Dose-dependent induction of transforming growth factor β (TGF-β) in the lung tissue of fibrosis-prone mice after thoracic irradiation
Dose-dependent induction of transforming growth factor β (TGF-β) in the lung tissue of fibrosis-prone mice after thoracic irradiation
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DOI：
10.1016/s0360-3016(00)00482-x
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发表时间：
2000-07-01
影响因子：
7
通讯作者：
Rübe, C
中科院分区：
文献类型：
--
作者：
Rübe, CE;Uthe, D;Rübe, C
目的：肺是胸部区域癌症放疗的主要剂量限制器官。辐射诱导的肺损伤在分子水平的发病机制仍不清楚。照射后即刻的细胞损伤被认为会导致细胞因子介导的多细胞相互作用，从而引发和促进纤维化组织反应。本研究的目的是在使用对纤维化敏感的C57BL/6小鼠的肺损伤模型中，评估辐射对转化生长因子β（TGF -β）基因表达的急性和长期影响。
方法和材料：分别用6戈瑞和12戈瑞对C57BL/6小鼠的胸部进行照射。在对应于潜伏期（照射后1、3、6、12、24、48、72小时和1周）、肺炎期（照射后2、4、8和16周）以及纤维化期开始（照射后21周）的时间点，处死接受处理的小鼠和假照射的对照小鼠。通过聚合酶链反应（PCR）、免疫组织化学和光学显微镜相结合的方法，对每个剂量和时间点的三只不同小鼠的肺组织进行分析。通过竞争性逆转录酶/聚合酶链反应（RT - PCR）对TGF -β的mRNA表达进行定量；通过免疫组织化学染色（碱性磷酸酶 - 抗碱性磷酸酶[APAAP]）确定TGF -β蛋白的细胞来源。将细胞因子在mRNA和蛋白水平的表达与组织病理学改变相关联。
结果：在用12戈瑞单剂量胸部照射后，肺组织中辐射诱导的TGF -β释放在最初几小时（照射后1、3和6小时）内就已明显，并在12小时后显著增加；随后（照射后48小时、72小时和1周）TGF -β表达下降至基础水平。在肺炎期开始时，照射介导的TGF -β释放刺激在2周和4周达到最大值。已发现潜伏期TGF -β mRNA水平升高与肺泡巨噬细胞的免疫组织化学染色相关。在肺炎的急性期，TGF -β免疫反应性增加最为显著。在整个观察期间，II型肺泡上皮细胞和成纤维细胞（除炎症细胞外）是TGF -β表达的重要来源。在组织病理学辐射损伤区域显著检测到TGF -β表达增加。在暴露于6戈瑞单剂量辐射后，肺组织仅显示出轻微的辐射介导的TGF -β mRNA反应。这种适度的上调在照射后6小时至48小时范围内。与6戈瑞胸部照射后仅有的轻微组织病理学变化相对应，在后期时间点对TGF -β mRNA水平的测量显示，与未处理的对照小鼠相比没有显著变化。
结论：本研究表明，在用12戈瑞胸部照射后，肺组织中TGF -β的表达出现急性和长期的增加。TGF -β在炎症细胞浸润和纤维化区域的主要定位表明该细胞因子参与了辐射诱导的肺纤维化的发病机制。应进行进一步的研究以探索其他细胞因子在辐射损伤发展中的作用。对肺纤维化潜在机制的更好理解最终可能导致在分子水平进行调节性干预以改变纤维化过程。（C）2000爱思唯尔科学公司
Purpose: The lung is the major dose-limiting organ for radiotherapy of cancer in the thoracic region. The pathogenesis of radiation-induced lung injury at the molecular level is still unclear. Immediate cellular damage after irradiation is supposed to result in cytokine-mediated multicellular interactions with induction and progression of fibrotic tissue reactions. The purpose of this investigation was to evaluate the acute and long-term effects of radiation on the gene expression of transforming growth factor beta (TGF-beta) in a model of lung injury using fibrosis-sensitive C57BL/6 mice.Methods and Materials: The thoraces of C57BL/6 mice were irradiated with 6 and 12 Gy, respectively. Treated and sham-irradiated control mice mere sacrificed at times corresponding to the latent period (1, 3, 6, 12, 24, 48, 72 hours and 1 week postirradiation), the pneumonic phase (2, 4, 8, and 16 weeks postirradiation), and the beginning of the fibrotic phase (2 1 weeks postirradiation), The lung tissue from three different mice per dosage and time point was analyzed by a combination of polymerase chain reaction (PCR), immunohistochemistry, and light microscopy. The mRNA expression of TGF-beta was quantified by competitive reverse transcriptase/polymerase chain reaction (RT-PCR); the cellular origin of the TGF-beta protein was identified by immunohistochemical staining (alkaline phosphatase-anti-alkaline phosphatase [APAAP]). The cytokine expression on mRNA and protein level was correlated with the histopathological alterations.Results: Following thoracic irradiation with a single dose of 12 Gy, radiation-induced TGF-beta release in lung tissue was appreciable already within the first hours (1, 3, and 6 hours postirradiation) and reached a significant increase after 12 hours; subsequently (48 hours, 72 hours, and 1 week postirradiation) the TGF-beta expression declined to basal levels. At the beginning of the pneumonic phase, irradiation-mediated stimulation of TGF-beta release reached maximal values at 2 and 4 weeks. The elevated levels of TGF-beta mRNA during the latent phase have been found to correlate with immunohistochemical staining of alveolar macrophages. The most striking increase in TGF-beta immunoreactivity was seen during the acute phase of pneumonitis. Throughout this observation period, type II pneumocytes and fibroblasts (apart from inflammatory cells) served as important sources of TGF-beta expression. Increased TGF-beta expression was detected prominently in regions of histopathologic radiation injury. After exposure to a single radiation dose of 6 Gy, the lung tissue revealed only a minor radiation-mediated TGF-beta mRNA response. The modest upregulation ranged from 6 hours to 48 hours after irradiation. Corresponding to the only minor histopathologic changes after thoracic irradiation with 6 Gy, measurement of TGF-beta mRNA levels during the later time points revealed no significant alterations in comparison to untreated control mice.Conclusions: This study demonstrates an acute and long-lasting increase in the expression of TGF-beta in lung tissue following thoracic irradiation with 12 Gy. The predominant localization of TGF-beta in areas of inflammatory cell infiltrates and fibrosis suggests involvement of this cytokine in the pathogenesis of radiation-induced pulmonal fibrosis. Further studies should be performed to explore the role of other cytokines in the development of radiation injury. An improved understanding of the underlying mechanisms of pulmonary fibrosis may eventually lead to modulatory intervention at the molecular level to modify the fibrotic process. (C) 2000 Elsevier Science Inc.