Blocking TG2 attenuates bleomycin-induced pulmonary fibrosis in mice through inhibiting EMT
Introduction
Lung fibrosis, highlighted with the parenchymal cell reduction and fiber connective tissue hyperplasia, progressively causes the structural destruction and function decrease of the lung tissue, which results in lung failure and severely threatens patients' health. Worldwide, tissue fibrosis is a main cause of major morbidity and mortality. Concerned statistic data indicates a high mortality in lung fibrosis patients, with only 2–5 years survival after the diagnosis of idiopathic pulmonary fibrosis and 5-year survival rate low to 20 %. Besides, the mechanism of the development of lung fibrosis is very complicated (Raghu et al., 2006). Previous reports have suggested that dysregulation of inflammatory factors (Goodwin and Jenkins, 2009; Xin et al., 2019), extracellular matrix density increase (Shinde et al., 2017), and oxidative stress (Yang et al., 2018) were involved in this process. In recent years, the role of epithelial-mesenchymal transition (EMT) in lung fibrosis was widely studied. EMT is a process by which epithelial cells lose their cell polarity and cell-cell adhesion, acquire migratory and invasive properties, and finally turn into mesenchymal cells. And most researches showed that alveolar epithelial cells underwent EMT during the development of lung fibrosis, leading to an increase of Interpulmonary fibers (Li et al., 2017; Ma et al., 2017; Rout-Pitt et al., 2018). Thus, EMT was thought to be pivotal to the occurrence and development of pulmonary fibrosis and finding the key targets regulating EMT is of great significance for the treatment of pulmonary fibrosis.
Transglutaminase is widely expressed in various tissues of mammals, among which Transglutaminase 2(TG2) has been most studied and its function is the most important. Recent studies showed that TG2 was involved in many diseases such as neurodegenerative diseases, autoimmune diseases, etc (Lai and Greenberg, 2013). Notably, TG2 was also reported to play an important role in the occurrence and development of pulmonary fibrosis (Olsen et al., 2011). However, it is still unknown whether it has a regulatory effect on EMT during the process of pulmonary fibrosis. Therefore, this study aimed to investigate the role of TG2 in pulmonary fibrosis and EMT in the bleomycin-treated mice and try to reveal its molecular mechanism.
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Animal experiments
6-8 weeks old male C57BL/6 mice were intraperitoneally injected with bleomycin (Sigma, product number BP971) at 1.5 mg/kg once every 2 days for 1 week. Lung tissues of the mice were collected for pathological examination at 1, 2 and 3 weeks after drug withdrawal to confirm the establishment of pulmonary fibrosis model.
18 mice were randomly divided into3 groups: #control group (intraperitoneal injection of normal saline at 10 mL/kg), #model group (intraperitoneal injection of bleomycin at 1.5
Pulmonary fibrosis was induced by bleomycin in mice
Mice were intraperitoneally injected with bleomycin at 1.5 mg/kg every 2 days for 1 week. Mouse lung tissues were then subjected to H&E staining and Masson staining at 1week, 2 weeks, and 3 weeks post bleomycin induction. H&E staining data showed that mice lungs appeared thick septa with infiltration of immune cells at 2nd week post bleomycin treatment, which came to the most significant at 3rd week (Fig. 1A, B). In addition, Masson staining results showed that no significant positive area was
Discussion
EMT is most studied in pulmonary fibrosis (Lai and Greenberg, 2013). After suffering hypoxia and inflammation, epithelial cells lose their cell polarity and cell-cell adhesion, gradually lose the epithelial morphology, and gain the characteristics of the mesenchymal cells (Higgins et al., 2007). During EMT process, the expression of epithelial marker E-cadherin decreases while those of mesenchymal markers such as α-SMA, Vimentin is upregulated, which induces cytoskeleton reorganization into
Declaration of Competing Interest
The authors declare that they have no competing interests.
Acknowledgements
This work was supported by the grants from National Natural Science Foundation of China (Grant number 81700078) and Natural Science Foundation of Jiangsu Province (Grant number BK20171172)
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These authors equally contributed to this article.