HuR/lincRNA152复合物在脓毒症免疫抑制中的调控机制

Impairment of TH1 cytokine-producing and antigen-presenting ability of monocytes is important contributing factors for sepsis-induced immunoparalysis. In our previous investigation, we found ouabain, as a specific Na+,K+-ATPase ligand, was able to restore the impaired TNF-α-producing activity in human monocytes, reverse immunoparalysis and improve survival, the underlying mechanism was that ouabain stimulated HuR nuclear export, which posttranscriptionally stabilized TNF-α, GM-CSF and IFN-γmRNA stability, and thereby reprogramming TH1 cytokine expression. Further analysis revealed that cytoplasm accumulation of HuR was relevant to ouabain-induced lincRNA152 production, lincRNA152 forms a complex with HuR. Taken these facts together, we presume that lincRNA152 plays an important role in the pathogenesis of sepsis-induced immunoparalysis. By interacting with HuR, lincRNA152 will prevent the re-entry of HuR into nucleus,fix the HuR onto the 3’-UTR of TH1 cytokines mRNA, and strengthen the effect of HuR on TH1 cytokines mRNA stabilization, finally increasing host immunity against invading microorganism. Investigation on the upregulation of lincRNA152 and interplays between lincRNA152 and HuR as well as the underlying mechanism provides novel insight into the pathological mechanism for sepsis.

单核细胞TH1细胞因子表达能力受抑与抗原提呈能力降低是脓毒症免疫麻痹的重要病理机制，但治疗药物缺乏。我们发现钠钾ATP酶配体哇巴因能恢复单核细胞TH1细胞因子表达能力，逆转脓毒症内毒素耐受，提高存活率，其机制在于哇巴因能刺激RNA 结合蛋白HuR出核，HuR转录后稳定TNFα、GM-CSF、IFN-γ mRNA，从而对TH1细胞因子表达重新编程。进一步研究发现HuR 胞浆内积聚与哇巴因诱导的lincRNA152表达相关，lincRNA152与HuR形成复合物。由此推测在脓毒症免疫抑制状态下，通过激活单核细胞lincRNA152表达，形成lincRNA152/HuR复合物将HuR滞留于胞浆中，并“锚定”HuR在TH1细胞因子mRNA 的3’非翻译区，可以稳定TH1细胞因子mRNA，增强机体清除病原微生物能力。研究lincRNA152/HuR精密调控细胞因子稳定性为脓毒症诊疗提供新的理论依据。

脓毒症发生率高、病情凶险、病死率高，对人类健康造成巨大威胁。免疫麻痹是重症脓毒症死亡的重要病因。利用HPLC-MS/MS检测方法，我们发现重症脓毒症病人血浆中内源性哇巴因大量表达。为探索作用机制，我们分别建立脓毒症炎症模型与能够模拟临床脓毒症免疫麻痹的小鼠盲肠穿孔结扎+沙门氏菌二次打击模型，发现哇巴因在脓毒症炎症模型上加速小鼠死亡，但在脓毒症免疫麻痹模型上却很显著性地改善小鼠存活率，由此推测内源性哇巴因具有调节脓毒症免疫状态新功能。为探索作用机制，我们发现哇巴因能对抗LPS诱导的单核细胞内毒素耐受。在脓毒症免疫麻痹状态下，LPS诱导的miR181d表达能促进TH1细胞因子（TNF-α、GM-CSF、IFN-γ）的快速降解，诱导内毒素耐受，使单核细胞对病原微生物的二次进攻失去响应。此时哇巴因的存在能刺激单核细胞核释放RNA结合蛋白HuR，HuR一方面与miR181d竞争在TNF-α mRNA 3’-UTR上结合位点；另一方面形成压力颗粒(stress granules)将TNF-α mRNA包裹在内,避免其被核酸酶降解,最终稳定TNF-α mRNA。通过诱导HuR出核，哇巴因也同时稳定了GM-CSF与IFN-γ等TH1炎症因子mRNA，最终实现在脓毒症免疫麻痹状态下对TH1细胞因子表达重新编程，逆转脓毒症免疫麻痹，增强机体在脓毒症免疫麻痹状态下抗病原微生物二次感染的能力。 进一步研究发现，哇巴因在调控单核细胞HuR出核方面存在新的作用机制，为了防止单核细胞HuR出核后再次进核，哇巴因能通过c-EBP/β激活LincRNA152表达，该长链非编码RNA能与HuR相互作用，象“绳子”一样将HuR“捆扎”，固定在TH1 细胞因子3’-UTR，从而增强HuR对TH1细胞因子mRNA稳定性。除了能调控TH1细胞因子mRNA稳定性，我们还发现哇巴因能激活免疫抑制状态下单核细胞HLA-DR的表达，增强单核细胞抗原提呈能力，而且哇巴因在调控HLA-DR表达的作用机制与IFN-γ有高度的相似性。因此我们推测内源性哇巴因在调节脓毒症免疫中具有干扰素样作用。