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中华危重症医学杂志(电子版) ›› 2018, Vol. 11 ›› Issue (01) : 3 -10. doi: 10.3877/cma.j.issn.1674-6880.2018.01.001

所属专题: 文献

论著

转化生长因子β1/Smads通路在百草枯中毒所致上皮-间充质转变和肺纤维化中的作用研究
吴逢选1, 张京臣1, 姜久昆1, 陆远强1,()   
  1. 1. 310003 杭州,浙江大学医学院附属第一医院急诊科
  • 收稿日期:2017-10-20 出版日期:2018-02-01
  • 通信作者: 陆远强
  • 基金资助:
    浙江省科技厅公益技术研究社会发展项目(2015C33146); 浙江省中医药(中西医结合)重点学科建设项目(2017-XK-A36)

Transforming growth factor-β1/Smads signaling pathway involves the development of paraquat-induced pulmonary endothlial-mesenchymal transition and fibrosis

Fengxuan Wu1, Jingchen Zhang1, Jiukun Jiang1, Yuanqiang Lu1,()   

  1. 1. Department of Emergency, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China
  • Received:2017-10-20 Published:2018-02-01
  • Corresponding author: Yuanqiang Lu
  • About author:
    Corresponding author: Lu Yuanqiang, Email:
引用本文:

吴逢选, 张京臣, 姜久昆, 陆远强. 转化生长因子β1/Smads通路在百草枯中毒所致上皮-间充质转变和肺纤维化中的作用研究[J/OL]. 中华危重症医学杂志(电子版), 2018, 11(01): 3-10.

Fengxuan Wu, Jingchen Zhang, Jiukun Jiang, Yuanqiang Lu. Transforming growth factor-β1/Smads signaling pathway involves the development of paraquat-induced pulmonary endothlial-mesenchymal transition and fibrosis[J/OL]. Chinese Journal of Critical Care Medicine(Electronic Edition), 2018, 11(01): 3-10.

目的

本研究通过观察百草枯诱导大鼠肺纤维化中转化生长因子β1(TGF-β1)/Smads信号通路的上皮-间充质转变(EMT)现象及相关蛋白的表达。

方法

将60只雄性大鼠分为对照组及百草枯组,每组各30只。百草枯组腹腔注射80 mg/kg百草枯建立中毒模型,对照组注射3 mL等渗NaCl溶液。分别在建模后1、3、7、14、21、28 d分批处死大鼠,比较两组大鼠肺组织湿干比重及羟脯氨酸水平,取左肺下叶予苏木素-伊红(HE)染色及Masson染色。评估肺组织病理学和肺纤维化程度,采用Western-blotting检测百草枯中毒后1、3、7、14、21、28 d肺组织中EMT相关指标和TGF-β1/Smads信号通路相关蛋白的表达。

结果

两组各时间点肺湿干比重及羟脯氨酸水平的比较,差异有统计学意义(F=9.772、22.541,P均< 0.001),且百草枯组湿干比重的比值在3、7、14、21、28 d均显著高于对照组(P均<0.05),而羟脯氨酸表达水平仅在14、21、28 d明显高于对照组(P均<0.05)。病理结果显示百草枯组大鼠中毒后1~7 d,炎症细胞浸润,肺泡间质增厚水肿,中毒后21 d和28 d可见肺泡壁明显增厚及胶原纤维明显增生,而对照组大鼠肺组织无明显改变。对照组与百草枯组各时间点TGF-β1、Smad2/3、p-Smad2/3、Smad4、p-Smad4、Smad7、p-Smad7、E-cad、α-平滑肌肌动蛋白(α-SMA)、Vimentin和成纤维细胞特异性蛋白1(FSP-1)表达水平的比较,差异均有统计学意义(F=10.685、7.381、7.878、10.743、14.575、17.791、33.200、14.453、10.849、25.415、26.263,P均< 0.001),且百草枯组TGF-β1、Smad2/3、p-Smad2/3、Smad4、p-Smad4、α-SMA、Vimentin及FSP-1表达水平逐渐升高,中毒后21 d达高峰(P均<0.05)。而Smad7、p-Smad7及E-cad表达水平逐渐降低,Smad7、p-Smad7水平在中毒后7 d达到谷底;E-cad表达水平在中毒后21 d达到谷底(P均<0.05)。

结论

TGF-β1/Smads信号通路可能通过调节EMT过程影响百草枯诱导的肺纤维化发生。

Objective

To observe the endothlial-mesenchymal transition (EMT) phenomenon and related protein expressions in transforming growth factor-β1 (TGF-β1)/Smads signaling pathway in lung tissues of the rats with paraquat-induced pulmonary fibrosis.

Methods

Sixty male SD rats were randomly divided into paraquat group (n=30) and control group (n=30). The paraquat poisoning model was established by intraperitoneally injecting with 80 mg/kg of paraquat saline solution in the paraquat group, while rats in the control group were injected with 3 mL of sterile saline. The rats were sacrificed in batches at 1, 3, 7, 14, 21, 28 d after modeling. The wet/dry ratio of left lung tissue and the expression of hydroxyproline were calculated between these two groups. The pathology and pulmonary fibrosis in the inferior lobe of left lung were assessed by hematoxylin-eosin (HE) staining and Masson staining. The expressions of related proteins in EMT and TGF-β1/Smads signaling pathway in lung tissues on days 1, 3, 7, 14, 21, 28 after paraquat poisoning were analyzed by Western-blotting.

Results

The wet/dry ratio and hydroxyproline both showed significant differences between two groups at different time points after modeling (F=9.772, 22.541, both P < 0.001). In the paraquat group, the wet/dry ratios at 3, 7, 14, 21, 28 d were much higher than those in the control group (all P<0.05), and the levels of hydroxyproline only increased at 14, 21, 28 d as compared with those in the control group (all P<0.05). Pathological findings indicated that inflammatory cell infiltrated, alveolar interstitium thickened in the paraquat group at 1, 3, 7 d, and alveolar walls thickened gradually and a large amount of collagen fibers proliferated at 21, 28 d after modeling. There was no significant change in lung tissue of the control group. The expressions of TGF-β1, Smad2/3, p-Smad2/3, Smad4, p-Smad4, Smad7, p-Smad7, E-cadherin (E-cad), α-smooth muscle actin (α-SMA), Vimentin and fibroblast specific protein-1 (FSP-1) between the two groups at each time point all showed significant differences (F=10.685, 7.381, 7.878, 10.743, 14.575, 17.791, 33.200, 14.453, 10.849, 25.415, 26.263, all P < 0.001). The expressions of TGF-β1, Smad2/3, p-Smad2/3, Smad4, p-Smad4, α-SMA, Vimentin and FSP-1 gradually increased and reached a peak at 21 d after paraquat poisoning, while the Smad7, p-Smad7 decreased and reached a bottom at 7 d in the paraquat group (all P<0.05). The expression of E-cad in the paraquat group reduced from 3 d to 21 d (all P<0.05).

Conclusion

The TGF-β1/Smads signaling pathway may affect the development of PQ-induced pulmonary fibrosis by regulating EMT process.

表1 两组大鼠建模后各时间点肺组织肺湿干比重的变化(±s
图1 两组大鼠建模后不同时间点的病理表现
表2 两组大鼠不同时间点羟脯氨酸水平变化的变化(mg/L,±s
图2 Western-blotting法测两组大鼠各时间点TGF-β1/Smads通道相关蛋白表达的比较
图3 两组大鼠各时间点上皮-间充质转变相关蛋白表达的比较
[1]
Lee EY, Hwang KY, Yang JO, et al. Predictors of survival after acute paraquat poisoning[J]. Toxicol Ind Health, 2002, 18 (18): 201-206.
[2]
Dinisoliveira RJ, Duarte JA, Sáncheznavarro A, et al. Paraquat poisonings: mechanisms of lung toxicity, clinical features, and treatment[J]. Crit Rev Toxicol, 2008, 38 (1): 13-71.
[3]
Shimada H, Hirai K, Simamura E, et al. Paraquat toxicity induced by voltage-dependent anion channel 1 acts as an NADH-dependent oxidoreductase[J]. J Biol Chem, 2009, 284 (42): 28642.
[4]
Huang M, Wang Y, Zhu L, et al. MAPK pathway mediates epithelial-mesenchymal transition induced by paraquat in alveolar epithelial cells[J]. Environ Toxicol, 2015, 31 (11): 1407-1414.
[5]
Zhu Y, Tan J, Xie H, et al. HIF-1α regulates EMT via the Snail and β-catenin pathways in paraquat poisoning-induced early pulmonary fibrosis[J]. J Cell Mol Med, 2016, 20 (4): 688.
[6]
Wang J, Zhu Y, Tan J, et al. Lysyl oxidase promotes epithelial-to-mesenchymal transition during paraquat-induced pulmonary fibrosis[J]. Mol Biosyst, 2015, 12 (2): 499-507.
[7]
Hu L, Yu Y, Huang H, et al. Epigenetic regulation of interleukin 6 by histone acetylation in macrophages and its role in paraquat-induced pulmonary fibrosis[J]. Front Immunol, 2016 (7): 696.
[8]
Xie H, Tan JT, Wang RL, et al. Expression and si-gnificance of HIF-1α in pulmonary fibrosis induced by paraquat[J]. Exp Biol Med (Maywood), 2013, 238 (9): 1062-1068.
[9]
Yamada A, Aki T, Unuma K, et al. Paraquat induces epithelial-mesenchymal transition-like cellular response resulting in fibrogenesis and the prevention of apoptosis in human pulmonary epithelial Cells[J]. PLoS One,2015, 10 (3): e0120192.
[10]
Han Y, Shen P, Chang W. Involvement of epithelial-to-mesenchymal transition and associated transforming growth factor-β/Smad signaling in paraquat-induced pulmonary fibrosis[J]. Mol Med Rep, 2015, 12 (6): 7979-7984.
[11]
Chen CY, Peng WH, Wu LC, et al. Luteolin ame-liorates experimental lung fibrosis both in vivo and in vitro: implications for therapy of lung fibrosis[J]. J Agric Food Chem, 2010, 58 (22): 11653-11661.
[12]
Song JS, Kang CM, Rhee CK, et al. Effects of el-astase inhibitor on the epithelial cell apoptosis in bleomycin-induced pulmonary fibrosis[J]. Exp Lung Res, 2009, 35 (10): 817.
[13]
Wang HR, Pan J, Shang AD, et al. Time-dependent haemoperfusion after acute paraquat poisoning[J]. Sci Rep, 2017, 7 (1): 2239.
[14]
Shang AD, Lu YQ. A case report of severe paraquat poisoning in an HIV-positive patient: an unexpected outcome and inspiration[J]. Medicine (Baltimore), 2015, 94 (8): e587.
[15]
Zhang Q, Wu WZ, Lu YQ, et al. Successful treatment of patients with paraquat intoxication: three case reports and review of the literature[J]. J Zhejiang Univ Sci B, 2012, 13 (5): 413-418.
[16]
Huang WD, Wang JZ, Lu YQ, et al. Lysine acetylsa-licylate ameliorates lung injury in rats acutely exposed to paraquat[J]. Chin Med J (Engl), 2011, 124 (16): 2496-2501.
[17]
Zhu Y, Wang J, Meng X, et al. A positive feedback loop promotes HIF-1α stability through miR-210-mediated suppression of RUNX3 in paraquat-induced EMT[J]. J Cell Mol Med, 2017, 21 (12): 3529-3539.
[18]
Fang Z, Zhang YZ, Cai T, et al. Expression of transf-orming growth factor-β1 and its receptors in peripheral blood of patients with immune thrombocytopenic purpura[J]. Zhongguo Shi Yan Xue Ye Xue Za Zhi, 2012, 20 (3): 664-666.
[19]
Ma J, Bishoff B, Mercer RR, et al. Role of epithelial-mesenchymal transition (EMT) and fibroblast function in cerium oxide nanoparticles-induced lung fibrosis[J]. Toxicol Appl Pharmacol, 2017 (323): 16-25.
[20]
Nirajan S, Lokendra C, Han MK, et al. Glutamine in-hibits CCl4 induced liver fibrosis in mice and TGF-β1 mediated epithelial-mesenchymal transition in mouse hepatocytes[J]. Food Chem Toxicol, 2016 (93): 129-137.
[21]
Kim H, Lee SW, Baek KM, et al. Continuous hypoxia attenuates paraquat-induced cytotoxicity in the human A549 lung carcinoma cell line[J]. Exp Mol Med, 2011, 43 (9): 494-500.
[22]
Qian J, Ye Y, Lv L, et al. FTY720 attenuates paraq-uat-induced lung injury in mice[J]. Int Immunopharmacol, 2014, 21 (2): 426-431.
[23]
Song X, Liu W, Xie S, et al. All-transretinoic acid ameliorates bleomycin-induced lung fibrosis by downregulating the TGF-β1/Smad3 signaling pathway in rats[J]. Lab Invest, 2013, 93 (11): 1219-1231.
[24]
Yu N, Sun YT, Su XM, et al. Melatonin attenuates TGFβ1-induced epithelial-mesenchymal transition in lung alveolar epithelial cells[J]. Mol Med Rep, 2016, 14 (6): 5567-5572.
[25]
Ding NH, Li JJ, Sun LQ. Molecular mechanisms and treatment of radiation-induced lung fibrosis[J]. Curr Drug Targets, 2013, 14 (11): 1347-1356.
[26]
Wang Y, Li X L, An GL, et al. SB203580 inhibits epithelialmesenchymal transition and pulmonary fibrosis in a rat silicosis model[J]. Toxicology Letters, 2016 (259): 28-34.
[27]
Tao Z, Wei Z, Min X, et al. Protective role of an-drographolide in bleomycin-induced pulmonary fibrosis in mice[J]. Int J Mol Sci, 2013, 14 (12): 23581.
[28]
Zhao YL, Zhu RT, Sun YL. Epithelial-mesenchymal transition in liver fibrosis[J]. Biomed Rep, 2016, 4 (3): 269-274.
[29]
Lovisa S, Zeisberg M, Kalluri R. Partial epithelial-to-mesenchymal transition and other new mechanisms of kidney fibrosis[J]. Trends Endocrinol Metab, 2016, 27 (10): 681.
[30]
Jeon YJ, Jung N, Park JW, et al. Epithelial-mesen-chymal transition in kidney tubular epithelial cells induced by globotriaosylsphingosine and globotriaosylceramide[J]. PLoS One, 2015, 10 (8): e0136442.
[31]
Zhou Y, He Z, Gao Y, et al. Induced pluripotent stem cells Inhibit bleomycin-induced pulmonary fibrosis in mice through suppressing TGF-β1/Smad-mediated epithelial to mesenchymal transition[J]. Front Pharmacol, 2016 (7): 430.
[32]
Hua XF, Li XH, Li MM, et al. Doxycycline attenuates paraquat-induced pulmonary fibrosis by downregulating the TGF-β signaling pathway[J]. J Thorac Dis, 2017, 9 (11): 4376-4386.
[33]
Chen CM, Chou HC, Hsu HH, et al. Transforming growth factor-beta1 upregulation is independent of angiotensin in paraquat-induced lung fibrosis[J]. Toxicology, 2005, 216 (2-3): 181-187.
[34]
Zhang Z, Ding L, Wu L, et al. Salidroside alleviates paraquat-induced rat acute lung injury by repressing TGF-β1 expression[J]. Int J Clin Exp Pathol, 2014, 7 (12): 8841.
[35]
Xie L, Zhou D, Xiong J, et al. Paraquat induce pulmonary epithelial-mesenchymal transition through transforming growth factor-α1-dependent mechanism[J]. Exp Toxicol Pathol, 2016, 68 (1): 69-76.
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