Epithelial-mesenchymal Transition In A549 Cells Research Articles

Addition of Polyphenols to Drugs: The Potential of Controlling "Inflammaging" and Fibrosis in Human Senescent Lung Fibroblasts In Vitro.

The combination of a polyphenol, quercetin, with dasatinib initiated clinical trials to evaluate the safety and efficacy of senolytics in idiopathic pulmonary fibrosis, a lung disease associated with the presence of senescent cells. Another approach to senotherapeutics consists of controlling inflammation related to cellular senescence or "inflammaging", which participates, among other processes, in establishing pulmonary fibrosis. We evaluate whether polyphenols such as caffeic acid, chlorogenic acid, epicatechin, gallic acid, quercetin, or resveratrol combined with different senotherapeutics such as metformin or rapamycin, and antifibrotic drugs such as nintedanib or pirfenidone, could present beneficial actions in an in vitro model of senescent MRC-5 lung fibroblasts. A senescent-associated secretory phenotype (SASP) was evaluated by the measurement of interleukin (IL)-6, IL-8, and IL-1β. The senescent-associated β-galactosidase (SA-β-gal) activity and cellular proliferation were assessed. Fibrosis was evaluated using a Picrosirius red assay and the gene expression of fibrosis-related genes. Epithelial-mesenchymal transition (EMT) was assayed in the A549 cell line exposed to Transforming Growth Factor (TGF)-β in vitro. The combination that demonstrated the best results was metformin and caffeic acid, by inhibiting IL-6 and IL-8 in senescent MRC-5 cells. Metformin and caffeic acid also restore cellular proliferation and reduce SA-β-gal activity during senescence induction. The collagen production by senescent MRC-5 cells was inhibited by epicatechin alone or combined with drugs. Epicatechin and nintedanib were able to control EMT in A549 cells. In conclusion, caffeic acid and epicatechin can potentially increase the effectiveness of senotherapeutic drugs in controlling lung diseases whose pathophysiological component is the presence of senescent cells and fibrosis.

The flavonoids from the fruits of Psoralea corylifolia and their potential in inhibiting metastasis of human non-small cell lung cancers

Nineteen flavonoids were isolated from the fruits of Psoralea corylifolia L., including a novel flavanol (3) and three novel isoflavones (12–14). Their chemical structures were unequivocally determined through comprehensive spectral data analysis. The anti-proliferative effect of the isolated flavonoids was assessed in vitro using the MTT assay. Molecular docking and ELISA were employed to determine the inhibitory effects of the active compounds on ALK5. Isobavachalcone was found to inhibit TGF-β1 induced EMT in A549 cells by Wound healing assay and Transwell chamber assay. Immunofluorescence assay and Western blot assay showed that IBC could inhibit cytoskeleton rearrangement, reduce the phosphorylation of ALK5, ERK, and Smad, down-regulate Snail expression, and up-regulate E-cadherin expression in TGF-β1 induced A549 cells, thereby exerting the potential inhibitory effects on epithelial-mesenchymal transition (EMT) process in A549 cells. The findings presented herein establish a fundamental basis for investigating the anti-proliferative and anti-metastatic properties of psoralen flavonoids in human non-small cell lung cancer.

Effect of WISP1 on paraquat-induced EMT

BackgroundParaquat (PQ) can induce pulmonary fibrosis (PF) by modulating epithelial-mesenchymal transition (EMT) of alveolar epithelial cells, but the molecular mechanism is unknown. In this paper, the role of Wnt-inducible signaling protein-1 (WISP1) in PQ-induced EMT was inspected. MethodsThe morphology, apoptosis, and mortality of A549 cells were observed through a microscope. The mRNA and protein levels of WISP1, E-cadherin, and Vimentin were confirmed by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blot. ResultsWith the increase of PQ concentration, the morphology of A549 cells was apparently changed, cell apoptosis and mortality were enhanced. Besides, the E-cadherin abundance was reduced (p < 0.01), however, WISP1 and Vimentin contents were boosted after PQ treatment (p < 0.01). With the increase of PQ treatment time, the epithelial index of cells first increased and then decreased. The expression of WISP1 gene increased significantly with the increase of PQ treatment time (p < 0.01). Silence of WISP1 abolished the effect of PQ treatment on E-cadherin and Vimentin levels (p < 0.01). Downregulation of WISP1 curbed morphology change and PQ-induced EMT in A549 cells. ConclusionKnockdown of WISP1 inhibited PQ-induced EMT in A549 cells. This conclusion might provide a new therapeutic target for PQ poisoning treatment.

Lipopolysaccharide (LPS) extracted from Bacteroides vulgatus effectively prevents LPS extracted from Escherichia coli from inducing epithelial‑mesenchymal transition.

Pathological epithelial‑mesenchymal transition (EMT) has been shown to fulfill a key role in the development and progression of a variety of lung diseases. It has been demonstrated that the inflammatory microenvironment is a decisive factor in inducing pathological EMT. Hexacylated lipopolysaccharide (LPS) [or proacylated lipopolysaccharide (P‑LPS), which functions as proinflammatory lipopolysaccharide] is one of the most effective Toll‑like receptor 4 (TLR4) agonists. Furthermore, the pentacylated and tetracylated form of lipopolysaccharide (or A‑LPS, which functions as anti‑inflammatory lipopolysaccharide) has been shown to elicit competitive antagonistic effects against the pro‑inflammatory activity of P‑LPS. At present, it remains unclear whether LPS extracted from Bacteroides vulgatus (BV‑LPS) can prevent LPS extracted from Escherichiacoli (EC‑LPS) from inducing pathological EMT. In the present study, A549 cells and C57BL/6 mice lung tissue were both induced by EC‑LPS (P‑LPS) and BV‑LPS (A‑LPS), either alone or in combination. The anticipated anti‑inflammatory effects of BV‑LPS were analyzed by examining the lung coefficient, lung pathology, A549 cell morphology and expression levels both of the inflammatory cytokines, IL‑1β, IL‑6 and TNF‑α and of the EMT signature proteins, epithelial cadherin (E‑cadherin), α‑smooth muscle actin (α‑SMA) and vimentin. In addition, the expression levels of TLR4, bone morphogenic protein and activin membrane‑bound inhibitor (BAMBI) and Snail were detected and the possible mechanism underlying how BV‑LPS may prevent EC‑LPS‑induced EMT was analyzed. The results obtained showed that the morphology of the A549 cells was significantly polarized, the lung index was significantly increased, the alveolar structure was collapsed and the expression levels of IL‑1β, IL‑6, TNF‑α, α‑SMA, vimentin, TLR4 and Snail in both lung tissue and A549 cells were significantly increased, whereas those of E‑cadherin and BAMBI were significantly decreased. Treatment with BV‑LPS in combination with EC‑LPS was found to reverse these changes. In conclusion, the present study demonstrated that BV‑LPS is able to effectively prevent EC‑LPS‑induced EMT in A549 cells and in mouse lung tissue and furthermore, the underlying mechanism may be associated with inhibition of the TLR4/BAMBI/Snail signaling pathway.

Mesenchymal Stem Cells Inhibit Epithelial-to-Mesenchymal Transition by Modulating the IRE1α Branch of the Endoplasmic Reticulum Stress Response.

Idiopathic pulmonary fibrosis (IPF) is the most common idiopathic interstitial lung disease, and it carries a poor prognosis due to a lack of efficient diagnosis methods and treatments. Epithelial-mesenchymal transition (EMT) plays a key role in IPF pathogenesis. Endoplasmic reticulum (ER) stress contributes to fibrosis via EMT-mediated pathways. Mesenchymal stem cell (MSC) transplantation is a promising treatment strategy for pulmonary fibrosis and ameliorates lung fibrosis in animal models via paracrine effects. However, the specific mechanisms underlying the effect of transplanted MSCs are not known. We previously reported that MSCs attenuate endothelial injury by modulating ER stress and endothelial-to-mesenchymal transition. The present study investigated whether modulation of ER stress- and EMT-related pathways plays essential roles in MSC-mediated alleviation of IPF. We constructed a A549 cell model of transforming growth factor-β1 (TGF-β1)-induced fibrosis. TGF-β1 was used to induce EMT in A549 cells, and MSC coculture decreased EMT, as indicated by increased E-cadherin levels and decreased vimentin levels. ER stress participated in TGF-β1-induced EMT in A549 cells, and MSCs inhibited the expression of XBP-1s, XBP-1u, and BiP, which was upregulated by TGF-β1. Inhibition of ER stress contributed to MSC-mediated amelioration of EMT in A549 cells, and modulation of the IRE1α-XBP1 branch of the ER stress pathway may have played an important role in this effect. MSC transplantation alleviated bleomycin (BLM)-induced pulmonary fibrosis in mice. MSC treatment decreased the expression of ER stress- and EMT-related genes and proteins, and the most obvious effect of MSC treatment was inhibition of the IRE1α/XBP1 pathway. The present study demonstrated that MSCs decrease EMT by modulating ER stress and that blockade of the IRE1α-XBP1 pathway may play a critical role in this effect. The current study provides novel insight for the application of MSCs for IPF treatment and elucidates the mechanism underlying the preventive effects of MSCs against pulmonary fibrosis.

Effects of daphnetin on the mechanism of epithelial-mesenchymal transition induced by HMGB1 in human lung adenocarcinoma cells (A549 cell line)

ABSTRACT As a cancer with the highest incidence in recent years, lung cancer is mainly composed of three diseases: non-small cell lung cancer, small cell lung cancer and neuroendocrine tumor. The morbidity and mortality of this malignant tumor are the highest in both male and female populations worldwide. In my country, lung cancer has become the most common cancer disease and the leading cause of cancer death, so it is extremely important to find lung cancer therapeutic targets. Based on previous studies, we speculated that the TLR4-Myd88-NFκB pathway may be involved in hmgb1-induced EMT in A549 cells, and daphnetin may also inhibit hmgb1-induced EMT through the TLR4-Myd88-NFκB pathway in A549 cells, but related studies have not linked it to hmgb1-induced EMT. Therefore, the innovation of this study is to test these two conjectures and analyze how daphnetin affects the epithelial-mesenchymal transition (EMT) mechanism induced by HMGB1 in human lung adenocarcinoma cells (A549 cell line), aiming at lung adenocarcinoma cells, foundation for clinical treatment. The proliferation rate and the migrating cell number presented an obvious decrease in the HMGB1+TLR4-shRNA group and the HMGB1+daphnetin group relative to the HMGB1 group (P < 0.0001). The intracellular expression of TLR4, Myd88, NFκB, vimentin and snail1 proteins were significantly decreased (P < 0.001), while that of E-cadherin presented a remarkable increase (P < 0.001) in the HMGB1+TLR4-shRNA and HMGB1+daphnetin group compared with the HMGB1 group. TLR4-MyD88-NFκB pathway is associated with HMGB1-induced EMT in A549 cells. Daphnetin had an inhibitory effect on HMGB1-induced EMT via the TLR4-Myd88-NF-κB pathway in A549 cells.

ABCB1 and ABCC1 Function during TGF-β-Induced Epithelial-Mesenchymal Transition: Relationship between Multidrug Resistance and Tumor Progression.

Multidrug resistance (MDR) and induction of metastasis are some of the puzzles encountered during cancer chemotherapy. The MDR phenotype is associated with overexpression of ABC transporters, involved in drug efflux. Metastasis originates from the epithelial-mesenchymal transition (EMT), in which cells acquire a migratory phenotype, invading new tissues. ABC transporters' role during EMT is still elusive, though cells undergoing EMT exhibit enhanced ABCB1 expression. We demonstrated increased ABCB1 expression but no change in activity after TGF-β-induced EMT in A549 cells. Moreover, ABCB1 inhibition by verapamil increased snail and fibronectin expression, an event associated with upregulation of ABCB1, evidencing coincident cell signaling pathways leading to ABCB1 and EMT-related markers transcription, rather than a direct effect of transport. Additionally, for the first time, increased ABCC1 expression and activity was observed after EMT, and use of ABCC1 inhibitors partially inhibited EMT-marker snail, although increased ABCC1 function translated into collateral sensibility to daunorubicin. More investigations must be done to evaluate the real benefits that the gain of ABC transporters might have on the process of metastasis. Considering ABCC1 is involved in the stress response, affecting intracellular GSH content and drug detoxification, this transporter could be used as a therapeutic target in cancer cells undergoing EMT.

Therapeutic potential of amitriptyline for paraquat-induced pulmonary fibrosis: Involvement of caveolin-1-mediated anti-epithelial-mesenchymal transition and inhibition of apoptosis

ObjectiveTreatment of pulmonary fibrosis caused by paraquat (PQ) poisoning remains problematic. Amitriptyline (AMT) has multiple pharmacological effects. Here we investigated the anti-fibrotic effect of AMT on PQ-induced pulmonary fibrosis and its possible mechanism. MethodsC57BL/6 mice were randomly divided into control, PQ, PQ + AMT and AMT groups. Histopathology of the lungs, blood gas analysis, and levels of hydroxyproline (HYP), transforming growth factor β1 (TGF-β1) and interleukin 17 (IL-17) were measured. The siRNA transfection inhibited caveolin-1 in A549 cells, which induced epithelial-mesenchymal transition (EMT) by PQ and followed intervention with AMT. E-cadherin, N-cadherin, α-smooth muscle actin (α-SMA) and caveolin-1 were studied by immunohistochemistry and western blot analysis. The apoptosis rate was measured by flow cytometry. ResultsCompared with the PQ group, the PQ + AMT group displayed mild pathological changes in pulmonary fibrosis, lower HYP, IL-17 and TGF- β1 levels in lung, but high TGF- β1 in serum. Levels of N-cadherin and α-SMA in the lungs were significantly decreased, but caveolin-1 was increased, while SaO2 and PaO2 levels were higher. Compared with the PQ group, the apoptosis rate, N-cadherin and α-SMA levels in A549 cells were significantly decreased after PQ treatment and high dose AMT intervention (p < 0.01). The expressions of E-cadherin, N-cadherin and α-SMA in the PQ-induced cells transfected with caveolin-1 siRNA or siControl RNA were significantly different (p < 0.01), but the apoptosis rate was unaltered. ConclusionAMT inhibited PQ-induced EMT in A549 cells and improved lung histopathology and oxygenation in mice by up-regulating caveolin-1.

ST3GAL5-catalyzed gangliosides inhibit TGF-β-induced epithelial-mesenchymal transition via TβRI degradation.

Epithelial-mesenchymal transition (EMT) is pivotal in the initiation and development of cancer cell metastasis. We observed that the abundance of glycosphingolipids (GSLs), especially ganglioside subtypes, decreased significantly during TGF-β-induced EMT in NMuMG mouse mammary epithelial cells and A549 human lung adenocarcinoma cells. Transcriptional profiling showed that TGF-β/SMAD response genes and EMT signatures were strongly enriched in NMuMG cells, along with depletion of UDP-glucose ceramide glucosyltransferase (UGCG), the enzyme that catalyzes the initial step in GSL biosynthesis. Consistent with this finding, genetic or pharmacological inhibition of UGCG promoted TGF-β signaling and TGF-β-induced EMT. UGCG inhibition promoted A549 cell migration, extravasation in the zebrafish xenograft model, and metastasis in mice. Mechanistically, GSLs inhibited TGF-β signaling by promoting lipid raft localization of the TGF-β type I receptor (TβRI) and by increasing TβRI ubiquitination and degradation. Importantly, we identified ST3GAL5-synthesized a-series gangliosides as the main GSL subtype involved in inhibition of TGF-β signaling and TGF-β-induced EMT in A549 cells. Notably, ST3GAL5 is weakly expressed in lung cancer tissues compared to adjacent nonmalignant tissues, and its expression correlates with good prognosis.

Study of paraquat-induced pulmonary fibrosis using biomimetic micro-lung chips

Paraquat (PQ) poisoning induces pulmonary fibrosis in vivo. The pathogenesis of pulmonary fibrosis is complex, which has prevented the development of specific treatments. Pulmonary fibrosis shows several characteristics including epithelial-mesenchymal transition (EMT), fibroblast activation, and extracellular matrix (ECM) deposition. To investigate pulmonary fibrosis, we designed a biomimetic multichannel micro-lung chip to imitate the in vivo interface between the lung epithelium and the lung interstitium. In our model, A549 (lung epithelial cells) and MRC-5 (fetal lung fibroblasts) cells were used to test the efficacy of our chip-based model. Rat tail type I collagen and hyaluronic acid were used to simulate ECM and to provide a 3D microenvironment. The micro-lung chips were cultured with PQ (0, 75, 150, 300, and 400 µM). The viability of A549 and MRC-5 cells significantly decreased with increasing PQ concentrations. There were significant changes in surfactant proteins C (SP-C), alpha smooth muscle actin protein (α-SMA), and vimentin protein levels during PQ-induced pulmonary fibrosis. SP-C levels were decreased in A549 cells, while those of α-SMA and vimentin were increased in A549 cells and MRC-5 cells treated with PQ in the micro-lung chip. We also designed a reference model without interaction between the lung epithelial cells and fibroblasts. Compared to the non-contact model, co-culturing A549 and MRC-5 cells in chips induced more severe EMT in A549 cells after treatment with 75 µM PQ and together defended against PQ-induced damage. Thus, our novel co-culture micro-lung chip that models the lung epithelium and interstitium may provide a new approach for studying lung fibrosis and will facilitate drug development.

Role of Nrf2 in Methotrexate-Induced Epithelial-Mesenchymal Transition in Alveolar A549 Cells.

Methotrexate (MTX) is known to induce serious lung diseases, such as pulmonary fibrosis. Although we demonstrated that MTX is associated with epithelial-mesenchymal transition (EMT), the underlying mechanism remains unclear. Nuclear factor erythroid 2-related factor 2 (Nrf2), an oxidative stress response regulator, is related to EMT induction. In the present study, we examined the association of Nrf2 with the MTX-induced EMT in the alveolar epithelial cell line A549. MTX treatment decreased the mRNA expression of heme oxidase-1 (HO-1), a target of Nrf2, which was inhibited by co-treatment with diethyl maleate (DEM), an Nrf2 activator. Additionally, the MTX-induced increase in reactive oxygen species (ROS) production was significantly suppressed by DEM. Furthermore, DEM decreased mRNA/protein expression levels of α-smooth muscle actin (SMA), a representative EMT marker, which were upregulated by MTX. Nuclear expression and localization of Nrf2 were suppressed by MTX treatment, which led to a decrease in Nrf2 activity. Finally, in Nrf2 knockdown cells, the MTX-induced enhancement of α-SMA mRNA/protein expression was not observed, indicating that downregulation of Nrf2 may play a critical role in the MTX-induced EMT in A549 cells. These results suggest that Nrf2-regulated transcriptional activity would be associated with the MTX-induced EMT induction.

Establishment and Preliminary Analysis of Lung Cancer Cell Line A549 with Stable MAP4 K4 Knockdown

To analyze the effect of knocking down MAP4 K4 expression on the proliferation and migration of cancer cells, and to explore its underlining molecular mechanisms. A stable knockdown MAP4 K4 cell line was constructed and the subcellular localization of the cells was determined with immunofluorescence, cell proliferation assay and cell migration assay. In addition, the effects of down-regulated MAP4 K4 expression were analyzed by examining the difference between the proliferation and migration of cancer cells in the knockdown group and those of the control group. MAP4 K4 was localized in focal adhesion and cell edges in A549 cells. Stable knockdown of MAP4 K4 expression induced cancer cells to grow in clusters and arrested the progression of the cell cycle and cell migration. Further analysis found that knocking down MAP4 K4 expression in A549 cells induced the accumulation of epithelial cell marker E-cadherin, and subsequently, the down-regulation of N-cadherin, a mesenchymal cell marker, thereby disrupting the "cadherin switch" and the epithelial-mesenchymal conversion. Then, the control group and the knockdown group both received the combined treatment of cisplatin at a final concentration of 5 μmol/L and paclitaxel at a final concentration of 20 nmol/L. The stably knocked down MAP4 K4 expressing cells showed significantly enhanced toxicity of chemotherapeutic drugs to cancer cells. The study shows that MAP4 K4 regulates the malignant phenotypes of cancer cells and chemoresistance by regulating "cadherin switch" to promote epithelial-mesenchymal transition in A549 cells.

CircCCT3 Enhances Invasion and Epithelial-Mesenchymal Transition (EMT) of Non-Small-Cell Lung Cancer (NSCLC) via the miR-107/Wnt/FGF7 Axis

Background CircRNAs play a role in a variety of biological processes, including tumorigenesis. circCCT3 has been shown to regulate cancer initiation and progression. Unfortunately, whether circCCT3 is involved in non-small-cell lung cancer (NSCLC) metastasis remains unclear. Methods Our study utilized RT-qPCR to examine gene expression levels. A transwell assay was used to measure invasion ability of cells. Starbase software and TargetScan software were used to predict target genes. Results circCCT3 knockdown attenuated invasion and epithelial-mesenchymal transition (EMT) of A549 and Calu-1 cells. miR-107 mimics could rescue circCCT3-induced invasion and EMT. Next, miR-107 mimics and circCCT3 knockdown suppressed Wnt3a and FGF7 expression. An miR-107 inhibitor promoted Wnt3a and FGF7 expressions. Finally, FGF7 greatly restored miR-107-inhibited invasion and EMT of A549 cells. Conclusion Here, we reveal a molecular mechanism circCCT3/miR-107/Wnt/FGF7 responsible for NSCLC metastasis.

Design and Synthesis of Novel PRMT1 Inhibitors and Investigation of Their Effects on the Migration of Cancer Cell.

Protein arginine methyltransferase 1 (PRMT1) can catalyze the protein arginine methylation by transferring the methyl group from S-adenosyl-L-methionine (SAM) to the guanidyl nitrogen atom of protein arginine, which influences a variety of biological processes including epithelial–mesenchymal transition (EMT) and EMT-mediated mobility of cancer cells. The upregulation of PRMT1 is involved in a diverse range of cancer, such as lung cancer, and there is an urgent need to develop novel and potent PRMT1 inhibitors. In this article, a series of 2,5-substituted furan derivatives and 2,4-substituted thiazole derivatives were designed and synthesized by targeting at the substrate arginine-binding site on PRMT1, and 10 compounds demonstrated significant inhibitory effects against PRMT1. Among them, the most potent inhibitor, compound 1r (WCJ-394), significantly affected the expression of PRMT1-related proteins in A549 cells and downregulated the expression of mesenchymal markers, by which WCJ-394 inhibited the TGF-β1-induced EMT in A549 cells and prevented the cancer cell migration. The current study demonstrated that WCJ-394 was a potent PRMT1 inhibitor, which could be used as the leading compound for further drug discovery.

Regulation of Epithelial-Mesenchymal Transition of A549 Cells by Prostaglandin D2.

Despite significant advances in diagnostic and operative techniques, lung cancer remains one of the most lethal malignancies worldwide. Since prostaglandins such as prostaglandin D2 (PGD2) is involved in various pathophysiological process, including inflammation and tumorigenesis, this study aims to investigate the role of PGD2 during the process of epithelial-mesenchymal transition (EMT) in A549 cells. A549 cells were stimulated with PGD2 and expression of EMT markers was analyzed by immunoblotting and immunofluorescence. EMT-related gene, Slug expression was evaluated using quantitative real-time polymerase chain reaction (qPCR). Migration and invasion abilities of A549 cells were determined in chemotaxis and Matrigel invasion assays, respectively. We also inhibited the TGF/Smad signaling pathway using a receptor inhibitor or silencing of TGF-β1 and TGFβ type I receptor (TGFβRI), and protein expressionwas assessed by immunoblotting and immunofluorescence. Here, we found that stimulation of A549 cells with PGD2 resulted in morphological changes into a mesenchymal-like phenotype under low serum conditions. Stimulation of A549 cells with PGD2 resulted in a significant reduction in proliferation, whereas invasion and migration were enhanced. The expression of E-cadherin was markedly downregulated, while Vimentin expression was upregulated after treatment of A549 cells with PGD2. Slug expression was markedly upregulated by stimulating A549 cells with PGD2, and stimulation of A549 cells with PGD2 significantly enhanced TGF-β1 expression, and silencing of TGF-β1 significantly blocked PGD2-induced EMT and Smad2 phosphorylation. In addition, PGD2-induced Smad2 phosphorylation and EMT were significantly abrogated by either pharmacological inhibition or silencing of TGFβRI. PGD2-induced expression of Slug and EMT were significantly augmented in low nutrient and low serum conditions. Finally, the subsequent culture of mesenchymal type of A549 cells under normal culture conditions reverted the cell's phenotype to an epithelial type. Given these results, we suggest that tumor microenvironmental factors such as PGD2, nutrition, and growth factors could be possible therapeutic targets for treating metastatic cancers.

Role of integrin α2 in methotrexate-induced epithelial-mesenchymal transition in alveolar epithelial A549 cells.

Methotrexate (MTX) is widely used to treat various diseases. However, it induces adverse reactions like serious lung injury, including pulmonary fibrosis. Increasing evidence suggests that epithelial-mesenchymal transition (EMT) in injured alveolar epithelium contributes to the development of the pathophysiological state of the lung. We demonstrated that MTX induced EMT in cultured alveolar epithelial cell lines. Integrin-mediated signaling is considered a significant factor in recognizing the EMT process. However, the relationship between MTX-induced EMT and integrin family members is poorly understood. In the present study, we aimed to clarify the role of integrin in MTX-induced EMT in A549 and NCI-H1299 (H1299) cells by focusing on the integrin alpha 2 (ITGA2) subunit, selected based on our microarray analysis. MTX treatment for 72h significantly increased the mRNA and cell surface expression of ITGA2 in both cell lines. However, this upregulation by MTX was suppressed by co-treatment with SB431542 and folic acid, which are inhibitors of MTX-induced EMT in A549 cells. The mRNA expression levels of EMT-related genes were more affected in the MTX-treated A549 cells with high ITGA2 expression than in those with low ITGA2 expression. Finally, E7820, an ITGA2 inhibitor, suppressed MTX-induced EMT-related phenotypic changes, such as morphology and mRNA and protein expression of α-smooth muscle actin, a representative EMT marker. These findings suggest that ITGA2 may play a key role in MTX-induced EMT in alveolar epithelial cells.

Interleukin-1β released from macrophages stimulated with indium tin oxide nanoparticles induces epithelial-mesenchymal transition in A549 cells

Indium tin oxide (ITO) nanoparticles triggered the release of IL-1β from macrophages, followed by the significant induction of epithelial-mesenchymal transition (EMT) in alveolar epithelial cells.

Ginsenoside CK Inhibits TGF-β-Induced Epithelial-Mesenchymal Transition in A549 Cell via SIRT1

Ginsenoside CK is the main metabolite of protopanaxadiol saponins in intestinal bacteria. Previous studies have shown that ginsenoside CK can affect many aspects of tumor development through a variety of mechanisms. However, few studies have reported the antimetastatic effects of ginsenoside CK in non-small-cell lung cancer (NSCLC). In this study, we explored the effect of ginsenoside CK on epithelial-mesenchymal transition (EMT) induced by TGF-β in A549 cells and the potential molecular mechanisms. Our data showed that ginsenoside CK effectively prevented TGF-β-induced EMT, as indicated by the upregulation of E-cadherin and downregulation of vimentin. Furthermore, ginsenoside CK inhibited the metastatic ability of A549 cells in the tail vein lung metastasis model of nude mice. Additionally, ginsenoside CK decreased the expression of silent information regulator 2 homolog 1 (SIRT1) in the inhibition of EMT induced by TGF-β. Moreover, the antimetastatic effect of ginsenoside CK was reversed by SIRT1 overexpression. Generally, our results indicated the antimetastatic effect and underlying mechanism of ginsenoside CK on TGF-β-induced EMT in A549 cells, suggesting that ginsenoside CK can be used as an effective antineoplastic agent.

Atractylodin Suppresses TGF-β-Mediated Epithelial-Mesenchymal Transition in Alveolar Epithelial Cells and Attenuates Bleomycin-Induced Pulmonary Fibrosis in Mice.

Idiopathic pulmonary fibrosis (IPF) is characterized by fibrotic change in alveolar epithelial cells and leads to the irreversible deterioration of pulmonary function. Transforming growth factor-beta 1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) in type 2 lung epithelial cells contributes to excessive collagen deposition and plays an important role in IPF. Atractylodin (ATL) is a kind of herbal medicine that has been proven to protect intestinal inflammation and attenuate acute lung injury. Our study aimed to determine whether EMT played a crucial role in the pathogenesis of pulmonary fibrosis and whether EMT can be utilized as a therapeutic target by ATL treatment to mitigate IPF. To address this topic, we took two steps to investigate: 1. Utilization of anin vitro EMT model by treating alveolar epithelial cells (A549 cells) with TGF-β1 followed by ATL treatment for elucidating the underlying pathways, including Smad2/3 hyperphosphorylation, mitogen-activated protein kinase (MAPK) pathway overexpression, Snail and Slug upregulation, and loss of E-cadherin. Utilization of an in vivo lung injury model by treating bleomycin on mice followed by ATL treatment to demonstrate the therapeutic effectiveness, such as, less collagen deposition and lower E-cadherin expression. In conclusion, ATL attenuates TGF-β1-induced EMT in A549 cells and bleomycin-induced pulmonary fibrosis in mice.

Andrographolide alleviates bleomycin-induced NLRP3 inflammasome activation and epithelial-mesenchymal transition in lung epithelial cells by suppressing AKT/mTOR signaling pathway.

BackgroundAndrographolide (Andro), a diterpenoid extracted from Andrographis paniculata, has been shown to attenuate pulmonary fibrosis in rodents; however, the potential mechanisms remain largely unclear. This study investigated whether and how Andro alleviates bleomycin (BLM)-induced NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome activation and epithelial-mesenchymal transition (EMT) in the lung epithelial cells.MethodsThe in vivo effects of Andro were evaluated in a rat model of BLM-induced pulmonary fibrosis. The roles of Andro in BLM-induced NLRP3 inflammasome activation, EMT and AKT/mTOR signaling were investigated using human alveolar epithelial A549 cells.ResultsWe found that Andro significantly alleviated pulmonary edema and histopathological changes, decreased weight loss, and reduced collagen deposition. Andro downregulated the levels of NLRP3, the adaptor molecule apoptosis-associated speck-like protein containing a CARD (ASC), and Caspase-1 in the lungs of BLM-treated rats, suggesting the inhibitory effect of Andro on NLRP3 inflammasome activation in vivo. Additionally, the symptoms of BLM-mediated EMT phenotype in the lung were also attenuated after Andro administration. In vitro, Andro also markedly inhibited BLM-induced NLRP3 inflammasome activation and EMT in A549 cells. Moreover, Andro inhibited BLM-induced phosphorylation of AKT and mTOR in A549 cells, suggesting that AKT/mTOR inactivation mediates Andro-induced effects on BLM-induced NLRP3 inflammasome activation and EMT.ConclusionsThese data indicate that Andro can reduce BLM-induced pulmonary fibrosis through suppressing NLRP3 inflammasome activation and EMT in lung epithelial cells via AKT/mTOR signaling pathway.