炎症抑制因子SIRT1通过调控PKM2核/浆分布阻抑代谢-炎症偶联的机制研究

Inflammation requires intensive metabolic support while modulation of the metabolic process is emerging as a novel approach to limit inflammatory injury. We have previously found that metabolic intervention such as glycolysis inhibition alleviated inflammation. In addition, our recent study has found that the nuclear translocation of pyruvate kinase M2 (PKM2) played crucial roles in inflammatory injury. The nuclear translocation of PKM2 impairs the generation of pyruvate kinase and upregulates the key enzymes in glycolysis pathway, it also stimulate the expression of inflammatory factors via modulating STAT3. The nuclear translocation of PKM2 is regarded as a critical event links metabolism and inflammation. The nucleocytoplasmic distribution of PKM2 could be regulated by post-translational modifications. We recently found that the activity of the inflammation repressor sirtuin 1 (SIRT1) was negatively correlated with the acetylation level and nuclear content of PKM2. Therefore, SIRT1 might broke the link between metabolism and inflammation via deacetylation of PKM2 and supression of its nuclear accumulation. To prove this hypothesis, we firstly plan to confirm that SIRT1 interacts with PKM2 and catalyses the deacetylation of PKM2. In the next step, we plan to confirm that PKM2 is crucial for the anti-inflammatory activity of SIRT1. And finally, the status of glycolysis and the activation nuclear pro-inflammatory signal would be investigated to reveal the downstream mechanisms underlying the repressive effects of SIRT1 on the link between metabolism and inflammation. This study would reveal a novel mechanism of inflammation regulation concerning the link between metabolism and inflammation, which might imply a new strategy for the intervention of inflammatory injury.

炎症需要高强度代谢支持，调控代谢已成为限制炎症损伤的新途径。我们前期发现抑制糖酵解等代谢干预可减轻炎症；近期又发现M2型丙酮酸激酶（PKM2）核转位在炎症损伤中发挥重要作用。PKM2核转位可阻断丙酮酸合成并上调糖酵解关键酶；也可通过STAT3等促进炎症因子表达，是偶联代谢与炎症的关键事件。PKM2核/浆分布受翻译后修饰的调节，我们新近发现炎症抑制因子SIRT1的活性与PKM2的乙酰化及胞核内含量负相关。因此，我们推测SIRT1可能通过去乙酰化PKM2降低其核聚集、阻断PKM2介导的代谢-炎症偶联，这可能是炎症调控新机制。为此，本项目拟首先探讨SIRT1可否结合并去乙酰化PKM2，随后证实PKM2是SIRT1抑制炎症的重要靶点，最后通过分析糖酵解状态和胞核内促炎信号激活状态揭示SIRT1阻抑代谢-炎症偶联的下游机制。本项目有望从代谢-炎症偶联角度揭示炎症调控新机制并为炎症损伤防治提供新启示。

沉默信息调节因子2相关酶1（sirtuin1, SIRT1）是体内的重要代谢感受器，对炎症具有显著的负调节作用。我们的研究发现，SIRT1激活剂干预可抑制内毒素诱导的致死性肺内及全身性炎症反应，减轻肺、肝、心、肾损伤，提升模型小鼠生存率；SIRT1激活剂干预可增强自噬并下调PKM2的水平，联合应用自噬抑制剂或溶酶体抑制剂可阻断SIRT1激活剂的抗炎保护效应及其对PKM2的下调作用；因此，SIRT1激活剂可通过促进PKM2的自噬性降解发挥抗炎作用。除总量变化外，PKM2可移位至胞核内，采用工具药抑制PKM2的核转位可显著减轻炎症损伤；PKM2的核转位伴随丙酮酸激酶活性降低和丙酮酸含量减少；补充丙酮酸衍生物也可减轻炎症损伤；信号通路抗体芯片广谱筛查及免疫印迹验证发现，抑制PKM2核转位或补充丙酮酸衍生物可显著抑制CDK1的抑制性磷酸化；联合应用CDK1抑制剂可阻断PKM2核转位抑制剂及丙酮酸衍生物的保护效应；因此，PKM2/丙酮酸可通过调节CDK1的活性影响炎症损伤的发生发展。丙酮酸可经LDHA的催化代谢为乳酸，抑制LDHA可减轻内毒素诱导的炎症损伤，其机制可能与活性氧的产生减少有关；采用靶向线粒体的抗氧化剂mitoQ、或NADPH氧化酶抑制剂也可减轻内毒素诱导的炎症损伤；因此，催化丙酮酸代谢转化的LDHA在炎症损伤中也发挥了重要作用。此外，转入胞核内的PKM2还可发挥蛋白激酶的作用，抑制PKM2的核转位可下调转录因子STAT3的磷酸化水平，这可能在炎症损伤及损伤后的细胞再生中发挥重要调节作用。总之，本研究揭示了SIRT1调节PKM2的新机制，揭示了PKM2作为代谢激酶在炎症损伤中发挥调控作用的新机制，也揭示了PKM2作为蛋白激酶在炎症损伤后细胞再生中的新作用。