Silica-induced Epithelial-mesenchymal Transition Research Articles

CircPVT1 promotes silica-induced epithelial-mesenchymal transition by modulating the miR-497-5p/TCF3 axis.

Epithelial-mesenchymal transition (EMT) is a vital pathological feature of silica-induced pulmonary fibrosis. However, whether circRNA is involved in the process remains unclear. The present study aimed to investigate the role of circPVT1 in the silica-induced EMT and the underlying mechanisms. We found that an elevated expression of circPVT1 promoted EMT and enhanced the migratory capacity of silica-treated epithelial cells. The isolation of cytoplasmic and nuclear separation assay showed that circPVT1 was predominantly expressed in the cytoplasm. RNA immunoprecipitation assay and RNA pull-down experiment indicated that cytoplasmic-localized circPVT1 was capable of binding to miR-497-5p. Furthermore, we found that miR-497-5p attenuated the silica-induced EMT process by targeting transcription factor 3 (TCF3), an E-cadherin transcriptional repressor, in the silica-treated epithelial cells. Collectively, these results reveal a novel role of the circPVT1/miR-497-5p/TCF3 axis in the silica-induced EMT process in lung epithelial cells. Once validated, this finding may provide a potential theoretical basis for the development of interventions and treatments for pulmonary fibrosis.

Matrix stiffness regulates α-TAT1-mediated acetylation of α-tubulin and promotes silica-induced epithelial-mesenchymal transition via DNA damage.

Silicosis is characterized by silica exposure-induced lung interstitial fibrosis and formation of silicotic nodules, resulting in lung stiffening. The acetylation of microtubules mediated by α-tubulin N-acetyltransferase 1 (α-TAT1) is a posttranslational modification that promotes microtubule stability in response to mechanical stimulation. α-TAT1 and downstream acetylated α-tubulin (Ac-α-Tub) are decreased in silicosis, promoting the epithelial-mesenchymal transition (EMT); however, the underlying mechanisms are unknown. We found that silica, matrix stiffening or their combination triggered Ac-α-Tub downregulation in alveolar epithelial cells, followed by DNA damage and replication stress. α-TAT1 elevated Ac-α-Tub to limit replication stress and the EMT via trafficking of p53-binding protein 1 (53BP1, also known as TP53BP1). The results provide evidence that α-TAT1 and Ac-α-Tub inhibit the EMT and silicosis fibrosis by preventing 53BP1 mislocalization and relieving DNA damage. This study provides insight into how the cell cycle is regulated during the EMT and why the decrease in α-TAT1 and Ac-α-Tub promotes silicosis fibrosis.This article has an associated First Person interview with the first authors of the paper.

MiR-34a-5p Attenuates EMT through targeting SMAD4 in silica-induced pulmonary fibrosis.

Silicosis is an incurable occupational disease, and its pathological feature is diffuse pulmonary fibrosis. Pulmonary epithelial‐mesenchymal transition (EMT) is one of the important events in the pathogenesis of silicosis. Previous studies found that abnormal expression of various microRNAs (miRNAs) involved in the development of lung fibrosis. However, their roles in silicosis have not been elucidated. To research the biological effects of miR‐34a in EMT process in silica‐induced lung fibrosis, we established the silicosis model in mouse and miR‐34a intervention in a cell model of TGF‐β1 stimulated lung epithelial cells (A549). The results showed that miR‐34a expression was down‐regulated in the fibrotic lung tissue after silica treatment, and it was similarly expressed in A549 cells stimulated by TGF‐β1. Meanwhile, silica‐induced EMT process can increase expression of two mesenchymal markers, α‐SMA and vimentin. Furthermore, overexpression miR‐34a markedly inhibited EMT stimulated by TGF‐β1. Mechanistically, SMAD4 was identified as the target of miR‐34a. SMAD4 levels decreased in mRNA and protein levels in A549 cells upon miR‐34a overexpression. In addition, the knockdown of SMAD4 blocked the EMT process. Taken together, miR‐34a regulated EMT, which might be partially realized by targeting SMAD4. Our data might provide new insight into treatment targets for silica‐induced pulmonary fibrosis.

Tanshinone IIA attenuates silica-induced pulmonary fibrosis via Nrf2-mediated inhibition of EMT and TGF-β1/Smad signaling

Silicosis is an occupational pulmonary fibrosis that is caused by inhalation of silica (SiO2), and there are no effective drugs to treat this disease. Tanshinone IIA (Tan IIA), a natural product, has been reported to possess antioxidant and anti-fibrotic properties in various diseases. The purpose of the current study was to examine Tan IIA's protective effects against silica-induced pulmonary fibrosis and to explore the underlying mechanisms. We found that in vivo treatment with Tan IIA significantly relieved silica-induced lung fibrosis in a silicosis rat model by histological and immunohistochemical analyses. Further, in vitro mechanistic investigations, mainly using western blot and immunofluorescence analyses, revealed that Tan IIA administration markedly inhibited the silica-induced epithelial-mesenchymal transition (EMT) and transforming growth factor-β1 (TGF-β1)/Smad signaling pathway and also reduced silica-induced oxidative stress and activated the nuclear factor erythroid 2-related factor-2 (Nrf2) signaling pathway in A549 and human bronchial epithelial (HBE) cells. Furthermore, through transfection with siRNA, we demonstrate that Nrf2 activation partially mediates the suppression effects of Tan IIA on EMT and TGF-β1/Smad signaling pathway activation induced by silica exposure, thus mediating the anti-fibrotic effects of Tan IIA against silica-induced pulmonary fibrosis. In our study, Tan IIA has been identified as a possible anti-oxidative and anti-fibrotic drug for silicosis.

MiR-let-7d attenuates EMT by targeting HMGA2 in silica-induced pulmonary fibrosis.

Silicosis is a serious occupational disease characterized by pulmonary chronic inflammation and progressive fibrosis. Epithelial-mesenchymal transition (EMT) of alveolar epithelial cells plays a vital role in silicosis. Recent studies discovered a variety of microRNAs (miRNAs) participating in fibrotic diseases. Here, we aimed to explore the function and mechanism of miRNA let-7d in the EMT process in silica-induced alveolar epithelial cells. To detect whether let-7d and its target HMGA2 were involved in silica-induced EMT, we established a silicosis mouse model and found that let-7d was down-regulated and HMGA2 was up-regulated in the silica-treated group. Then we applied an in vitro co-culture system to imitate the EMT process in A549 cells after silica treatment. The down-regulation of let-7d and up-regulation of HMGA2 were also observed in vitro. The knockdown of HMGA2 significantly inhibited the silica-induced EMT. Furthermore, we found that overexpression of let-7d could reduce the expression of HMGA2 and consequently inhibited the silica-induced EMT, whereas inhibition of let-7d increased the expression of HMGA2 and promoted the silica-induced EMT. In conclusion, let-7d negatively regulated silica-induced EMT and inhibited silica-induced pulmonary fibrosis, which might be partially realized by directly binding to HMGA2. Our data suggested that miRNA let-7d might have a potential protective effect in the fibrotic process and become a new therapeutic target for silicosis or other fibrotic diseases.

MicroRNA-29b Mediates Lung Mesenchymal-Epithelial Transition and Prevents Lung Fibrosis in the Silicosis Model

Lung epithelial-mesenchymal transition (EMT) plays an important role in silicosis fibrosis. The reverse process of EMT is mesenchymal-epithelial transition (MET), which is viewed as an anti-EMT therapy and is a good target toward fibrosis. MicroRNAs (miRNAs) have emerged as potent regulators of EMT and MET programs, and, hence, we tested the miRNA expression using microarray assay and investigated their roles in silica-induced EMT in lung epithelial cells. We found that miRNA-29b (miR-29b) was dynamically downregulated by silica and influenced the promotion of MET in RLE-6TN cells. Furthermore, delivery of miR-29b to mice significantly inhibited silica-induced EMT, prevented lung fibrosis, and improved lung function. Together, our results clearly demonstrated that miR-29b acted as a novel negative regulator of silicosis fibrosis-inhibited lung fibrosis, probably by promoting MET and by suppressing EMT in the lung. These findings may represent a new potential therapeutic target for treating silicosis fibrosis.

MicroRNA-29b as a Potentially Therapeutic Molecular Via Binding to SP-1 in Silica Induced EMT and Pulmonary Fibrosis

Workers expose to crystalline silica dust could develop into silicotic fibrosis, which is characterized by diffused lung fibrosis and silicon nodules. Up to now, scholars have verified that epithelial-mesenchymal transition (EMT) is an important source of fibroblasts. During the process of EMT, epithelial and mesenchymal markers change significantly, in which transcription factors also play a vital role. Emerging evidence illustrates that microRNA-29b (miR-29b), which is one type of the non-coding small RNAs, mediates various pathogenic organ fibrotic processes by regulating the target gene transcriptionally. However, the function and mechanism of miR-29b that participates in silica-exposed pulmonary fibrosis remain unclear. In the present study, we found that silica induced EMT and pulmonary fibrosis, and the expression of miR-29b was downregulated significantly. This was performed not only in mice with silica induced EMT and MLE-12 cells treated with silica but also in the serum from workers, who had been diagnosed with the silicosis (phase I and II). In silica-induced EMT and lung fibrosis, the changes of epithelial markers (ZO-1, SP-C) and mesenchymal markers (a-SMA) were identified by RT-qPCR or Western blot, and the increasing expression of SP-1 and CTGF were also observed. Next, we fully illustrated that the down-regulation of miR-29b was associated with the up-regulation of SP-1 and CTGF by using luciferase assays. Further experiments verified that the overexpression of miR-29b inhibited silica-induced EMT in vitro. Moreover, the agomiR-29b blocked silica-induced EMT and regulated the levels of SP-1 and CTGF in mice. Our study elucidates that miR-29b have a therapeutic effect on silica induced EMT and lung fibrosis via mediation of SP-1. Funding Statement: This work was supported by grants from the National Natural Science Foundation of China (No. 81273047, 91643114, 81472958, 81703197 and 81602832) and Key Projects of Science and Technology Program by Beijing Municipal Education Commission (No. KZ201610025020). Declaration of Interests: The authors declare that there are no conflicts of interest. Ethics Approval Statement: The research program was approved by the Research Ethics Committee of Capital Medical University. All protocols were completed according to approved guidelines.

The interplay between TGF-β/SMAD and BMP/SMAD signaling pathways in the epithelial mesenchymal transition of A549 cells induced by silica

The epithelial–mesenchymal transition (EMT) is a phenotype transdifferentiation of epithelial into mesenchymal cells and contributes to pulmonary fibrotic disease. SMAD-dependent pathway has been reported to play a key role in the multiple fibrotic diseases. We hypothesized that TGF-β/SMAD signaling could cross-interact with BMP/SMAD signaling pathways in silica-induced EMT in A549 cells. We investigated that the ability of silica-induced EMT in A549 cells, and this process was significantly inhibited by SB431542 through up-regulation of Vimentin, α-SMA and collagen type I expression and down-regulation of E-cadherin expression. Whereas BMP/SMAD inhibition using LDN193189 enhanced EMT. In addition, we also demonstrated that SB431542 could enhance BMP/SMAD signaling pathways in silica-induced EMT and vice versa. Therefore, our study provides evidence that the TGF-β/SMAD pathway was a crucial regulator in silica-induced EMT and that SB431542 could prevent the EMT. More importantly, we have identified that the interplay of TGF-β/SMAD and BMP/SMAD pathways in silica-induced EMT in A549 cells.

Bone morphogenetic protein 7 attenuates epithelial-mesenchymal transition induced by silica.

The epithelial-mesenchymal transition (EMT) is a critical process in the pulmonary fibrosis. It has been reported that bone morphogenetic protein 7 (BMP-7) was able to reverse EMT in proximal tubular cells. Therefore, we test the hypothesis that EMT contributes to silica-induced pulmonary fibrosis and BMP-7 inhibits EMT in silica-induced pulmonary fibrosis. Progressive silica-induced pulmonary fibrosis in the rat was used as a model of silicosis. Epithelial and mesenchymal markers were measured from rat fibrotic lungs. Then the effects of BMP-7 on the EMT were further confirmed in A549 cells. There are increases of vimentin as a mesenchymal marker and decreases of E-cadherin as an epithelial marker in the silica-exposed rat lungs, which is in agreement with the A549 cells data. However, BMP-7 treatment significantly reduced expression of vimentin in the rat pulmonary fibrosis model and in A549 cells. In conclusion, EMT contributes to silica-induced pulmonary fibrosis. Meanwhile, the treatment of BMP-7 can inhibit silica-induced EMT in vitro and in vivo.

Roles of Rho/Rock Signaling Pathway in Silica-induced Epithelial-mesenchymal Transition in Human Bronchial Epithelial Cells

ObjectiveTo investigate the roles of Rho/Rock signaling pathway in silica-induced Epithelial-mesenchymal transition (EMT) in human bronchial epithelial cells (BEC) in vitro. MethodsHuman BEC were incubated with silica with various concentrations for indicated times. Cell viability was assayed by MTT test. Morphologic Changes were observed by microscope. Mesenchymal marker α-smooth muscle actin (α-SMA), vimentin (Vim), and epithelial marker E-cadherin (E-cad) were analyzed by Western Blot. The pull-down assay was used to measure Rho activity. In the prevention experiments, the specific inhibitor for Rho effector ROCK (Y27632) was used to inhibit the activity of Rho. ResultsHuman BEC stimulated with silica were converted from a “cobblestone” epithelial structure into an elongated fibroblast-like shape structure. Incubation of human BEC with silica induced de novo expression of α-SMA and Vim, and loss of E-cad. Also, silica treatment resulted in Rho activation in human BEC. Y27632 up-regulated the E-cad expression but attenuated α-SMA and Vim expression in silica-stimulated cells. ConclusionThe activation of Rho/ROCK signaling pathways is most likely involved in Silica-induced EMT in human bronchial epithelial cells.

Silica-induced epithelial-mesenchymal transition (EMT) in human bronchial epithelial cells

Silica-induced epithelial-mesenchymal transition (EMT) in human bronchial epithelial cells