Role of Protein Kinase C in Advanced Glycation End Products-induced Epithelial-Mesenchymal Transition in Renal Proximal Tubular Epithelial Cells

The role of protein kinase C （PKC） activation in advanced glycation end products （AGEs）-induced epithelial-mesenchymal transition in renal proximal tubular epithelial cells was investigated. HKC cells were divided into three groups： normal group, AGE-BSA group （100 mg/L AGE-BSA） and AGE-BSA＋PKC inhibitor （10 μmol/L chelerythrine chloride） group. PKC activity was measured by PKC assay kit. The expression of Vimentin, and phosphorylated β-catenin was detected by using Western blotting, and the content of TGF-β1 was examined by ELISA method. The intracellular disposition of Vimentin was observed by fluorescence microscopy. As compared with normal group, PKC activity was increased significantly in AGE-BSA group. The expression of Vimentin, phosphorylated β-catenin, and TGF-β1 was enhanced significantly in AGE-BSA group. The expression of Vimentin, phosphorylated β-catenin, and TGF-β1 was significantly blocked by chelerythrine chloride. High expression of Vimentin, phosphorylated β-catenin, and TGF-β1 induced by AGE-BSA may be mediated via the activation of PKC signal transduction pathway.

Neural Wiskott-Aldrich syndrome protein(N-WASP)promotes distant metastasis in pancreatic ductal adenocarcinoma via activation of LOXL2

Pancreatic ductal adenocarcinoma(PDAC)is one of the most aggressive solid malignancies.A specific mechanism of its metastasis has not been established.In this study,we investigated whether Neural Wiskott-Aldrich syndrome protein(N-WASP)plays a role in distant metastasis of PDAC.We found that N-WASP is markedly expressed in clinical patients with PDAC.Clinical analysis showed a notably more distant metastatic pattern in the N-WASP-high group compared to the N-WASP-low group.N-WASP was noted to be a novel mediator of epithelialmesenchymal transition(EMT)via gene expression profile studies.Knockdown of N-WASP in pancreatic cancer cells significantly inhibited cell invasion,migration,and EMT.We also observed positive association of lysyl oxidase-like 2(LOXL2)and focal adhesion kinase(FAK)with the N-WASP-mediated response,wherein EMT and invadopodia function were modulated.Both N-WASP and LOXL2 depletion significantly reduced the incidence of liver and lung metastatic lesions in orthotopic mouse models of pancreatic cancer.These results elucidate a novel role for N-WASP signaling associated with LOXL2 in EMT and invadopodia function,with respect to regulation of intercellular communication in tumor cells for promoting pancreatic cancer metastasis.These findings may aid in the development of therapeutic strategies against pancreatic cancer.

Epithelial-mesenchymal transition mediated tumourigenesis in the gastrointestinal tract

Epithelial-mesenchymal transition (EMT) is a highly conserved process that has been well characterised in embryogenesis. Studies have shown that the aberrant activation of EMT in adult epithelia can promote tumour metastasis by repressing cell adhesion molecules,including epithelial (E)-cadherin. Reduced intracellular adhesion may allow tumour cells to disseminate and spread throughout the body. A number of transcription proteins of the Snail superfamily have been implicated in EMT. These proteins have been shown to be over-expressed in advanced gastrointestinal (GI) tumours including oesophageal adenocarcinomas,colorectal carcinomas,gastric and pancreatic cancers,with a concomitant reduction in the expression of E-cadherin. Regulators of EMT may provide novel clinical targets to detect GI cancers early,so that cancers previously associated with a poor prognosis such as pancreatic cancer can be diagnosed before they become inoperable. Furthermore,pharmacological therapies designed to inhibit these proteins will aim to prevent local and distant tumour invasion.

The role of mechanical stretch and TGF-β2 in epithelial-mesenchymal transition of retinal pigment epithelial cells

AIM: To explore the effects and mechanisms of mechanical stress and transforming growth factor-beta2(TGF-β2) on epithelial-mesenchymal transition(EMT) in cultured human retinal pigment epithelial(RPE) cells. METHODS: Human RPE cells were inoculated on BioF ex 6-well plates and RPE cells received 0, 1, 2, 3, or 4 mild stretch injuries delivered 3h apart after 24h of culture. The device of mechanical stress parameters were set to sine wave, frequency 1 Hz, stretch strength 20%. For treatment with TGF-β2, when the inoculated RPE cells in 6-well plates were around 60% confluent, serum was reduced to 0 for 12h and recombinant human TGF-β2(0, 1, 5, 10 ng/mL)was added for 48h. α-SMA, Vimentin and N-Cadherin, fibronectin proteins expressions were detected by Western blotting, confocal cell immunofluorescence and quantitative real-time polymerase chain reaction(q RT-PCR). Then we detected the change of mi RNA-29b and ascertained the changes of phosphatidylinositol 3-kinase-serine threonine protein kinase(PI3K/Akt) pathway after RPE cells were stretched by the device of mechanical stress and induced by TGF-β2 by Western blotting, confocal cell immunofluorescence and qR T-PCR. RESULTS: Mechanical stress induce EMT and activate the PI3K/Akt pathway in ways that lead to the EMT process. TGF-β2 induce RPE cells EMT and in a certain range and TGF-β2 decrease the miR NA-29b expression in RPE cells, and the inhibitory effect is more obvious with the increase of TGF-β2 concentration. CONCLUSION: Our findings are crucial steps in determining the critical roles of the PI3K/Akt signaling pathway and mi RNA-29b in pathogenesis of proliferative vitreoretinopathy(PVR) which may be a potential target for preventing or treating PVR.

EDIL3 depletion suppress epithelial-mesenchymal transition of lens epithelial cells via transforming growth factor β pathway

AIM： To study the effect of discoidin I-like domaincontaining protein 3（EDIL3） depletion on the proliferation and epithelial-mesenchymal transition（EMT） in human lens epithelial cells（LECs）. METHODS： RNA interference was used to inhibit the expression of EDIL3 in human LECs in vitro. The morphology of cells was observed using an inverted microscope. Cell proliferation was assessed using Ed U kit. Cell migration was investigated using Transwell chamber and EMT of LECs was assessed using confocal microscope and Western blotting. The transforming growth factor β（TGFβ） pathway was investigated using Western blotting. RESULTS： The data showed that silencing EDIL3 expression changed LECs morphology and suppressed LECs proliferation（P〈0.05） and migration（P〈0.01）. Furthermore, the result of Western blotting showed that EDIL3 depletion reduced the expression of α-smooth muscle actin（α-SMA）（P〈0.001） and vimentin（P〈0.01）, while increased the expression of E-cadherin（P〈0.001）. EDIL3 depletion could suppress the phosphorylation of Smad2（P〈0.01） and Smad3（P〈0.01） and the activation of exracellular signal regulated kinase（ERK）（P〈0.05）. CONCLUSION： The findings indicate that EDIL3 might participate in the proliferation and EMT in LECs via TGFβ pathway and may be a potential therapeutic target for the treatment of posterior capsule opacification.

Effects of chimeric intron on the epithelial-mesenchymal transformation of non-small cell lung cancer PC-9

Objective:To investigate the effects of Intron on the EMT capability of non-small cell lung cancer cell line PC-9.Methods:Firstly,using the psiCHECK-2 plasmid as a basic framework to construct the recombinant plasmid of psiCHECK-2-Intron dual-luciferase reporter gene;secondly,the psiCHECK-2-Intron and psiCHECK-2 were transfected into PC-9 cells respectively.The migration and invasion abilities of PC-9 cells were analyzed by Matrigel assay.The expression changes of EMT related hallmarks,including N-cadherin,β-catenin and snail,were detected by qRT-PCR and Western Blotting.Results:Compared with the control group,the migration and invasion abilities of PC-9 cells in Intron group significantly decreased(p<0.001).The expression of N-cadherin,β-catenin and snail also down-regulated(p<0.001).Conclusion:The introns could inhibit the EMT of PC-9 cells.

Inhibition of EGFR attenuates EGF-induced activation of retinal pigment epithelium cell via EGFR/AKT signaling pathway

AIM:To explore the effect of epidermal growth factor receptor(EGFR)inhibition by erlotinib and EGFR siRNA on epidermal growth factor(EGF)-induced activation of retinal pigment epithelium(RPE)cells.METHODS:Human RPE cell line(ARPE-19 cells)was activated by 100 ng/mL EGF.Erlotinib and EGFR siRNA were used to intervene EGF treatment.Cellular viability,proliferation,and migration were detected by methyl thiazolyl tetrazolium(MTT)assay,bromodeoxyuridine(BrdU)staining assay and wound healing assay,respectively.EGFR/protein kinase B(AKT)pathway proteins and N-cadherin,α-smooth muscle actin(α-SMA),and vimentin were tested by Western blot assay.EGFR was also determined by immunofluorescence staining.RESULTS:EGF treatment for 24h induced a significant increase of ARPE-19 cells’viability,proliferation and migration,phosphorylation of EGFR/AKT proteins,and decreased total EGFR expression.Erlotinib suppressed ARPE-19 cells’viability,proliferation and migration through down regulating total EGFR and AKT protein expressions.Erlotinib also inhibited EGF-induced an increase of proliferative and migrative ability in ARPE-19 cells and clearly suppressed EGF-induced EGFR/AKT proteins phosphorylation and decreased expression of N-cadherin,α-SMA,and vimentin proteins.Similarly,EGFR inhibition by EGFR siRNA significantly affected EGF-induced an increase of cell proliferation,viability,and migration,phosphorylation of EGFR/AKT proteins,and up-regulation of N-cadherin,α-SMA,and vimentin proteins.CONCLUSION:Erlotinib and EGFR-knockdown suppress EGF-induced cell viability,proliferation,and migration via EGFR/AKT pathway in RPE cells.EGFR inhibition may be a possible therapeutic approach for proliferative vitreoretinopathy(PVR).

Saponin Ⅰ from Shuitianqi(Rhizoma Schizocapasae Plantagineae) inhibits metastasis by negatively regulating the transforming growth factor-β1/Smad7 network and epithelial-mesenchymal transition in the intrahepatic metastasis Bagg's Albino/c mouse model

OBJECTIVE:To examine the influence of SaponinⅠfrom Shuitianqi(Rhizoma Schizocapasae Plantagineae)(SSPHⅠ)on hepatocellular carcinoma(HCC)metastasis,and to elucidate the underlying mechanism.METHODS:The intrahepatic metastasis Bagg's Albino/c(BALB/c)mouse model was established with human hepatocellular carcinomas(HepG2)cells,then treated with normal saline(once per day),cisplatin(2 mg/kg,once every 2 d),and SSPHⅠ(25,50,and 75 mg/kg,once per day).Then,we assessed alterations in the hepatic pathology and target protein expressions in the intrahepatic metastasis BALB/c mouse model using a series of molecular biology techniques.RESULTS:Based on our analysis,SSPHⅠsignificantly alleviated hepatocyte necrosis and tumor cells infiltration.Moreover,SSPHⅠsuppressed extracellular matrix(ECM)degradation and angiogenesis via a decrease in matrix etalloproteinase-2(MMP-2),MMP-9,CD31,CD34,and vascular endothelial growth factor(VEGF)levels.Furthermore,SSPHⅠrepressed invasion and metastasis by suppressing the transforming growth factor-β1(TGF-β1)/Smad7 axis and epithelial-mesenchymal transition(EMT),as evidenced by the scarce TGF-β1,Ncadherin,and Vimentin expressions,and elevated Smad7 and E-cadherin expressions.CONCLUSION:The SSPHⅠ-mediated negative regulation of the TGF-β1/Smad7 axis and EMT are critical for the inhibition of HCC invasion and metastasis.

MiR-3653 blocks autophagy to inhibit epithelial-mesenchymal transition in breast cancer cells by targeting the autophagy-regulatory genes ATG12 and AMBRA1

Background:Metastasis is the main cause of tumor-associated death and mainly responsible for treatment failure of breast cancer.Autophagy accelerates tumor metastasis.In our work,we aimed to investigate the possibility of microRNAs(miRNAs)which participate in the regulation of autophagy to inhibit tumor metastasis.Methods:MiRNA array and comprehensive analysis were performed to identify miRNAs which participated in the regulation of autophagy to inhibit tumor metastasis.The expression levels of miR-3653 in breast cancer tissues and cells were detected by quantitative real-time polymerase chain reaction.In vivo and in vitro assays were conducted to determine the function of miR-3653.The target genes of miR-3653 were detected by a dual luciferase reporter activity assay and Western blot.The relationship between miR-3653 and epithelial-mesenchymal transition(EMT)was assessed by Western blot.Student’s t-test was used to analyze the difference between any two groups,and the difference among multiple groups was analyzed with one-way analysis of variance and a Bonferroni post hoc test.Results:miR-3653 was downregulated in breast cancer cells with high metastatic ability,and high expression of miR-3653 blocked autophagic flux in breast cancer cells.Clinically,low expression of miR-3653 in breast cancer tissues(0.054±0.013 vs.0.131±0.028,t=2.475,P=0.014)was positively correlated with lymph node metastasis(0.015±0.004 vs.0.078±0.020,t=2.319,P=0.023)and poor prognosis(P<0.001).miR-3653 ameliorated the malignant phenotypes of breast cancer cells,including proliferation,migration(MDA-MB-231:0.353±0.013 vs.1.000±0.038,t=16.290,P<0.001;MDA-MB-468:0.200±0.014 vs.1.000±0.043,t=17.530,P<0.001),invasion(MDA-MB-231:0.723±0.056 vs.1.000±0.035,t=4.223,P=0.013;MDA-MB-468:0.222±0.016 vs.1.000±0.019,t=31.050,P<0.001),and colony formation(MDA-MB-231:0.472±0.022 vs.1.000±0.022,t=16.620,P<0.001;MDA-MB-468:0.650±0.040 vs.1.000±0.098,t=3.297,P=0.030).The autophagy-associated genes autophagy-related gene 12(ATG12)and activating molecule in beclin 1-regulated autophagy protein 1(AMBRA1)are target genes of miR-3653.Further studies showed that miR-3653 inhibited EMT by targeting ATG12 and AMBRA1.Conclusions:Our findings suggested that miR-3653 inhibits the autophagy process by targeting ATG12 and AMBRA1,thereby inhibiting EMT,and provided a new idea and target for the metastasis of breast cancer.

Secreted protein acidic and rich in cysteine(SPARC)induces epithelial-mesenchymal transition,enhancing migration and invasion,and is associated with high Gleason score in prostate cancer

Secreted protein acidic and rich in cysteine(SPARC)is a matricellular protein highly expressed in bone tissue that acts as achemoattractant factor promoting the arrival of prostate cancer(PCa)cells to the bone marrow.However,the contribution of SPARCduring the early stages of tumor progression remains unclear.In this study,we show that SPARC is highly expressed in PCa tissueswith a higher Gleason score.Through stable knockdown and overexpression of SPARC in PC3 and LNCaP cells,respectively,here wedem on strate that en doge nous SPARC induces the epithelial-mesenchymal tran sition(EMT),decreasing E-cadheri n and cytokeratin18 and increasing N-cadheri n and vime ntin.Moreover,SPARC in duces the expression of EMT regulatory tran scription factors Snailfamily transcriptional repressor 1(Snail),Snail family transcriptional repressor 2(Slug),and zinc finger E-box binding homeobox 1(Zeb1).In addition,SPARC knockdown in PC3 cells decreases migration and invasion in vitro,without modifying cell proliferation.Our results indicate that SPARC might facilitate tumor progression by modifying the cellular phenotype in cancer cells.

Inhibition of Nonsmall Cell Lung Cancer Cell Migration by Protein Arginine Methyltransferase 1-small Hairpin RNA Through Inhibiting Epithelial-mesenchymal Transition,Extracellular Matrix Degradation, and Src Phosphorylation In Vitro

Background：Protein arginine methyltransferases 1 （PRMT1） is over-expressed in a variety of cancers,including lung cancer,and is correlated with a poor prognosis of tumor development.This study aimed to investigate the role of PRMT1 in nonsmall cell lung cancer （NSCLC） migration in vitro.Methods：In this study,PRMT1 expression in the NSCLC cell line A549 was silenced using lentiviral vector-mediated short hairpin RNAs.Cell migration was measured using both scratch wound healing and transwell cell migration assays.The mRNA expression levels of matrix metalloproteinase 2 （MMP-2） and tissue inhibitor ofmetalloproteinase 1,2 （TIMP l,2） were measured using quantitative real-time reverse transcription-polymerase chain reaction.The expression levels of protein markers for epithelial-mesenchymal transition （EMT） （E-cadherin,N-cadherin）,focal adhesion kinase （FAK）,Src,AKT,and their corresponding phosphorylated states were detected by Western blot.Results：Cell migration was significantly inhibited in the PRMT1 silenced group compared to the control group.The mRNA expression of MMP-2 decreased while TIMP 1 and TIMP2 increased significantly.E-cadherin mRNA expression also increased while N-cadherin decreased.Only phosphorylated Src levels decreased in the silenced group while FAK or AKT remained unchanged.Conclusions：PRMT1-small hairpin RNA inhibits the migration abilities of NSCLC A549 cells by inhibiting EMT,extracellular matrix degradation,and Src phosphorylation in vitro.

Expression and significance of E-cadherin,N-cadherin,transforming growth factor-β1 and Twist in prostate cancer

Objective:To study the expression of E-cadherin,N-cadherin,TGF-|3 1 and Twist protein and investigate its significance in the occurrence and development of prostate cancer.Methods:The expression of E-cadherin,N-cadherin,TGF-β1 and Twist protein in 59 prostate cancer tissues and 21 adjacent tissues were detected by immunohistochemical SABC staining,and the correlation with clinicopathological features was analyzed.Results:Positive rates of E-cadherin,N-cadherin,TGF-β1 and Twist were 32.2%,54.2%,71.2%and 74.6%,respectively,in prostate cancer tissues and 85.7%,9.52%,19.0%and 9.52%,respectively,in cancer—adjacent tissues,with significant differences between the two groups(P<0.05).The reduced expression of E-cadherin was related to the differentiation of prostate cancer tissues and PSA level,but was not associated with clinical stage,lymph node metastasis,bony metastasis and age.The increased expression of N-cadherin,TGF-β1 and Twist was related to the differentiation of prostate cancer tissues,clinical stage,lymph node metastasis,bony metastasis,but not to age.The difference in positive expression of N-cadherin and TGF-β1 was significant between PSA≤20μg/L group and PSA>20μg/L group,but the positive expression of Twist was not significant between groups.The expression of E-cadherin was highly negatively correlated with that of N-cadherin and also highly negatively correlated with that of Twist The expression of TGF-β1 was correlated with those of E-cadherin,N-cadherin and Twist.Conclusions:The reduced expression of E-cadherin,abnormal expression of N-cadherin,transformation form E-cadherin to N-cadherin and the increased expression of TGF-β1 and Twist play an important role in the occurrence and development of prostate cancer.

MAWBP and MAWD inhibit proliferation and invasion in gastric cancer

AIM: To investigate role of putative mitogen-activated protein kinase activator with WD40 repeats (MAWD)/ MAWD binding protein (MAWBP) in gastric cancer (GC). METHODS: MAWBP and MAWD mRNA expression level was examined by real-time reverse transcriptasepolymerase chain reaction and semi-quantitative polymerase chain reaction in six GC cell lines. Western blotting was used to examine the protein expression levels. We developed GC cells that stably overexpressed MAWBP and MAWD, and downregulated expression by RNA interference assay. Proliferation and migration of these GC cells were analyzed by 3-(4,5-dimethyl2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT), soft agar, tumorigenicity, migration and transwell assays. The effect of expression of MAWBP and MAWD on transforming growth factor (TGF)-β1-induced epithelialmesenchymal transition (EMT) was examined by transfection of MAWBP and MAWD into GC cells. We detected the levels of EMT markers E-cadherin, N-cadherin and Snail in GC cells overexpressing MAWBP and MAWD by Western blotting. The effect of MAWBP and MAWD on TGF-β signal was detected by analysis of phosphorylation level and nuclear translocation of Smad3 using Western blotting and immunofluorescence. RESULTS: Among the GC cell lines, expression of endogenous MAWBP and MAWD was lowest in SGC7901 cells and highest in BGC823 cells. MAWBP and MAWD were stably overexpressed in SGC7901 cells and knocked down in BGC823 cells. MAWBP and MAWD inhibited GC cell proliferation in vitro and in vivo . MTT assay showed that overexpression of MAWBP and MAWD suppressed growth of SGC7901 cells (P < 0.001), while knockdown of these genes promoted growth of BGC823 cells (P < 0.001). Soft agar colony formation experiments showed that overexpression of MAWBP and MAWD alone or together reduced colony formation compared with vector group in SGC7901 (86.25±8.43, 12.75±4.49, 30±6.41 vs 336.75±22.55, P < 0.001), and knocked-down MAWBP and MAWD demonstrated opposite effects (131.25±16.54, 88.75±11.12, 341.75±22.23 vs 30.25±8.07, P < 0.001). Tumorigenicity experiments revealed that overexpressed MAWBP and MAWD inhibited GC cell proliferation in vivo (P < 0.001). MAWBP and MAWD also inhibited GC cell invasion. Transwell assay showed that the number of traverse cells of MAWBP, MAWD and coexpression group were more than that in vector group (84±16.57, 98.33±9.8, 29±16.39 vs 298±11.86, P < 0.001). Coexpression of MAWBP and MAWD significantly decreased the cells traversing the matrix membrane. Conversely, knocked-down MAWBP and MAWD correspondingly promoted invasion of GC cells (100.67±14.57, 72.66±8.51, 330.67±20.55 vs 27±11.53, P < 0.001). More importantly, coexpression of MAWBP and MAWD promoted EMT. Cells that coexpressed MAWBP and MAWD displayed a pebble-like shape and tight cell-cell adhesion, while vector cells showed a classical mesenchymal phenotype. Western blotting showed that expression of E-cadherin was increased, and expression of N-cadherin and Snail was decreased when cells coexpressed MAWBP and MAWD and were treated with TGF-β1. Nuclear translocation of p-Smad3 was reduced by attenuating its phosphorylation. CONCLUSION: Coexpression of MAWBP and MAWD inhibited EMT, and EMT-aided malignant cell progression was suppressed.

S100A8 promotes epithelial-mesenchymal transition and metastasis under TGF-β/USF2 axis in colorectal cancer

Background:The transforming growth factor-β(TGF-β)pathway plays a pivotal role in inducing epithelial-mesenchymal transition(EMT),which is a key step in cancer invasion and metastasis.However,the regulatory mechanism of TGF-βin inducing EMT in colorectal cancer(CRC)has not been fully elucidated.In previous studies,it was found that S100A8 may regulate EMT.This study aimed to clarify the role of S100A8 in TGF-β-induced EMT and explore the underlying mechanism in CRC.Methods:S100A8 and upstream transcription factor 2(USF2)expression was detected by immunohistochemistry in 412 CRC tissues.Kaplan-Meier survival analysis was performed.In vitro,Western blot,and migration and invasion assays were performed to investigate the effects of S100A8 and USF2 on TGF-β-induced EMT.Mouse metastasis models were used to determine in vivo metastasis ability.Luciferase reporter and chromatin immunoprecipitation assay were used to explore the role of USF2 on S100A8 transcription.Results:During TGF-β-induced EMT in CRC cells,S100A8 and the transcription factor USF2 were upregulated.S100A8 promoted cell migration and invasion and EMT.USF2 transcriptionally regulated S100A8 expression by directly binding to its promoter region.Furthermore,TGF-βenhanced the USF2/S100A8 signaling axis of CRC cells whereas extracellular S100A8 inhibited the USF2/S100A8 axis of CRC cells.S100A8 expression in tumor cells was associated with poor overall survival in CRC.USF2 expression was positively related to S100A8 expression in tumor cells but negatively related to S100A8-positive stromal cells.Conclusions:TGF-βwas found to promote EMT and metastasis through the USF2/S100A8 axis in CRC while extracellular S100A8 suppressed the USF2/S100A8 axis.USF2 was identified as an important switch on the intracellular and extracellular S100A8 feedback loop.

Wulong Xiaozheng Wan medicated serum inhibits epithelial-mesenchymal transition in human gastric carcinoma cell line BGC823 by modulation of transforming growth factor-β1/Smad signaling

OBJECTIVE:To evaluate the effect of Wulong Xiaozheng Wan medicated serum on the epithelial-mesenchymal transition (EMT) of BGC823 cell induced by transforming growth factor-β,(TGF-β,) and to explore its mechanism.METHODS:EMT model of BGC823 was stimulated by TGF-β1.Wulong Xiaozheng Wan medicated serum and LY-364947 were used as intervention.The proliferation and adhesion of BGC823 were detected by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide and flow cytometry was used to detect the apoptosis.The invasion and migration were detected by Transwell.The level of matrix metalloproteins was detected by enzyme-linked immunosorbent assay.The expressions of related proteins and mRNA of EMT marker and TGF-β1/Smad signal pathway were detected by Western blot and reverse transcription-polymerase chain reaction.RESULTS:Compared with the TGF-β1 group,Wulong Xiaozheng Wan medicated serum could inhibit the ability of proliferation,heterogeneous adhesion,invasion,and migration.It also promotes apoptosis and homotypic adhesion in BGC823,with a dose-dependent manner.Meanwhile,Wulong Xiaozheng Wan medicated serum could regulate the expression of related proteins and mRNA of TGF-β1/Smad signaling pathway,and inhibit the expressions of EMT transcription factors and EMT markers.CONCLUSION:Wulong Xiaozheng Wan medicated serum inhibited epithelial-mesenchymal transition by down-regulated the expression of TβRI and the activation of TGF-β1/Smad signaling pathway.

The loss-of-function mutations and down-regulated expression of ASB3 gene promote the growth and metastasis of colorectal cancer cells

Background: Ankyrin repeat and SOCS box protein 3(ASB3) is a member of ASB family and contains ankyrin repeat sequence and SOCS box domain. Previous studies indicated that it mediates the ubiquitination and degradation of tumor necrosis factor receptor 2 and is likely involved in inflammatory responses. However, its effects on oncogenesis are unclear. This study aimed to investigate the effects of ASB3 on the growth and metastasis of colorectal cancer(CRC).Methods: We used next?generation sequencing or Sanger sequencing to detect ASB3 mutations in CRC specimens or cell lines, and used real?time quantitative polymerase chain reaction, Western blotting, and immunohistochemical or immunofluorescence assay to determine gene expression. We evaluated cell proliferation by MTT and colony for?mation assays, tested cell cycle distribution by flow cytometry, and assessed cell migration and invasion by transwell and wound healing assays. We also performed nude mouse experiments to evaluate tumorigenicity and hepatic metastasis potential of tumor cells.Results: We found that ASB3 gene was frequently mutated(5.3%) and down?regulated(70.4%) in CRC cases. Knock?down of endogenous ASB3 expression promoted CRC cell proliferation, migration, and invasion in vitro and facilitated tumorigenicity and hepatic metastasis in vivo. Conversely, the ectopic overexpression of wild?type ASB3, but not that of ASB3 mutants that occurred in clinical CRC tissues, inhibited tumor growth and metastasis. Further analysis showed that ASB3 inhibited CRC metastasis likely by retarding epithelial?mesenchymal transition, which was characterized by the up?regulation of β?catenin and E?cadherin and the down?regulation of transcription factor 8, N?cadherin, and vimentin.Conclusion: ASB3 dysfunction resulted from gene mutations or down?regulated expression frequently exists in CRC and likely plays a key role in the pathogenesis and progression of CRC.

CtBP家族在肿瘤中的相关研究进展

肿瘤的发生、发展是个多因素,多步骤,多阶段的复杂过程,其中涉及到了多个因素的相互作用,羧基末端结合蛋白(c—terminal—bindingprotein,CtBP)在进化上保守,主要发挥转录辅抑制因子的功能。研究发现CtBP与多个肿瘤相关基因的转录调控有关,促进肿瘤细胞的迁移/侵袭/转移, 激活WNT通路,参与肿瘤细胞的上皮-间充质转化(EMT),肿瘤细胞“Warburg效应”, 促进肿瘤干细胞自我更新等多个过程,在肿瘤的发生、发展过程中起到了重要作用。因此,CtBP的研究对于揭示肿瘤的发生、发展,以及肿瘤的治疗,预防具有重要意义,本文就近年来CtBP肿瘤相关的研究作一综述。

Apelin aggravates the migration and invasion of non-small cell lung cancer cells via YAP1

Background:Apelin,an endogenous ligand of G-protein coupled receptor(GPCR),is a secreted peptide involved in the development of various tumors.However,the relationship between apelin and non-small cell lung cancer(NSCLC)is not quite clear.This study was designed to investigate the effect and mechanism of apelin on cell proliferation,migration and invasion of NSCLC cells.Methods:Twelve NSCLC specimens were collected for hematoxylin-eosin(HE)staining and immunohistochemistry analyses.Cell proliferation was examined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)and cell migration and invasion were assessed using wound-healing and transwell assays.The subcellular location of yes associated protein 1(YAP1)in A549 cells was determined by immunofluorescence.The mRNA and protein levels in NSCLC tissues and cell lines were measured by qRT-PCR and western blot,respectively.Results:Apelin was upregulated in tumor tissues compared with the adjacent tissues.Apelin promoted proliferation,migration,and invasion of A549 and H460 cells,which was reversed by competitive apelin receptor(APJ)antagonist ML221.Additionally,apelin upregulated YAP1 expression,whereas silence of YAP1 by small interfering RNA(siRNA)attenuated apelin-induced cell proliferation,migration and invasion and suppressed epithelial-mesenchymal transition progression.Conclusion:Apelin promotes NSCLC cells proliferation,migration,and invasion by modulating YAP1 and might be a potential therapeutic target for NSCLC treatment.

Change of HOXB7 gene expression in colon cancer lesions and its correlation with the pathological characteristics of tumor

Objective: To study the change of homeobox protein 7 (HOXB7) gene expression in colon cancer lesions and its correlation with the pathological characteristics of tumor. Methods: The patients with colon cancer who underwent radical resection in Tangshan Hospital of Traditional Chinese Medicine between March 2015 and February 2018 were selected as the research subjects, and right amount of colon cancer lesions and lesions adjacent to carcinoma were collected to measure the expression of HOXB7 gene, cell proliferation-related genes Notch1, Hes1, P53, ASPP2 and WTX as well as invasion-related genes CAT-D, TMEM16a, CBX7, ZEB1, E-cadherin and N-cadherin. Results: HOXB7, Notch1, Hes1, CAT-D, TMEM16a, CBX7, ZEB1 and N-cadherin mRNA expression in colon cancer lesions were higher than those in adjacent lesions whereas P53, ASPP2, WTX and E-cadherin mRNA expression were lower than those in adjacent lesions;the mRNA of HOXB7 gene in colon cancer was positively correlated with the mRNA expression of Notch1, Hes1, CAT-D, TMEM16a, CBX7, ZEB1 and N-cadherin, and negatively correlated with the mRNA expression of P53, ASPP2, WTX and E-cadherin. Conclusion: The high expression of HOXB7 gene in the colon cancer lesions can enhance the pathological characteristics of tumor proliferation and invasion.

Relationship of GPRC6A and PKCzeta expression levels in prostate cancer lesions with cell proliferation and EMT

Objective: To study the relationship of GPRC6A and PKCzeta expression levels in prostate cancer lesions with cell proliferation and epithelial-mesenchymal transition (EMT). Methods:Patients with prostate cancer and patients with benign prostatic hyperplasia who received surgical resection in Huizhou Third People's Hospital between June 2014 and March 2017 were selected, right amount of prostate cancer tissue and tissue adjacent to carcinoma was collected from patients with prostate cancer, and the expression of GPRC6A, PKCzeta, cell proliferation genes and EMT genes in lesions were detected. Results: GPRC6A, Survivin, SRSF1, Bcl-xl, N-cadherin and Vimentin expression in prostate cancer lesions and adjacent lesions were significantly higher than those in benign prostatic hyperplasia lesions while PKCzeta, Caspase-3, Caspase-9, Apaf-1, E-cadherin and CK5/6 expression were significantly lower than those in benign prostatic hyperplasia lesions;Survivin, SRSF1 and Bcl-xl expression in prostate cancer lesions with lower PKCzeta expression were significantly higher than those in prostate cancer lesions with higher PKCzeta expression while Caspase-3, Caspase-9 and Apaf-1 expression were significantly lower than those in prostate cancer lesions with higher PKCzeta expression;E-cadherin and CK5/6 expression in prostate cancer lesions with lower GPRC6A expression were significantly higher than those in prostate cancer lesions with higher GPRC6A expression while N-cadherin and Vimentin expression were significantly lower than those in prostate cancer lesions with higher GPRC6A expression. Conclusion:Highly expressed GPRC6A and lowly expressed PKCzeta in prostate cancer lesions can promote cell EMT and proliferation respectively.