正常小鼠血清对小鼠放射性肺损伤的影响

Effect of normal mouse serum on radiation-induced lung injury in mice

  • 摘要:
    目的 探讨正常小鼠血清(NMS)对小鼠放射性肺损伤(RILI)的影响。
    方法 通过全肺13 Gy 照射建立放射性肺炎模型和右肺17 Gy 照射建立放射性肺纤维化小鼠模型,将24只小鼠按分层随机法分为对照组、血清组、照射组和照射+血清组,每组6只。照射后,血清组和照射+血清组小鼠经尾静脉注射100 μl NMS,对照组和照射组小鼠经尾静脉注射100 μl生理盐水,每隔1天注射1次,共计8次,照射后1.5个月和7个月取材检测相关指标。小鼠血清和肺组织中的炎症因子水平采用酶联免疫吸附法检测;肺组织形态学变化和胶原沉积水平通过苏木精-伊红和Masson染色检测;肺中纤维化相关基因的表达水平通过定量逆转录聚合酶链反应检测;肺中纤维化相关蛋白的表达水平通过免疫组织化学和Western Blot检测。计量资料的2组间比较采用t检验。
    结果 与照射组相比,照射+血清组小鼠13 Gy全肺照射1.5个月后的肺脏器系数降低(0.766±0.076)%对(0.649±0.050)%,且差异有统计学意义(t=2.626,P<0.05)。与照射组相比,照射+血清组小鼠肺组织上清液中与肺炎相关炎症因子,转化生长因子(TGF)β、肿瘤坏死因子(TNF)α、白细胞介素(IL)1α、IL-6水平下降(457.100±46.900) pg/ml对(382.500±24.700) pg/ml、(426.200±56.250) pg/ml对(349.600±41.830) pg/ml、(16.680±1.015) pg/ml对(14.550±0.523) pg/ml、(40.650±4.345) pg/ml对(32.830±3.640) pg/ml,同时血清中TGF-β、TNF-α、IL-1α、IL-6的水平亦下降(482.800±41.190) pg/ml对(401.900±53.980) pg/ml、(492.300±38.690) pg/ml对(403.500±39.990) pg/ml、(18.740±2.172) pg/ml对(15.830±1.028) pg/ml、(41.920±2.865) pg/ml对(32.980±3.193) pg/ml,差异均有统计学意义 (t=2.464~5.104,均P<0.05)。与照射组相比,照射+血清组小鼠右肺17 Gy照射7个月后肺组织中胶原沉积和形态学的病理表征显著改善,肺组织中纤维化相关基因Mmp2、Fibronectin、Mcp1、TNF-α、Col1a1、Mmp13(1.862±0.456对1.240±0.245、5.314±2.022对2.681±1.115、3.123±0.573对1.608±0.283、1.975±0.365对1.192±0.352、2.646±0.551对1.473±0.677、2.492±0.444对1.136±0.475)及蛋白Col1a1和α-SMA(2.732±0.612对1.433±0.282、1.888±0.174对1.220±0.372)的表达水平下降,差异均有统计学意义(t=2.550~5.236,均P<0.05)。
    结论 NMS可有效减轻小鼠放射性肺炎和放射性肺纤维化的程度,对小鼠放射性肺损伤有一定的治疗作用,及早对早期放射性肺炎进行干预可有效减轻延缓放射性肺纤维化的进展。

     

    Abstract:
    Objective To investigate the effects of normal mouse serum (NMS) on radiation-induced lung injury (RILI) in mice.
    Methods Radiation pneumonitis models were established by whole-lung irradiation with 13 Gy, and radiation-induced pulmonary fibrosis models were established by right-lung irradiation with 17 Gy in mice. Twenty-four mice were assigned through stratified randomization into control, serum, irradiation, and irradiation+serum groups, with 6 mice in each group. Mice in the serum and irradiation+serum groups received intravenous injections of 100 μl of NMS via the tail vein after irradiation, whereas mice in the control and irradiation groups received 100 μl of normal saline via the same route. Injections were administered every other day for a total of eight doses. Tissue and blood samples were collected at 1.5 and 7 months post-irradiation. Levels of inflammatory factors in mice serum and lung tissue were quantified through enzyme-linked immunosorbent assay kit. Lung morphological changes and collagen deposition were assessed through hematoxylin–eosin and Masson′s trichrome staining. The expression levels of fibrosis-related genes were detected by quantitative reverse transcription polymerase chain reaction, whereas the expression levels of corresponding fibrosis-associated proteins were evaluated through immunohistochemistry and Western Blot analysis. The comparison between two groups of measurement data was conducted using t-test.
    Results The lung organ coefficient in the irradiation+serum group was significantly lower than that in the irradiation group ((0.766±0.076) % vs. (0.649±0.050) %) 1.5 months after 13 Gy whole-lung irradiation, and the difference was statistically significant (t=2.626, P<0.05). The levels of pneumonia-related inflammatory factors, transforming growth factor-β (TGF-β), tumor necrosis factor-α (TNF-α), interleukin (IL)-1α, and IL-6 in the supernatant of mice lung tissue decreased ((457.100±46.900) pg/ml vs. (382.500±24.700) pg/ml; (426.200±56.250) pg/ml vs. (349.600±41.830) pg/ml; (16.680±1.015) pg/ml vs. (14.550±0.523) pg/ml; (40.650±4.345) pg/ml vs. (32.830±3.640) pg/ml); the levels of TGF-β, TNF-α, IL-1α, and IL-6 in the serum also decreased ((482.800±41.190) pg/ml vs. (401.900±53.980) pg/ml; (492.300±38.690) pg/ml vs. (403.500±39.990) pg/ml; (18.740±2.172) pg/ml vs.(15.83±1.028) pg/ml; (41.920±2.865) pg/ml vs. (32.980±3.193) pg/ml), and the differences were statistically significant (t=2.464–5.104; all P<0.05). Furthermore, compared with the irradiation group, the pathological manifestations of collagen deposition and morphological alterations in the right lung tissue of the irradiation+serum group were markedly improved 7 months after 17 Gy irradiation. The expression levels of fibrosis-related genes, including Mmp2, fibronectin, Mcp1, TNF-α, Col1a1, and Mmp13 (1.862±0.456 vs. 1.240±0.245; 5.314±2.022 vs. 2.681±1.115; 3.123±0.573 vs. 1.608±0.283; 1.975±0.365 vs. 1.192±0.352; 2.646±0.551 vs. 1.473±0.677; 2.492±0.444 vs. 1.136±0.475) and the expression levels of fibrosis-related proteins Col1a1 and α-SMA (2.732±0.612 vs. 1.433±0.282; 1.888±0.174 vs. 1.220±0.372) were significantly decreased, and the differences were statistically significant (t=2.550–5.236; all P<0.05).
    Conclusions NMS can effectively alleviate the severity of radiation pneumonia and radiation pulmonary fibrosis in mice, demonstrating a certain therapeutic effect for RILI. Early intervention during the early phase of radiation pneumonitis may effectively reduce and delay the progression of radiation-induced pulmonary fibrosis.

     

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