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.

CENPM基因表达下调的前列腺癌PC3细胞增殖、凋亡、侵袭能力及EMT表型变化

目的观察着丝粒蛋白M(CENPM)基因表达下调对前列腺癌PC3细胞增殖、凋亡、侵袭能力及上皮—间充质转化(EMT)表型的影响。方法体外培养人前列腺癌PC3细胞并分为实验组和对照组,实验组转染CENPM shRNA,对照组转染阴性对照shRNA。采用CCK-8法检测细胞增殖能力并计算增殖率,采用流式细胞术检测细胞凋亡情况并计算凋亡率,采用Transwell实验检测细胞侵袭能力,采用Western blotting法检测EMT相关蛋白E-cadherin、Vimentin、N-cadherin。结果实验组细胞增殖率、侵袭能力及Vimentin、N-cadherin蛋白表达低于对照组,细胞凋亡率及E-cadherin蛋白表达高于对照组(P均<0.05)。结论CENPM基因表达下调可降低前列腺癌PC3细胞的增殖和侵袭能力,抑制EMT,促进细胞凋亡。

PLCD3调控EMT进程促进结直肠癌细胞的增殖、侵袭和迁移

目的:研究磷脂酶PLCD3在结直肠癌组织和细胞中的表达情况和对结直肠癌细胞增殖、侵袭和迁移的影响及发生机制。方法:采用免疫组化分析PLCD3在结直肠癌组织中的表达,利用Kaplan-Meier Plotter数据库得知预后情况。使用RT-qPCR技术验证了PLCD3在人永生化结肠上皮细胞NCM460和结直肠癌细胞的表达水平,利用基因工具干预SW620和SW480细胞。采用克隆形成实验、CCK8增殖实验、划痕实验和Transwell实验来探究PLCD3对结直肠癌细胞增殖、迁移和侵袭的影响。运用Western blot验证EMT相关蛋白的变化。结果:PLCD3在结直肠癌组织中的表达高于癌旁组织(P<0.05),PLCD3高表达与预后生存率低密切相关(P<0.001)。敲低PLCD3抑制了SW620细胞增殖、侵袭和迁移,N-cadherin、MMP2、MMP9相对表达量显著降低(均P<0.01),E-cadherin表达水平显著升高(P<0.001)。过表达PLCD3促进了SW480细胞增殖、侵袭和迁移,N-cadherin、MMP2、MMP9相对表达量显著升高(均P<0.001),E-cadherin表达水平显著降低(P<0.0001)。结论:PLCD3在结直肠癌中表达上调,PLCD3可能通过调控EMT进程促进结直肠癌细胞的增殖、侵袭和迁移。

Induction of epithelial-mesenchymal transition (EMT) in human hepatocellular carcinoma after radiotherapy

Objective: Epithelial-mesenchymal transition (EMT) is a critical early event for the invasion and metastasis of many carcinomas. In the present study, we examined EMT markers in the residual cancer cells of hepatocellular carcinoma (HCC) after radiotherapy. Methods: Eight patients with large HCC who underwent hepatectomy with preoperative radiothera- py were studied. The expressions of E-cadherin and vimentin were determined immunohistochemically in the residual cancer cells of HCC following radiotherapy, and also in the pre-radiotherapy biopsy cancer cells. Results: Histological analysis showed that some residual cancer cells of HCC displayed an elongated spindle or fibroblast-like shape. The expression of E- cadherin was markedly reduced or negative in the spindle residual cancer cells, but the expression of vimentin significantly in- duced. However, the above changes were not found in the pre-radiotherapy biopsy cancer cells. Conclusion: EMT is induced in the residual cancer cells of HCC following radiotherapy, which may facilitate the systemic dissemination of cancer cells.

间充质干细胞来源外泌体对HDM刺激的气道上皮细胞炎症及EMT的影响

目的:探究间充质干细胞来源外泌体(MSC-EXO)对屋尘螨(HDM)刺激的气道上皮细胞炎症及上皮-间质转化(EMT)的影响。方法:从人脐带中分离MSCs(hUCMSCs),经过分选和鉴定后分离提取MSC-EXO。透射电镜和Western blot检测外泌体形态及标志蛋白CD63、CD81、TSG101表达。将MSC-EXO与HDM刺激的16HBE细胞共培养后,观察细胞形态变化;MTT检测细胞增殖能力;流式细胞术检测细胞凋亡率;qRT-PCR检测细胞炎症因子TNF-α和IL-6 mRNA水平;荧光探针法检测细胞活性氧(ROS)水平;Western blot检测EMT相关蛋白E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)、波形蛋白(VIM)、α平滑肌肌动蛋白(α-SMA)表达。结果:hUCMSC中CD73、CD90和CD105表达呈阳性,CD34、CD45和HLA-DR表达呈阴性;电子透射显微镜下MSC-EXO呈囊泡状,直径30~200 nm,表达CD63、CD81、TSG101。与对照组相比,HDM组16HBE细胞变长、变尖呈长梭形,细胞凋亡率、ROS、TNF-α和IL-6 mRNA水平、N-cadherin、VIM、α-SMA表达显著升高,E-cadherin表达显著降低(P<0.05);与HDM组相比,50、100μg/ml EXO组细胞形态接近正常细胞,细胞凋亡率、ROS、TNF-α和IL-6 mRNA水平、N-cadherin、VIM、α-SMA表达明显降低,E-cadherin表达明显升高(P<0.05)。结论:MSC-EXO可减轻HDM诱导的气道上皮细胞炎症,抑制EMT过程。

Molecular detection of epithelial-mesenchymal transition markers in circulating tumor cells from pancreatic cancer patients:Potential role in clinical practice

AIM To evaluate the clinical properties of three subpopulations of circulating tumor cells(CTCs) undergoing epithelial-mesenchymal transition(EMT) in pancreatic ductal adenocarcinoma(PDAC) patients.METHODS We identified CTCs for expression of the epithelial cell marker cytokeratin or epithelial cell adhesion molecule(EpCAM)(E-CTC), the mesenchymal cell markers vimentin and twist(M-CTC), or both(E/M-CTC) using the CanPatrol system. Between July 2014 and July 2016, 107 patients with PDAC were enrolled for CTC evaluation. CTC enumeration and classification were correlated with patient clinicopathological features and outcomes.RESULTS CTCs were detected in 78.5% of PDAC patients. The number of total CTCs ranged from 0 to 26 across all 107 patients, with a median value of six. CTC status correlated with lymph node metastasis, TNM stage, distant metastasis, blood lymphocyte counts, and neutrophil-to-lymphocyte ratio(NLR). Kaplan-Meier survival analysis showed that patients with ≥ 6 total CTCs had significantly decreased overall survival and progression-free survival compared with patients with < 6 total CTCs. The presence of M-CTCs was positively correlated with TNM stage(P < 0.01) and distant metastasis(P < 0.01). Additionally, lymphocyte counts and NLR in patients without CTCs were significantly different from those in patients testing positive for each CTC subpopulation(P < 0.01).CONCLUSION Classifying CTCs by EMT markers helps to identify the more aggressive CTC subpopulations and provides useful evidence for determining a suitable clinical approach.

Inflammatory microenvironment contributes to epithelial-mesenchymal transition in gastric cancer

Gastric cancer(GC) is the fifth most common malignancy in the world. The major cause of GC is chronic infection with Helicobacter pylori(H. pylori). Infection with H. pylori leads to an active inflammatory microenvironment that is maintained by immune cells such as T cells, macrophages, natural killer cells, among other cells. Immune cell dysfunction allows the initiation and accumulation of mutations in GC cells, inducing aberrant proliferation and protection from apoptosis. Meanwhile, immune cells can secrete certain signals, including cytokines, and chemokines, to alter intracellular signaling pathways in GC cells. Thus, GC cells obtain the ability to metastasize to lymph nodes by undergoing the epithelial-mesenchymal transition(EMT), whereby epithelial cells lose their epithelial attributes and acquire a mesenchymal cell phenotype. Metastasis is a leading cause of death for GC patients, and the involved mechanisms are still under investigation. In this review, we summarize the current research on how the inflammatory environment affects GC initiation and metastasis via EMT.

MiR-19a promotes epithelial-mesenchymal transition through PI3K/AKT pathway in gastric cancer

AIM: To investigate the mechanism by which mi R-19 a is up-regulated in gastric cancer(GC), which plays an oncogenic role.METHODS: In the present study, we investigated the role of mi R-19 a in gastric tissues as well as two GC cell lines. In vivo, we detected the basal expression level of mi R-19 a using real-time reverse transcription-PCR(RTPCR), and the relevance between expression of mi R-19 a and clinicopathological information was analyzed.In vitro, mi R-19 a was ectopically expressed using overexpression and knock-down strategies.RESULTS: Overexpression of mi R-19 a was significantly associated with metastasis of GC and inferior overall prognosis. However, no significant correlation was f o u n d b e t w e e n m i R- 1 9 a e x p r e s s i o n a n d o t h e r characteristics such as age, gender, tobacco, alcohol or tumor size. Cell proliferation, migration and invasion assays showed that overexpression of mi R-19 a promoted the proliferation, migration and invasion, and that overexpression of mi R-19 a promoted the epithelialmesenchymal transition through activating the PI3K/AKT pathway. Blocking the PI3K/AKT pathway could cancel the effect of mi R-19 a.CONCLUSION: All together, our results suggest that mi R-19 a could be used as a promising therapeutic target in the treatment of GC.

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.

Re-evaluating the role of epithelial-mesenchymal-transition in cancer progression

Epithelial-mesenchymal transition（EMT） and mesenchymal-epithelial transition（MET） are essential for embryonic development and also important in cancer progression. In a conventional model, epithelial-like cancer cells transit to mesenchymal-like tumor cells with great motility via EMT transcription factors; these mesenchymallike cells migrate through the circulation system, relocate to a suitable site and then convert back to an epithelial-like phenotype to regenerate the tumor. However, recent findings challenge this conventional model and support the existence of a stable hybrid epithelial/mesenchymal（E/M） tumor population. Hybrid E/M tumor cells exhibit both epithelial and mesenchymal properties, possess great metastatic and tumorigenic capacity and are associated with poorer patient prognosis. The hybrid E/M model and associated regulatory networks represent a conceptual change regarding tumor metastasis and organ colonization. It may lead to the development of novel treatment strategies to ultimately stop cancer progression and improve disease-free survival.

Transcriptional Factor Snail Mediates Epithelial-Mesenchymal Transition in Human Bronchial Epithelial Cells Induced by Silica

Epithelial-mesenchymal transition （EMT） plays an important role in fibrotic diseases. We have previously showed that silica induces EMT in human bronchial epithelial cells （BECs）; however, the underlying mechanism of silica-induced EMT is poorly understood. In the present study, we investigated the role of Snail in silica-induced EMT in human BECs in vitro. Human BECs were treated with silica at various concentrations and incubation times. Then MTr assay, western blot, electrophoretic mobility shift assay （EMSA）, and small interfering RNA （siRNA） transfection were performed. We found that silica increased the expression and DNA binding activity of Snail in human BECs. SNAI silica-induced expression siRNA upregulated the siRNA inhibited the of Snail. Moreover, SNAI expression of epithelial marker E-cadherin, but attenuated the expression of mesenchymal marker a-smooth muscle actin and vimentin in silica-stimulated cells. These results suggest that Snail mediates the silica-induced EMT in human BECs.

Strategies for Synchronous and Multiple Metastatic Liver Tumors Designed from Epithelial-Mesenchymal Transition Concept

At some point in the natural course of colorectal cancer up to 50% of patients will develop metastasis to the liver and it is one of the most critical effects for patient prognosis. The incidence of synchronous liver metastasis has been detected at around 20% - 25%, but the optimal timing of surgical resection remains controversial. Neoadjuvant chemotherapy has also been found to be beneficial not only for initially unresectable but also resectable synchronous metastases. Then, traditional surgical strategies of hepatic resection in accordance with past chemotherapeutic regimens have been used decreasingly over the past several years. This review will primarily discuss treatments in association with the recent developed chemotherapeutic regimens and surgical procedure from the clinical data and the concept for epithetlial-mesenchymal transition, which has recently been studied to elucidate mechanisms of the liver metastatic process.

MiR-663a Inhibits Radiation-Induced Epithelium-to-Mesenchymal Transition by Targeting TGF-β1

Objective miR-663 a has been reported to be downregulated by X-ray irradiation and participates in radiation-induced bystander effect via TGF-β1.The goal of this study was to explore the role of mi R-663 a during radiation-induced Epithelium-to-mesenchymal transition(EMT).Methods TGF-β1 or IR was used to induce EMT.After mi R-663 a transfection,cell migration and cell morphological changes were detected and the expression levels of mi R-663 a,TGF-β1,and EMT-related factors were quantified.Results Enhancement of cell migration and promotion of mesenchymal changes induced by either TGF-β1 or radiation were suppressed by mi R-663 a.Furthermore,both X-ray and carbon ion irradiation resulted in the upregulation of TGF-β1 and downregulation of mi R-663 a,while the silencing of TGF-β1 by mi R-663 a reversed the EMT process after radiation.Conclusion Our findings demonstrate an EMT-suppressing effect by mi R-663 a via TGF-β1 in radiationinduced EMT.

Silencing Neuropilin 1 gene reverses TGF-β1-induced epithelial mesenchymal transition in HGC-27 gastric cancer cell line

Objective The aim of this study was to determine Neuropilin 1(NRP1)contribution to transforming growth factorβ1(TGF-β1)-induced epithelial mesenchymal transition(EMT)of HGC-27 gastric cancer cells and study its mechanism.Methods In this study,TGF-β1 was used to induce EMT in HGC-27 cells.Further,these cells were stably transfected with siRNA targeting NRP1.Wound healing and transwell assays were used to measure cell migration and invasion,respectively.NRP1 and EMT markers were measured using quantitative real time reverse transcription polymerase chain reaction and western blotting.Results Exposure of TGF-β1 conferred a fibroblastic-like shape to cancer cells and significantly increased the expression of NRP1 in HGC-27 cells.TGF-β1 subsequently promoted migration and invasion of HGC-27 cells.Furthermore,silencing NRP1 inhibited the invasion and migration of TGF-β1-induced cells undergoing EMT.Conclusion Silencing NRP1 can inhibit cell migration,invasion,and metastasis and reverse the TGF-β1-induced EMT process of gastric cancer.

Effect of Rapamycin on TGF-β_1-induced epithelial-mesenchymal transition in LoVo colonic adenocarcinoma cells

Objective： To investigate the effect of Rapamycin on epithelial-mesenchymal transition（EMT） of LoVo colonic adenocarcinoma cells in vitro. Methods：Cultured LoVo colonic adenocarcinoma cells were divided into three groups： negative control group, EMT-inducing group（TGF-β1） and EMT-interfering group（TGF-β1 plus Rapamycin）. E-cadherin expression in LoVo cells was detected by Western Blot, while the expression of vimentin was evaluated through immunocytochemistry. The Snail mRNA in LoVo cells was examined by RT- PCR. Results：TGF-β1 induced LoVo cell switching from polygonal to spindle-shaped. TGF-β1 enhanced the expression of vimentin, but lowered the level of E-cadhefin. In contrast, Rapamycin impaired the transition induced by TGF-β1. Rapamycin dramatically abrogated TGF-β1-induced vimentin expression and restored E-cadherin expression in LoVo cells. Rapamycin significantly repressed the upregulation of Snail mRNA expression induced by TGF-β1. Conclusion：Rapamycin dramatically abrogated TGF-β1 induced Snail mRNA expression in LoVo cells, hence inhibiting EMT of these cells in vitro.

The roles of micro RNAs and epithelial-mesenchymal transition in colorectal cancer metastasis

Colorectal cancer(CRC) is the second most common cause of cancer death worldwide. Distant metastasis is the major cause of death in patients with CRC. During progression to metastasis in which malignant cells disseminate from the primary tumor to seeding other organs, a multistep process is involved. Cancer cells proliferate, invade microenvironment, enter into the blood circulation, then survive and colonize into distant organs. Micro RNAs(mi RNAs) and epithelial-mesenchymal transition(EMT) are key regulators and mechanism in tumorigenesis and cancer metastasis. We review the roles of EMT and micro RNAs, especially EMT related micro RNAs in the metastatic pathway of CRC. Micro RNAs provide us a set of potential therapeutic applications and molecular target for CRC.

白藜芦醇通过Hippo-YAP信号通路在TGF-β1诱导胃癌细胞EMT过程中的作用及其机制

目的探讨白藜芦醇(RSVL)通过Hippo-YAP信号通路在TGF-β1诱导胃癌细胞上皮间充质转化(EMT)过程中的作用及其机制。方法选取胃癌细胞SGC-7901为研究对象,首先将其随机分为空白组、RSVL低剂量组(5μM)、RSVL中剂量组(10μM)、RSVL高剂量组(20μM),通过细胞增殖(MTT)实验确定RSVL浓度,然后对细胞进行转染,分为si-NC组、TGF-β1+10μM RSVL组、TGF-β1+si-YAP组、TGF-β1+pcDNA3.1-YAP组、TGF-β1+10μM RSVL+pcDNA3.1-YAP组。采用MTT、细胞侵袭(Transwell)、划痕实验,分别检测细胞的增殖、侵袭和迁移;采用蛋白质印迹法(WB)和荧光定量PCR检测E-钙粘连蛋白(E-cadherin)、神经型钙黏附蛋白(N-cadherin)、波形蛋白(Vimentin)、Snali 1、HIP-PO/Yes相关蛋白(YAP)和mRNA。其次,选取24只裸鼠,将其分为模型组、RSVL组(10μM)、RSVL+pcDNA3.1组、RSVL+pcDNA3.1-YAP组,每组各六只。计算小鼠肿瘤的重量和体积;采用免疫组化检测Ki67蛋白。结果RSVL对细胞增殖有抑制作用(P<0.05);与si-NC组相比,其余各组侵袭细胞数、迁移率较低(P<0.05);与si-NC组相比,其余各组E-cadherin蛋白及mRNA表达较高,N-cadherin、Vimentin、Snali 1蛋白及mRNA以及YAP蛋白表达较低(P<0.05);与Model组相比,其余各组小鼠肿瘤的重量和体积均较低(P<0.05);与Model组比较,其余各组Ki67染色强度均显著减弱。结论RSVL可以通过抑制YAP相关通路的激活,来发挥抑制肿瘤细胞SGC-7901上皮间充质转化、增殖、迁移、侵袭的作用。

低氧微环境通过TGFBI调控Wnt/β-catenin通路介导胰腺癌化疗耐药及机制研究

目的:研究低氧微环境通过TGFBI调控胰腺癌耐药的作用及分子机制。方法:以CCK-8方法检测细胞增殖;以Western blotting技术检测蛋白表达水平;以ImageJ软件分析蛋白灰度值;RNAi技术用于敲减TGFBI基因。结果:TGFBI在Panc-1细胞中表达比正常细胞增强;低氧促进胰腺癌细胞Panc-1增殖并减弱顺铂对Panc-1的抑制作用,而高氧抑制Panc-1细胞增殖并加强顺铂的杀伤作用;低氧促进TGFBI表达及EMT行为;低氧通过TGFBI调控Panc-1细胞增殖及顺铂的杀伤作用;低氧通过TGFBI调控Wnt/β-catenin信号,进而促进EMT行为。结论:低氧微环境通过增强TGFBI表达促进胰腺癌细胞增殖及耐药;低氧微环境通过TGFBI激活Wnt/β-catenin信号通路,促进EMT标志分子表达。