卵巢癌组织中UHRF1蛋白的表达及其与卵巢癌细胞增殖和侵袭的关系

目的:研究卵巢癌组织中泛素样含PHD和环指域1(Ubiquitin-like with PHD and ring finger domains 1,UHRF1)蛋白的表达及UHRF1对卵巢癌细胞增殖、侵袭的影响。方法:选取卵巢癌组织和癌旁正常组织,采用蛋白印迹法(Western blot)检测其UHRF1的蛋白表达。体外培养卵巢癌SKOV-3细胞株,分别转染UHRF1的si RNA和阴性对照si RNA,采用CCK-8检测细胞活力,Transwell检测细胞侵袭能力,荧光定量聚合酶链式反应法(FQ-PCR)检测Cyclin D1、CDK6、MMP2和MMP9的m RNA表达。结果:卵巢癌组织中UHRF1蛋白表达水平显著高于癌旁正常组织(P0.05);与转染阴性对照si RNA的SKOV-3细胞相比,转染UHRF1的si RNA的SKOV-3细胞活力明显降低、侵袭细胞数目明显减少(P0.05),且细胞中Cyclin D1、CDK6、MMP2和MMP9基因的m RNA表达水平显著降低(P0.05)。结论:UHRF1蛋白在卵巢癌组织中呈高表达状态,且可促进卵巢癌细胞的增殖和侵袭。     

SPOP对卵巢癌细胞生物学行为的影响及其分子机制的研究

探讨人卵巢癌组织中斑点状蛋白(SPOP)的表达及对其卵巢癌细胞生物行为学的影响及相关机制。采用免疫组化技术分别检测正常卵巢组织10例、卵巢癌组织90例的组织芯片中SPOP的表达;体外培养永生化人卵巢癌细胞株OVCAR-3、SKOV3及永生化的人卵巢表皮上皮细胞株HOSE,RT-PCR、WB检测3种细胞株SPOP的表达。用慢病毒稳定转染OVCAR-3、HOSE细胞株,构建过表达、敲低SPOP的细胞株。流式细胞仪检测转染后细胞株的凋亡,CCK8检测转染后细胞株的增殖情况,Transwell小室检测转染后细胞株的迁移侵袭情况。WB检测转染后细胞株PI3K-Akt通路相关蛋白的表达情况。组织学免疫组化评分显示卵巢癌组织染色明显高于正常卵巢组织。细胞学实验显示,成功构建过表达、敲低SPOP的OVCAR-3细胞,过表达组、过表达空载组、敲低组、敲低空载组细胞凋亡及增殖能力无明显差异;各组细胞迁移及侵袭结果与空载组有明显差异,且差异有统计学意义(p0.05)。过表达组细胞P-GSK3β(Ser9)表达水平明显低于敲低组(p0.05),MMP-9表达水平明显高于敲低组(p0.05)。SPOP在卵巢癌组织中表达上升,且具有促进卵巢癌细胞迁移侵袭的作用,其调节机制可能与p-GSK3β(Ser9)通路相关。     

Nucleostemin基因在卵巢上皮性肿瘤的表达及与病理分型的关系

目的：研究核干细胞因子Nucleostemin（NS）基因在卵巢上皮性肿瘤中的表达,探讨其与肿瘤病理分型的关系。方法：采用RT-PCR及Western blot检测36例卵巢癌组织手术标本,32例卵巢良性上皮肿瘤组织手术标本,12例正常卵巢组织标本中Nu-cleostemin基因及相应蛋白的表达,采用分组对照的方法对比3组样本中NS基因及蛋白的表达情况,并进行相对定量研究。采用统计学方法检测NS基因的表达是否与临床病理分级及血清CA125存在关联。结果：①卵巢癌组织中NS的阳性表达率显著高于良性肿瘤组织及正常卵巢组织;②卵巢癌组织中,淋巴结转移组NS的表达水平高于未转移组;③临床分期Ⅲ期组的表达水平高于ⅠB期组;④中、低分化组的表达水平高于高分化组。结论：卵巢癌组织中存在NS基因的高表达,其表达量与组织类型无关,而与临床TNM分期及组织分级正相关。     

WT-1、Smac蛋白在上皮性卵巢癌中的表达和临床意义研究

目的:探讨WT-1和Smac蛋白在上皮性卵巢癌中的表达及其临床意义。方法:应用免疫组织化学染色方法检测40例正常卵巢组织,40例卵巢上皮性良性肿瘤组织,60例全面分期手术治疗的上皮性卵巢癌组织中WT-1、Smac蛋白的表达,并分析WT-1、Smac蛋白的表达与上皮性卵巢癌临床病理特征的相关性及二者之间的相关性。结果:WT-1蛋白在上皮性卵巢癌组织中的表达明显高于正常卵巢组织或卵巢上皮性良性肿瘤组织(P0.05);Smac蛋白在上皮性卵巢癌组织中表达明显低于正常卵巢组织或卵巢上皮性良性肿瘤组织(P0.05)。上皮性卵巢癌组织中WT-1、Smac蛋白的表达与肿瘤临床分期、组织分化程度、淋巴结有无转移均显著相关(P0.05)。且上皮性卵巢癌中WT-1、Smac蛋白的表达呈明显负相关性(r=-0.35,P0.05)。结论:WT-1蛋白高表达或Smac蛋白低表达可能在上皮性卵巢癌的发生、发展中发挥重要作用,检测WT-1、Smac蛋白的表达有助于上皮性卵巢癌恶性程度的判断和预后评估。     

MicroRNA-141 在上皮性卵巢癌患者组织和血清中的表达 及临床意义的探讨

目的:探讨微小RNA-141(miR-141)在卵巢癌患者组织和血清中的表达情况并初步探讨其作为肿瘤标记物早期诊断上皮性卵巢癌的可行性。方法:采用实时荧光定量逆转录聚合酶链反应(real-time RT-PCR)检测16例上皮性卵巢癌患者和4例正常人卵巢组织及血清标本中miR-141的表达;检测4例良性卵巢肿瘤血清中miR-141的表达。结果:miR-141在卵巢癌患者组织中相对表达量分别为(72.846±76.671)显著高于正常人(2.869±3.201)(P<0.05);miR-141在卵巢癌患者血清中相对表达量(31.581±52.885)显著高于良性卵巢肿瘤患者(0.668±1.196)和正常人(1.690±1.697)(P<0.05),后两组间表达无差异(P>0.05);miR-141表达随卵巢癌临床分期的进展呈上升趋势(P<0.01),在无淋巴结转移组明显高于有淋巴结转移组(P<0.05),与组织分级和CA125的升高无关(P>0.05)。在组织中miR-141表达水平与上皮性卵巢癌临床病理特征均未见明显差异(P>0.05)。结论:miR-141可能在上皮性卵巢癌的发生发展中发挥癌基因的作用;miR-141用于检测上皮性卵巢癌敏感性和特异度较高,有望成为上皮性卵巢癌早期诊断的新指标。     

Runx3蛋白在卵巢上皮性癌中的表达及临床意义

目的:探讨Runx3蛋白在卵巢癌的发生发展、浸润转移和化疗耐药中的作用,为以Runx3作为分子治疗靶点的抗肿瘤基因治疗提供理论依据。方法:运用组织芯片技术联合免疫组化法检测27例卵巢癌、20例正常卵巢组织、30例卵巢良性肿瘤中抑癌基因Runx3的表达情况,分析其与卵巢癌临床病理特征的关系,并进行临床随访分析Runx3基因与卵巢癌化疗耐药的关系。结果:1.Runx3蛋白在卵巢癌组织中的表达明显低于正常卵巢组织及卵巢良性肿瘤中的表达,两两比较,差异有统计学意义(P0.05);Runx3蛋白表达与肿瘤的临床分期和病理学分级有关,差异有显著性(P0.05);与肿瘤的组织学类型、患者年龄、是否绝经、及有无腹水形成无关(P0.05)。2.Runx3蛋白在化疗敏感组中高表达。结论:Runx3蛋白低表达对卵巢癌的发生发展及化疗耐药中起重要作用。     

DKK3在上皮性卵巢癌中表达的初步探索

目的:探讨DKK3表达与卵巢上皮性癌的关系。方法:采用免疫组化法检测不同卵巢组织DKK3蛋白表达水平,其中88例卵巢上皮性癌组织(以下简称卵巢癌组织)、39例卵巢良性肿瘤组织及46例正常卵巢组织。结果:DKK3在正常卵巢组织、卵巢良性肿瘤及卵巢癌组织的表达有明显差异,差异有统计学意义(P0.05)。而DKK3表达与上皮性卵巢癌的临床病理分期及分化程度无关(P0.05)。结论:DKK3表达水平下降与卵巢癌有关,DKK3可作为预测卵巢癌的指标。     

胰腺癌上调因子在卵巢癌中水平变化及其与TLR4/MyD88通路的关系

该文探讨了胰腺癌上调因子(PAUF)在卵巢癌中的表达水平,及其与TLR4/MyD88信号通路在卵巢癌中的关系。选取2014年10月至2016年5月在我院就诊的卵巢癌患者共217例,纳入卵巢癌组,另随机选取同期诊断为卵巢良性肿瘤的45例患者纳入卵巢良性肿瘤组和52例健康人群纳入对照组。采用免疫组织化学染色法检测受试者卵巢组织中的PAUF、TLR4和MyD88表达水平。比较三组PAUF、TLR4和MyD88的表达水平;分析PAUF和TLR4、MyD88之间存在的相关性;分析卵巢癌患者中影响PAUF、TLR4和MyD88表达水平的因素。结果显示,卵巢癌患者组织中的PAUF、TLR4和MyD88水平均显著高于良性肿瘤组和对照组(P0.05);卵巢癌组织中PAUF与TLR4表达水平呈正相关(r=0.521,P0.001),且PAUF与MyD88表达也成正相关(r=0.581, P0.001);卵巢癌组织PAUF水平与肿瘤种类、FIGO分期和肿瘤分化程度有关(P0.05),但与患者年龄无关(P0.05);而癌组织中TLR4水平只与肿瘤的分化程度有关(P0.05), MyD88表达水平只与癌症种类有关(P0.05)。由此说明,卵巢癌患者组织中的PAUF表达水平显著升高,与TLR4和MyD88表达成正相关。PAUF可能通过参与TLR4/MyD88信号通路,参与了卵巢癌的发生发展。     

特异AT序列结合蛋白1在卵巢癌的表达及临床病理学意义

目的:检测上皮性卵巢癌中特异AT序列结合蛋白1(Special AT-rich sequence-bindingprotein 1,SATB1)的表达,探讨SATB1表达与卵巢癌的发生,进展及转移的关系及意义.方法:以实时荧光定量RT-PCR和免疫组化方法检测91例上皮性卵巢癌组织、13例卵巢交界性囊腺瘤及8例正常卵巢组织中SATB1mRNA和蛋白的表达,分析SATB1基因表达与临床病理参数的相关性.结果:上皮性卵巢癌组织和交界性囊腺瘤组织中SATB1 mRNA表达量分别为正常卵巢组织的6.97倍和5.70倍,差异有统计学意义(P＜0.05).SATB1 mRNA表达水平随着卵巢癌分期升高而增加:Ⅰ期,Ⅱ期和Ⅲ期分别为正常卵巢组织的5.31倍,6.67倍和8.04倍(P＜0.05),并且在淋巴结转移组表达明显高于无转移组(P＜0.05).免疫组化也显示SATB1蛋白在上皮性卵巢癌中表达高于正常卵巢组织,差异具有统计学意义(P＜0.05).结论:SATB1的表达水平可能与上皮性卵巢癌发生,进展及转移有关.     

miR-550a-3靶向NFIC表达调控肺癌细胞HCC827增殖、迁移和侵袭

[目的]探讨miR-550a-3靶向NFIC表达调控肺癌细胞HCC827增殖、迁移和侵袭作用。[方法]检测40例肺癌患者癌组织和癌旁组织中miR-550a-3和NFIC mRNA表达,并分析NFIC表达与肺癌患者病理特征的相关性。按Lipofectamine 2000方法分别将miR-NC、miR-550a-3 mimics和miR-550a-3inhibitor转染到HCC827细胞,48h后检测细胞增殖、迁移和侵袭情况,同时检测细胞中miR-550a-3和NFIC mRNA表达,并检测细胞中NFIC、E-cadherin和N-cadherin蛋白表达。[结果]肺癌组织中miR-550a-3表达水平(1.83±0.19)高于癌旁组织(1.00±0.15),NFIC mRNA表达水平(0.62±0.14)低于癌旁组织(1.00±0.10)(P<0.05);以miR-550a-3 mRNA均数为标准,将肺癌患者分为高表达组(22例)和低表达组(18例),miR-550a-3与病理类型和TNM分期相关(P<0.05),与年龄、性别、是否吸烟和肿瘤直径不相关(P>0.05)...     

Chk1反义寡核苷酸联合顺铂抑制卵巢癌侵袭转移的分子机制

目的:探究Chk1反义寡核苷酸(CHK1-ASODN)单独或联合顺铂(DDP)对卵巢癌细胞系SKOV-3侵袭转移能力的影响,并阐明其可能的分子机制。方法:体外培养人卵巢癌细胞系SKOV-3,CHK1-ASODN单独或联合DDP处理48 h后,划痕实验检测细胞迁移能力;Transwell实验检测细胞侵袭能力;显微镜下观察细胞上皮或间质表型特征;Western blot及实时定量PCR技术分别检测上皮间质转化(EMT)特异性标志物(E-cadherin、N-cadherin)以及EMT关键调控分子ZEB1的蛋白及m RNA的表达水平。结果:与对照组相比较,CHK1-ASODN单独或联合DDP均能显著抑制SKOV-3细胞的迁移及侵袭(P0.05);细胞表现为间质化表型;E-cadherin的表达显著升高(P0.05),而N-cadherin的表达则显著降低(P0.05);ZEB1的表达显著降低(P0.05)。结论:CHK1-ASODN单独或联合DDP下调ZEB1的表达进而逆转EMT可能是其抑制卵巢癌侵袭转移的重要机制之一。     

Cosmc基因在卵巢癌中生物学功能的研究

目的:明确Cosmc基因在卵巢癌细胞中的生物学功能。方法:本研究采用慢病毒转染技术,在卵巢癌细胞A2780和SKOV3中过表达Cosmc基因,MTT实验、细胞凋亡实验以及Transwell侵袭实验对过表达Cosmc后卵巢癌细胞在生长、凋亡、侵袭等方面的影响。结果:慢病毒转染卵巢癌细胞A2780和SKOV-3后,Cosmc的蛋白表达显著提高;MTT结果显示,与空载体对照组相比,过表达Cosmc的卵巢癌细胞生长能力显著减弱;流式细胞术结果表明Cosmc过表达的卵巢癌细胞凋亡数较空载体对照组细胞显著增加;Transwell实验显示Cosmc过表达的细胞侵袭和迁移能力较空载体对照组显著减少。结论:卵巢癌细胞系中过表达Cosmc基因能抑制卵巢癌细胞的生长和侵袭并促进细胞凋亡。     

Stem Cell-Like Gene Expression in Ovarian Cancer Predicts Type II Subtype and Prognosis

Although ovarian cancer is often initially chemotherapy-sensitive, the vast majority of tumors eventually relapse and patients die of increasingly aggressive disease. Cancer stem cells are believed to have properties that allow them to survive therapy and may drive recurrent tumor growth. Cancer stem cells or cancer-initiating cells are a rare cell population and difficult to isolate experimentally. Genes that are expressed by stem cells may characterize a subset of less differentiated tumors and aid in prognostic classification of ovarian cancer. The purpose of this study was the genomic identification and characterization of a subtype of ovarian cancer that has stem cell-like gene expression. Using human and mouse gene signatures of embryonic, adult, or cancer stem cells, we performed an unsupervised bipartition class discovery on expression profiles from 145 serous ovarian tumors to identify a stem-like and more differentiated subgroup. Subtypes were reproducible and were further characterized in four independent, heterogeneous ovarian cancer datasets. We identified a stem-like subtype characterized by a 51-gene signature, which is significantly enriched in tumors with properties of Type II ovarian cancer; high grade, serous tumors, and poor survival. Conversely, the differentiated tumors share properties with Type I, including lower grade and mixed histological subtypes. The stem cell-like signature was prognostic within high-stage serous ovarian cancer, classifying a small subset of high-stage tumors with better prognosis, in the differentiated subtype. In multivariate models that adjusted for common clinical factors (including grade, stage, age), the subtype classification was still a significant predictor of relapse. The prognostic stem-like gene signature yields new insights into prognostic differences in ovarian cancer, provides a genomic context for defining Type I/II subtypes, and potential gene targets which following further validation may be valuable in the clinical management or treatment of ovarian cancer.     

Glucocorticoid regulation of SLIT/ROBO tumour suppressor genes in the ovarian surface epithelium and ovarian cancer cells

The three SLIT ligands and their four ROBO receptors have fundamental roles in mammalian development by promoting apoptosis and repulsing aberrant cell migration. SLITs and ROBOs have emerged as candidate tumour suppressor genes whose expression is inhibited in a variety of epithelial tumours. We demonstrated that their expression could be negatively regulated by cortisol in normal ovarian luteal cells. We hypothesised that after ovulation the locally produced cortisol would inhibit SLIT/ROBO expression in the ovarian surface epithelium (OSE) to facilitate its repair and that this regulatory pathway was still present, and could be manipulated, in ovarian epithelial cancer cells. Here we examined the expression and regulation of the SLIT/ROBO pathway in OSE, ovarian cancer epithelial cells and ovarian tumour cell lines. Basal SLIT2, SLIT3, ROBO1, ROBO2 and ROBO4 expression was lower in primary cultures of ovarian cancer epithelial cells when compared to normal OSE (P<0.05) and in poorly differentiated SKOV-3 cells compared to the more differentiated PEO-14 cells (P<0.05). Cortisol reduced the expression of certain SLITs and ROBOs in normal OSE and PEO-14 cells (P<0.05). Furthermore blocking SLIT/ROBO activity reduced apoptosis in both PEO-14 and SKOV-3 tumour cells (P<0.05). Interestingly SLIT/ROBO expression could be increased by reducing the expression of the glucocorticoid receptor using siRNA (P<0.05). Overall our findings indicate that in the post-ovulatory phase one role of cortisol may be to temporarily inhibit SLIT/ROBO expression to facilitate regeneration of the OSE. Therefore this pathway may be a target to develop strategies to manipulate the SLIT/ROBO system in ovarian cancer.     

Gambogic Acid Sensitizes Ovarian Cancer Cells to Doxorubicin Through ROS-Mediated Apoptosis

Ovarian cancer is one human malignancy which has response portly to doxorubicin. The anti-cancer activity of gambogic acid has been tested in in vitro and in vivo studies. In this study, we showed that gambogic acid, a natural compound, could potentiate the anticancer activity of doxorubicin in ovarian cancer through ROS-mediated apoptosis. Platinum-resistant human ovarian cancer cell line (SKOV-3) was treated with gambogic acid, doxorubicin, or the combination of both to investigate cell proliferation and apoptosis. We found that the combination of gambogic acid and doxorubicin causes synergistic loss of cell viability in SKOV-3 cells and this synergistic effect correlated with increased cellular ROS accumulation. Moreover, in vivo results showed that gambogic acid and doxorubicin combination resulted in a synergistic suppressing effect on tumor growth in ovarian cancer mice model. Taken together, the results suggested that doxorubicin in combination with gambogic acid might provide a promising therapeutic strategy to enhance chemosensitivity of ovarian cancer to doxorubicin.     

shRNA沉默卵巢癌细胞SKOV-3的IDO基因表达在体外对NK细胞杀伤作用敏感性的影响

目的通过shRNA沉默吲哚胺2,3-双加氧酶（indoleamine2,3-dioxygenase,IDO）基因表达,研究IDO表达在体外对NK细胞杀伤能力的作用。方法 shRNA沉默IDO基因表达质粒和空白质粒分别稳定转染至人卵巢癌细胞SKOV-3,应用Western blot检测IDO在SKOV-3、SKOV-3/Mock和SKOV-3/shIDO细胞中的表达情况,用MTT试剂盒检测3组肿瘤细胞体外生长速度和对NK细胞杀伤作用的敏感性。结果 IDO蛋白在SKOV-3/shIDO细胞中表达被抑制,在SKOV-3和SKOV-3/Mock细胞中有表达。3组肿瘤细胞体外生长曲线比较差异无统计学意义（P〉0.05）。SKOV-3/shIDO细胞存活的百分比明显低于其他2组对照（SKOV-3和SKOV-3/Mock）细胞,差异有统计学意义（P〈0.05）。结论本研究应用shRNA沉默IDO基因表达质粒稳定转染卵巢癌细胞SKOV-3,获得IDO无表达卵巢癌细胞SKOV-3/shIDO,结果显示抑制IDO表达对卵巢癌细胞体外生长速度无影响,但可增强卵巢癌细胞SKOV-3对NK细胞杀伤作用的敏感性。因此,IDO可以作为卵巢癌基因治疗的潜在新靶点,shRNA沉默IDO基因表达可以作为卵巢癌治疗的新方法。     

Impact of Human Cytomegalovirus Infection and its Immune Response on Survival of Patients with Ovarian Cancer

Human cytomegalovirus (HCMV) has been detected in various types of tumors. We studied the prevalence of HCMV in ovarian cancer and its relation to clinical outcome. Paraffin-embedded tissues obtained prospectively from 45 patients with ovarian cancer and 30 patients with benign ovarian cystadenoma were analyzed for expression of HCMV immediate-early protein (IE) and HCMV tegument protein (pp65) by immunohistochemistry. Plasma was analyzed for HCMV serology. HCMV-IgG levels were higher in patients with ovarian cancer or benign cystadenoma than in age-matched controls (P?=?.002, P?<?.0001, respectively). HCMV IgM was detected in 12% of ovarian cancer patients and 3% of patients with benign tumors but was absent in controls. In patients with ovarian cancer, higher IgG levels were associated with better outcomes (P?=?.04). Extensive HCMV-IE protein expression was detected in 75% of ovarian cancers and 26% of benign tumors; pp65 was detected in 67% of ovarian cancers and 14% of benign tumors. A higher grade of HCMV infection was associated with higher stage of disease. Extensive HCMV-pp65 expression was associated with shorter median overall survival than focal expression (39 versus 42.5?months, P?=?.03). At study closure, 58% of ovarian cancer patients with focal pp65 expression were alive versus 27% of patients with extensive pp65 expression (P?=?.03). Thus, HCMV proteins are detected at different levels in ovarian tumors and benign cystadenomas. Ovarian cancer patients with focal HCMV-pp65 expression in their tumors and high IgG levels against HCMV lived longer, highlighting a need for in-depth studies of the oncomodulatory role of HCMV in ovarian cancer.     

趋化因子受体CCR3在上皮性卵巢癌中的表达及临床意义

目的:探讨趋化因子受体CCR3在上皮性卵巢癌组织中的表达情况,以及其与卵巢癌临床病理特征的关系。方法:收集上皮性卵巢癌组织、良性上皮性卵巢肿瘤组织以及正常卵巢组织标本各30例,采用多聚腺苷酸加尾实时荧光定量反转录聚合酶链反应[poly(A)-RT-qPCR]检测其CCR3的表达,并分析上皮性卵巢癌组织中CCR3的表达与患者临床病理特征之间的关系。结果:上皮性卵巢癌组织中CCR3的表达显著高于良性卵巢肿瘤组和正常卵巢组,差异有统计学意义(P0.05);且上皮性卵巢癌组织中CCR3的表达与患者的分期、组织分级及淋巴结转移均显著相关(P0.05)。结论:CCR3在上皮性卵巢癌组织中呈高表达,且在上皮性卵巢癌的发生和发展过程中均起着十分重要的作用。