组蛋白甲基转移酶SETD2通过HIF-1α甲基化介导脓毒症时单核-巨噬细胞功能障碍的机制研究

Dysfunction of monocytes/macrophages plays an vital role in immunosuppression during sepsis. However, the underlying mechanism remains unclear. HIF-1α, a "trigger point" of metabolism, plays an essential role in metabolic reprogramming of phenotype and function of macrophages. Methylation, a new kind of modification of HIF-1α was discovered recently in tumor research. Methylation of HIF-1α plays an important role in tumor proliferation and metastasis via mediate the degradation of HIF-1α. However, the methylation of HIF-1α has not been reported in sepsis. Our preliminary experiment found that the methylation of HIF-1α increased in endotoxin-tolerant macrophages, and the expression of SETD2, a member of histone methyltransferase, increased significantly in peripheral blood monocytes in patients with severe sepsis, which is correlated with the severity of disease. We also found that SETD2 negatively regulates macrophage function via HIF-1α in vitro. So we propose hypothesis that SETD2 may mediate the dysfunction of macrophage in sepsis via HIF-1α methylation-mediated cellular reprogramming. The project was designed to investigate our hypothesis in septic patients, mice, as well as in vitro cultured cells. We would investigate the expression of SETD2 in monocyte during sepsis, and the effect of SETD2 on monocyte/macrophage function. Whether the regulation was mediated through methylation of HIF-1α, and the molecular mechanism of degradation HIF-1α after methylation by SETD2 would also be investigated. The present project may contribute to the further understanding of the metabolic regulation and its influence in pathophysiology of sepsis. The results of this project will provide the experimental evidence for identifying a therapeutic target for sepsis in future.

单核-巨噬细胞(Mφ)功能障碍是脓毒症免疫抑制的关键环节之一。HIF-1α可通过能量代谢重编程调控Mφ的表型及功能。甲基化是新发现的HIF-1α修饰方式，但HIF-1α甲基化对脓毒症Mφ调控作用不明。我们前期研究显示内毒素耐受Mφ中HIF-1α甲基化增强，而筛查常见甲基转移酶发现脓毒症免疫抑制患者单核细胞SETD2表达改变最显著，且与病情严重程度呈正相关。敲减SETD2后内毒素耐受Mφ功能显著改善，这一效应依赖于HIF-1α存在。据此，我们推测SETD2可能通过HIF-1α甲基化调控细胞代谢重编程而介导脓毒症时Mφ功能障碍。本研究拟从临床、细胞和实验动物层面研究脓毒症时SETD2表达改变及其对Mφ功能的调控作用，探讨该调控是否通过HIF-1α甲基化介导，并探索SETD2甲基化HIF-1α后介导其蛋白降解的分子机制。本研究将有助于进一步揭示脓毒症免疫抑制的机制，为脓毒症的防治提供新靶点。

单核-巨噬细胞(Mφ)功能障碍是脓毒症免疫抑制的关键环节之一。HIF-1α可通过能量代谢重编程调控Mφ的表型及功能。SETD2是组蛋白甲基转移酶成员之一，可特异性催化组蛋白H3第36位赖氨酸（H3K36）的三甲基化，在转录延伸、RNA剪接和DNA损伤修复等过程中发挥关键调节作用。我们前期研究显示内毒素耐受Mφ中HIF-1α甲基化增强，而筛查常见甲基转移酶发现脓毒症免疫抑制患者单核细胞SETD2表达改变最显著，且与病情严重程度呈正相关。敲减SETD2后内毒素耐受Mφ功能显著改善，这一效应依赖于HIF-1α存在。据此，我们推测SETD2可能通过HIF-1α甲基化调控细胞代谢重编程而介导脓毒症时Mφ功能障碍。本研究拟从临床、细胞和实验动物层面研究探索脓毒症时SETD2的表达及其对单核-巨噬细胞功能的调控作用、该调控作用是否通过影响HIF-1α介导，并进一步探索其分子机制。本研究将有助于进一步揭示脓毒症免疫抑制的机制，为脓毒症的防治提供新靶点。我们的研究显示，SETD2可抑制LPS 诱导的 M1 巨噬细胞极化和糖酵解，以及HIF-1α的上调。SEDT2可抑制巨噬细胞中HIF-1启动子的活性，从而下调HIF-1α水平。SETD2对巨噬细胞的功能和糖酵解具有负调控作用，这一调控作用是通过HIF-1α介导的。随后的小鼠在体试验显示SETD2过表达可改善CLP小鼠存活率并减轻其肺损伤。因此，本项目主要在离体和在体水平：明确SETD2可抑制脓毒症时巨噬细胞的糖酵解和向 M1极化；SETD2对于脓毒症时巨噬细胞的调控是通过调控 HIF-1α 转录实现的，SETD2对脓毒症小鼠具有一定的保护作用。该项目资助并标注该项目的SCI 论文1篇，正在投稿中。共有3名硕士研究生参与该项目，在该项目支持下，协助其导师培养其中2 名研究生顺利毕业。