Bone morphogenetic protein-6 suppresses TGF-β_(2)-induced epithelial-mesenchymal transition in retinal pigment epithelium

AIM:To evaluate the effect of bone morphogenetic protein-6(BMP-6)on transforming growth factor(TGF)-β_(2)-induced epithelial-mesenchymal transition(EMT)in retinal pigment epithelium(RPE).METHODS:Adult retinal pigment epithelial cell line(ARPE-19)were randomly divided into control,TGF-β_(2)(5μg/L),and BMP-6 small interfering RNA(siRNA)group.The cell morphology was observed by microscopy,and the cell migration ability were detected by Transwell chamber.The EMT-related indexes and BMP-6 protein levels were detected by Western blotting.Furthermore,a BMP-6 overexpression plasmid was constructed and RPE cells were divided into the control group,TGF-β_(2)+empty plasmid group,BMP-6 overexpression group,and TGF-β_(2)+BMP-6 overexpression group.The EMT-related indexes and extracellular regulated protein kinases(ERK)protein levels were detected.RESULTS:Compared with the control group,the migration of RPE cells in the TGF-β_(2) group was significantly enhanced.TGF-β_(2) increased the protein expression levels ofα-smooth muscle actin(α-SMA),fibronectin and vimentin but significantly decreased the protein levels of E-cadherin and BMP-6(P<0.05)in RPE.Similarly,the migration of RPE cells in the BMP-6 siRNA group was also significantly enhanced.BMP-6 siRNA increased the protein expression levels ofα-SMA,fibronectin and vimentin but significantly decreased the protein expression levels of E-cadherin(P<0.05).Overexpression of BMP-6 inhibited the migration of RPE cells induced by TGF-β_(2) and prevented TGF-β_(2) from affecting EMT-related biomarkers(P<0.05).CONCLUSION:BMP-6 prevents the EMT in RPE cells induced by TGF-β_(2),which may provide a theoretical basis for the prevention and treatment of proliferative vitreoretinopathy.

Correction to “Interleukin-34 promotes the proliferation and epithelial-mesenchymal transition of gastric cancer cells”

Correction to“Interleukin-34 promotes the proliferation and epithelialmesenchymal transition of gastric cancer cells”.In this article,we found the following error in Figure 3A:The panel image"24 h,sh-RNA1"in the AGS cells wound healing assay was incorrectly inserted during the preparation of the submission;the correct figure is provided in this correction.

BMSC-derived Exosomes Ameliorate Peritoneal Dialysis-associated Peritoneal Fibrosis via the Mir-27a-3p/TP53 Pathway

Objective:Peritoneal fibrosis(PF)is the main cause of declining efficiency and ultrafiltration failure of the peritoneum,which restricts the long-term application of peritoneal dialysis(PD).This study aimed to investigate the therapeutic effects and mechanisms of bone marrow mesenchymal stem cells-derived exosomes(BMSC-Exos)on PF in response to PD.Methods:Small RNA sequencing analysis of BMSC-Exos was performed by second-generation sequencing.C57BL/6J mice were infused with 4.25%glucose-based peritoneal dialysis fluid(PDF)for 6 consecutive weeks to establish a PF model.A total of 36 mice were randomly divided into 6 groups:control group,1.5%PDF group,2.5%PDF group,4.25%PDF group,BMSC-Exos treatment group,and BMSC-Exos+TP53 treatment group.Reverse transcription quantitative polymerase chain reaction(RT-qPCR)was performed to measure the expression level of miR-27a-3p in BMSC-Exos and peritoneum of mice treated with different concentrations of PDF.HE and Masson staining were performed to evaluate the extent of PF.The therapeutic potential of BMSC-Exos for PF was examined through pathological examination,RT-qPCR,Western blotting,and peritoneal function analyses.Epithelial-mesenchymal transition(EMT)of HMrSV5 was induced with 4.25%PDF.Cells were divided into control group,4.25%PDF group,BMSC-Exos treatment group,and BMSC-Exos+TP53 treatment group.Cell Counting Kit-8 assay was used to measure cell viability,and transwell migration assay was used to verify the capacity of BMSC-Exos to inhibit EMT in HMrSV5 cells.Results:Small RNA sequencing analysis showed that miR-27a-3p was highly expressed in BMSC-derived exosomes compared to BMSCs.The RT-qPCR results showed that the expression of miR-27a-3p was upregulated in BMSC-Exos,but decreased in PD mice.We found that PF was glucose concentration-dependently enhanced in the peritoneum of the PD mice.Compared with the control mice,the PD mice showed high solute transport and decreased ultrafiltration volume as well as an obvious fibroproliferative response,with markedly increased peritoneal thickness and higher expression ofα-SMA,collagen-I,fibronectin,and ECM1.The mice with PD showed decreased miR-27a-3p.Peritoneal structural and functional damage was significantly attenuated after BMSC-Exos treatment,while PF and mesothelial damage were significantly ameliorated.Additionally,markers of fibrosis(α-SMA,collagen-I,fibronectin,ECM1)and profibrotic cytokines(TGF-β1,PDGF)were downregulated at the mRNA and protein levels after BMSC-Exos treatment.In HMrSV5 cells,BMSC-Exos reversed the decrease in cell viability and the increase in cell migratory capacity caused by high-glucose PDF.Western blotting and RT-qPCR analysis revealed that BMSC-Exos treatment resulted in increased expression of E-cadherin(epithelial marker)and decreased expression ofα-SMA,Snail,and vimentin(mesenchymal markers)compared to those of the 4.25%PDF-treated cells.Importantly,a dual-luciferase reporter assay showed that TP53 was a target gene of miR-27a-3p.TP53 overexpression significantly reversed the decreases in PF and EMT progression induced by BMSC-Exos.Conclusion:The present results demonstrate that BMSC-Exos showed an obvious protective effect on PD-related PF and suggest that BMSC-derived exosomal miR-27a-3p may exert its inhibitory effect on PF and EMT progression by targeting TP53.

Effects of epidermal growth factor on transforming growth factor-beta1-induced epithelial-mesenchymal transition and potential mechanism in human corneal epithelial cells

AIM: To evaluate the effects of epidermal growth factor(EGF) on transforming growth factor-beta1(TGF-β1)-induced epithelial-mesenchymal transition(EMT) in human corneal epithelial cells(HCECs). METHODS: HCECs were cultured and treated with TGF-β1 for establishing the model of EMT in vitro. Biological effect of EGF on TGF-β1-induced EMT was evaluated. Proteins and m RNAs expression changes of E-cadherin, N-cadherin and Fibronectin(EMT-relative markers) after TGF-β1 or TGF-β1 combined EGF treatment were detected by Western blot and RT-PCR, respectively. Viability and migration of HCECs were measured by CCK-8, transwell cell migration assay and cell scratch wound healing assay. Activation of Smad2, ERK, p38, JNK and Akt signaling pathways were evaluated by Western blot. Inhibitors of relevant signaling pathways were added to the HCECs to explore the key signal mechanism.RESULTS: With treatment of TGF-β1 only, three EMTrelative proteins and m RNA expression showed that EMT up-regulated in a concentration-dependent and time-dependent manner, with significantly decreasing cell viability(TGF-β1≥5 ng/m L, P<0.05) and increasing cell migration(TGF-β1≥5 ng/m L, P<0.01). The phosphorylation of Smad2 and p38 was a key process of TGF-β1-induced EMT. Meanwhile, EMT-relative proteins and m RNA expression showed that EGF inhibited TGF-β1-indued EMT, with significantly increasing cell viability(EGF≥10 ng/m L, P<0.01). It was noteworthy that EGF significantly enhanced cell migration although EMT was inhibited(EGF≥10 ng/m L, P<0.01), and the blockage of p38(by SB202190, a p38 inhibitor) was a potential mechanism of this phenomenon. CONCLUSION: EGF inhibits TGF-β1-induced EMT via suppressive p38, and promotes cells proliferation and migration in a non-EMT process by inhibiting p38 pathway.

Prostate tumor neuroendocrine differentiation via EMT: The road less traveled

The long-standing challenge in the treatment of prostate cancer is to overcome therapeutic resistance during progression to lethal disease.Aberrant transforming-growth factor-b(TGF-b)signaling accelerates prostate tumor progression in a transgenic mouse model via effects on epithelial-mesenchymal transition(EMT),and neuroendocrine differentiation driving tumor progression to castration-resistant prostate cancer(CRPC).Neuroendocrine prostate cancer(NEPC)is highly aggressive exhibiting reactivation of developmental programs associated with EMT induction and stem cell-like characteristics.The androgen receptor(AR)is a critical driver of tumor progression as well as therapeutic response in patients with metastatic CRPC.The signaling interactions between the TGF-β mechanistic network and AR axis impact the EMT phenotypic conversions,and perturbation of epithelial homeostasis via EMT renders a critical venue for epithelial derived tumors to become invasive,acquire the neuroendocrine phenotype,and rapidly metastasize.Combinations of microtubule targeting taxane chemotherapy and androgen/AR targeting therapies have survival benefits in CRPC patients,but therapeutic resistance invariability develops,leading to mortality.Compelling evidence from our group recently demonstrated that chemotherapy(cabazitaxel,second line taxane chemotherapy),or TGF-β receptor signaling targeted therapy,caused reversion of EMT to mesenchymal-epithelial transition and tumor re-differentiation,in in vitro and in vivo prostate cancer models.In this review,we discuss the functional contribution of EMT dynamic changes to the development of the neuroendocrine phenotypedthe newly characterized pathological feature of prostate tumors in the context of the tumor microenvironment-navigated cell lineage changes and the role of this neuroendocrine phenotype in metastatic progression and therapeutic resistance.

Salivary Fusobacterium nucleatum serves as a potential diagnostic biomarker for gastric cancer

BACKGROUND As one of the most common tumors,gastric cancer(GC)has a high mortality rate,since current examination approaches cannot achieve early diagnosis.Fusobacterium nucleatum(Fn)primarily colonized in the oral cavity,has been reported to be involved in the development of gastrointestinal tumor.Until now,little is known about the relationship between salivary Fn and GC.AIM To determine whether salivary Fn could be a biomarker to diagnose GC and explore the influence of Fn on GC cells.METHODS The abundance of Fn in saliva was quantified by droplet digital polymerase chain reaction in 120 GC patients,31 atrophic gastritis(AG)patients,35 non-AG(NAG)patients,26 gastric polyp(GP)patients,and 20 normal controls(NC)from Qilu Hospital of Shandong University from January 2019 to December 2020.Receiver operating characteristic(ROC)curve analysis was performed to evaluate the diagnostic value of Fn as well as traditional serum tumor markers,including carcinoembryonic antigen(CEA),carbohydrate antigen(CA)19-9,and CA72-4.Transwell assay and wound-healing assay were conducted to assess the influence of Fn infection on GC cells.The expression of epithelial-mesenchymal transition(EMT)markers was detected using western blot assay.RESULTS We found that the level of salivary Fn in GC patients was significantly increased compared with those in AG,NAG,and GP patients and NC(P<0.001).ROC curve analysis showed a favorable capability of Fn(73.33% sensitivity;82.14%specificity;area under the curve:0.813)in GC diagnosis,which was superior to that of CEA,CA19-9,CA72-4,ferritin,and sialic acid.The Fn level in saliva of GC patients was increased as the TNM stage increased.GC patients with lymph node metastasis had higher Fn levels than those without metastasis.Both transwell and wound-healing assays indicated that Fn infection promoted the migration and invasion of GC cells.Western blot analysis showed that Fn infection decreased the expression of E-cadherin and increased the expressions of N-cadherin,vimentin,and snail.CONCLUSION Fn abundance in saliva could be used as a promising biomarker to diagnose GC,and Fn infection could promote GC metastasis by accelerating the EMT process.

LncRNA cancer susceptibility 20 regulates the metastasis of human gastric cancer cells via the miR-143-5p/MEMO1 molecular axis

BACKGROUND Gastric cancer(GC)is considered as one of the most widespread malignancies.Emerging evidence has shown that lncRNAs can function as important oncogenes or tumor suppressors during GC progression.AIM To investigate the effect and mechanism of lncRNA cancer susceptibility 20(CASC20)in the proliferation and metastasis of GC cells.METHODS Data mining and clinical samples were used to evaluate the expression of CASC20 in GC and adjacent tissues.CASC20 was down-regulated in GC cells by shortinterfering RNA.Cell proliferation was evaluated by CCK-8 assay,and cell migration and invasion were detected by wound healing and Transwell assays.The expressions of proteins related to epithelial-mesenchymal transition were detected by western blot assay.RESULTS The expression of CASC20 was increased in GC tumor tissues and various GC cell lines.High CASC20 expression was correlated with a high risk of lymphatic metastasis and poor prognosis in GC patients.In vitro assays showed that silencing CASC20 reduced cell proliferation,migration,and invasion in GC cells.Mechanistic studies revealed that CASC20 exhibits oncogenic functions by regulating MEMO1 expression through competitive endogenous binding to miR-143-5p,leading to induction of epithelial-mesenchymal transition.CONCLUSION Our findings indicate that CASC20 serves as a tumor promoter by regulating metastasis in GC via the miR-143-5p/MEMO1 axis.CASC20 may be a potential therapeutic target for GC.

Inhibition of bromodomain-containing protein 4 enhances the migration of esophageal squamous cell carcinoma cells by inducing cell autophagy

BACKGROUND Esophageal squamous cell carcinoma(ESCC),the predominant type of esophageal cancer,has a 5-year survival rate less than 20%.Although the cause of poor prognosis is the high incidence and mortality of ESCC,the high rate of metastasis after esophageal cancer surgery is the main cause of death after the surgery.Bromodomain-containing protein 4(BRD4),an epigenetic reader of chromatinacetylated histones in tumorigenesis and development,plays an essential role in regulating oncogene expression.BRD4 inhibition and BRD4 inhibition-based treatment can potentially suppress ESCC growth.However,the effects and mechanisms of action of BRD4 on ESCC cell migration remain unclear.AIM To explore the effect of BRD4 on cell migration of ESCC in vitro and its possible molecular mechanism.METHODS Human ESCC cell lines KYSE-450 and KYSE-150 were used.The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay was performed to examine cell proliferation,and the transwell migration assay was conducted to test ESCC cell migration.JQ1,a BRD4 inhibitor,was applied to cells,and BRD4 siRNA was transfected into ESCC cells to knockdown endogenous BRD4.GFP-RFP-LC3 adenovirus was infected into ESCC cells to evaluate the effect of JQ1 on autophagy.Western blotting was performed to determine the protein levels of BRD4,E-cadherin,vimentin,AMP-activated protein kinase(AMPK),and p-AMPK.RESULTS BRD4 was either downregulated by small interfering RNA or pretreated with JQ1 in ESCC cells,leading to increased tumor migration in ESCC cells in a dose-and time-dependent manner.Inhibition of BRD4 not only significantly suppressed cell proliferation but also strongly increased cell migration by inducing epithelial-mesenchymal transition(EMT).The protein expression of vimentin was increased and E-cadherin decreased in a dose-dependent manner,subsequently promoting autophagy in KYSE-450 and KYSE-150 cells.Pretreatment with JQ1,a BRD4 inhibitor,inhibited BRD4-induced LC3-II activation and upregulated AMPK phosphorylation in a dosedependent manner.Additionally,an increased number of autophagosomes and autolysosomes were observed in JQ1-treated ESCC cells.The autophagy inhibitor 3-methyladenine(3-MA)reversed the effects of BRD4 knockdown on ESCC cell migration and blocked JQ1-induced cell migration.3-MA also downregulated the expression of vimentin and upregulation E-cadherin.CONCLUSION BRD4 inhibition enhances cell migration by inducing EMT and autophagy in ESCC cells via the AMPK-modified pathway.Thus,the facilitating role on ESCC cell migration should be considered for BRD4 inhibitor clinical application to ESCC patients.

Understanding the biology of urothelial cancer metastasis

Management of unresectable urothelial cancer(UC)has been a clinical challenge for decades.While drug resistance is a key issue,precise understanding of biology of UC metastasis is another challenge for the improvement of treatment outcome of UC patients.Introduction of the cell biology concepts including epithelial-mesenchymal transition(EMT)and cancer stemness seems to explain UC metastasis.Molecular genetics based on gene expression profiling,next generation sequencing,and explosion of non-coding RNA world has opened the door to intrinsic molecular subtyping of UC.Next steps include,based on the recently accumulated understanding,the establishment of novel disease models representing UC metastasis in various experimental platforms,particularly in vivo animal systems.Indeed,novel knowledge molecular genetics has not been fully linked to the modeling of UC metastasis.Further understanding of bladder carcinogenesis is needed particularly with regard to cell of origin related to tumor characteristics including driver gene alterations,pathological differentiations,and metastatic ability.Then we will be able to establish better disease models,which will consequently lead us to further understanding of biology and eventually the development of novel therapeutic strategies for UC metastasis.

Adenomyosis-derived extracellular vesicles endow endometrial epithelial cells with an invasive phenotype through epithelial-mesenchymal transition

Extracellular vesicles from highly metastatic tumor cells have been shown to mediate epithelial-mesenchymal transition(EMT)-related events in recipient cells.In endometrial epithelial cells,EMT processes are known to be involved in the development of adenomyosis.We aimed to investigate whether adenomyosis-derived extracellular vesicles(AMEVs)are able to induce an EMT process in endometrial epithelial cells.In this study,AMEVs were isolated from patients with adenomyosis and characterized by transmission electron microscopy,Western blot,and nanoparticle tracking.Primary endometrial epithelial cells(EECs)were derived from normal endometrium tissues from patients with leiomyoma and co-cultured with AMEVs in vitro.AMEV uptake was examined by fluorescence confocal microscopy.The invasion of EECs was confirmed by Transwell assay.Immunohistochemistry,Western blot,and qRT-PCR were performed on EECs to illustrate the expression levels of cytokeratin 19,E-cadherin,vimentin,and zinc finger E-boxbinding homeobox 1(ZEB1).The results indicated that the cellular fluorescence intensity gradually increased after 48 h of co-culture,but decreased after 72 h.After co-culturing with AMEVs for 72 h,EECs expressed significantly lower levels of cytokeratin 19 and E-cadherin,and significantly higher levels of vimentin and ZEB1.Together these results demonstrated that AMEVs induce an EMT process and enhance the invasion of EECs.These changes may contribute to the pathogenesis and progression of adenomyosis.

Metabolic reprogram associated with epithelial-mesenchymal transition in tumor progression and metastasis

Epithelial-mesenchymal Transition(EMT)is a de-differentiation program that imparts tumor cells with the phenotypic and cellular plasticity required for drug resistance,metastasis,and recurrence.This dynamic and reversible events is governed by a network of EMT-transcription factors(EMT-TFs)through epigenetic regulation.Many chromatin modifying-enzymes utilize metabolic intermediates as cofactors or substrates;this suggests that EMT is subjected to the metabolic regulation.Conversely,EMT rewires metabolic program to accommodate cellular changes during EMT.Here we summarize the latest findings regarding the epigenetic regulation of EMT,and discuss the mutual interactions among metabolism,epigenetic regulation,and EMT.Finally,we provide perspectives of how this interplay contributes to cellular plasticity,which may result in the clinical manifestation of tumor heterogeneity.

Tetratricopeptide repeat domain 36 deficiency mitigates renal tubular injury by inhibiting TGF-β1-induced epithelialemesenchymal transition in a mouse model of chronic kidney disease

The damage of proximal tubular epithelial cells(PTECs)is considered a central event in the pathogenesis of chronic kidney disease(CKD)and deregulated repair processes of PTECs result in epithelialemesenchymal transition(EMT),which in turn aggravates tubular injury and kidney fibrosis.In this study,we firstly revealed that the reduction of TTC36 is associated with unilateral ureteral obstruction(UUO)-induced CKD;besides,ablation of TTC36 attenuated tubular injury and subsequent EMT in UUO-treated mice kidneys.Consistently,TTC36 overexpression promoted EMT in TGF-β1-induced HK2 cells.Moreover,TTC36 elevated the protein expression of CEBPB,which was involved in the regulation of TGF-β/SMAD3 signaling,and augmented SMAD3 signaling and downstream genetic response were reduced by CEBPB silencing.Collectively,our results uncovered that TTC36 deficiency plays a protective role in tubular injury and renal fibrosis triggered by UUO;further,TTC36 overexpression exacerbated TGF-β/SMAD3 signaling via elevating the stability of SMAD3 and CEBPB,suggesting that TTC36 inhibition may be a potential strategy in the therapy of obstructive nephropathy.