二甲双胍对放射性肺损伤的防护作用及其机制研究

Radiation-induced lung injury (RILI) is the common complication with poor prognosis in thoracic irradiation. Thus, active treatment is clinically important for the prevention of RILI. Local inflammatory reaction is one of the main mechanisms of RILI. However, the exact mechanism remains unclear, and an ideal drug is still lacking. Local inflammatory reaction is mainly mediated by high mobility group box-1 (HMGB1) signal, and metformin (MET)，an ancient hypoglycemic drug, can reduce local inflammatory reaction. Our previous study first found that MET treatment significantly prolonged survival time of mice irradiated by 20Gy X-ray. The protective effect on lung was obviously observed. We also found that MET reduced HMGB1 release in the blood of irradiated mice. Thus, we put forward a hypothesis that MET protects radiation-induced RILI through HMGB1 signaling pathway. A radiation-induced lung injury model was developed to investigate the molecular biological mechanisms underlying the pathogenesis of RILI and the effect of MET on this signaling pathway. Molecular biological techniques, such as RT-PCR, western-blot, were used in vivo and MD experiment to determine the therapeutic effects of MET on RILI and HMGB1 signaling pathway. Furthermore, computer-aided design was used to find the interactive sites between MET and HMGB1. The combination of experiment and theory in this study is helpful to elucidate the pathogenesis of RILI and provide a new approach to the treatment of RILI using ancient medications.

放射性肺损伤是胸部肿瘤放疗的主要并发症，患者生存质量严重受其影响，因此早期防治具有重要意义。局部炎症反应是导致放射性肺损伤的主要机制，但作用机理尚不清楚，且缺乏有效治疗手段。高迁移率族蛋白1(HMGB1)信号传导途径是介导炎症反应的主要介质，我们的前期研究发现降糖药二甲双胍对放射性肺损伤有一定的防护作用，能明显提高致死剂量照射小鼠的生存率，且能有效抑制血液中照射所引起的HMGB1释放。据此我们提出二甲双胍可能通过抑制HMGB1介导的炎症反应从而减轻放射性肺损伤发生和发展的假说。为验证此假说，本研究从组织细胞和分子水平探讨HMGB1信号传导在放射性肺损伤的发生和发展中作用，并采用计算机辅助设计进一步验证二甲双胍与HMGB1相互作用的活性位点。本项目采用实验和理论双向结合反复验证，研究结果将有助于阐明放射性肺损伤的发病机制及二甲双胍的作用机理，为传统药物防治放射性肺损伤提供新的思路和理论依据。

放射性治疗是肺癌、乳腺癌、食道癌等胸部肿瘤患者的主要治疗手段。然而，因肺对辐射中度敏感，放射性肺损伤是胸部肿瘤放疗的常见并发症，它不仅严重影响肿瘤患者的正常治疗和生活质量，也是限制放疗应用最主要的因素。因此,寻找有效防治放射性肺损伤的药物具有非常重要的临床意义。本课题采用6 MV的X线直线加速器进行单次全胸15Gy照射C57BL/6小鼠，成功建立了小鼠的肺损伤动物模型，通过生物信息学分析、RT- PCR、Western blot、免疫组化、Elisa等检测手段，从动物、组织、分子水平探讨了二甲双胍在放射性肺损伤发生、发展中的作用。结果发现：二甲双胍通过抑制炎症因子IL-6、TNF-α 、HMGB1的释放、调节HMGB1-TLR4炎症信号通路和Collagen I /TGF-β1/Smad2/3纤维化信号通路、抑制改善脂质过氧化来调节铁死亡，从而改善了小鼠的放射性肺炎和肺的纤维化。此外，课题在研究过程中还发现了二甲双胍可通过调节肠道内乳杆菌和毛螺菌科细菌的丰度，进而改善放射性肺纤维化等。综上所述：HMGB1是二甲双胍发挥辐射防护作用的关键靶点；以HMGB1/TLR4和Collagen I /TGF-β1/Smad2/3信号通路为线索，可进一步探究二甲双胍对辐照小鼠放射性肺炎和肺纤维化的调控网络，另外二甲双胍还可调节肠道菌群改善放射性肺损伤。本课题从二甲双胍的新视角揭示了对放射性肺损伤的防护作用及相关机制，为临床放射性肺损伤防治提供了理论依据。