Expression of ezrin protein and phosphorylated ezrin in pelvic endometriotic lesions

Abstract Objective: To evaluate the expression of Ezrin and Phosphorylated Ezrin (Phospho-Ezrin) in endometriosis lesions and its relation to the menstrual cycle phase, stage of endometriosis, histological classification, and clinical symptoms. Material and methods: The authors conducted a retrospective study, with endometriotic lesions collected from women with endometriosis (n = 57) who underwent laparoscopy from 2017 to 2018. The expression of Ezrin and Phosphorylated Ezrin proteins was analyzed by immunohistochemistry. Results: All the endometriotic lesions contained immunostaining for Ezrin in the glands. Phosphorylated Ezrin was expressed in the stroma of all endometriotic lesions. There was no difference in the Ezrin and Phosphorylated Ezrin's expression in the retrocervical, ovarian, superficial, and intestinal lesions in the same patient. Dysmenor-rhea, dyspareunia, acyclic pain, infertility, and dysuria were similar in the three groups of Ezrin staining. There was an inversely proportional relationship between severe dyschezia and Ezrin's intensity, being 66.7% of Ezrin 1 (weak intensity), 36.7 Ezrin 2 (moderate intensity), and 10.0% of Ezrin 3 (p = 0.013). Regarding Phospho-Ezrin there wasn't a significant difference between all the analyzed variables. Histological classification and menstrual cycle phase had also no significant difference between Ezrin and Phospho-Ezrin immunostaining. Conclusion: Ezrin protein and Phospho-Ezrin can be considered important markers to elucidate the mechanisms related to migration and attachment of endometriotic lesions. It is still unclear if Ezrin and Phospho-Ezrin are a cause or consequence of endometriosis. Further studies comparing different types of lesions and eutopic endometrium are necessary to elucidate the role of these proteins in the pathogenesis of endometriosis. HIGHLIGHTS The implantation of endometrial cells in the pelvic cavity has been related to some factors such as a receptive environment that allows the implantation and proliferation of these cells. Several studies have shown the participation of the Ezrin protein in the process of invasion of malignant cells. The expression of Ezrin and its activated form was observed in endometriotic lesions providing great evidence that these proteins can play an important role in the migration and attachment of endometriotic lesions.

Ezrin Promotes the Proliferation, Migration, and Invasion of Ovarian Cancer Cells / 生物医学与环境科学（英文）

Objective@#The underlying mechanism of Ezrin in ovarian cancer (OVCA) is far from being understood. Therefore, this study aimed to assess the role of Ezrin in OVCA cells (SKOV3 and CaOV3) and investigate the associated molecular mechanisms.@*Methods@#We performed Western blotting, reverse transcription-quantitative polymerase chain reaction, MTT, cell colony, cell wound healing, transwell migration and invasion, RhoA and Rac active pull down assays, and confocal immunofluorescence experiments to evaluate the functions and molecular mechanisms of Ezrin overexpression or knockdown in the proliferation and metastasis of OVCA cells.@*Results@#The ectopic expression of Ezrin significantly increased cell proliferation, invasiveness, and epithelial-mesenchymal transition (EMT) in OVCA cells. By contrast, the knockdown of endogenous Ezrin prevented OVCA cell proliferation, invasiveness, and EMT. Lastly, we observed that Ezrin can positively regulate the active forms of RhoA rather than Rac-1 in OVCA cells, thereby promoting robust stress fiber formation.@*Conclusion@#Our results indicated that Ezrin regulates OVCA cell proliferation and invasiveness by modulating EMT and induces actin stress fiber formation by regulating Rho-GTPase activity, which provides novel insights into the treatment of the OVCA.

Identification of ezrin gene transcriptional regulatory region and its gRNA target site in pancreatic cancer cells / 医学研究生学报

Objectiveezrin gene is overexpressed in pancreatic cancer, and its upstream sequence plays an important role in gene expression. This study intends to knock out ezrin transcriptional regulatory region and identify its gRNA target sites for gene editing in pancreatic cancer cells.MethodsThe reporter gene expression vectors carrying the upstream segment of ezrin gene were transiently transfected into Panc-1 cells. The ezrin transcriptional regulatory regions were identified by double luciferase reporter gene detection system. Then, the online software was utilized to predict the gRNA target sites located at the upstream and downstream of ezrin transcriptional regulatory region. Two recombinant plasmids pX459-sgRNA-L and pX458-sgRNA-R contained these two sequences were constructed for gene editing. Moreover, in order to identify the targeted knockout of ezrin transcriptional regulatory region, the recombinant plasmids were co-transfected into Panc-1 cells, and the genome DNA contained gRNA target sites were amplified, subcloned and sequenced. Finally, Panc-1 cells transfected with recombinant plasmids were preliminary sorted using puromycin treatment. The cell proliferation was detected by water-soluble tetrazolium salt method.ResultsLuciferase data showed that ezrin gene fragment -1297/-1186 enhanced the transcriptional activity of SV40 promoter and ezrin promoter in Panc-1 cells. Subclonal sequencing data revealed that the recombinant plasmids carrying the gRNA target sequence of ezrin transcriptional regulatory region were co-transfected into Panc-1 cells could trigger the genomic DNA fragments, which located between gRNA-L and gRNA-R target sites. Cell proliferation assay showed that the proliferation was significantly inhibited after transfection.ConclusionThe targeted knockout of ezrin transcriptional regulatory region was achieved and the inhibition of Panc-1 cell proliferation may be related to this knockout.

Expression of survivin, fibronectin-1, VEGF and ezrin in thyroid tumors and their relationship with tumor pathological characteristics / 中国肿瘤生物治疗杂志

@# Objective: To investigate the expression of survivin, fibronectin-1, vascular endothelial growth factor (VEGF) and ezrin in thyroid tumors and their relationship with the pathological characteristics of thyroid tumors. Methods: Ninety patients with thyroid tumors admitted to the third affiliated hospital of Zunyi Medical University and the first hospital during Oct. 2016 and Oct. 2018 were selected as the observation group. Seventy-five patients with normal thyroid confirmed by pathology in the same period were selected as the control group. The protein levels of survivin, fibronectin-1, VEGF and ezrin were detected by immunohistochemical method. Results: The positive rates of survivin, fibronectin-1, VEGF and Ezrin in the control group were 2.67%, l4.00%, 1.33% and 1.33%, which were lower than 97.78%, 96.67%, 93.33% and 95.56% in the observation group, respectively (all P<0.05 or P<0.01). The expressions of survivin, fibronectin-1, VEGF and ezrin were significantly correlated with TNM staging, tumor diameter, extrathyroid invasion and lymphatic metastasis (all P<0.05). Conclusion: Survivin, fibronectin-1, VEGF and ezrin proteins are all involved in the occurrence and development of thyroid tumors. The combined detection of these four indicators is of great significance in the diagnosis, treatment and prognosis of thyroid tumors.

Ezrin enhancer knockout inhibits the proliferation and migration of human esophageal carcinoma Eca-109 cells / 中国肿瘤生物治疗杂志

@#Objective: To investigate the effects of ezrin enhancer knockout on ezrin gene expression, cell proliferation and migration of human esophageal carcinoma Eca-109 cells. @*@#Methods: The CRISPR/Cas9 recombinant plasmids targeting upstream/downstream of human ezrin enhancer were co-transfected into human esophageal carcinoma Eca-109 cells, and the cell line Eca-C2 with ezrin enhancer knockout was screened by purinomycin. Then the expression levels of ezrin mRNAand protein in Eca-C2 cells were detected by Real-time quantitative PCR (qPCR) and Western blotting, respectively; The expression levels of MAPK-pathway-related proteins were detected by protein array technology; and the effects of ezrin enhancer knockout on the proliferation and migration of Eca-C2 cells were analyzed by WST-1 method and wound-healing assay, respectively. @*@# Results:The human esophageal carcinoma cell line Eca-C2 with stable ezrin enhancer knockout was established successfully. Compared with control cells, the mRNA and protein expressions of ezrin in Eca-C2 cells were significantly reduced (all P<0.05).Among the 17 detected MAPK pathway related proteins in Eca-C2 cells, 9 proteins (AKT, CREB, GSK3b, MKK6, mTOR, P38, P53, P70S6K and RSK1) were down-regulated, and the cell proliferation and migration were significantly inhibited (all P<0.05).@*@# Conclusion: ezrin enhancer knockout can significantly inhibit the cell proliferation and migration of human esophageal carcinoma Eca-109 cells.

Effect of apoptosis and proliferation of endometrial cancer HEC-1B cell by siRNA down-regulating expression of Ezrin / 中国综合临床

Objective To observe the changes of cell cycle,apoptosis and proliferation of endometrial cancer cells after the expression and down-regulation of Ezrin in endometrial cancer cells and to explore whether Ezrin may be a candidate gene for targeted therapy.Methods Endometrial cancer cells were from Shanghai Institute of Cell Research,of Chinese Academy of Medical Sciences in February 2017 and divided into blank control group and siEzrin group according to the intervention methods.Western blot and qRT-PCR was used to detect the expression of Ezrin protein and mRNA in endometrial cell lines.Small interfering RNA (siRNA) was used to transfect HEC-1B cell and down-regulate Ezrin.Cell cycle and apoptosis were detected by flow cytometry.MTT assay was used to detect multiplication.Results Western blot showed that Ezrin protein was expressed in Ishikawa (31.742 ± 5.832)、HEC-1A (16.326 ± 3.135)、HEC-1B(17.636±4.426) and KLE(14.862±5.109) and qRT-PCR showed that mRNA was expressed in Ishikawa (2.513±0.725),HEC-1A (1.655±0.692),HEC-1B (3.237±0.411) and KLE (0.962±0.235) cell lines,and expressed highest in HEC-1B cells (F=6.173,P＜0.05;F=7.042,P＜0.05).Flow cytometry assay showed that compared with blank control group less cells stayed in G1 phase and G2 phase,more stayed in S phase (t=3.118,P＜0.05;t=5.435,P＜0.05;t=3.332,P＜0.05).The apoptotic rate of HEC-1B cells increased from (9.84 ± 2.37) ％ to (17.64 ± 5.96) ％ (t =8.963,P ＜ 0.01) after Ezrin was downregulated.MTT assay showed that the proliferation of HEC-1B cells in 72 h and 96 h siEzrin transfection group was lower than that in blank control group (t =3.209,P＜ 0.05;t =3.726,P＜ 0.05).Conclusion Down-regulating of Ezrin may promote more endometrial cancer cells stay in S phase and promote apoptosis,inhibit proliferation,Ezrin may become target candidate gene in target therapy.

Inhibitory effect of nvp-bez235 on proliferation and migration of hepatocellular carcinoma cell hepg2 and its mechanism / 中国药理学通报

Aim To evaluate the efficacy of NVP-BEZ235 on the proliferation and migration of HepG2 human hepatocellular carcinoma cell in vitro, and to investigate the molecular mechanism. Methods The cytotoxicity of NVP-BEZ235 on HepG2 cells treated with NVP-BEZ235(0, 25, 50, 100, 250, 500 nmol·L-1) was detected by MTT assay. The ability of NVP-BEZ235 to modulate HepG2 cells proliferation and apoptosis was detected by morphology assay and Hoechst 33342 staining. The motility of NVP-BEZ235 treatment on HepG2 cells was determined by wound healing assay and Transwell assay.The expression levels of the related protein including PI3K/Akt/mTOR signal pathway, epithelial-mesenchymal transition(EMT) process and Six1/Ezrin signal axis were detected by Western blot. The expression levels of EMT markers were also detected by immunofluorescence staining. Results The cell viability significantly decreased in HepG2 cells treated with NVP-BEZ235 compared with control group, which showed a dose-dependent manner. NVP-BEZ235 could induce apoptosis and inhibit HepG2 cell migration. E-cadherin protein expression significantly increased; however, the expressions of p-AktSer473, p-S6Thr389, p-4E-BP1Thr37/46, Vimentin, Snail, Six1 and p-EzrinTyr353 decreased in NVP-BEZ235 treated HepG2 cells compared with those in control group. Conclusions NVP-BEZ235 could effectively suppress the proliferation and migration of HepG2 cells, which may be related to the down-regulation of p-AktSer473, p-S6Thr389, p-4E-BP1Thr37/46 and modulating both Six1/Ezrin signal axis and the EMT process by NVP-BEZ235 treatment.

Advances in biological functions of Ezrin and its effects on tumor metastasis / 中国药理学通报

As a member of the ERM (Ezrin/Radixin/Moesin) protein family, Ezrin is widely distributed in the body. Ezrin acts as a "scaffold" participating in anchorage and interacting between plasma membrane and cytoskeleton. Its special subcel-lular localization is critical for many complex cell processes. Increasing evidence suggests that the abnormal expression, phosphorylation and localization of Ezrin would affect tumor progression. The influence of Ezrin on the morphology of tumor cells during metastasis has gradually attracted the attention of researchers. Further investigations that focus on the mechanism of Ezrin' s influence on different stages of tumor metastasis will be gradually elucidated. In this article, we review the biological functions of Ezrin and its research progress in tumor metastasis, and explain the mechanism of Ezrin-mediated tumor metastasis. It is proposed that strategies targeting Ezrin for tumor metastasis treatment are a promising way to achieve great success in clinic.

Effect of Yishen Tongluo Formula on kidney protection and podocyte skeleton related proteins in rats with membranous nephropathy / 中草药

Objective To explore the kidney protection and possible mechanism of Yishen Tongluo Formula (YTF) in rats with membranous nephropathy (MN). Methods A total of 60 SD healthy male rats were randomly divided into 10 for normal group and 50 for MN rat model group. The MN rat model was established by tail iv cationic bovine serum albumin (C-BSA). After successful modeling, they were randomly divided into model group, benazepril group, and YTF groups at low, medium, and high doses (6.61, 13.22, and 26.44 g/kg). Rats in each group were ig administrated once daily for continuous four weeks according to the corresponding dose. At the end of administration, the 24 h urine total protein (UTP), total cholesterol (TC), total protein (TP), albumin (ALB), blood urea nitrogen (BUN), and serum creatinine (Scr) levels were measured. Immunofluorescence was used to detect the deposition of IgG immune complexes in renal tissue. The glomerular basement membrane and podocyte morphology were observed under electron microscope. Immunohistochemistry and qRT-PCR were used to detect the expression of cytoskeleton-related proteins ezrin and synaptopodin in rat kidneys. Results Compared with the model group, the UTP and TC levels in the rats in each treatment group decreased significantly (P < 0.01), and the TP and ALB levels increased significantly (P < 0.01). The middle and high dose groups of YTF were similar to the benazepril group, and the effect was better than the low dose group of YTF. There was no significant difference in the BUN and Scr among the groups. Compared with the control group, the expression of ezrin and synaptopodin mRNA in the kidney of the model group was significantly decreased (P < 0.01). Compared with the model group, the expression of ezrin and synaptopodin mRNA in the podocytes of different treatment groups was increased in different degrees (P < 0.01). The expression levels of ezrin and synaptopodin mRNA in the kidney of rats in the middle and high dose groups of YTF were similar to those in the benazepril group, which were higher than that in the low dose group of YTF. Conclusion YTF has a therapeutic effect on membranous nephropathy in rats. The mechanism may be related to the inhibition of the degradation of podocyte skeleton related proteins ezrin and synaptopodin and the maintenance of the integrity of the podocyte skeleton and foot process.

The expressions and clinical significance of Ezrin and PTEN protein in intrahepatic cholangiocarcinoma / 中华肝胆外科杂志

Objective To study the expressions and the clinicopathologic significance of Ezrin and PTEN protein in intrahepatic cholangiocarcinoma.Methods The ICC tissues (the ICC group) and the corresponding para-carcinoma tissues (the control group) were collected from 50 patients with intrahepatic cholangiocarcinoma (IHC) to detect the expressions of Ezrin and PTEN protein by using SP immunohistochemistry (IHC).The clinicopathologic parameters were analyzed.Results The positive expression rates of Ezrin were 78.0％ (39/50) and 46.0％ (23/50) in the ICC group and the control group,respectively.The difference in expression between the two groups was statistically significance (P＜0.05).The expression of Ezrin in the ICC group was highly related to tumor size,differentiation grade,TNM stage,and lymphatic metastasis (P＜0.05),but it was not related to age,gender,serum CA19-9 level,hepatitis B virus infection,intrahepatic duct calculus (P＞0.05).The positive expression rates of PTEN was 46.0％ (23/50) and 88.0％ (44/50) in the ICC group and the control group,respectively.The difference in expression between the two groups was statistically significance (P＜0.05).The expression of PTEN in the ICC group was highly related to differentiation grade,TNM stage,and lymphatic metastasis (P＜0.05),but it was not related to age,gender,serum CA19-9 level,hepatitis B virus infection,tumor size,intrahepatic duct calculus (P＞0.05).There was a negative correlation between the expression of Ezrin and PTEN protein in ICC (r=-0.382,P＜0.01).Conclusion The abnormal and negative correlation between the expression of Ezrin and PTEN protein in ICC may play an important role in invasion and metastasis of ICC.

Ezrin promotes metastasis of lung cancer via L1CAM signaling pathway / 重庆医学

Objective To investigate the molecular mechanism of Ezrin in promoting human lung cancer metastasis.Methods The expression of Ezrin in lung cancer cell lines was detected by using RT-PCR and western blot;the cell scratch test was used to detect the effect of Ezrin on the migration ability of lung cancer cells;the mechanism of Ezrin for regulating L1CAM was detected by western blot.Results The western blot detection results showed that compared with the low metastatic lung cancer cell line H460 and EBC-1,the expression of Ezrin protein the high metastatic lung cancer cell line 95D and PC9 were higher (P＜0.05).After silencing the expression of Ezrin in 95D cells (siEzrin-95D) by using the genetic method,the cell scratch test showed that the migration ability of siEzrin-95D cells was significantly weakened compared with 95D cells (P＜0.05).Compared with 95D cells and H460 cells,after genetically silencing the Ezzin expression in 95D and H460 cells,the L1CAM expression was significantly down-regulated (P＜0.05).Conclusion Ezrin promotes lung cancer metastasis possibly by regulating the expression of L1CAM.

Overexpression of ezrin and galectin-3 as predictors of poor prognosis of cervical cancer

The aim of this study was to explore the correlation of ezrin and galectin-3 expressions with prognosis in cervical cancer. The immunohistochemical method was applied to detect ezrin and galectin-3 expressions in normal cervix tissues (n=30), cervicitis tissues (n=28), cervical intraepithelial neoplasia (CIN) tissues (classified as I-III, n=89), and cervical carcinoma tissues (n=84). Follow-up was conducted for 5 to 78 months to analyze the correlation of protein expressions with prognosis. Ezrin and galectin-3 expressions in cervical cancer were significantly higher than in normal cervix, cervicitis and CIN (all P<0.05), and expressions in CIN were significantly higher than in normal cervix and cervicitis (both P<0.05). The expressions of ezrin and galectin-3 were both related with histological grade, deep myometrial invasion and lymph node metastasis (all P<0.05). Spearman analysis showed that ezrin expression was positively correlated with galectin-3 expression in cervical cancer (r=0.355, P<0.05). The survival rate of patients with high expressions of ezrin and galectin-3 was significantly lower than those with low expressions of proteins (both P<0.05). The expressions of ezrin and galectin-3, histological grade, depth of stromal invasion, and lymph node metastasis are risk factors affecting the survival rate of patients with cervical cancer. The expressions of ezrin and galectin-3 were correlated with the development of cervical cancer, and overexpressions of those proteins were indicative of poor prognosis in patients with cervical cancer.

Experimental research of miR-132 inhibits proliferation and induces apoptosis of ovarian cancer via Ezrin / 中国免疫学杂志

Objective:To explore the biological function of miR-132 in ovarian cancer and the target. Methods: 22 cases ovarian cancer tissue and non-tumor tissue adjacent were collected,the expression of miR-132 in tumor tissue and non-tumor tissue, normal ovarian epithelial cells and ovarian cancer cell were detected by RT-PCR. The normal ovarian epithelial cells which the expression of miR-132 maximum or minimum were chosen, and they were divided into two groups, respectively with transfection of negative control plasmid ( NC) and miR-132 mimic plasmid. The expression of miR-132 after transfection was detected by RT-PCR,the cell proliferation and cell apoptosis were detected by CCK-8 method and flow cytometry instrument respectively,the expression of Ezrin protein was detected by Western blot. Results:The expression of miR-132 in tumor tissue was significantly lower than the tumor tissue adjacent,the expression of miR-132 in ovarian cancer cell lines was significantly lower than normal ovarian epithelial cells, the differences were statistically significant (P<0. 05). The SKOV3 cell lines was chosed for gene transfection,compared with NC group, transfection with miR-132 mimic plasmid could significantly reduce cell proliferation, increase cell apoptosis, the difference had statistical significance ( P<0. 05 ) . Western blot results showed that up-regulation miR-132 significantly increased the Ezrin protein expression in ovarian cancer SKOV3 cells ( P<0. 05 ) . Conclusion: In ovarian cancer, miR-132;inhibits proliferation and induces apoptosis of ovarian cancer via Ezrin,it may be a tumor suppressor gene.

Expression of signal-induced proliferation-associated gene 1, Ezrin and E-cadherin proteins in epithelial ovarian carcinoma and their clinical significances / 肿瘤研究与临床

Objective To investigate the expression of signal-induced proliferation-associated gene 1 (SIPA1), Ezrin and E-cadherin (E-cad), and their relationship with clinical patterns in epithelial ovarian carcinoma.Methods Immunohistochemistry was used to detect the expression of SIPA1, Ezrin and E-cad in normal ovarian tissue, benign epithelial ovarian tumor, borderline epithelial ovarian tumor and epithelial ovarian carcinoma,respectively. Results The positive rate of SIPA1 expression was 44.2 % (23/52), 64.5 %(20/31), 93.3 % (28/30) and 100.0 % (15/15) in epithelial ovarian carcinoma, borderline epithelial ovarian tumor, benign epithelial ovarian tumor, and normal ovarian tissue, respectively, and there was a statistical difference (χ2 = 29.159, P= 0.000). The corresponding rates were 57.7 % (30/52), 61.3 % (19/31), 90.0 %(27/30) and 93.3 % (14/15) for the positive rate of Ezrin expression (χ2= 14.555, P= 0.002), as well as for 23.1 % (12/52), 58.1 % (18/31), 86.7 % (26/30) and 0 (0/15) for the positive rate of E-cad expression, respectively (χ2= 45.731, P= 0.000). In patients with epithelial ovarian carcinoma, the expression of SIPA1 was correlated with tumor differentiation (χ2=3.895, P=0.048), but not with histological type and clinical stage (all P>0.05). The expression of Ezrin was not correlated with histological type, tumor differentiation and clinical stage (all P>0.05). There was a positive correlation between expression of E-cad and SIPA1, Ezrin in epithelial ovarian carcinoma, respectively (r= 0.339, P= 0.014; r= 0.284, P= 0.041), but no correlation between the expression of SIPA1 and Ezrin (r= 0.214, P= 0.128). Conclusions SIPA1, Ezrin and E-cad play important roles in the occurrence and development of epithelial ovarian carcinoma. They cooperate in the progression and their combined detection can better evaluate the prognosis of epithelial ovarian carcinoma.

RhoA/mDia1 pathway involved in the expression of p-ERM in the pulmonary micro-vascular endothelial cell induced by lipopolysaccharide / 中华急诊医学杂志

Objective To investigate the possibility of the involvement of RhoA/mDia1 pathway to cause the expression of phosphorylate ezrin-radixin-moesin (p-ERM) in rat pulmonary micro-vascular endothelial cell (PMVEC) after the stimulation of lipopolysaccharide (LPS).Methods The specimens of lung tissue were taken from healthy male SPF grade SD rat with 100-120 g body weight which was purchased from the laboratory animal center of Anhui province.After culture,the PMVECs were randomly divided into dose-dependent groups (0,0.1,1,10 μg/mL LPS added in PMVECs and cultured for30 min,n =8 in each),time-dependent groups (10 μg/mL LPS added to PMVECs cultured for 0,15,30,60,120 min,n =8 in each) and intervention group (n =8).In the intervention group,PMVECs were cultured with 1 μg/mL C3 transferase in serum free media for 240 min,followed by treatment with 10 μg/mL LPS for 30 min.Meanwhile,two control groups in serum-free DMEM medium were made by adding 10.μg/mL LPS to PMVECs and 1 μg/mL C3 transferase to PMVECs respectively cultured for 30 min (n =8 in each).Western blot was used to detect the level of p-ERM,ERM and mDia1.Data were analyzed with SPSS 16.0 software,while one way analysis of variance (ANOVA) was used to compare multiple sets of variables,the intergroup comparisons were analyzed by the least-significant-difference (LSD) tests,with P ＜0.05 for the statistically significant difference.Results ERM,p-ERM and mDia1 were presented in rat PMVEC.Stimulation with LPS up-regulated p-ERM,mDia1 in a dose-dependent manner:LPS [0 μg/mL LPS group:(0.520±0.101),0.1 μg/mL LPS group:(0.657 ±0.092),1 μg/mL LPS group:(0.891 ±0.167),10 μg/mL LPS group:(1.227 ±0.106);0 μg/mL group vs.0.1 μg/mL group,P ＞0.05;the rest P ＜0.01];and mDia1 [0 μg/mL LPS group:(0.200 ±0.102),0.1 μg/mL LPS group:(0.430 ±0.121),1 μg/mL LPS group:(0.603 ±0.154),10 μg/mL LPS group:(0.887 ±0.204);0.1 μg/mL group vs.1 μg/mL group,P ＞ 0.05;the rest P ＜ 0.05].In time-dependent group,the level of p-ERM protein increased at 15 min (0.670 ±0.149),peaked at 30 min (1.175 ±0.103),then decreased,at 60 min (0.959 ±0.189),90 min (0.842 ±0.129),but kept at higher level at 120 min (0.767 ±0.097) than that in control group (0.471 ±0.157,15 min group vs.120 min group,60 min group vs.90 min group and 90 min group vs.120 min group,P ＞ 0.05;the rest P ＜ 0.05);and the level of mDia1 increased at 15 min (0.779 ±0.035),peaked at 30 min (0.889 ±0.036) then decreased at 60 min (0.648 ±0.019),90 min (0.582 ±0.068),but kept at higher level at 120 min (0.526 ±0.059) than that in control group (0.284±0.118,all P ＜ 0.01).C3 transferase caused a marked attenuation of LPS induced p-ERM expression [control group:(0.339 ± 0.069);C3 + LPS group:(0.438 ± 0.07);C3 control group:(0.352 ± 0.071);LPS control group:(0.634 ± 0.191),C3 + LPS group vs.LPS control group,P =0.01],as the same in mDia1 [control group:(0.449 ±0.122);C3 + LPS group:(0.380 ±0.148);C3 control group:(0.404 ±0.164);LPS control group:(0.622 ±0.174),C3 + LPS group vs.LPS control group,P ＜ 0.01].Conclusions LPS could up-regulated the expression of p-ERM protein,and inhibition of RhoA/mDia1 signal pathway by C3 transferase could down-regulated the p-ERM levels.

A previously unidentified deletion in G protein‑coupled receptor 143 causing X‑linked congenital nystagmus in a Chinese family.

Background: Congenital nystagmus (CN) is characterized by conjugated, spontaneous, and involuntary ocular oscillations. It is an inherited disease and the most common inheritance pattern is X‑linked CN. In this study, our aim is to identify the disease‑causing mutation in a large sixth‑generation Chinese family with X‑linked CN. Methods: It has been reported that mutations in four‑point‑one, ezrin, radixin, moesin domain‑containing 7 gene (FRMD7) and G protein‑coupled receptor 143 gene (GPR143) account for the majority patients of X‑linked nystagmus. We collected 8 ml blood samples from members of a large sixth‑generation pedigree with X‑linked CN and 100 normal controls. FRMD7 and GPR143 were scanned by polymerase chain reaction (PCR)‑based DNA sequencing assays, and multiplex PCR assays were applied to detect deletions. Results: We identified a previously unreported deletion covering 7 exons in GPR143 in a Chinese family. The heterozygous deletion from exon 3 to exon 9 of GPR143 was detected in all affected males in the family, while it was not detected in other unaffected relatives or 100 normal controls. Conclusions: This is the first report of molecular characterization in GPR143 gene in the CN family. Our results expand the spectrum of GPR143 mutations causing CN and further confirm the role of GPR143 in the pathogenesis of CN.

Clinical Significance of Ezrin Associated Subcellular Localization of E-cadherin Expression in Lymph Nodes Positive Breast Cancer / 中国医科大学学报

Objective To investigate the relationship between Ezrin expression and subcellular localization of E?cadherin(E?cad),and explore the clinical significance of this relationship to pathological features such as lymph nodes metastasis in breast cancer. Methods Ninety four cases of breast cancer tissue samples with lymph node metastasis were collected. The expression of Ezrin and E?cad was detected by immunohistochemi?cal method. Results The positive rates of and E?cad and Ezrin were respectively 45.7%and 58.5%in 94 nodes positive breast cancer,containing membranal expression of E?cad(E?cadm)in 20 cases and cytoplasmic expression of E?cad(E?cadc)in 23 cases;the frequency of E?cadc positive staining was significantly higher in Ezrin(+)tissues than that in Ezrin(-)tissues(P=0.025);E?cad expression level was significantly lower in TNMⅡ?Ⅲstage cases(P=0.001),and Ezrin expression(P=0.036)and E?cadc(P=0.013)was significantly increased in bigger cases;com?pared with E?cad(+),Ezrin(-),E?cadm tissues,the number of lymph node metastasis in E?cad(-)(P=0.011),Ezrin(+)(P=0.002),E?cadc (P=0.020)tissues were increased significantly;in the order of E?cad(+)/Ezrin(-),E?cad(-)/Ezrin(-),E?cad(+)/Ezrin(+),and E?cad(-)/Ezrin(+),the number of lymph node metastasis was increased significantly(P<0.001);similarly,in the order of E?cadm/Ezrin(-),E?cadc/Ezrin(-),E?cadm/Ezrin(+),and E?cadc/Ezrin(+),the number of lymph node metastasis was increased significantly(P=0.007). Conclusion Ezrin may regulate the subcellular localization of E?cad in metastatic breast cancer ,which may affect the course of breast cancer and promote the metastasis of lymph nodes.

Knockdown of ezrin suppresses the migration and angiogenesis of human umbilical vein endothelial cells in vitro / 华中科技大学学报（医学）（英德文版）

Progressive tumor growth is dependent on angiogenesis. The mechanisms by which endothelial cells (ECs) are incorporated to develop new blood vessels are not well understood. Recent studies reveal that the ezrin radixin moesin (ERM) family members are key regulators of cellular activities such as adhesion, morphogenetic change, and migration. We hypothesized that ezrin, one of the ERM family members, may play important roles in ECs organization during angiogenesis, and new vessels formation in preexisting tissues. To test this hypothesis, in this study, we investigated the effects of ezrin gene silencing on the migration and angiogenesis of human umbilical vein endothelial cells (HUVECs) in vitro. HUVECs were transfected with plasmids with ezrin-targeting short hairpin RNA by using the lipofectamine-2000 system. Wound assay in vitro and three-dimensional culture were used to detect the migration and angiogenesis capacity of HUVECs. The morphological changes of transfected cells were observed by confocal and phase contrast microscopy. Our results demonstrated that the decreased expression of ezrin in HUVECs significantly induced the morphogenetic changes and cytoskeletal reorganization of the transfected cells, and also reduced cell migration and angiogenesis capacity in vitro, suggesting that ezrin play an important role in the process of HUVECs migration and angiogenesis.

Correlation and Clinical Significance of Ezrin and β-catenin Protein Expression in Sporadic Breast Cancer / 中国医科大学学报

Objective To explore the clinical significance of Ezrin and β?catenin in breast cancer. Methods Immunohistochemical staining method was adopted to detect Ezrin and β?catenin protein expression level in 145 cases of breast cancer tissues,and their correlation with clinical data and prognosis of breast cancer was analyzed. Results Ezrin was expressed in 70 cases(48.3%),β?catenin was expressed in 82 cases (56.6%),and there was significantly negative correlation(r=0.267,P=0.001). The higher histologic grade of breast cancer,the higher expres?sion level of Ezrin(P=0.007),and the lower expression level of β?catenin(P<0.001). Ezrin expression level was increased significantly(P=0.027),but β?catenin expression level was reduced significantly(P=0.011)in lymph node positive breast cancer tissue. Ezrin expression was sig?nificantly correlated with shorter overall survival(P=0.004)and disease free survival(P=0.017)of breast cancer patients,but β?catenin expres?sion was significantly correlated with longer overall survival(P<0.001)and disease free survival(P=0.001)of breast cancer patients. However , Ezrin and β?catenin were not the independent risk factors of breast cancer patients as determined by multivariate Cox regression. Conclusion Ez?rin was significantly negative correlated with β?catenin in breast cancer. They play a role in the progression and poor prognosis of breast cancer , which can be used as breast cancer treatment targets.

The expression and significance of Ezrin in human triple negative breast cancer / 西安交通大学学报(医学版)

Objective To study the expression and significance of Ezrin in triple negative breast cancer tissues .Methods We selected 102 cases ,including 24 ones of triple negative breast cancer ,58 ones of non‐triple negative breast cancer ,and 20 of benign breast disease .The expression of Ezrin in all the specimens was detected by SP immunohistochemistry .We observed whether there was any difference between the positive expression rates of Ezrin in the three groups . We also analyzed the correlation between Ezrin expression and clinicopathologic parameters of triple negative breast cancer .Results The positive expression rate of Ezrin in groups of triple negative breast cancer , non‐triple negative breast cancer , and benign breast disease was 15 .00% ,48 .28% and 75 .00% ,respectively . The difference between the three groups differed significantly ( P 0 .05 ) . Conclusion Ezrin is highly expressed in triple negative breast cancer tissues ;therefore , it can be used as an important indicator of poor prognosis of triple negative breast cancer .