正常小鼠血清通过抑制focal adhesion信号通路减轻小鼠放射性肺炎

Normal mouse serum alleviates radiation pneumonitis in mice by inhibiting the focal adhesion signaling pathway

摘要

目的 探讨正常小鼠血清（NMS）对放射性肺炎的治疗作用及可能机制。 方法 建立胸腔照射诱导的放射性肺炎模型，将小鼠分为对照组、静脉注射血清组、照射组和照射后静脉注射血清组。注射血清组小鼠在照射后立即静脉注射正常小鼠血清100 μL，对照组小鼠注射100 μL 生理盐水，隔天注射1次，共注射8次。照射后15 d取材，HE染色检测肺组织形态学变化，ELISA检测小鼠肺组织和血清中炎症因子肿瘤坏死因子-α（TNF-α）、转化生长因子-β（TGF-β）、白细胞介素-1α（IL-1α）、白细胞介素-6（IL-6）水平；流式细胞术检测肺组织内淋巴细胞比例变化。外泌体miRNA高通量测序探索处理后小鼠的信号通路变化，qRT-PCR检测免疫相关基因的表达水平，使用Western blotting检测黏着斑通路talin-1、tensin 2、FAK、vinculin、α-actinin和paxillin蛋白的表达。 结果 与照射组相比，照射后注射血清组小鼠肺脏器系数、血清及肺组织上清液中炎症因子TNF-α、TGF-β、IL-1α、IL-6水平显著降低（P<0.05），CD45⁺、CD4⁺、T_reg淋巴细胞在小鼠肺组织中的浸润程度显著下降（P<0.05）；肺中Egfr和Pik3cd的mRNA和蛋白表达水平显著下调，talin-1、tensin 2、FAK、vinculin、α-actinin和paxillin蛋白的表达水平也显著降低（P<0.05）。 结论 正常小鼠血清通过抑制focal adhesion信号通路关键蛋白的表达减轻电离辐射诱发的小鼠放射性肺炎。

Abstract

Objective To evaluate the therapeutic effect of normal mouse serum on radiation pneumonitis in mice and explore the possible mechanism. Methods Mouse models of radiation pneumonitis induced by thoracic radiation exposure were given intravenous injections of 100 μL normal mouse serum or normal saline immediately after the exposure followed by injections once every other day for a total of 8 injections. On the 15th day after irradiation, histopathological changes of the lungs of the mice were examined using HE staining, the levels of TNF-α, TGF-β, IL-1α and IL-6 in the lung tissue and serum were detected using ELISA, and the percentages of lymphocytes in the lung tissue were analyzed with flow cytometry. High-throughput sequencing of exosome miRNA was carried out to explore the changes in the signaling pathways. The mRNA expression levels of the immune-related genes were detected by qRT-PCR, and the protein expressions of talin-1, tensin2, FAK, vinculin, α-actinin and paxillin in the focal adhesion signaling pathway were detected with Western blotting. Results In the mouse models of radiation pneumonitis, injections of normal mouse serum significantly decreased the lung organ coefficient, lowered the levels of TNF-α, TGF-β, IL-1α and IL-6 in the serum and lung tissues, and ameliorated infiltration of CD45⁺, CD4⁺ and T_reg lymphocytes in the lung tissue (all P<0.05). The expression levels of Egfr and Pik3cd genes at both the mRNA and protein levels and the protein expressions of talin-1, tensin2, FAK, vinculin, α‑actinin and paxillin were all significantly down-regulated in the mouse models after normal mouse serum treatment. Conclusion Normal mouse serum ameliorates radiation pneumonitis in mice by inhibiting the expressions of key proteins in the Focal adhesion signaling pathway.

关键词

正常小鼠血清 / 放射性肺炎 / 外泌体 / 粘着斑途径

Key words

normal mouse serum / radiation pneumonitis / exosomes / focal adhesion pathway

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