Proliferation Of Ovarian Cells Research Articles

Exosomal miRNA-17-5p derived from human umbilical cord mesenchymal stem cells improves ovarian function in premature ovarian insufficiency by regulating SIRT7.

Premature ovarian insufficiency (POI) is clinically irreversible in women aged over 40 years. Although numerous studies have demonstrated satisfactory outcomes of mesenchymal stem cell therapy, the underlying therapeutic mechanism remains unclear. Exosomes were collected from the culture medium of human umbilical cord mesenchymal stem cells (hUMSCs) and assessed by electron microscopy and Western blot (WB) analysis. Then, exosomes were added to the culture medium of cyclophosphamide (CTX)-damaged human granulosa cells (hGCs), and the mixture was injected into the ovaries of CTX-induced POI model mice before detection of antiapoptotic and apoptotic gene expression. Next, the microRNA expression profiles of hUMSC-derived exosomes (hUMSC-Exos) were detected by small RNA sequencing. The ameliorative effect of exosomal microRNA-17-5P (miR-17-5P) was demonstrated by miR-17-5P knockdown before assessment of ovarian phenotype and function, reactive oxygen species (ROS) levels and SIRT7 expression. Finally, SIRT7 was inhibited or overexpressed by RNA interference or retrovirus transduction, and the protein expression of PARP1, γH2AX, and XRCC6 was analyzed. The ameliorative effect of hUMSC-Exos on POI was validated. Our results illustrated that hUMSC-Exos restored ovarian phenotype and function in a POI mouse model, promoted proliferation of CTX-damaged hGCs and ovarian cells, and alleviated ROS accumulation by delivering exosomal miR-17-5P and inhibiting SIRT7 expression. Moreover, our findings elucidated that miR-17-5P repressed PARP1, γH2AX, and XRCC6 by inhibiting SIRT7. Our findings suggest a critical role for exosomal miR-17-5P and its downstream target mRNA SIRT7 in hUMSC transplantation therapy. This study indicates the promise of exosome-based therapy for POI treatment.

CiRS-126 inhibits proliferation of ovarian granulosa cells through targeting the miR-21-PDCD4-ROS axis in a polycystic ovarian syndrome model.

Polycystic ovarian syndrome (PCOS) is considered to be one of the most prevalent endocrine disorders affecting women of reproductive age. CiRS-126, an innovative circular microRNA, has previously been proven to be a promising miR-21 sponge. However, a proper understanding of the impact of ciRS-126 on PCOS is needed. Circular RNA (CiRS) profiles were initially evaluated in ovarian cortex samples obtained from 18 women with PCOS as well from 9 women without PCOS. Insulin-induced ovarian granulosa cells isolated from mice were utilized for the functional study. CiRS microarray analysis and quantitative real-time PCR indicated that ciRs-126 expression was downregulated while miR-21 expression was upregulated in PCOS samples and insulin-induced granulosa cells as compared with non-PCOS samples and non-insulin-induced granulosa cells. Furthermore, ectopic overexpression of ciRS-126 was associated with a reduction in proliferation and increased apoptosis in insulin-treated granulosa cells. Meanwhile, bioinformatic prediction and the results of the dual-luciferase reporter assay indicated the presence of consecutive binding in the ciRS-126-miR-21-programmed cell death protein 4 (PDCD4) axis. Moreover, overexpression of miR-21 blocked ciRS-126 repression of proliferation and triggered the death of insulin-induced granulosa cells. Excessive PDCD4 expression counteracted the influence of miR-21 on cell death and proliferation. The data indicated that PDCD4 played a regulatory role in ROS generation, which is reportedly involved in apoptosis. Therefore, ciRS-126 reduction in PCOS granulosa cells targeted the miR-21-PDCD4 axis to reduce proliferation and promote apoptosis. CiRS-126 shows potential as a promising predictor of clinical outcome as well as a therapeutic target in PCOS.

Effects of Ochratoxin A exposure on DNA damage in porcine granulosa cells in vitro

Ochratoxin A (OTA), a feed mycotoxin, tends to impair the reproductive performance of animals. Our previous studies have demonstrated that OTA exposure inhibits porcine ovarian granulosa cell (GC) proliferation and induces their apoptosis, but the underlying toxic mechanism is still uncertain. In this study, we explored the OTA exposure on porcine GCs in vitro and found that OTA exposure inhibited the proliferation of porcine GCs and arrested cell cycle of GCs in the G2/M phase. The results based on RNA-Seq revealed that 20 μM and 40 μM OTA exposure increase DNA damage of porcine GCs in vitro. The differentially expressed genes (DEGs) of 40 μM OTA exposure were enriched in the pathways of mismatch repair, nucleotide excision repair and homologous recombination in DNA replication compared with control group and 20 μM OTA exposure group. Meanwhile, OTA exposure increased the expression levels of DNA double-strand breaks (DSBs) gene γ-H2AX, and DNA repair related genes, such as BRCA1, XRCC1, PARP1, and RAD51. Above all, our research revealed that OTA might exert deleterious effects on porcine ovarian GCs, influencing DNA repair-related biological processes and causing DNA damage response.

LncRNA TONSL-AS1 regulates miR-490-3p/CDK1 to affect ovarian epithelial carcinoma cell proliferation

BackgroundLncRNA TONSL-AS1 has been characterized as a critical player in gastric cancer. By analyze the TCGA dataset, we observed the upregulation of TONSL-AS1 in ovarian epithelial carcinoma (EOC). We therefore investigated the involvement of TONSL-AS1 in EOC.MethodsThe differential expression of TONSL-AS1 in EOC was first explored by analyzing the TCGA dataset. The effects of overexpression of TONSL-AS1 and miR-490-3p on the expression of CDK1 mRNA and protein in OVCAR3 cells were evaluated by qPCR and western blot, respectively. CCK-8 assay was performed to investigate the effects of overexpression of TONSL-AS1, miR-490-3p and CDK1 on proliferation of OVCAR3 cells.ResultsWe observed that TONSL-AS1 was upregulated in EOC tumor tissues from EOC patients, and its high expression level was correlated with poor survival. Dual luciferase assay and RNA interaction prediction showed the direct interaction between TONSL-AS1 and miR-490-3p. However, overexpression of miR-490-3p did not affect the expression of TONSL-AS1. Instead, overexpression of TONSL-AS1 resulted in the upregulation of CDK1, a target of miR-490-3p, in EOC cells. Overexpression of TONSL-AS1 and CDK1 resulted in increased proliferation rate of EOC cells. Overexpression of miR-490-3p played an opposite role and reduced the effects of overexpression of TONSL -AS1 and CDK1.ConclusionsTherefore, TONSL-AS1 may regulate miR-490-3p/CDK1 to affect EOC cell proliferation.

Interleukin-1β and TNF-α systems in ovarian follicles and their roles during follicular development, oocyte maturation and ovulation

Tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) are cytokines that are involved in the development, proliferation and apoptosis of ovarian follicular cells in domestic mammals. The expression of these cytokines in various follicular compartments, depending on the stage of follicle development, demonstrates their involvement in the control of primordial follicle growth up to the preovulatory stage. The mechanism of action of these factors depends on the presence of their receptors that transduce their biological actions. This review shows the expression sites of TNF-α, IL-1β and their receptors in ovarian follicles, and discusses the mechanism of action of these cytokines during follicle development, oocyte maturation and ovulation in domestic animals.

Downregulation of lncRNA ZFAS1 and upregulation of microRNA-129 repress endocrine disturbance, increase proliferation and inhibit apoptosis of ovarian granulosa cells in polycystic ovarian syndrome by downregulating HMGB1

ObjectiveThe objective was to find the role of long-non-coding RNA zinc finger antisense 1 (lncRNA ZFAS1)/microRNA (miR)-129/high-mobility group box protein 1 (HMGB1) axis in polycystic ovary syndrome (PCOS). MethodsOvarian granulosa cells from non-PCOS patients and PCOS patients were collected, and HMGB1, miR-129 and lncRNA ZFAS1 expression were detected. Ovarian granulosa cells were transfected with si-ZFAS1 or miR-129 mimics to verify their roles in P4 and E2 secretion, and the biological functions of ovarian granulosa cells. ResultsLncRNA ZFAS1 and HMGB1 were elevated, while miR-129 was down-regulated in ovarian granulosa cells of PCOS patients. Down-regulated lncRNA ZFAS1 or overexpressed miR-129 could decrease HMGB1 expression, increase P4 and E2 secretion, promote proliferation activity while inhibit apoptosis of ovarian granulosa cells in PCOS. ConclusionLncRNA ZFAS1 could bind to miR-129 to promote HMGB1 expression, thereby affecting the endocrine disturbance, proliferation and apoptosis of ovarian granulosa cells in PCOS.

Dioscin Ameliorates Polycystic Ovary Syndrome by Inhibiting PI3K/Akt Pathway-Mediated Proliferation and Apoptosis of Ovarian Granulosa Cells

To understand the mechanism underlying Dioscin inhibition of polycystic ovary syndrome, we have examined its effects on ovarian granulosa cells from letrozole-treated rats. To this end, Western blot was utilized to determine changes in the levels of Bcl-2, cleaved caspase-3, caspase-3, and phosphatidylinositol 3-kinase (PI3K)/protein kinase B pathway after Dioscin treatment in letrozole-treated rats. Dioscin ameliorated polycystic ovary syndrome by reducing the serum level of testosterone and increasing progesterone levels. It also inhibited proliferation and induced apoptosis of ovarian granulosa cells in the rat model by decreasing the level of Bcl-2 and elevating cleaved caspase-3. Western blot analysis revealed that Dioscin suppressed the PI3K/Akt pathway by inhibiting p-AKT/AKT. SC79, a p-AKT/AKT activator, reversed the effects of Dioscin on the proliferation and apoptosis of ovarian granulosa cells. In conclusion, Dioscin might present a novel therapeutic opportunity for patients with polycystic ovary syndrome.

Türkiye’de yetiştirilen dört keçi populasyonunda Sinir Büyüme Faktörü (NGF) geninde SacII ve MboII polimorfizmlerinin belirlenmesi

Nerve Growth Factor Gene (NGF) has important functions in the maintenance of viability and proliferation of ovarian cells. Recent studies on caprine NGF gene revealed two SNPs (determined by SacII/C291A and MboII/A705G polymorphisms) related with body length and litter size in goats. This study aimed to determine SacII and MboII polymorphisms of NGF in native Turkish goat populations including Turkish Hair (HAI), Kabakulak (KBK), Honamlı (HNM) and Norduz (NRD) by using PCR-RFLP. Amplificated PCR fragments of 808 were digested with SacII, while amplificated PCR fragments of 319 bp were digested with MboII restriction enzyme in order to detect different genotypes of NGF gene. The “A” allele frequency ranged from 0.500 (HAI, HNM and KBK) to 0.950 (NRD), while the “C” allele frequency ranged from 0.025 (NRD) to 0.500 (HAI, HNM and KBK) in NGF/SacII polymorphism. The “A” allele frequency ranged from 0.213 (HAI) to 1.000 (NRD), while the “G” allele frequency ranged from 0.000 (NRD) to 0.787 (HAI) in NGF/MboII polymorphism. Deviation from HW equilibrium was significant in HNM goat population (P < 0.05). In this study, polymorphisms of caprine NGF gene in native Turkish goat populations were revealed for the first time. The results obtained from this study showed that NGF/SacII polymorphisms could be used for body length in NRD population while NGF/MboII polymorphisms could be used for litter size in HAI, HNM and KBK populations in MAS studies.

Circular RNA expression profiling in the fetal side of placenta from maternal polycystic ovary syndrome and circ_0023942 inhibits the proliferation of human ovarian granulosa cell.

Circular RNAs (circRNAs) are widely expressed noncoding RNAs which play important roles in various processes. The present study aimed to explore the effect of maternal PCOS on the expression of circRNAs in fetus and assessed the potential role of circRNA in human ovarian granulosa cell proliferation. Total RNA was extracted from the fetal side of placental tissues from maternal PCOS (n = 3) and healthy puerpera (n = 3) for circRNA microarray. Real-time reverse transcriptase quantitative PCR (RT-qPCR) was used to validate the microarray data in fetal side of placental tissues from puerpera with (n = 18) and without (n = 30) PCOS. Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis were applied to predict the functions and pathways of circ_0023942 host genes. The circRNA-miRNA-mRNA network was constructed through bioinformatics prediction. Circ_0023942 overexpression vector was transiently transfected into human ovarian granulosa cell lines KGN and COV434. Cell proliferation was detected by cell counting kit-8. The protein expression level was determined by western blot. Compared with healthy puerpera, 14 circRNAs were significantly upregulated and 101 circRNAs were significantly downregulated in the fetal side of placenta from maternal PCOS according to the microarray data. Six differentially expressed circRNAs were selected for validation by RT-qPCR, and the expression patterns of circ_0023942, circ_0002151, circ_0001274, and circ_0008514 were consistent with the microarray data. Circ_0023942 was chosen for further investigation. GO and KEGG analysis predicted that circ_0023942 participated in the regulation of developmental process and the MAPK signaling pathway. Seven miRNAs were predicted to be the targets of circ_0023942. Overexpression of circ_0023942 inhibited human ovarian granulosa cell proliferation and suppressed the expression of CDK-4. Maternal PCOS impairs circ_0023942 expression in fetus. Overexpression of circ_0023942 inhibits human ovarian granulosa cell proliferation possibly via regulating CDK-4.

Two insulin receptors coordinate oogenesis and oviposition via two pathways in the green lacewing, Chrysopa pallens

Insulin signalling in insects, as in mammals, regulates various physiological functions, such as reproduction. However, the molecular mechanism by which insulin signals orchestrate ovarian stem cell proliferation, vitellogenesis, and oviposition remains elusive. Here, we investigate the functions of the phosphoinositide 3-kinase (PI3K)-serine/threonine kinase (Akt) pathway, GTPase Ras/mitogen-activated protein kinase (MAPK) pathway, and their downstream messengers in a natural predator, Chrysopa pallens, by the RNAi method. When C. pallens vitellogenin gene 1 (CpVg1) expression was knocked down, the follicle maturation was arrested and total fecundity was reduced. Silencing C. pallens insulin receptor 1 (CpInR1) suppressed Vg transcription and reduced egg mass and hatching rate. Depletion of C. pallens insulin receptor 2 (CpInR2) transcripts lowered Vg transcript level, hampered ovarian development and decreased reproductive output. Knockdown of C. pallens Akt (CpAkt) and C. pallens extracellular-signal-regulated kinase (Cperk) caused phenotypes similar to those caused by knockdown of CpInR2. Disruption of C. pallens transcription factor forkhead box O (CpFoxO) expression caused no significant effects on ovarian development, but sharply impaired total fecundity. Interference with the expression of C. pallens target of rapamycin (CpTor) gene and C. pallens cAMP-response element binding protein (CpCreb) gene led to a down-regulation of Vg transcription, blocking of ovariole growth, and decrease in egg quality. These results suggested the two CpInRs orchestrate oogenesis and oviposition via two signalling pathways to guarantee natural reproduction in the green lacewing, C. pallens.

Adiponectin is secreted by theca layer cells isolated from chicken ovarian follicles.

Adiponectin, an adipokine hormone, influences glucose utilization, insulin sensitivity and energy homeostasis by signaling through two distinct receptors, ADIPOR1 and ADIPOR2. We previously reported that adiponectin and its receptors are expressed in several organs, including testes in chicken. We report herein that adiponectin gene is expressed exclusively in theca layer while ADIPOR1 and ADIPOR2 genes are expressed in granulosa and theca layers of all preovulatory and prehierarchical follicles of the chicken ovary. Estradiol and/or progesterone treatment of sexually immature chickens significantly altered expression of adiponectin and ADIPOR1 in the ovary. Using anti-chicken adiponectin-, ADIPOR1-, or ADIPOR2- antibodies, adiponectin-immunoreactive (ir) cells were found exclusively in the theca layer, and ADIPOR1-ir and ADIPOR2-ir cells were found both in theca and granulosa layers. Theca layer cells dispersed from preovulatory and prehierarchical follicles were found to synthesize and secrete a 720 kDa heavy molecular weight (HMW) isoform of adiponectin in vitro. Recombinant chicken adiponectin (rcADN) expressed in eukaryotic cells under serum-free conditions comprised primarily of the HMW isoform. Treatment of granulosa cells dispersed from 9 to 12 mm preovulatory follicle and 6 to 8 mm prehierarchical follicle with rcADN or an adiponectin receptor agonist, adipoRon, increased pERK and pACC abundance. In addition, both rcADN and adipoRon were found to significantly decrease the expression of steroidogenic acute regulatory protein gene expression in granulosa cells of preovulatory and prehierarchical follicles. In conclusion, adiponectin secreted by theca cell layer is identical in mass to circulating adiponectin. Systemic and/or theca-derived adiponectin is likely to affect proliferation, metabolism, and steroidogenesis of ovarian follicular cells.

The direct effects of gonadotropin-releasing hormone on proliferation of granulosa cells and development of follicles in goose

ABSTRACT 1. The study objectives were to determine the direct effects of gonadotropin-releasing hormone (GnRH) on the proliferation of ovarian granulosa cells (GCs) and the development of follicles in geese (Anser cygnoides) by colorimetry and ethynyl-2′-deoxyuridine (EdU) cell proliferation assays, in which primary GCs were treated with different concentrations of GnRH agonist (alarelin acetate) and an antagonist (cetrorelix acetate). Differently expressed genes (DEGs) were identified by RNA-sequencing and validated by quantitative reverse transcription polymerase chain reaction (RT–qPCR) and Western blotting. 2. The EdU assays showed that the proliferation of GCs was affected by the GnRH agonist and antagonist in a dose-dependent manner. The effect of treatment on cell proliferation was statistically significant at the concentrations of 10−5 mol/l alarelin and 1 mg/l cetrorelix acetate. A total of 134 DEGs (76 downregulated and 58 upregulated for alarelin treatment) and 226 DEGs (90 downregulated and 136 upregulated for cetrorelix) were identified by RNA-sequencing analysis, respectively. Enrichment analysis indicated that DEGs were enriched in the GO terms of cell–cell signalling and cell junctions. The pathways that regulate the development of follicles were identified, including the biological progress of cAMP accumulation, ovulation cycle and vasculature that are essential to follicular selection. 3. The results suggested that GnRH might directly regulate GC proliferation via autocrine or paracrine pathways related to cell junctions. In particular, it was confirmed that the mRNA and protein expression levels of the oestrogen receptor 2 (ESR2) gene, a negative transcription factor involved in follicular maturation and ovulation, were affected by GnRH agonist or antagonist in GCs. 4. In conclusion, GnRH might play an important role in follicular development by changing the expression of genes that participate in cAMP accumulation, ovulation cycle and cell junctions in ovarian GCs.

RETRACTED: lncRNA PVT1/MicroRNA-17-5p/PTEN Axis Regulates Secretion of E2 and P4, Proliferation, and Apoptosis of Ovarian Granulosa Cells in PCOS

Recently, the roles of microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) were identified in polycystic ovary syndrome (PCOS). In the present study, we investigated the role of the lncRNA PVT1/miR-17-5p/PTEN axis in PCOS ovarian granulosa cells. Expression of PVT1, miR-17-5p and PTEN in PCOS ovarian granulosa cells and follicular fluid was detected, and homeostatic model assessment of insulin resistance (HOMA-IR) and the levels of fasting plasma glucose (FPG), fasting insulin (FINS), and sex hormones were assessed. Then, the proliferation, apoptosis, and colony formation ability of ovarian granulosa cells were evaluated. The binding relationship between PVT1 and miR-17-5p as well as the target relationship between miR-17-5p and PTEN were determined by bioinformatics analysis, luciferase activity assay, RNA-induced silencing complex assay, and RNA pull-down assay. The levels of sex hormone-binding globulin and follicle-stimulating hormone were abated and the levels of luteinizing hormone, testosterone, FINS, FPG, and HOMA-IR were increased in PCOS serum. PVT1 and PTEN were overexpressed and miR-17-5p was reduced in PCOS ovarian granulosa cells and follicular fluid. Overexpressed miR-17-5p and inhibited PVT1 could decelerate apoptosis while accelerating colony formation ability and proliferation of ovarian granulosa cells in PCOS. Moreover, overexpression of PVT1 and reduced miR-17-5p could reverse these results. There existed target relation among PVT1, miR-17-5p, and PTEN, and PVT1 could inhibit miR-17-5p, thereby elevating PTEN. Our study suggests that inhibited PVT1 and overexpressed miR-17-5p result in downregulation of PTEN and promotion of cell proliferation, as well as inhibition of apoptosis of ovarian granulosa cells in PCOS.

Non‐coding RNA LOXL1-AS1 exhibits oncogenic activity in ovarian cancer via regulation of miR‐18b‐5p/VMA21 axis

Long non-coding RNAs (lncRNAs) exert critical effects in the process of malignant cancers and lncRNA LOXL1 Antisense RNA 1 (LOXL1-AS1) has been demonstrated to be a pro-oncogene in multiple tumor types. In the current study, we illuminated the precise roles of LOXL1-AS1 in the development of ovarian cancer. LOXL1-AS1 is significantly overexpressed in ovarian carcinoma tissue compared with adjacent non‐cancerous sample. The luciferase reporter gene assay reveals the relationship between LOXL1-AS1 and miR‐18b‐5p, miR‐18b‐5p and its target gene, Vacuolar ATPase Assembly Factor VMA21 (VMA21). Transfection of LOXL1-AS1 siRNA or miR‐18b‐5p mimics inhibits the growth and aggressive phenotypes of SKOV3 and OVCAR3 cell. Furthermore, miR-18b-5p suppresses ovarian carcinoma cell proliferation and metastasis by targeting VMA21 and LOXL1-AS1 regulates ovarian carcinoma cell growth and metastasis through sponging miR‐18b‐5p. These findings suggest that lncRNA LOXL1-AS1 promotes ovarian cancer cell growth, migratory and invasiveness via modulating miR‐18b‐5p/VMA21 axis.

RETRACTED ARTICLE: MiR-140 targets RAP2A to enable the proliferation of insulin-treated ovarian granulosa cells

BackgroundWe elucidated the role of specific MicroRNAs (miRNAs) in the development of polycystic ovary syndrome (PCOS) and explained the changes in the proliferation of granulosa cells. Excised ovarian cortex specimens were collected for miRNA profiling analysis (n = 20 PCOS females and 5 non-PCOS females). Insulin-treated ovarian granulosa cells isolated from mice were used for mechanical studies.ResultsHigh miR-140 expression was observed in PCOS samples and insulin-treated granulosa cells compared to that in non-PCOS and unstimulated cells, respectively. However, the Ras-related protein Rap-2a precursor (RAP2A) was downregulated in in PCOS. MTT assay and EdU staining showed that an miR-140 inhibitor attenuated viability in insulin-treated granulosa cells; cell viability increased with miR-140 overexpression. Reduced expression of miR-140 and the expression of the miR-140 mimic resulted in marked cell apoptosis, as evidenced by the results of PI flow cytometry and Annexin V-FITC; miR-140 overexpression results in downregulated RAP2A expression, and the miR-140 mimic directly bound to the RAP2A 3′-UTR, causing increase in RAP2A levels in insulin-treated granulosa cells; RNA-mediated silencing of RAP2A in insulin-treated granulosa cells restored cell proliferation and apoptosis to normal levels. Phosphorylated AKT was found to be negatively regulated through cross-talk between miR-140 and RAP2A.ConclusionsIn conclusion, PCOS ovarian cortex specimens and insulin-treated granulosa cells showed elevated expression of miR-140, which could lead to increased proliferation and reduced apoptosis of cells by targeting RAP2A. This study may pave the way for future research on the properties of granulosa cells in PCOS.

Pleuromutilin Inhibits Proliferation and Migration of A2780 and Caov-3 Ovarian Carcinoma Cells and Growth of Mouse A2780 Tumor Xenografts by Down-Regulation of pFAK2.

BackgroundPleuromutilin is a natural tricyclic, derived from the fungus, Pleurotus mutilus. This study aimed to investigate the effects of pleuromutilin on migration and proliferation of A2780 and Caov-3 human ovarian carcinoma cells and the growth of A2780 tumor xenografts in mice and the molecular mechanisms involved.Material/MethodsA2780 and Caov-3 human ovarian carcinoma cells were cultured with and without 40, 160, and 200 μM of pleuromutilin. The Edu fluorescence assay, the wound-healing assay, and Matrigel were used to measure A2780 and Caov-3 cell proliferation, migration, invasion, and adhesion in vitro, respectively. Western blot measured protein levels of FAK, p-FAK, MMP-2, and MMP-9. A2780 cells were injected subcutaneously into mice to determine the effects of pleuromutilin on the growth of tumor xenografts.ResultsPleuromutilin significantly reduced A2780 and Caov-3 cell proliferation at 48 h in a dose-dependent manner (P<0.05), and at 200 μM, pleuromutilin reduced cell proliferation by 21.43% and 23.65%, respectively. Treatment of A2780 cells with pleuromutilin significantly reduced cell migration, invasion, and adhesion and the expression of p-FAK, MMP-2, and MMP-9 compared with untreated controls. In the mouse tumor xenograft model, treatment with pleuromutilin significantly reduced tumor size compared with the untreated group and inhibited tumor metastasis to the intestine, spleen, and peritoneal cavity.ConclusionsIn A2780 and Caov-3 human ovarian carcinoma cells, pleuromutilin inhibited cell proliferation, migration, invasion, and adhesion in a dose-dependent manner, and reduced tumor growth and metastases in a mouse A2780 cell tumor xenograft model.

Puncturevine (Tribulus terrestris L.) affects the proliferation, apoptosis, and ghrelin response of ovarian cells

The objective of our study was to examine the direct effects of the medicinal plant Tribulus terrestris L. (puncturevine) on the basic functions of ovarian cells, including their proliferation, apoptosis, and response to the physiological hormonal stimulator ghrelin. In the first series of experiments, porcine ovarian granulosa cells were cultured with or without puncturevine extracts at concentrations of 0, 1, 10, or 100 μg/ml. In the second series of experiments, these cells were cultured with ghrelin at concentrations of 0, 1, 10, or 100 ng/ml, either alone or in combination with puncturevine (10 μg/ml). The expression levels of the proliferation marker PCNA and the apoptosis marker bax were analyzed via quantitative immunocytochemical methods. Puncturevine was found to stimulate the accumulation of both proliferation and apoptotic markers. Additionally, ghrelin alone could promote the proliferation and apoptosis of ovarian cells. The presence of puncturevine reversed ghrelin-stimulated apoptosis and instead induced apoptotic inhibition. However, puncturevine did not modify the proliferation-inducing effect of ghrelin. These observations demonstrated that (1) puncturevine directly promotes cell proliferation and apoptosis, turnover, of ovarian cells; (2) ghrelin is involved in the regulation of ovarian cell apoptosis and proliferation, consistent with existing evidence; (3) puncturevine antagonizes and even reverses the effects of the hormonal regulator, ghrelin, on ovarian cell apoptosis, but not proliferation; and (4) puncturevine affects not only the basic functions of ovarian cells but also their responses to upstream hormonal regulators.

Persistent endocrine-disrupting chemicals found in human follicular fluid stimulate IGF1 secretion by adult ovarian granulosa cell tumor spheroids and thereby increase proliferation of non-cancer ovarian granulosa cells

Insulin-like growth factor-1 (IGF1) is a hormone involved in cell proliferation. We previously showed that IGF1 directly stimulates cell proliferation in granulosa cell tumors (GCTs). Estrogen regulates IGF1 expression in several reproductive organs including the uterus and ovaries. This study aimed to investigate the effects of a mixture of endocrine-disrupting chemicals (EDCs) on secretion of IGF1 by COV434 and KGN cells, which have been used as in vitro models of juvenile and adult GCTs, respectively. The EDC mixture contained perfluorooctanoate, perfluorooctane sulfonate, 2,2-dichlorodiphenyldichloroethylene, hexachlorobenzene, and polychlorinated biphenyl 153, which are persistent hormonally active environmental toxicants present in ovarian follicular fluid (FF). Expression and secretion of IGF1 were significantly higher in GTCs than in HGrC1 human non-cancer granulosa cells (with the profile HGrC1 < COV434 < KGN). Treatment with the EDC mixture as well as individual test compounds significantly increased IGF1 secretion in KGN cells. Moreover, IGFBP3 gene expression in KGN cells was downregulated after treatment with the EDC mixtures. The estrogen receptor alpha pathway was involved in this effect. Conditioned medium of KGN cells treated with the EDC mixture increased proliferation of HGrC1 human non-cancer granulosa cells. These results indicate that the mixture of EDCs found in FF increases secretion of IGF1 by KGN cells and thus indirectly contributes to progression of adult GCTs, and increases proliferation of non-cancer granulosa cells.

MiR-29 promotes ovarian carcinoma cell proliferation through the PTEN pathway

The objective of this study was to investigate the role of miR-29 in ovarian carcinoma progression. Cell proliferation was measured with a cell counter. mRNA expression of miR-29, phosphatase and tensin homolog (PTEN), and proliferating cell nuclear antigen (PCNA) was measured by RT-PCR. Protein expression was detected by Western blot. The results demonstrated that miR-29 expression was upregulated in ovarian carcinoma and that miR-29 promoted cell proliferation. Consistently, miR-29 silencing decreased ovarian carcinoma cell proliferation. Mechanistically, we found PTEN expression was decreased following miR-29 overexpression and PTEN expression was increased following miR-29 silencing. Importantly, overexpression of PTEN was sufficient to inhibit ovarian carcinoma cell proliferation, demonstrating a key role for PTEN downstream of miR-29. Therefore, our study highlights the miR-29-PTEN pathway as a critical mediator of ovarian cancer cell proliferation, providing a new therapeutic target for patients with ovarian carcinoma.

KNDC1 Is a Predictive Marker of Malignant Transformation in Borderline Ovarian Tumors.

BackgroundFew screening markers for malignant transformation in borderline ovarian tumors (BOT) have been clearly established. The kinase noncatalytic C-lobe domain containing 1 (KNDC1), a brain-specific Ras guanine nucleotide exchange factor, negatively regulates dendrite growth. However, the biological role and underlying mechanism of KNDC1 in human cancers, including ovarian cancer (OC), remain unknown.MethodsGene chip screening was used to detect the expression of KNDC1 mRNA in normal ovarian tissues, BOT tissues, and OC tissues. And results were further validated by RT-qPCR, Western blotting and immunohistochemistry. KNDC1 overexpression and knockdown ovarian cancer cells were established to study the possible pathways that KNDC1 was involved. The effects of KNDC1 on the malignant behaviors of ovarian tumors were also investigated both in vitro and in vivo.ResultsWe observed that the expression of KNDC1 mRNA and KNDC1 protein in OC was significantly downregulated compared with BOT. Subsequent investigation revealed that knockdown of KNDC1 enhanced the proliferation of ovarian cancer cells in vitro via induction of ERK1/2 phosphorylation, whereas reinforcing the expression of KNDC1 attenuated the ERK1/2 activity. Similarly, knockdown of KNDC1 also promoted cell proliferation in vivo. Survival analysis showed that lower KNDC1 predicted a poor progression-free survival (PFS) for patients.ConclusionCollectively, we conclude that KNDC1 might function as a tumor suppressor in ovarian tumors, inhibiting the proliferation of ovarian cells by suppressing ERK1/2 activity and hindering the malignant transformation of BOT.