Pathogenic role of PDE4 in hepatic stellate cell activation and TGF signaling

不同病因的肝损伤导致涉及肝星状细胞活化的创伤愈合过程 （HSC）。然而，持续的肝细胞损伤和炎症导致不受控制的激活， HSC的增殖和肝纤维化的发展导致肝硬化甚至肝细胞癌。 尽管取得了一些进展，但我们在理解人权进程所涉机制方面仍然存在差距。 HSC从静止表型到活化表型的转化。最近，我们发现， 磷酸二酯酶4（PDE 4）亚家族的酶在胆汁淤积的发展中起致病作用 肝损伤和纤维化。值得注意的是，PDE 4不存在于静止的HSC中，并且在 体外活化。此外，PDE 4特异性抑制剂咯利普兰有效地减弱SMA、胶原蛋白 表达和伴随的形态学变化。PDE 4是cAMP中最大的亚家族。 水解PDE，其严格调节细胞cAMP的水平。cAMP通过其效应分子 蛋白激酶A（PKA）和cAMP直接激活的交换蛋白（EPAC），已被证明可以降低 调节非肝细胞中细胞因子诱导的纤维化基因。因此，我们假设诱导PDE 4 表达和活性通过降低cAMP-PKA/EPAC活性在HSC活化中起因果作用， 促进纤维化信号传导。我们推测在HSC活化过程中，启动子相关的表观遗传 变化和翻译后修饰在调节PDE 4表达中起重要作用， 活动我们还推测，PDE 4抑制将恢复PKA/EPAC活性，并减弱TGF β-Smad 通过以下途径传递信号：（i）相关MAPK的失活;和（ii）去抑制过氧化物酶体增殖物激活受体（PPAR），导致减少 SMA和Col 1A 1的表达。重要的是，抑制PDE 4活性可能是一种重要的治疗方法， 肝纤维化的治疗方法本提案的具体目标是：1）确定政策制定专家第四届会议在 调控HSC中的纤维化信号传导; 2）确定启动子相关的表观遗传修饰 有助于HSC活化过程中PDE 4同种型的诱导;和3）确定翻译后 在HSC活化过程中与PDE 4同种型功能相关的修饰（PTM）。重要的是，这一结果 COBRE资助的项目将为体内转化研究提供原理证明和机制依据 (R01)探讨PDE 4靶向治疗肝纤维化的策略。

Liver injury of different etiologies leads to a wound healing process involving activation of hepatic stellate cells (HSCs). However, an ongoing hepatocyte injury and inflammation results in an uncontrolled activation and proliferation of HSCs and development of hepatic fibrosis leading to cirrhosis and even hepatocellular cancer. Despite the advances made, gaps remain in our understanding of the mechanisms involved in the process of HSC transformation from quiescent to activated phenotype. Recently, we discovered that the phosphodiesterase 4 (PDE4) subfamily of enzymes play a pathogenic role in the development of cholestatic liver injury and fibrosis. Notably, PDE4s are not present in quiescent HSCs and are rapidly induced upon activation in vitro. Further, the PDE4 specific inhibitor, rolipram, effectively attenuates ¿SMA, collagen expression and accompanying morphological changes in HSCs. PDE4 is the largest sub-family among cAMP- hydrolyzing PDEs, which tightly regulate the levels of cellular cAMP. cAMP, through its effector molecules protein kinase A (PKA) and Exchange Protein directly Activated by cAMP (EPAC), has been shown to down- regulate cytokine induced fibrogenic genes in non-hepatic cells. Hence, we hypothesize that induction of PDE4 expression and activity plays a causal role in HSC activation by decreasing cAMP-PKA/EPAC activities and promoting fibrogenic signaling. We postulate that during HSC activation, promoter associated epigenetic changes and post-translational modifications play a significant role in the regulation of PDE4 expression and activity. We also postulate that PDE4 inhibition will restore PKA/EPAC activities and attenuate TGF¿-Smad signaling through: (i) inactivation of relevant MAPKs; and (ii) de-repressing PPAR¿ leading to decreased expression of ¿SMA and Col1A1. Importantly, inhibition of PDE4 activity may be a significant therapeutic approach for liver fibrosis. The specific aims of this proposal are to: 1) Determine the role of PDE4 in the regulation of fibrogenic signaling in HSCs; 2) Determine promoter-associated epigenetic modifications contributing to the induction of PDE4 isoforms during HSC activation; and 3) Determine the post-translational modifications (PTMs) relevant for PDE4 isoform function during HSC activation. Importantly, the results of this COBRE-funded project will provide proof-of-principle and mechanistic rationale for in vivo translational studies (R01) to examine PDE4 targeted strategies for prevention and treatment of hepatic fibrosis.