作者：张旭涛，张迁，王瀚，王斌，刘峰舟，吴侃，阮柏，张敏，王小成

单位：

1.710032陕西省西安市，空军军医大学航空航天医学系航空航天临床医学中心 2.710032陕西省西安市，空军军医大学西京医院空勤科 通信作者：王小成，E-mail：wxcnose@126.com

Units：

1.Center of Clinical Aerospace Medicine, Air Force Medical University Aerospace Medicine School, Xi'an 710032, China 2.Department of Aviation Medicine, Air Force Medical University Xijing Hospital, Xi'an 710032, China Corresponding author: WANG Xiaocheng, E-mail: wxcnose@126.com

关键词：

肺纤维化; 放射性肺纤维化; 丝氨酸/精氨酸蛋白激酶1; 巨噬细胞;

Keywords：

Pulmonary fibrosis; Radiation-induced pulmonary fibrosis; SRPK1; Macrophages

CLC：

DOI：

10.12114/j.issn.1008-5971.2022.00.170

Funds：

陕西省重点研发计划项目（2021SF-253，2021SF-146）；陕西省自然科学基础研究计划项目（2019JQ-702）

摘要：

目的 分析丝氨酸/精氨酸蛋白激酶1(SRPK1)在放射性肺纤维化(RIPF)中的作用及其机制。方法2021年3月至2022年2月,将30只2月龄C57BL/6J雌性小鼠随机分为对照组、模型组和干预组,各10只。对照组小鼠不进行干预;模型组和干预组小鼠均构建RIPF模型,建模成功后干预组小鼠腹腔注射SRPK1抑制剂——SRPIN340,模型组小鼠给予相同剂量的PBS;两组均干预16周,其中模型组小鼠死亡5只,干预组死亡3只。干预结束后收集各组小鼠全肺组织及肺泡灌洗液,采用HE染色观察小鼠肺组织结构,Masson染色观察小鼠肺组织细胞外基质(ECM)沉积情况,羟脯氨酸试剂盒检测小鼠肺组织羟脯氨酸水平,ELISA检测小鼠肺泡灌洗液中IL-6、IL-8、IL-13、转化生长因子β(TGF-β)1水平,免疫荧光法检测小鼠肺组织CD45+T淋巴细胞计数,免疫组化法检测小鼠肺组织精氨酸酶1(Arg-1)、SRPK1、α平滑肌肌动蛋白(α-SMA)表达水平。结果 HE染色结果显示,模型组小鼠肺组织出现明显的损伤和结构紊乱,肺泡壁增厚扭曲,可见大量病灶形成;干预组小鼠肺组织出现轻微损伤,肺组织结构较模型组完整,肺泡壁增厚现象减少。Masson染色结果显示,模型组小鼠肺组织中出现大量ECM沉积;与模型组相比,干预组小鼠肺组织ECM沉积明显减少。模型组、干预组小鼠肺组织羟脯氨酸水平高于对照组(P<0.05);干预组小鼠肺组织羟脯氨酸水平低于模型组(P<0.05)。模型组、干预组小鼠肺泡灌洗液中IL-6、IL-8、IL-13、TGF-β1水平高于对照组(P<0.05);干预组小鼠肺泡灌洗液中IL-6、IL-8、IL-13、TGF-β1水平低于模型组(P<0.05)。模型组、干预组小鼠肺组织CD45+T淋巴细胞计数高于对照组(P<0.05);干预组小鼠肺组织CD45+T淋巴细胞计数低于模型组(P<0.05)。模型组小鼠肺组织Arg-1、SRPK1、α-SMA表达水平高于对照组(P<0.05);干预组小鼠肺组织Arg-1、SRPK1表达水平高于对照组、低于模型组,α-SMA表达水平低于模型组(P<0.05)。结论 SRPK1可促进RIPF的发生发展,其可能机制为SRPK1促进肺组织内巨噬细胞向M2型极化,激活成纤维细胞,进而促进肺纤维化。

Abstract：

【Abstract】 Objective To analyze the role and mechanism of serine/arginine protein kinase 1 (SRPK1) in radiationinduced pulmonary fibrosis (RIPF) . Methods From March 2021 to February 2022, 30 2-month-old C57BL/6J female micewere randomly divided into control group, model group and intervention group, with 10 mice in each group. The mice in the controlgroup were not intervened; the mice in the model group and the intervention group were constructed with RIPF model. Aftersuccessful modeling, the mice in the intervention group were intraperitoneally injected with SRPK1 inhibitor SRPIN340, and themice in the model group were given the same dose of PBS. Both groups were intervened for 16 weeks, of which 5 mice died inthe model group and 3 mice died in the intervention group. After intervention, the whole lung tissue and bronchoalveolar lavagefluid of mice in each group were collected. HE staining was used to observe the structure of mouse lung tissue, Masson stainingwas used to observe the deposition of extracellular matrix (ECM) in mouse lung tissue, hydroxyproline kit was used to detect thelevel of hydroxyproline in mouse lung tissue, ELISA was used to detect the levels of IL-6, IL-8, IL-13 and transforming growthfactor β (TGF-β) 1 in mouse bronchoalveolar lavage fluid, immunofluorescence method was used to detect the count of CD45 + Tlymphocytes in mouse lung tissue, and immunohistochemical method was used to detect the expression levels of arginase 1 (Arg-1) , SRPK1 and α-smooth muscle actin (α-SMA) in mouse lung tissue.Results The results of HE staining showed that thelung tissue of the model group showed obvious damage and structural disorder, the alveolar wall thickened and distorted, and alarge number of lesions were formed; the lung tissue of the intervention group was slightly damaged, the lung tissue structure wasmore complete than that of the model group, and the thickening of alveolar wall was reduced. Masson staining showed that a largeamount of ECM deposition appeared in the lung tissue of the model group; compared with the model group, the ECM deposition inthe lung tissue of the intervention group was significantly reduced. The level of hydroxyproline in lung tissue of model group andintervention group was higher than that of control group (P < 0.05) ; the level of hydroxyproline in the lung tissue of the interventiongroup was lower than that of the model group (P < 0.05) . The levels of IL-6, IL-8, IL-13 and TGF-β1 in the bronchoalveolarlavage fluid of the model group and the intervention group were higher than those of the control group (P < 0.05) ; the levels of IL-6, IL-8, IL-13 and TGF-β1 in the bronchoalveolar lavage fluid of mice in the intervention group were lower than those in themodel group (P < 0.05) . The counts of CD45 + T lymphocytes in the lung tissue of the mice in the model group and the interventiongroup were higher than those in the control group (P < 0.05) ; the counts of CD45 + T lymphocytes in the lung tissues of the mice inthe intervention group were lower than those in the model group (P < 0.05) . The expression levels of Arg-1, SRPK1 andα-SMAin the lung tissue of mice in the model group were higher than those in the control group (P < 0.05) ; the expression levels of Arg-1 and SRPK1 in the lung tissue of mice in the intervention group were higher than those in the control group and lower than thosein the model group, and the expression level of α-SMA was lower than that in the model group (P < 0.05) . Conclusion SRPK1can promote the occurrence and development of RIPF, and the possible mechanism is that SRPK1 can promote the polarization ofmacrophages to M2 type in lung tissue, activate fibroblasts, and then promote pulmonary fibrosis.

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