| Objective: Silicosis is an irreversible occupational lung disease.In addition to traditional industries,there is a large amount of free silica that is closely related to the incidence of silicosis in contemporary industries.For example,in the production of jeans,artificial stone countertops,jewelry polishing,etc.,the failure to fully understand and prevent the risk factors of these industries has led to the reappearance of silicosis worldwide.Common features of silicosis are alveolar epithelial cell damage and proliferation,persistent inflammation,accumulation of extracellular matrix(ECM)in the interstitial tissue,eventually leading to permanent deformation and fibrosis,reducing the quality of life.Therefore,breakthrough from multiple angles is very important to find the mechanism of silicosis fibrosis.In recent,studies have shown that silica stimulation can transform epithelial cells into myofibroblasts by means of epithelial-mesenchymal transition(EMT).It is the main terminal effector cell in fibrosis and plays a fibro genic role.In this study,we established experimental silicosis mouse models to observe the occurrence of EMT and fibrosis in the lung tissue,and the regulatory role of galectin-3(Gal-3)in this process.Then,the Gal-3 inhibitor TD139 was given to find out the specific mechanism of Gal-3 regulating fibrosis,so as to provide new targets for the study of silicosis fibrosis.Methods: In this study,KM mice used to establish experimental subjects.The method of non-exposed intratracheal instillation was used,respectively instilled silica suspension or saline.Firstly,there were two time points,14 days and 28 days,including saline group and silica group.After 28 days,Gal-3 inhibitor TD139 was administered.There were four groups: saline+vehicle group,saline+TD139 group,silica+vehicle group,and silica+TD139 group,killed on 42 and 56 days.We used alveolar lavage fluid(BALF)inflammatory cell counting method,H&E staining,and Masson staining to detect pulmonary pathological changes.Western Blot technology was applied to detect the expression of EMT-related proteins,including epithelial marker E-cadherin,interstitial marker Vimentin,?-SMA,and protein Gal-3,in animal models at various time points.Immunohistochemistry was applied to detect the distribution of Gal-3 and act-?-catenin in lung tissue.Western Blot also was used to detect the expression of act-?-catenin,?-catenin,GSK-3?,and P-GSK-3.Results: 1.Silica expose can induce EMT and fibrosis in mice.The count of inflammatory cells in BALF showed that the number of inflammatory cells in silica group were higher than saline group both at 14 and 28 days,with the statistically significant.H&E staining of lung paraffin sections showed that in 14,28 days,mice in silica group had a significant inflammatory response,the complete alveolar structure was destroyed with alveolar wall thickening.There was no significant change in the saline group.Masson trichrome staining showed that no significant blue collagen deposits appeared in silica and saline groups in 14.In 28 days,a small amount of blue collagen was deposited in silica group but there was no significant change in the saline group.Western Blot results showed that compared with the control group,in 14 days,silica entered the lung tissue without causing changes in EMT-related proteins;in 28 days,the expression of E-cadherin decreased,and the expression of Vimentin and ?-SMA increased,which was statistically significant.In 14 days and 28 days,compared with saline group,the expression of Gal-3 protein was increased in silica group.At the same time,immunohistochemical showed the same results.2.Inhibition of Gal-3 can alleviates silica induced EMT and fibrosis in mice.After the treatment with TD139 and compared with saline+vehicle group,H&E results found that there was only slight inflammation but no obvious alveolar structural damage in saline+TD139 group.Compared with saline+vehicle group and saline+TD139 group,the lungs of silica+vehicle group and silica+TD139 group in 42,56 days appeared obvious fibrous nodules with infiltration of inflammatory cells,thickening of alveolar walls and small blood vessel walls.However,the silicon nodules became smaller and the degree of fibrosis was reduced in the silica+TD139 group.Masson trichrome staining showed at the 42,56 days,there were no significant blue collagen distribution in saline+vehicle group and saline+TD139 group;in contrast,there appeared obvious collagen distribution in silica+vehicle group and silica+TD139 group.Compared with silica+vehicle group,the blue collagen deposition was significantly reduced,and the alveolar cavity was partially fused in silica+ TD139 group.In addition,the results of Western blot showed that,TD139 can effectively inhibit the expression of Gal-3.And,compared with silica+vehicle group,the expression of epithelial marker protein E-cadherin was significantly increased,and the expression of mesenchymal marker proteins Vimentin and ?-SMA decreased after Gal-3 inhibition,which was statistically significant.3.Gal-3 promotes lung EMT and fibrosis by regulating the GSK-3?/?-catenin signaling pathway.At 14,28,42,and 56 days,compared with the saline or saline+vehicle group,the expression of act-?-catenin and p-GSK-3? protein in silica or silica+vehicle group increased with statistical significance.While the expression of total GSK-3? and ?-catenin did not change.After administration of TD139,in 42 and 56 days,in silica+TD139 group the expression of act-?-catenin and p-GSK-3? protein was significantly reduced,and the expression of total GSK-3? and ?-catenin did not change compared with silica+vehicle group.Conclusion: 1.Silica expose can induce EMT and fibrosis in KM mice.2.Inhibition of Gal-3 can alleviates silica induced EMT and fibrosis.3.Gal-3 promotes silica induced EMT and fibrosis by regulating the GSK-3?/?-catenin signaling pathway. |
PDF Full Text Request