromatin-modifying enzymes. Pilot data also reveal that inhibition of HSP90AA1 reduces alcohol mediated induction of glucose transporter (SLC2a1/GLUT1) and hexokinase II, two important mediators of glycolysis, suggesting a role for HSP90 in metabolic programming of M1 macrophages in ALD. On the other hand, HSP90B1/GP96 induced preferably in liver macrophages promotes inflammation via glycolysis and its inhibition induces GRP78+ATF3+Trem2+ restorative/reparative macrophages resolving ALD. We hypothesize that alcohol induced HSP90AA1 facilitates macrophage aerobic glycolysis via epigenetic mechanisms whereas HSP90B1/GP96 and ER stress facilitate macrophage glycolysis via HIF1α in ALD. Inhibition of HSP90 in ALD induces ATF3+Trem2+ reparative macrophages using OXPHOS facilitating protection. The specific Aims of this proposal are - 1) To identify HSP90AA1 mediated epigenetic and metabolic programming of macrophages in murine ALD in vivo and human AH PBMCs in vitro. 2) To characterize the significance of macrophage-specific HSP90B1/GP96 in macrophage metabolism and assess significance of ATF3 and TREM2 in macrophages conferring protection from liver injury. Collectively, during the next project period our studies will identify novel metabolic and epigenetic mechanisms regulated by HSP90 and characterize reparative macrophages crucial in resolution of ALD. Studying these pathways in mouse ALD model and human AH patient monocytes will provide basis for future clinical development of HSP90 in ALD.