Oestrogen‐induced angiogenesis promotes adenomyosis by activating the Slug‐VEGF axis in endometrial epithelial cells

Adenomyosis is an oestrogen‐dependent disease characterized by the invasion of endometrial epithelial cells into the myometrium of uterus, and angiogenesis is thought to be required for the implantation of endometrial glandular tissues during the adenomyotic pathogenesis. In this study, we demonstrate that compared with eutopic endometria, adenomyotic lesions exhibited increased vascularity as detected by sonography. Microscopically, the lesions also exhibited an oestrogen‐associated elevation of microvascular density and VEGF expression in endometrial epithelial cells. We previously reported that oestrogen‐induced Slug expression was critical for endometrial epithelial–mesenchymal transition and development of adenomyosis. Our present studies demonstrated that estradiol (E2) elicited a Slug‐VEGF axis in endometrial epithelial cells, and also induced pro‐angiogenic activity in vascular endothelial cells. The antagonizing agents against E2 or VEGF suppressed endothelial cells migration and tubal formation. Animal experiments furthermore confirmed that blockage of E2 or VEGF was efficient to attenuate the implantation of adenomyotic lesions. These results highlight the importance of oestrogen‐induced angiogenesis in adenomyosis development and provide a potential strategy for treating adenomyosis through intercepting the E2‐Slug‐VEGF pathway.     

TIPE1 suppresses invasion and migration through down‐regulating Wnt/β‐catenin pathway in gastric cancer

Epithelial–mesenchymal transition (EMT) plays an important role in the invasiveness and metastasis of gastric cancer. Therefore, identifying key molecules involved in EMT will provide new therapeutic strategy for treating patients with gastric cancer. TIPE1 is a newly identified member of the TIPE (TNFAIP8) family, and its contributions to progression and metastasis have not been evaluated. In this study, we found that the levels of TIPE1 were significantly reduced and inversely correlated with differentiation status and distant metastasis in primary gastric cancer tissues. We further observed overexpression of TIPE1 in aggressive gastric cancer cell lines decreased their metastatic properties both in vitro and in vivo as demonstrated by markedly inhibiting EMT and metastasis of gastric cancer cells in nude mice. Consistently, gene silencing of TIPE1 in well‐differentiated gastric cancer cell line (AGS) inhibited these processes. Mechanistically, we found that TIPE1‐medicated Wnt/β‐catenin signalling was one of the critical signal transduction pathways that link TIPE1 to EMT inhibition. Importantly, TIPE1 dramatically restrained the expression and activities of MMP2 and MMP9 which are demonstrated to promote tumour progression and are implicated in EMT. Collectively, these findings provide new evidence for a better understanding of the biological activities of TIPE1 in progression and metastasis of gastric cancer and suggest that TIPE1 may be an innovative diagnostic and therapeutic target of gastric cancer.     

Nintedanib reduces ventilation‐augmented bleomycin‐induced epithelial–mesenchymal transition and lung fibrosis through suppression of the Src pathway

Mechanical ventilation (MV) used in patients with acute respiratory distress syndrome (ARDS) can increase lung inflammation and pulmonary fibrogenesis. Src is crucial in mediating the transforming growth factor (TGF)‐β1‐induced epithelial–mesenchymal transition (EMT) during the fibroproliferative phase of ARDS. Nintedanib, a multitargeted tyrosine kinase inhibitor that directly blocks Src, has been approved for the treatment of idiopathic pulmonary fibrosis. The mechanisms regulating interactions among MV, EMT and Src remain unclear. In this study, we suggested hypothesized that nintedanib can suppress MV‐augmented bleomycin‐induced EMT and pulmonary fibrosis by inhibiting the Src pathway. Five days after administrating bleomycin to mimic acute lung injury (ALI), C57BL/6 mice, either wild‐type or Src‐deficient were exposed to low tidal volume (V_T) (6 ml/kg) or high V_T (30 ml/kg) MV with room air for 5 hrs. Oral nintedanib was administered once daily in doses of 30, 60 and 100 mg/kg for 5 days before MV. Non‐ventilated mice were used as control groups. Following bleomycin exposure in wild‐type mice, high V_T MV induced substantial increases in microvascular permeability, TGF‐β1, malondialdehyde, Masson's trichrome staining, collagen 1a1 gene expression, EMT (identified by colocalization of increased staining of α‐smooth muscle actin and decreased staining of E‐cadherin) and alveolar epithelial apoptosis (P < 0.05). Oral nintedanib, which simulated genetic downregulation of Src signalling using Src‐deficient mice, dampened the MV‐augmented profibrotic mediators, EMT profile, epithelial apoptotic cell death and pathologic fibrotic scores (P < 0.05). Our data indicate that nintedanib reduces high V_T MV‐augmented EMT and pulmonary fibrosis after bleomycin‐induced ALI, partly by inhibiting the Src pathway.     

Up‐regulation of glycolysis promotes the stemness and EMT phenotypes in gemcitabine‐resistant pancreatic cancer cells

Cancer stem cells (CSCs) and epithelial–mesenchymal transition (EMT)‐type cells are considered as underlying causes of chemoresistance, tumour recurrence and metastasis in pancreatic cancer. We aimed to describe the mechanisms – particularly glycolysis – involved in the regulation of the CSC and EMT phenotypes. We used a gemcitabine‐resistant (GR) Patu8988 cell line, which exhibited clear CSC and EMT phenotypes and showed reliance on glycolysis. Inhibition of glycolysis using 2‐deoxy‐D‐glucose (2‐DG) significantly enhanced the cytotoxicity of gemcitabine and inhibited the CSC and EMT phenotypes in GR cells both in vitro and in vivo. Intriguingly, the use of the reactive oxygen species (ROS) scavenger N‐acetylcysteine (NAC) restored the CSC and EMT phenotypes. H₂O₂ produced changes similar to those of 2‐DG, indicating that ROS were involved in the acquired cancer stemness and EMT phenotypes of GR cells. Moreover, doublecortin‐like kinase 1 (DCLK1), a pancreatic CSC marker, was highly expressed and regulated the stemness and EMT phenotypes in GR cell. Both 2‐DG and H₂O₂ treatment suppressed DCLK1 expression, which was also rescued by NAC. Together, these findings revealed that glycolysis promotes the expression of DCLK1 and maintains the CSC and EMT phenotypes via maintenance of low ROS levels in chemoresistant GR cells. The glycolysis‐ROS‐DCLK1 pathway may be potential targets for reversing the malignant behaviour of pancreatic cancer.     

lncRNA‐POIR promotes epithelial–mesenchymal transition and suppresses sorafenib sensitivity simultaneously in hepatocellular carcinoma by sponging miR‐182‐5p

Sorafenib (SOR) resistance remains a major obstacle in the effective treatment of hepatocellular carcinoma (HCC). A number of long noncoding RNAs (lncRNAs) are responsible for this chemoresistance. This study aimed to reveal the essential function of a recently defined lncRNA, lncRNA‐POIR, in the epithelial–mesenchymal transition (EMT) and SOR sensitivity of HCC cells. SOR‐induced cytotoxicity was analyzed via cell counting kit‐8 and ethynyl‐2'‐deoxyuridine incorporation assays, whereas immunoblotting and confocal immunofluorescence were used to determine the expression levels of EMT markers. Furthermore, loss‐ or gain‐of‐function approaches were used to demonstrate the role of lncRNA‐POIR/miR‐182‐5p on EMT and SOR sensitivity in HCC. The direct interaction between lncRNA‐POIR and miR‐182‐5p was verified using a luciferase reporter assay. We found that knockdown of lncRNA‐POIR sensitized HCC cells to SOR and simultaneously reversed EMT. As expected, miR‐182‐5p was confirmed as the downstream target of lncRNA‐POIR. Moreover, miR‐182‐5p overexpression clearly reversed EMT and promoted SOR‐induced cytotoxicity in representative HCC cells, whereas miR‐182‐5p downregulation played a contrasting role; miR‐182‐5p knockdown abolished the modulatory effects of lncRNA‐POIR siRNA on EMT and SOR sensitivity. Together, these pieces of data suggest that lncRNA‐POIR promotes EMT progression and suppresses SOR sensitivity simultaneously by sponging miR‐182‐5p. Thus, we proposed a compelling rationale for the use of lncRNA‐POIR as a promising predictor of SOR response and as a potential therapeutic target for HCC treatment in the future.     

Silencing of Prrx1b suppresses cellular proliferation,migration, invasion and epithelial–mesenchymal transition in triple‐negative breast cancer

Triple‐negative breast cancer (TNBC) is a highly aggressive tumour subtype associated with poor prognosis. The mechanisms involved in TNBC progression remains largely unknown. To date, there are no effective therapeutic targets for this tumour subtype. Paired‐related homeobox 1b (Prrx1b), one of major isoforms of Prrx1, has been identified as a new epithelial–mesenchymal transition (EMT) inducer. However, the function of Prrx1b in TNBC has not been elucidated. In this study, we found that Prrx1b was significantly up‐regulated in TNBC and associated with tumour size and vascular invasion of breast cancer. Silencing of Prrx1b suppressed the proliferation, migration and invasion of basal‐like cancer cells. Moreover, silencing of Prrx1b prevented Wnt/β‐catenin signaling pathway and induced the mesenchymal‐epithelial transition (MET). Taken together, our data indicated that Prrx1b may be an important regulator of EMT in TNBC cells and a new therapeutic target for interventions against TNBC invasion and metastasis.     

miR‐200c/Bmi1 axis and epithelial–mesenchymal transition contribute to acquired resistance to BRAF inhibitor treatment

Resistance to BRAF inhibitors (BRAFi) is one of the major challenges for targeted therapies for BRAF‐mutant melanomas. However, little is known about the role of microRNAs in conferring BRAFi resistance. Herein, we demonstrate that miR‐200c expression is significantly reduced whereas miR‐200c target genes including Bmi1, Zeb2, Tubb3, ABCG5, and MDR1 are significantly increased in melanomas that acquired BRAFi resistance compared to pretreatment tumor biopsies. Similar changes were observed in BRAFi‐resistant melanoma cell lines. Overexpression of miR‐200c or knock‐down of Bmi1 in resistant melanoma cells restores their sensitivities to BRAFi, leading to deactivation of the PI3K/AKT and MAPK signaling cascades, and acquisition of epithelial–mesenchymal transition‐like phenotypes, including upregulation of E‐cadherin, downregulation of N‐cadherin, and ABCG5 and MDR1 expression. Conversely, knock‐down of miR‐200c or overexpression of Bmi1 in BRAFi‐sensitive melanoma cells activates the PI3K/AKT and MAPK pathways, upregulates N‐cadherin, ABCG5, and MDR1 expression, and downregulates E‐cadherin expression, leading to BRAFi resistance. Together, our data identify miR‐200c as a critical signaling node in BRAFi‐resistant melanomas impacting the MAPK and PI3K/AKT pathways, suggesting miR‐200c as a potential therapeutic target for overcoming acquired BRAFi resistance.     

AEG‐1 induces gastric cancer metastasis by upregulation of eIF4E expression

Gastric cancer is the third leading cause of cancer‐related deaths worldwide, and patients with lymph node, peritoneal and distant metastasis have a poor prognosis. Overexpression of Astrocyte‐elevated gene‐1 (AEG‐1) has been reported to be correlated with the progression and metastasis of gastric cancer. However, its mechanisms are quite unclear. In this study, we found that elevated expression of AEG‐1 was correlated with metastasis in human gastric cancer tissues. Moreover, gain‐ or loss‐of‐function of AEG‐1, respectively, promoted or suppressed epithelial–mesenchymal transition (EMT), migration and invasion of gastric cancer cells. AEG‐1 positively regulated eIF4E, MMP‐9 and Twist expression. Manipulating eIF4E expression by transfection of overexpression constructs or siRNAs partially eliminated AEG‐1‐regulated EMT, cell migration and invasion. In addition, overexpression or knockdown of eIF4E promoted or suppressed EMT, cell migration and invasion in parallel with upregulation of MMP‐9 and Twist expression, while manipulating eIF4E expression partially abrogated AEG‐1‐induced MMP‐9 and Twist. Finally, silencing of AEG‐1 expression not only inhibited tumour growth in parallel with downregulation of eIF4E, MMP‐9 and Twist expression in a xenograft nude mouse model, but also suppressed lymph node and peritoneal metastasis of gastric cancer in an orthotopic nude mouse model. These findings suggest that AEG‐1 promotes gastric cancer metastasis through upregulation of eIF4E‐mediated MMP‐9 and Twist, which provides new diagnostic markers and therapeutic targets for cancer metastasis.     

Interplay between Trx‐1 and S100P promotes colorectal cancer cell epithelial–mesenchymal transition by up‐regulating S100A4 through AKT activation

We previously reported a novel positive feedback loop between thioredoxin‐1 (Trx‐1) and S100P, which promotes the invasion and metastasis of colorectal cancer (CRC). However, the underlying molecular mechanisms remain poorly understood. In this study, we examined the roles of Trx‐1 and S100P in CRC epithelial‐to‐mesenchymal transition (EMT) and their underlying mechanisms. We observed that knockdown of Trx‐1 or S100P in SW620 cells inhibited EMT, whereas overexpression of Trx‐1 or S100P in SW480 cells promoted EMT. Importantly, S100A4 and the phosphorylation of AKT were identified as potential downstream targets of Trx‐1 and S100P in CRC cells. Silencing S100A4 or inhibition of AKT phosphorylation eliminated S100P‐ or Trx‐1‐mediated CRC cell EMT, migration and invasion. Moreover, inhibition of AKT activity reversed S100P‐ or Trx‐1‐induced S100A4 expression. The expression of S100A4 was higher in human CRC tissues compared with their normal counterpart tissues and was significantly correlated with lymph node metastasis and poor survival. The overexpression of S100A4 protein was also positively correlated with S100P or Trx‐1 protein overexpression in our cohort of CRC tissues. In addition, overexpression of S100P reversed the Trx‐1 knockdown‐induced inhibition of S100A4 expression, EMT and migration and invasion in SW620 cells. The data suggest that interplay between Trx‐1 and S100P promoted CRC EMT as well as migration and invasion by up‐regulating S100A4 through AKT activation, thus providing further potential therapeutic targets for suppressing the EMT in metastatic CRC.     

DEK‐targeting aptamer DTA‐64 attenuates bronchial EMT‐mediated airway remodelling by suppressing TGF‐β1/Smad,MAPK and PI3K signalling pathway in asthma

This study is to investigate the inhibitory effects and mechanisms of DEK‐targeting aptamer (DTA‐64) on epithelial mesenchymaltransition (EMT)‐mediated airway remodelling in mice and human bronchial epithelial cell line BEAS‐2B. In the ovalbumin (OVA)‐induced asthmatic mice, DTA‐64 significantly reduced the infiltration of eosinophils and neutrophils in lung tissue, attenuated the airway resistance and the proliferation of goblet cells. In addition, DTA‐64 reduced collagen deposition, transforming growth factor 1 (TGF‐β1) level in BALF and IgE levels in serum, balanced Th1/Th2/Th17 ratio, and decreased mesenchymal proteins (vimentin and α‐SMA), as well as weekend matrix metalloproteinases (MMP‐2 and MMP‐9) and NF‐κB p65 activity. In the in vitro experiments, we used TGF‐β1 to induce EMT in the human epithelial cell line BEAS‐2B. DEK overexpression (ovDEK) or silencing (shDEK) up‐regulated or down‐regulated TGF‐β1 expression, respectively, on the contrary, TGF‐β1 exposure had no effect on DEK expression. Furthermore, ovDEK and TGF‐β1 synergistically promoted EMT, whereas shDEK significantly reduced mesenchymal markers and increased epithelial markers, thus inhibiting EMT. Additionally, shDEK inhibited key proteins in TGF‐β1‐mediated signalling pathways, including Smad2/3, Smad4, p38 MAPK, ERK1/2, JNK and PI3K/AKT/mTOR. In conclusion, the effects of DTA‐64 against EMT of asthmatic mice and BEAS‐2B might partially be achieved through suppressing TGF‐β1/Smad, MAPK and PI3K signalling pathways. DTA‐64 may be a new therapeutic option for the management of airway remodelling in asthma patients.     

Polydatin alleviated radiation‐induced lung injury through activation of Sirt3 and inhibition of epithelial–mesenchymal transition

Radiation‐induced lung injury (RILI) is one of the most common and fatal complications of thoracic radiotherapy. It is characterized with two main features including early radiation pneumonitis and fibrosis in later phase. This study was to investigate the potential radioprotective effects of polydatin (PD), which was shown to exert anti‐inflammation and anti‐oxidative capacities in other diseases. In this study, we demonstrated that PD‐mitigated acute inflammation and late fibrosis caused by irradiation. PD treatment inhibited TGF‐β1‐Smad3 signalling pathway and epithelial–mesenchymal transition. Moreover, radiation‐induced imbalance of Th1/Th2 was also alleviated by PD treatment. Besides its free radical scavenging capacity, PD induced a huge increase of Sirt3 in culture cells and lung tissues. The level of Nrf2 and PGC1α in lung tissues was also elevated. In conclusion, our data showed that PD attenuated radiation‐induced lung injury through inhibiting epithelial–mesenchymal transition and increased the expression of Sirt3, suggesting PD as a novel potential radioprotector for RILI.     

Exosomal miR‐663b targets Ets2‐repressor factor to promote proliferation and the epithelial–mesenchymal transition of bladder cancer cells

Exosomes circulating in biological fluids have the potential to be utilized as cancer biomarkers and are associated with cancer progression and metastasis. MicroRNA (miR)‐663b has been found to be elevated in plasma from patients with bladder cancer (BC). However, the functional role of exosomal miR‐663b in BC processes remains unknown. Here, we isolated exosomes from plasma and found that the miR‐663b level was elevated in exosomes from plasma of patients with BC compared with healthy controls. Exosomal miR‐663b from BC cells promoted cell proliferation and epithelial–mesenchymal transition. Moreover, exosomal miR‐663b targeted Ets2‐repressor factor and acted as a tumor promoter in BC cells. Taken together, our findings suggested that exosomal miR‐663b is a promising potential biomarker and target for clinical detection and therapy in BC.