Normal Ovarian Cells Research Articles

The role of R-spondin 1 through activating Wnt/β-catenin in the growth, survival and migration of ovarian cancer cells

Aberrant activation of the Wnt/β-catenin has been shown to promote progression in various cancers, including ovarian cancer. However, the molecular mechanisms involved in Wnt/β-catenin activation are not well elucidated. In the work, we identify that R-spondin 1 is an upstream regulator in Wnt/β-catenin pathway to promote growth, survival and migration in ovarian cancer cells. We observe the upregulation of transcript and protein levels of R-spondin 1 in ovarian cancer cell lines and tissues compared to normal counterparts. R-spondin 1 upregulation via genetic (overexpression) and pharmacological (recombinant protein) approaches facilitates growth and migration of normal ovarian cells. R-spondin 1 downregulation via siRNA knockdown decreases proliferation and migration, and induces apoptosis in ovarian cancer cells. In addition, recombinant R-spondin 1 protects ovarian cancer cell against chemotherapy whereas R-spondin 1 knockdown sensitizes ovarian cancer cell response to chemotherapy. Importantly, increased β-catenin activities and mRNA expression levels of Wnt/β-catenin-targeted genes are detected in normal ovarian cells overexpressing R-spondin 1. In contrast, R-spondin 1 inhibition suppresses Wnt/β-catenin signaling in ovarian cancer cells. We further identify that R-spondin 1 regulates ovarian cancer biological activities via activating Wnt/β-catenin. Our work is the first to highlight the critical roles of R-spondin 1 in ovarian cancer progression and chemoresistance. Our work also provides a proper understanding on the regulation of Wnt/β-catenin pathway in ovarian cancer.

Effect of testosterone on the expression of PPARγ mRNA in PCOS patients.

Expression of peroxisome proliferator-activated receptor γ (PPARγ) mRNA in ovarian granulosa cells of patients with polycystic ovary syndrome (PCOS) were explored. Ovarian granulosa cells were extracted from 5 patients with PCOS and 30 normal controls. Expression of PPARγ mRNA in granulosa cells of the 5 PCOS patients (observation group) and 5 normal controls (control group) was detected by RT-qPCR. The remaining 25 cases of normal human ovarian granulosa cells were cultured in vitro for 48 h, followed by cell culture for another 24 h with different concentrations of testosterone, insulin (INS), and rosiglitazone (RGZ). After that, expression of PPARγ mRNA was detected by RT-qPCR. Relative expression level of PPARγ mRNA in the observation group was significantly lower than that in the control group. Compared with testosterone concentration at 10-2 mol/ml, testosterone concentration at 10-3 mol/ml significantly reduced the expression level of PPARγ mRNA. When the INS concentration was 10-5 mol/ml, relative expression level of PPARγ mRNA was significantly higher than that of the control group (P<0.01). Relative expression level of PPARγ mRNA was significantly higher than that of the control group when INS concentration was 10-4 mol/ml. When the concentration of RGZ was 10-4 mol/ml, the relative expression level of PPARγ mRNA was significantly higher than that of the control group (P<0.01). When concentration of RGZ was 10-3 mol/ml, expression level of PPARγ mRNA was significantly increased comparing to that under a RGZ concentration of 10-4 mol/ml or that of the control group (P<0.01). Appropriate concentrations of testosterone can inhibit the expression of PPARγ mRNA in ovarian granulosa cells, and certain concentrations of INS and RGZ can induce the expression of PPARγ mRNA in ovarian granulosa cells. Abnormal expression of PPARγ mRNA in ovarian granulosa cells may be related to the mechanism of PCOS.

Knockdown of NRSF inhibits cell proliferation of ovarian cancer via activating Hippo pathway

AimsOvarian cancer is the most leading cause of deaths among gynecologic malignancies, and Neuron-restrictive silencer factor (NRSF) can be upregulated or downregulated according to the type of tumor. However, the expression and function of NRSF in ovarian cancer is still unknown. Main methodsExpression of NRSF in normal ovary and ovarian cancer cells were evaluated by quantitative PCR (qPCR). NRSF expression in normal ovary and ovarian cancer tissue samples were examined by qPCR, western blotting and immunohistochemistry (IHC). MTT, colony formation, anchorage-independent growth assay were applied to examine the effect of NRSF on ovarian cancer cell proliferation. Bromodeoxyuridine (BrdUrd) labeling and flow cytometry assay were carried out to investigate the role of NRSF on cell cycle of ovarian cancer cells. Luciferase reporter assay and western blotting, immunofluorescence labeling were devoted to explore the mechanism by which NRSF contributes to proliferation of ovarian cancer cells. Key findingsThe results demonstrated that NRSF is significantly upregulated in ovarian cancer cells and tissues and negatively related with the survival of patients with ovarian cancer, and knockout of NRSF inhibit proliferation of ovarian cancer cells. Further analysis showed that NRSF can influence G1/S transition of cell cycle via regulating the transcription of Hippo pathway. SignificanceHerein, our study suggest that NRSF is associated with the progression of ovarian cancer, and NRSF may be a valuable early detection marker of ovarian cancer and inhibiting NRSF expression may be an effective method to treat ovarian cancer.

High LIN28A and PLK4 co‑expression is associated with poor prognosis in epithelial ovarian cancer.

Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy. LIN28 homolog A (LIN28A) is a RNA-binding protein, which serves a fundamental role in cell development and pluripotency. Polo-like kinase 4 (PLK4) is a member of the polo-like kinase family, which primarily takes part in the mitotic regulation. Overexpression of LIN28A has been demonstrated in ovarian cancer; however, the expression of PLK4 and the correlation between the expression of LIN28A and PLK4 in EOC has not been discussed. In the present study, the mRNA and protein levels of LIN28A and PLK4 were evaluated by reverse transcription-quantitative polymerase chain reaction and immunohistochemistry in ovarian tissues of patients. Results demonstrated significantly increased expression in EOC compared with benign epithelial ovarian tumors. High expression of LIN28A and PLK4 was detected at the advanced pathological stage. Furthermore, PLK4 expression was positively correlated with LIN28A (r=0.555; P=0.039). The median survival analysis of patients with EOC with LIN28A and PLK4 double positive expression was 14 months, compared with 30 months in single positive and 60 months in double negative patients by Kaplan-Meier analysis (P<0.05). The expressions of LIN28A and PLK4 was elevated in different EOC cell lines compared to with a normal ovarian cell line. The 293T cells transfected with LIN28A plus a PLK4 plasmid were the fastest-growing group. These results suggest that co-expression of LIN28A and PLK4 may be associated with poor prognosis of EOC and could serve as promising prognostic biomarkers and therapeutic targets in EOC. LIN28A and PLK4 may be used along with traditional morphological and clinical characteristics for predicting prognosis.

LncRNA MEG3 impacts proliferation, invasion, and migration of ovarian cancer cells through regulating PTEN.

We investigated the expressions of lncRNA MEG3 and PTEN in ovarian cancer tissues and their effects on cell proliferation, cycle and apoptosis of ovarian cancer. Expression levels of MEG3 in ovarian cancer cell lines and normal ovarian cell lines were detected by qRT-PCR. Cell viability was detected by MTT assay. Cell apoptosis and cell cycle distribution were measured by flow cytometry. Cell invasion capability was tested by transwell assay. Cell migration capacity was tested by wound healing. The xenograft model was constructed to explore the effect of lncRNA MEG3 on ovarian cancer in vivo. Compared with normal ovarian cells, expression levels of MEG3 and PTEN were relatively lower in ovarian cancer cells. There was a positive correlation between the expression of PTEN and the expression of MEG3. Enhanced expression level of PTEN suppressed SKOV3 cell proliferation, increased cell apoptosis rate, and decreased cell invasion and migration. LncRNA MEG3 and PTEN were down-regulated in ovarian cancer cells. LncRNA MEG3 regulated the downstream gene PTEN in ovarian cancer cells to prohibit cell proliferation, promote apoptosis and block cell cycle progression.

Poly r(C) Binding Protein 1 Regulates Posttranscriptional Expression of the Ubiquitin Ligase TRIM56 in Ovarian Cancer.

Our earlier work has shown that the E3 ligase TRIM56 messenger RNA (mRNA) level and vimentin protein expression followed an inverse correlation in ovarian carcinoma patients; however, the regulatory mechanisms underlying TRIM56 expression is unclear. Steady state expression of TRIM56 mRNA expression in the normal ovarian cell line Moody and ovarian cancer cell lines SKOV-3, A2780, and Caov-3 were not significantly different; however, TRIM56 protein expression was significantly lower in the ovarian cancer cell lines compared to the Moody cell line. Polysome profiling showed significant increase in translation of TRIM56 messenger RNA in the Moody cells compared to the SKOV-3 cells. We performed RNA-affinity pulldown using biotinylated TRIM56 5 'and 3'-UTR and postnuclear extracts from Moody and SKOV-3 cells. Whereas no notable difference was observed in affinity pull-down obtained with the 5'-UTR, there was obvious difference in protein binding patterns with the 3'-UTR. Mass spectrometry was used to determine the most differentially binding protein as poly r (c) binding protein 1 (PCBP1). PCBP1 expression and binding to the 3'-UTR was both higher in SKOV-3 cells compared to the Moody cells. Silencing of TRIM56 in Moody cells cause an increase in in vitro migration and invasion, and a similar effect was mimicked by overexpression of PCBP1. Conversely, silencing of PCBP1 or overexpression of TRIM56 in SKOV-3 cells significantly decreased in vitro migration and invasion. In xenograft assays, SKOV-3 cells stably overexpressing shRNA targeting PCBP1 decreased metastasis, whereas shRNA-targeting TRIM56 potentiated detection of metastatic lesions, compared to the parental SKOV-3 cells themselves. Taken together our results reveal a yet undefined posttranscriptional regulatory mechanism underlying low expression of TRIM56 in ovarian cancer. © 2018 IUBMB Life, 71(1):177-182, 2019.

MicroRNA-200a promotes proliferation and invasion of ovarian cancer cells by targeting PTEN.

We investigate whether microRNA-200a could regulate proliferation and invasion of ovarian cancer cells, thereby participating in the occurrence and development of ovarian cancer. We also explore the specific mechanism of microRNA-200a in regulating ovarian cancer. Expression level of microRNA-200a in ovarian cancer tissues and paracancerous tissues were detected by quantitative Real-time polymerase chain reaction (qRT-PCR). The regulatory effects of microRNA-200a on proliferation and invasion of ovarian cancer cells were examined by Cell counting kit-8 (CCK-8) and cell invasion assay, respectively. Dual-luciferase reporter gene assay was performed to confirm the binding relationship between microRNA-200a and PTEN (phosphatase and tensin homolog deleted on chromosome ten). The regulatory role of microRNA-200a in PTEN expression was accessed by Western blot. Rescue experiments were conducted to assess whether microRNA-200a regulated proliferation and invasion of ovarian cancer cells by inhibiting PTEN expression. MicroRNA-200a expression in ovarian cancer tissues was significantly higher than that of paracancerous tissues. Besides, microRNA-200a was also overexpressed in ovarian cancer cell lines than that of normal ovarian cells. Overexpression of microRNA-200a promoted the proliferative and invasive abilities of SKOV3 and OVCAR3 cells. Dual-luciferase reporter gene assay showed that microRNA-200a could directly degrade PTEN. Overexpression of PTEN in SKOV3 and OVCAR3 cells partially reversed the increased cell proliferation and invasion induced by overexpressed microRNA-200a. Overexpressed microRNA-200a promoted the proliferative and invasive abilities of ovarian cancer cells, which might be related to the targeted regulation of PTEN expression.

Girinimbine Inhibits the Proliferation of Human Ovarian Cancer Cells In Vitro via the Phosphatidylinositol-3-Kinase (PI3K)/Akt and the Mammalian Target of Rapamycin (mTOR) and Wnt/β-Catenin Signaling Pathways.

BackgroundWorldwide, ovarian cancer is increasing in prevalence and has a high mortality rate. Girinimbine is a carbazole alkaloid isolated from Murraya koenigii (the curry tree) and is used in Chinese herbal medicine. The aim of this study was to evaluate the effects of girinimbine on cell proliferation, cell migration, and apoptosis in human ovarian cancer cells in vitro.Material/MethodsA human ovarian cancer cell line panel, which included SKOV3 cells and the SV40 immortalized normal human ovarian cell line, were treated with increasing doses of girinimbine. Cell proliferation was evaluated using the MTT assay. Confocal immunofluorescence using 4′,6-diamidino-2-phenylindole (DAPI), Annexin-V, and propidium iodide (PI) were used to measure cell apoptosis. Cell migration and invasion were determined by transwell assays. Protein expression was determined by Western blot.ResultsGirinimbine inhibited SKOV3 ovarian cancer cell proliferation in a dose-dependent manner. The half maximal inhibitory concentration (IC50) of grinimbine was 15 μM for the SKOV3 cells, and 120 μM for the SV40 cells. Grinimbine treatment resulted in apoptosis of SKOV3 cells, from 2.2% in untreated cells to 58.8% at a dose of 30 μM, which was associated with an increase in the Bax/Bcl-2 ratio. Girinimbine inhibited cell migration and invasion of the SKOV3 cancer cells in vitro and inhibited the PI3K/AKT/mTOR and Wnt/β-catenin signaling pathways.ConclusionsGirinimbine, a carbazole alkaloid used in Chinese herbal medicine, inhibited the proliferation and cell migration of human ovarian cancer cells in vitro, in a dose-dependent manner, via the PI3K/Akt/mTOR and Wnt/β-catenin signaling pathways.

Characterization of ovarian cancer cells and tissues by Fourier transform infrared spectroscopy

BackgroundOvarian cancer is the most lethal of gynecological malignancies. Fourier Transform Infrared (FTIR) spectroscopy has gradually developed as a convenient, inexpensive and non-destructive technique for the study of many diseases. In this study, FTIR spectra of normal and several heterogeneous ovarian cancer cell lines as well as ovarian cancer tissue samples were compared in the spectral region of 4000 cm− 1 - 600 cm− 1.MethodsCell samples were collected from human ovarian surface epithelial cell line (HOSEpiC) and five ovarian cancer cell lines (ES2, A2780, OVCAR3, SKOV3 and IGROV1). Validation spectra were performed on normal and cancerous tissue samples from 12 ovarian cancer patients. FTIR spectra were collected from a NICOLET iN10 MX spectrometer and the spectral data were analyzed by OMNIC 8.0 software.ResultsSpectral features discriminating malignant tissues from normal tissues were integrated by cell line data and tissue data. In particular changes in cancerous tissues, the decrease in the amount of lipids and nucleic acids were observed. Protein conformation and composition were also altered in some cancer cells. The band intensity ratio of 1454/1400 was higher in normal cells/tissues and lower in cancer cells/tissues.ConclusionThe spectral features revealed the important molecular characteristics about ovarian cancer cells/tissues. These findings demonstrate the possible diagnostic use of FTIR spectroscopy, providing the research model and evidences, and supporting the future study on more tissue samples to establish a data bank of spectra features for the possible discrimination of ovarian cancers.

LINC01116 promotes the progression of epithelial ovarian cancer via regulating cell apoptosis.

To explore the biological function of LINC01116 in epithelial ovarian cancer (EOC) and its underlying mechanism. The expression level of LINC01116 in 60 EOC tissues, 30 normal ovarian tissues and EOC cells were detected by qRT-PCR (quantitative real-time polymerase chain reaction). Disease-free survival (DFS) and overall survival (OS) of enrolled EOC patients were recorded. The correlation between LINC01116 expression, DFS and OS of EOC patients was analyzed using ROC curve. Influencing factors for DFS and OS were analyzed by univariable and multivariable Cox regression model. For in vitro experiments, the effect of LINC01116 knockdown on proliferation, invasion and apoptosis of EOC cells were detected by CCK-8 (cell counting kit-8), transwell assay and flow cytometry, respectively. Protein expressions of apoptosis-related genes in EOC cells transfected with pc-DNA-LINC01116 or si-LINC01116 were detected by Western blot. LINC01116 was overexpressed in EOC tissues than that of paracancerous tissues. DFS and OS in EOC patients with higher expression of LINC01116 were remarkably shorter than those with lower expression. FIGO (International Federation of Gynecology and Obstetrics) clinical stage and LINC01116 expression were the independent factors that affected DFS and OS of EOC patients. Besides, LINC01116 expression was positively correlated to the diagnostic sensitivity of EOC patients. In vitro experiments found that LINC01116 overexpression promoted proliferation and invasion of EOC cells. Overexpressed LINC01116 resulted in upregulated Bcl-2, and downregulated cleaved Caspase-3 and cleaved Caspase-9 in EOC cells. Overexpressed LINC01116 promotes EOC progression via increasing proliferation and migration, and inhibiting cell apoptosis.

Abstract 1923: Identification and validation of Integrin-Linked Kinase (ILK) signaling as a therapeutic target for ovarian cancer

Abstract Ovarian cancer affects approximately 25,000 women and contributes to around 14,000 deaths in the United States each year. Although significant advances have been made in the discovery of molecular diagnostics and therapeutics for other cancers, ovarian cancer still lacks reliable diagnostic and prognostic markers as well as appropriate targeted therapeutics. A major reason for the lagging development of targeted therapeutics is the molecular heterogeneity of high grade serous ovarian cancer and the lack of better understanding of driver mutations and pathogenesis. Although gene expression data sets are available, most of them lack appropriate pair-matched controls to define the alterations that result in the transformation of normal ovarian cells to highly proliferative cancerous cells. We compared differential gene expression of high-grade treatment-naïve ovarian cancer tissue samples to matched-pair adjacent benign ovarian tissue specimens from 24 patients. A significant upregulation of the integrin-linked kinase (ILK) pathway was identified in 22 of the 24 cancer specimens, identifying a potential player that could contribute to the transformation of normal ovarian cells to cancerous cells. To determine the therapeutic potential of ILK inhibition, ILK expression was reduced using siRNA in SKOV3 cells, which resulted in decreased cell proliferation. This is consistent with prior studies demonstrating reduced adhesion to and invasion of collagen gels and organotypic meso-mimetic ovarian cancer cultures with down-regulating ILK using siRNA resulted in. We then assessed the ability of a targeted agent, compound 22, a small molecule ILK-selective inhibitor, to inhibit proliferation in various ovarian cancer cell lines with different phenotypic characteristics (SKOV3, OVCAR3, OVCAR8, OV90). Compound 22 reduced the phosphorylation of Akt, a downstream target of ILK, and also inhibited cellular proliferation. Our initial findings validate ILK as a potential therapeutic target for molecular inhibition in ovarian cancer and warrants further investigation. Citation Format: Michael A. Ulm, Suriyan Ponnusamy, Adam C. ElNaggar, Ramesh Narayanan. Identification and validation of Integrin-Linked Kinase (ILK) signaling as a therapeutic target for ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1923.

CircHIPK3 is upregulated and predicts a poor prognosis in epithelial ovarian cancer.

CircRNAs have been recently identified as important regulators in tumors biological functions. However, the clinical significance of circHIPK3 in epithelial ovarian cancer (EOC) remains unknown. The expression of circHIPK3 in EOC tumor tissues and adjacent noncancerous tissues was analyzed by quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). The association between circHIPK3 expression and clinicopathological factors was analyzed by using Chi-square test. Kaplan-Meier method and log-rank test were used to analyze the association of circHIPK3 expression with disease-free survival (DFS) and overall survival (OS) time of EOC patients. Univariate and multivariate Cox analysis was also performed. We found that circHIPK3 was higher expressed in EOC tissues and cells compared to adjacent normal tissue and ovarian epithelium cell line, respectively. Higher circHIPK3 expression associated with lymph node invasion, FIGO stage, and worse DFS and OS of patients. Moreover, multivariate Cox analysis showed that higher circHIPK3 was an independent predictor of DFS and OS in EOC patients. Thus, circHIPK3 may be a novel biomarker for predicting EOC prognosis.

Piperine functions as a tumor suppressor for human ovarian tumor growth via activation of JNK/p38 MAPK-mediated intrinsic apoptotic pathway

Piperine, a kind of natural alkaloid found in the fruit of black (Piper nigrum Linn) and long (Piper longum Linn), has shown antitumor activities toward various cancer cell lines. However, the antitumor effects of Piperine on ovarian cancer and the underlying mechanism are not fully elucidated. Our result showed that Piperine reduced the cell viability of A2780 cells in a concentration and time-dependent manner, but has not any effect on normal ovarian cells. Flow cytometric analysis revealed that Piperine suppressed cells proliferation via induction of apoptosis, which was followed by release of mitochondrial cytochrome c to cytosol, activation of caspase-3 and -9, as well as cleaved PARP. Moreover, Western blot results confirmed that Piperine (8, 16, and 20 μM) decreased phosphorylation of JNK and p38 MAPK in A2780 cells. In addition, caspase-3 inhibitor (Z-DEVD-FMK), caspase-9 inhibitor (Z-LEDH-FMK), JNK-inhibitor (SP600125), or p38 MAPK inhibitor (SB203580) could abate the apoptosis induced by Piperine (20 μM) treatment, while caspase-8 inhibitor (Z-IETD- FMK) exhibited no inhibitory effect on the induction of apoptosis in A2780 cells. These results provide the first evidence for the anticancer potential of Piperine in ovarian cancer cells, partially via JNK/p38 MAPK-mediated intrinsic apoptotic pathway.

Early transcriptional response of human ovarian and fallopian tube surface epithelial cells to norepinephrine

Evidence from human and animal studies suggests that chronic behavioral stress and resulting activation of the sympathetic nervous system may influence initiation and progression of tumors. However, the underlying mechanisms for these observations are poorly understood. The purpose of this study is to explore the effects of adrenergic signaling on cell line models derived from normal cells presumed to originate epithelial ovarian cancers. Here we explored the effects of the stress-related hormone, norepinephrine, on the transcriptional program of normal immortalized ovarian (iOSE) and fallopian tube (iFTSEC) surface epithelial cells. Analysis of RNA-Seq data of treated and untreated cells revealed a significant overlap between the responses in iOSE and iFTSEC cells. Most genes modulated by norepinephrine in ovarian and fallopian tube epithelial cells are already expressed in normal ovarian and fallopian tissue and cells. For several genes, expression changes were reflected at the protein level. Genes in immune-related and developmental pathways were enriched in the set of genes modulated by norepinephrine. We identified HOXA5, SPIB, REL, SRF, SP1, NFKB1, MEF2A, E2F1, and EGR1 transcription factor binding sites to be highly enriched in our dataset. These data represent the early transcriptional response to norepinephrine in cells postulated to originate epithelial ovarian cancer.

Genes Predisposed to DNA Hypermethylation during Acquired Resistance to Chemotherapy Are Identified in Ovarian Tumors by Bivalent Chromatin Domains at Initial Diagnosis.

Bivalent chromatin domains containing both active H3K4me3 and repressive H3K27me3 histone marks define gene sets poised for expression or silencing in differentiating embryonic stem (ES) cells. In cancer cells, aberrantly poised genes may facilitate changes in transcriptional states after exposure to anticancer drugs. In this study, we used ChIP-seq to characterize genome-wide positioning of H3K4me3- and H3K27me3-associated chromatin in primary high-grade serous ovarian carcinomas and in normal ovarian surface and fallopian tube tissue. Gene sets with proximal bivalent marks defined in this manner were evaluated subsequently as signatures of systematic change in DNA methylation and gene expression, comparing pairs of tissue samples taken from patients at primary presentation and relapse following chemotherapy. We found that gene sets harboring bivalent chromatin domains at their promoters in tumor tissue, but not normal epithelia, overlapped with Polycomb-repressive complex target genes as well as transcriptionally silenced genes in normal ovarian and tubal stem cells. The bivalently marked genes we identified in tumors before chemotherapy displayed increased promoter CpG methylation and reduced gene expression at relapse after chemotherapy of ovarian cancer. Overall, our results support the hypothesis that preexisting histone modifications at genes in a poised chromatin state may lead to epigenetic silencing during acquired drug resistance.Significance: These results suggest epigenetic targets for intervention to prevent the emergence of cancer drug resistance. Cancer Res; 78(6); 1383-91. ©2018 AACR.

CTD-2020K17.1, a Novel Long Non-Coding RNA, Promotes Migration, Invasion, and Proliferation of Serous Ovarian Cancer Cells In Vitro.

BackgroundOvarian cancer is the most lethal malignant tumor of the female reproductive system, and the metastasis is one of the major factors that contribute to the poor outcome of patients with OC. Accumulating evidence indicates that lncRNAs are expressed and play important regulatory roles in ovarian cancer.Material/MethodsAberrant lncRNAs in primary ovarian cancer tissues (POCTs) and paired omental metastasis tissues (OMTs) of patients with HGSOC were studied via lncRNA microarray. Real-time PCR was performed to examine CTD-2020K17.1 expression in HGSOC tissues from 38 patients, a normal ovarian surface epithelium cell line, and 4 ovarian cancer cell lines. Additionally, Transwell assays, wound healing assays, CCK-8 proliferation assays, and flow cytometry were used to explore the biological function of CTD-2020K17.1 in ovarian cancer cells. Finally, Western blot analysis was used to verify the potential target gene of CTD-2020K17.1.ResultsA novel lncRNA named CTD-2020K17.1 was identified via microarray analysis. Expression of CTD-2020K17.1 was significantly increased in OMTs and in 4 ovarian cancer cell lines compared with POCTs (P<0.05) or normal ovarian surface epithelial cell line (P<0.05). Moreover, CTD-2020K17.1 overexpression promoted migration, invasion, and proliferation of ovarian cancer cells, and CTD-2020K17.1 regulated the expression of CARD11.ConclusionsCTD-2020K17.1 is significantly upregulated in OMTs and ovarian cancer cell lines. It can promote the migration, invasion, and proliferation of ovarian cancer cells, and CARD11 is regulated by CTD-2020K17.1.

Daidzein exerts anticancer activity towards SKOV3 human ovarian cancer cells by inducing apoptosis and cell cycle arrest, and inhibiting the Raf/MEK/ERK cascade.

Ovarian cancer is the main cause of gynecological cancer‑associated mortality around the world. Despite initial responses to chemotherapy, frequent relapse occurs. Daidzein is an important flavonoid and has been shown to exhibit a diversity of pharmacological properties, including antimicrobial and anticancer activities. However, information on the anticancer activity of daidzein against ovarian cancer remains limited. Therefore, the present study evaluated the anticancer activity of daidzein against a panel of human ovarian cancer cell lines and one normal ovarian cell line (Moody). The results revealed that daidzein exhibited potent anticancer activity against SKVO3 cells with a half‑maximal inhibitory concentration (IC50) of 20µM. However, it exhibited comparatively lower activity against normal ovarian Moody cells, which had an IC50 of 100µM. Daidzein induced morphological changes in SKOV3 cells and mitochondrial apoptosis, as evident from DAPI, AO/EB and AnnexinV/propidium iodide staining. This was associated with the upregulation of B‑cell lymphoma 2‑associated X protein, cytochromec, cleaved caspase‑3and‑9, and cleaved poly (ADP‑ribose) polymerase. Daidzein also triggered G2/M cell arrest through the downregulation of pCdc25c, Cdc25c, pCdc2, Cdc2 and cyclinB1. The effect of daidzein on the migration of SKOV3 cells was also determined, the results of which indicated that daidzein inhibited cell migration in a concentration‑dependent manner and was coupled with concomitant decrease in the expression of matrix metalloproteinase (MMP)‑2 and‑9. Additionally, daidzein‑inhibited cell growth was simultaneous with suppression of the expression of phosphorylated mitogen‑activated protein kinase kinase and phosphorylated extracellular signal‑regulated kinase. The present study also examined whether daidzein exerts similar activity against SKOV3 cells in nude mouse xenograft models and it was revealed that daidzein considerably reduced the tumorigenesis invivo, indicative of the potential for daidzein as a lead molecule in the development of ovarian cancer chemotherapy.

MiR-1182 inhibited metastasis and proliferation of ovarian cancer by targeting hTERT.

To investigate the potential effect of miR-1182 on the development of ovarian cancer and the relevant mechanism. The expression levels of miR-1182 were detected in ovarian cancer tissues and cells (SKOV3) comparing with corresponding adjacent normal tissues and normal ovarian cell (IOSE80). Luciferase assay was performed to evaluate the interaction between miR-1182 and hTERT. The effects of the miR-1182/hTERT axis on SKOV3 cells were determined by subsequent experiments including cell proliferation, expression level of hTERT, the invasion and metastasis detection. miR-1182 was found repressed in ovarian cancer tissues and we got the same results at the cellular level. In order to research potential target of miR-1182, we checked it in three publicly available algorithms, TargetScan, miRDB and microRNA. We found that hTERT is a direct target of miR-1182, and luciferase assays confirmed our hypothesis. The subsequent experiments showed that decreased expression of hTERT resulting from the up-regulation of miR-1182 could decelerate cell proliferation, invasion, and metastasis in ovarian cancer cells. Our research discovers the suppressor function of miR-1182 in ovarian cancer by targeting hTERT, revealing that miR-1182/hTERT axis may be a potential therapeutic target for the treatment of ovarian cancer.

MiR-144 Potentially Suppresses Proliferation and Migration of Ovarian Cancer Cells by Targeting RUNX1.

BackgroundOvarian cancer (OC) is one of the most common malignant diseases of the female reproductive system worldwide. Evidence has shown that microRNAs are involved in the development of ovarian cancer. miR-144, one of these microRNAs, has been found have upregulated expression in various human malignancies. The present study aimed to investigate the role miR-144 in ovarian cancer cell lines and to elucidate the mechanism involved.Material/MethodsHuman ovarian cancer cell lines (SKOV3/OVCAR3) and a normal ovarian cell line (IOSE80) were used to identify the miR-144 expression though qRT-PCR method. SKOV3/OVCAR3 cells were transfected with miR-144 mimics by Lipofectamine, and the proliferation, migration, and invasion ability of these cells were detected by MTT assay, wound healing assay, and Transwell assays, respectively. MMP2 and MMP9 expression were detected at mRNA and protein levels. The results of dual luciferase reporter assay confirmed that miR-144 could down-regulate RUNX1 expression level. Finally, the expression of runt-related transcription factor 1 (RUNX1) was examined using qRT-PCR and Western blot analysis.ResultsOur results demonstrate that the expression level of miR-144 was downregulated in SKOV3/OVCAR3 compared to IOSE80, and we found that miR-144 suppresses the proliferation and migration of ovarian cancer cells. Moreover, RUNX1 was predicted and confirmed to be a target of miRNA-144. Additionally, after 48-h transfection with miR-144 mimics, the expression of RUNX1 was downregulated in OC cells.ConclusionsmiR-144 mimics can inhibit the proliferation and migration of ovarian cancer cells though regulating the expression of RUNX1.

Rab23 promotes the cisplatin resistance of ovarian cancer via the Shh-Gli-ABCG2 signaling pathway.

As a novel member of the Rab GTPase family, the role of Rab23 has been reported in multiple types of tumor. However, to the best of our knowledge, the role of Rab23 in ovarian cancer (OC) has not yet been reported. In the present study, immunohistochemistry analysis demonstrated that Rab23 was upregulated in OC tissue; survival analysis indicated that Rab23 expression was associated with a reduced overall survival (OS) rate and disease-free survival (DFS) time. In vitro experiments also demonstrated the increased expression of Rab23 in the OC cells lines, A2780 and SKOV-3, compared with in the normal ovarian cell line, IOSE80. Following the silencing of ABCG2 in SKOV-3 cells, ATP-binding cassette sub-family G member 2 (ABCG2) expression was significantly downregulated both at the RNA and protein levels. The cisplatin (DDP) IC50 declined from 43.09±7.12 µmol/l in control cells to 26.46±5.38 µmol/l in SKOV-3 cells with silenced Rab23. In contrast, in A2780 cells overexpressing Rab23 (A2780-Rab23), ABCG2 expression was significantly upregulated and the DDP IC50 increased from 27.42±6.54 µmol/l in control cells to 45.92±5.23 µmol/l in A2780-Rab23. Investigation into the potential molecular mechanisms for this revealed that the expression of sonic hedgehog (Shh) and Gli family zinc finger 1 (Gli1) was increased in A2780-Rab23 cells, whereas silencing Rab23 in SKOV-3 cells significantly inhibited the expression of Shh and Gli1. The Gli1 inhibitor GANT-61 significantly abrogated the increased ABCG2 expression in A2780-Rab23 cells. Furthermore, the DDP IC50 in A2780-Rab23 cells decreased significantly following the silencing of ABCG2 expression; the IC50 declined from 51.66±8.32 µmol/l in A2780-Rab23 cells to 25.61±6.17 µmol/l in A2780-Rab23 cells with silenced ABCG2. Collectively, the results indicate that Rab23 promotes the DDP resistance of OC cells via the Shh-Gli1-ABCG2 pathway, providing the proof of principle for the further investigation of drug resistance therapy targeting Rab23.