Lu Zhonghua, Ma Yan, Zhang Shuyu, Liu Fenju, Wan Meizhen, Luo Judong
Department of Radiotherapy, Changzhou Tumor Hospital, Soochow University, Changzhou, Jiangsu 213001, P.R. China.
Jiangsu Provincial Key Laboratory of Radiation Medicine and Protection, School of Radiation Medicine and Protection, Soochow University, Suzhou, Jiangsu 215123, P.R. China.
Mol Med Rep. 2015 Mar;11(3):2055-61. doi: 10.3892/mmr.2014.2923. Epub 2014 Nov 11.
In the present study, a human transforming growth factor‑β1 (TGF‑β1) small interfering RNA (siRNA) plasmid vector (TGF‑β1‑siRNA) was constructed to investigate its effects on the proliferation and differentiation of human lung fibroblasts in vitro and its interference effects on radiation‑induced lung injury in vivo. Reverse transcription quantitative polymerase chain reaction and enzyme linked immunosorbent assay revealed that the mRNA and protein expression of TGF‑β1 in the HFL‑I cells were inhibited by TGF‑β1‑siRNA and flow cytometry demonstrated a significant increase in apoptosis of the HFL‑I cells. Adult, female, specific‑pathogen‑free C57BL/6 mice were used in the in vivo animal investigations and were randomly divided into the four following groups: control without any treatment, radiation alone, radiation followed by empty vector transfection and radiation followed by TGF‑β1‑siRNA vector transfection. Hematoxylin and eosin and Van‑Gieson staining revealed that certain radiation‑induced histopathological changes of the lung, including inflammation, edema, the density of surface pulmonary interstitial collagen fibers in the alveolar septum, TGF‑β1‑positive reactions in alveolar epithelial cells and pulmonary interstitial macrophages were less marked in the mice transfected with TGF‑β1‑siRNA compared with the mice without transfection or those transfected with empty vectors. The serum levels of TGF‑β1 levels in the irradiated mice increased significantly at four weeks and peaked at eight weeks after radiation, compared with the control. Serum levels of TGF‑β1 in the irradiated mice transfected with TGF‑β1‑siRNA also increased gradually and a significant difference was observed compared with those irradiated without transfection. The mRNA expression levels of TGF‑β1 in the mice transfected with TGF‑β1‑siRNA were markedly lower compared with those of the other radiation groups. The present study suggested that the TGF‑β1‑siRNA vector reduced the activity of TGF‑β1 by downregulating the mRNA expression of TGF‑β1 and thereby effectively suppressing inflammatory reactions and defending against radiation‑induced lung injury.
在本研究中,构建了一种人转化生长因子-β1(TGF-β1)小干扰RNA(siRNA)质粒载体(TGF-β1-siRNA),以研究其对人肺成纤维细胞体外增殖和分化的影响以及对体内辐射诱导的肺损伤的干扰作用。逆转录定量聚合酶链反应和酶联免疫吸附测定显示,TGF-β1-siRNA抑制了HFL-I细胞中TGF-β1的mRNA和蛋白表达,流式细胞术表明HFL-I细胞的凋亡显著增加。成年雌性无特定病原体C57BL/6小鼠用于体内动物研究,并随机分为以下四组:未进行任何处理的对照组、单纯辐射组、辐射后空载体转染组和辐射后TGF-β1-siRNA载体转染组。苏木精-伊红染色和Van-Gieson染色显示,与未转染或转染空载体的小鼠相比,转染TGF-β1-siRNA的小鼠中某些辐射诱导的肺部组织病理学变化,包括炎症、水肿、肺泡间隔表面肺间质胶原纤维密度、肺泡上皮细胞和肺间质巨噬细胞中的TGF-β1阳性反应,均不那么明显。与对照组相比,照射小鼠血清中TGF-β1水平在辐射后4周显著升高,并在8周达到峰值。转染TGF-β1-siRNA的照射小鼠血清中TGF-β1水平也逐渐升高,与未转染照射的小鼠相比观察到显著差异。转染TGF-β1-siRNA的小鼠中TGF-β1的mRNA表达水平明显低于其他辐射组。本研究表明,TGF-β1-siRNA载体通过下调TGF-β1的mRNA表达降低了TGF-β1的活性,从而有效抑制炎症反应并抵御辐射诱导的肺损伤。