miR-150/Foxp1调控髓源抑制细胞扩增与活化在脓毒症免疫抑制中的作用

Immunosuppression and the resultant increased susceptibility to secondary infection are major causes of sepsis-related death. Myeloid derived suppressor cells (MDSC) are heterogeneous cell population with potent suppressive activity to T cell function. Expansion and activation of MDSC play a crucial role in the development of immunosuppression in sepsis. Recent reports have shown that the phenotype of MDSC evolves from a proinflammatory to an anti-inflammatory, more immature and more potent suppressive activity to T cell function. However, the underlying mechanisms remain elusive. In our preliminary research, we identified a panel of differentially expressed microRNAs, including miR-150, miR-146a, miR-192, in MDSC from the spleen of a murine cecal ligation and punture (CLP) model. The expressions of these microRNAs were further verified by qRT-PCR. The results revealed that miR-150 expression was progressively down-regulated in MDSC of CLP septic mice as compared to that of control mice. Meanwhile, expression of Foxp1, a target gene of miR-150, was significantly up-regulated in MDSC of CLP septic mice. In the present study, we hypothesize that miR-150 modulate the expansion and activation of MDSC in CLP septic mice through tageting Foxp1 expression, thereafter plays a critical role in the development and progression of immunosuppression in sepsis. In the present proposal, we will conduct the following researches: 1. to verify the efffect of miR-150 on the expression of Foxp1 in MDSC in sepsis; 2. to investigate the role of miR-150/Foxp1 in the expansion and activation of MDSC, and the effect on development and progression of immunosuppression in sepsis. 3. to determine the underlying signal transduction pathway of miR-150/Foxp1 in the expansion and activation of MDSC in sepsis. The successful fulfillment of this proposal will gain insight into the mechanisms underlying the development of sepsis-induced immunosuppression, and help to explore novel therapeutics for the treatment of sepsis, which in turn, contributes to improving the prognosis of these patients.

免疫抑制和继发感染是目前脓毒症患者重要死亡原因。髓源抑制细胞（MDSC）是近年新发现的调节性免疫细胞亚群，在脓毒症免疫抑制中起关键作用。MDSC表型脓毒症病程进展而演变，抗炎和免疫抑制活性增强，但其机制尚不清楚。申请者前期研究通过microRNA芯片筛选和验证，发现脓毒症MDSC中miR-150表达显著下调，其靶基因Foxp1表达上调；体外实验发现抑制miR-150表达可促进MDSC扩增，并增强其免疫抑制活性。在此基础上，本项目拟：（1）鉴定脓毒症MDSCs中miR-150靶向调节Foxp1表达；（2）研究miR-150/Foxp1对脓毒症MDSC扩增与活化中的影响，及其在脓毒症免疫抑制中的作用；（3）阐明miR-150/Foxp1通过何种信号通路调控脓毒症MDSC扩增与活化。研究结果将加深对脓毒症免疫抑制机制的认识，为确定miR-150/Foxp1作为脓毒症免疫调理治疗靶点提供实验依据。

脓毒症是严重创伤、大手术和感染的常见并发症，治疗困难。免疫抑制和继发感染是脓毒症患者院内死亡的主要原因。申请者前期研究发现，miR-150/FoxP1信号可能是调控脓毒症脓毒症髓源抑制细胞（MDSCs）增殖与活化的重要机制，在脓毒症免疫抑制中发挥重要作用。本研究主要研究内容：1、鉴定脓毒症MDSC中miR-150靶向调控Foxp1表达；2、研究miR-150/Foxp1对脓毒症MDSC扩增与活化的影响；3、研究miR-150/Foxp1在脓毒症免疫抑制发生中的作用。.主要研究结果：1）小鼠CLP脓毒症模型中MDSCs的比例和分群变化：随着脓毒症病程的进展，CLP小鼠脾脏和骨髓中MDSCs的数量逐渐增加，于7d达到最高峰。2）CLP小鼠脾脏和骨髓中miR-150和Foxp1的表达：脓毒症小鼠M-MDSC和PMN-MDSC中，miR-150在CLP术后7天降至最低；骨髓和脾脏中的Foxp1均显著增高。Foxp1的表达变化与脾脏、骨髓中MDSCs数量的变化一致，也与miR-150的变化一致。Luciferase双荧光素酶报告基因检测，证实miR-150对Foxp1有直接调控作用。3）miR-150/Foxp1对MDSCs增殖分化及小鼠免疫功能的一些的影响：尾静脉注射miR-150的慢病毒，提高小鼠体内miR-150表达水平，显著抑制MDSCs增殖与活化，改善实验组小鼠T细胞功能，提高小鼠生存率。4）临床脓毒症患者外周血清中miR-150/FOXP1的变化：临床研究发现，创伤性脓毒症患者外周血中miR-150的表达水平则明显低于非脓毒症患者及健康对照。5）miR-150通过ARG1途径抑制MDSCs的功能：蛋白组学检测，发现精氨酸酶（ARG1）蛋白的表达显著下降。.以上研究成果加深了对脓毒症免疫抑制发生机制的认识，为确定miR150-FoxP1为脓毒症免疫抑制治疗靶点提供了实验依据。