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OBJECTIVES@#Polycystic ovary syndrome (PCOS) is one of the most common endocrine diseases in women with reproductive age, which is associated with hyperandrogenism, insulin resistance, and ovulatory dysfunction. Progesterone receptor membrane component 1 (PGRMC1) can mediate progesterone to inhibit the apoptosis of ovarian granulosa cells and the growth of follicles, and to induce glucolipid metabolism disorder in ovarian granulosa cells, which is closely related to the occurrence and development of PCOS. This study aims to determine the expression of PGRMC1 in serum, ovarian tissue, ovarian granulosa cells, and follicular fluid in PCOS patients and non-PCOS patients, analyze the value of PGRMC1 in diagnosis and prognosis evaluation of PCOS, and investigate its molecular mechanism on ovarian granulosa cell apoptosis and glucolipid metabolism.@*METHODS@#A total of 123 patients were collected from the Department of Obstetrics and Gynecology in Guangdong Women and Children Hospital (hereinafter referred to as "our hospital") from August 2021 to March 2022 and divided into 3 groups: a PCOS pre-treatment group (n=42), a PCOS treatment group (n=36), and a control group (n=45). The level of PGRMC1 in serum was detected by enzyme linked immunosorbent assay (ELISA). The diagnostic and prognostic value of PGRMC1 was evaluated in patients with PCOS by receiver operating characteristic (ROC) curve. Sixty patients who underwent a laparoscopic surgery from the Department of Obstetrics and Gynecology in our hospital from January 2014 to December 2016 were collected and divided into a PCOS group and a control group (n=30). The expression and distribution of PGRMC1 protein in ovarian tissues were detected by immunohistochemical staining. Twenty-two patients were collected from Reproductive Medicine Center in our hospital from December 2020 to March 2021, and they divided into a PCOS group and a control group (n=11). ELISA was used to detect the level of PGRMC1 in follicular fluid; real-time RT-PCR was used to detect the expression level of PGRMC1 mRNA in ovarian granulosa cells. Human ovarian granular cell line KGN cells were divided into a scrambled group which was transfected with small interfering RNA (siRNA) without interference and a siPGRMC1 group which was transfected with specific siRNA targeting PGRMC1. The apoptotic rate of KGN cells was detected by flow cytometry. The mRNA expression levels of PGRMC1, insulin receptor (INSR), glucose transporter 4 (GLUT4), very low density lipoprotein receptor (VLDLR), and low density lipoprotein receptor (LDLR) were determined by real-time RT-PCR.@*RESULTS@#The serum level of PGRMC1 in the PCOS pre-treatment group was significantly higher than that in the control group (P<0.001), and the serum level of PGRMC1 in the PCOS treatment group was significantly lower than that in the PCOS pre-treatment group (P<0.001). The areas under curve (AUC) of PGRMC1 for the diagnosing and prognosis evaluation of PCOS were 0.923 and 0.893, respectively, and the cut-off values were 620.32 and 814.70 pg/mL, respectively. The positive staining was observed on both ovarian granulosa cells and ovarian stroma, which the staining was deepest in the ovarian granulosa cells. The average optical density of PGRMC1 in the PCOS group was significantly increased in ovarian tissue and ovarian granulosa cells than that in the control group (both P<0.05). Compared with the control group, the PGRMC1 expression levels in ovarian granulosa cells and follicular fluid in the PCOS group were significantly up-regulated (P<0.001 and P<0.01, respectively). Compared with the scrambled group, the apoptotic rate of ovarian granulosa cells was significantly increased in the siPGRMC1 group (P<0.01), the mRNA expression levels of PGRMC1 and INSR in the siPGRMC1 group were significantly down-regulated (P<0.001 and P<0.05, respectively), and the mRNA expression levels of GLUT4, VLDLR and LDLR were significantly up-regulated (all P<0.05).@*CONCLUSIONS@#Serum level of PGRMC1 is increased in PCOS patients, and decreased after standard treatment. PGRMC1 could be used as molecular marker for diagnosis and prognosis evaluation of PCOS. PGRMC1 mainly localizes in ovarian granulosa cells and might play a key role in regulating ovarian granulosa cell apoptosis and glycolipid metabolism.
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Criança , Gravidez , Humanos , Feminino , Síndrome do Ovário Policístico , Apoptose , Células da Granulosa , Metabolismo dos Lipídeos , Proteínas de Membrana , Receptores de ProgesteronaRESUMO
Objective To explore the effect and mechanism of insulin-like growth factor 2 ( IGF2 ) on the proliferation of human ovarian granulosa cells ( KGN ). Methods KGN cells cultured in vitro and treated with different concentrations of IGF2 were divided into control group and IGF2 group (25 μg/L, 50 μg/L, 100 μg/L), and then cells were divided into control group, 100 μg/L IGF2 group, LY294002 group, and IGF2 +LY294002 group after intervened the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway with LY294002. MTS and 5-ethynyl-2'- deoxyuridine (EdU) method was used to detect the effect of IGF2 on KGN cell proliferation, and enzyme linked immunosorbent assay was used to detect the contents of estrogen and progesterone in cell culture supernatant. The expressions of insulin like growth factor 1 receptor (IGF1R), protein kinase B (Akt), phosphorylated protein kinase B(p-Akt) and CYP19A1 protein in each group were detected by Western blotting. Results With the concentration gradient of IGF2, the proliferation rate of KGN cells and the secretion of estrogen and progesterone gradually increased. The cell proliferation rate and hormone level in the group treated with lOOfig/L IGF2 were the highest (P<0.01), while the PI3K/Akt signaling pathway was inhibited, and the cell proliferation rate and hormone secretion decreased significantly (P<0.01). The protein expression levels of IGF1R, p-Akt and CYP19A1 in different concentration groups increased significantly (P<0.05). While the expression of the above proteins were affected by intervened the PI3K/Akt signaling pathway. Compared with the control group, the protein expression of IGF1R and p-Akt increased significantly in IGF2 group and IGF2 +LY294002 group(P<0.01), CYP19A1 increased significantly in IGF2 group(P<0.01), the protein expression of p-Akt and CYP19A1 decreased significantly in LY294002 group (P<0.05), there was no significant difference in the protein expression of IGF 1R. Compared with the IGF2 group, the protein expression of p-Akt and CYP19A1 decreased in IGF2 +LY294002 group (P<0.01), there was no statistically significant difference in the protein expression of IGF1R, and the expression levels of IGF1R, p-Akt and CYP19A1 were significantly reduced in LY294002 group (P<0.01). Conclusion IGF2 may promote the proliferation and secretion of human ovarian granulosa cells through the PI3K/Akt signaling pathway mediated by IGF1R.
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Background: CPEB is considered as an RNA-binding protein first identified in Xenopus oocytes. Although CPEB1 was involved in the growth of oocyte, its role in goat follicular granulosa cell has not been fully elucidated. To clarify the functions of this gene in goat follicular granulosa cells, CPEB1-overexpressing vector and interference vector were structured and transfected into follicular granulosa cells from Jiangsu native white goats of Nantong city, Jiangsu Province, China. The expression levels of differentiation-related genes including CDK1, Cyclin B1, and C-mos were determined 24 h after administration of CPEB1 by quantitative real-time polymerase chain reaction and Western blotting methods. Results: The expression levels of CDK1, Cyclin B1, and C-mos were significantly upregulated after overexpression and significantly downregulated after interference with CPEB1. Conclusions: The CPEB1 gene expression could affect the transcription of genes related to early cleavage divisions, which provided a reference for further research on its role in the growth and maturation of oocytes.
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Animais , Feminino , Oócitos , Fatores de Transcrição/genética , Cabras/genética , Transfecção , Fertilização in vitro , Expressão Gênica , Western Blotting , Reação em Cadeia da Polimerase/métodos , Proteínas de Ligação a RNA , Transferência Embrionária , Gado , Fluorescência , Células da GranulosaRESUMO
Objective To compare the MRI features of ovarian thecoma-fibroma and adult-type ovarian granulosa cell tumor (OGCT),and to investigate the differential value of MRI.Methods 13 patients (8 cases of ovarian thecoma-fibroma,5 cases of adult-type OGCT)confirmed by pathology were collected,in which MRI findings were analyzed retrospectively.Results 13 lesions were all unilateral.The shape of more lesions appeared round or oval,and huge lesions were lobulated.Most tumors were dominant solid and well-defined borders.The length-diameter of ovarian thecoma-fibromas ranged from 23 to 268 mm,with the mean diameter of (124±84)mm;on T1WI the solid part of tumors showed equal signal in 5 cases,slight lower signal in 3 cases,on T2WI all tumors were heterogeneous signal,6 lesions appeared equal signal with obscure nebulous high signal;all of 8 lesions were enhanced slightly;4 cases were accompanied with pelvic effusion and other type tumors of the ovarian.The length-diameter of adult-type OGCT tumors ranged from 27 to 110 mm, with the mean diameter of (84±33)mm;the solid part of lesions showed slight lower signal on T1WI and high signal on T2WI,the honeycomb and fissure cystic areas were found in all tumors,4 cases were accompanied with hemorrhage;all tumors were enhanced from moderately to highly,and delay contrast enhancement;the cases were accompanied with endometrial hyperplasia in 3 cases,endometrial carcinoma in 1 case.The non-enhanced cystic areas of two groups of tumors showed lower signal on T1WI and high signal on T2WI. Conclusion There are different characteristics between ovarian thecoma-fibroma and adult-type OGCT on MRI findings,which are helpful to the differential diagnosis.
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Objective To study the CT and MRI features of ovarian granulosa cell tumor(OGCT).Methods The clinical and imaging data of 12 cases with OGCT proved by pathology were analyzed retrospectively(10 cases were performed with CT,5 cases were performed with MRI,and 3 cases were performed with both CT and MRI).Results All lesions were solitary.The tumors presented round,oval, or lobulated with clear boundary.In 12 cases, 3 cases were multiple cystic masses with uneven thickness of cyst walls and septation.Mild enhancement of the cyst walls and septation was showed after contrast enhancement.7 cases were cystic and solid masses with multiple cysts, typically with sponge-like pattern,and most cases showed slightly to moderately enhancement.2 cases were solid mass with septation,1 case showed significant enhancement,and 1 case showed slightly enhancement.6 of 12 cases presented enlargement of the utreus and endometrium thickening,1 case was associated with endometrial carcinoma.Conclusion OGCT has characteristic CT and MRI features, and CT and MRI examinations combined with estrogen abnormality are helpful for diagnosis.
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Objective The androgen signaling pathway is involved in the regulation of early follicular growth and follicular atresia, and sex hormone binding globulin (SHBG) is an important factor in regulating the level of local ovarian androgen.This study was to investigate the effects of androgen on the expression of SHBG in ovarian granulosa cells.Methods Human ovarian granule cancer cells were cultured with 0 nmol/L dihydrotestosterone (DHT), 500 nmol/L DHT, or 500 nmol/L DHT + 60 μmol/L Flutamide.The expression of SHBG was detected by Western blot and immunofluorescence staining.Hyperandrogenism (HA) was induced in 6 SD rats by subcutaneous injection of dehydroepiandrosterone (DHEA) and another 6 rats were included in the vehicle control group.At 35 days after modeling, all the rats were sacrificed for measurement of the level of SHBG in the blood from the inferior caval vein by ELISA, the expression of SHBG in the ovarian granulosa cells by immunohistochemistry, and expressions of androgen receptor (AR) and SHBG in the liver by Western blot.Results At 24 hours after modeling, the expression of AR was significantly upregulated in the 300, 400, and 500 nmol/L DHT groups as compared with the 0 nmol/L DHT group (1.06±0.02, 1.61±0.11, and 2.38±0.14 vs 1.06±0.03, P<0.05), and so was that of SHBG, increasing in a concentration-dependent manner, most significantly at 500 nmol/L (P<0.01).Both the expressions of AR and SHBG proteins remarkably elevated in the 500 nmol/L DHT group in comparison with the 0 nmol/L DHT group (P<0.01), but markedly downregulated as compared with the 500 nmol/L DHT + 60 μmol/L Flutamide group (P<0.05).Immunofluorescence staining showed that DHT promoted while the addition of Flutamide inhibited the expressions of AR and SHBG.Immunohistochemical staining of the ovarian tissue revealed a high level of SHBG in the HA rats.Compared with the control group, the HA animals exhibited a significantly decreased expression of serum SHBG (4.80±0.35 vs 2.41±0.14, P<0.01) and that in the liver, but a markedly increased level of the AR protein (P<0.05).Conclusion Activation of the androgen signaling pathway by DHT can promote the expression of SHBG in rat ovarian granulosa cells.
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Objective To study the effect of intedeukin 6 (IL-6) on the expression of androgen receptor (AR) in porcine ovarian granulosa cells(GC) and to explore its role of in the polycystic ovary syndrome(PCOS) pathogenesis.Methods Different concentrations of IL-6 (10,100 and 1 000 ng/L) were treated with GC.Expression of androgen receptor (AR) mRNA and protein in GC were detected by real-time quantitative RT-PCR and Western blot.Results The mRNA and protein expressions of AR in GC were upregulated in groups with 100 and 1 000 ng/L IL-6 at 72 h.Conclusion IL-6 can improve androgen activity.This process may explain the function of chronic subclinical inflammation in PCOS pathogenesis.
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OBJECTIVE: Ovarian granulosa cell tumors are rare malignancies with a relatively favorable prognosis. However, patients still suffer from disease-related mortality. Therefore, the prognostic factors should be clarified. The purpose of this study was to investigate the clinical and pathologic characteristics related with disease recurrence and mortality in adult type ovarian granulosa cell tumors. METHODS: Eighty surgically staged patients with granulosa cell ovarian tumor treated at the Hacettepe University Hospital between 1982 and 2006 were retrospectively reviewed. Clinical and pathological characteristics were analyzed. RESULTS: Granulosa cell ovarian tumors accounted for 4.3% of malignant ovarian neoplasms. Mean age was 47.6 years. The most common presenting symptom was abnormal uterine bleeding (53.7%). Endometrial pathology was detected in 51.2% of patients preoperatively. Seventy percent of patients were diagnosed at stage I, and 53.8% of patients received adjuvant treatment. Mean follow-up was 67.5 months. Overall 5-year and 10-year survival was 91% and 86%, respectively. Mean survival was 147.1 months. Recurrence rate was 11.2%. In univariate analysis, advanced stage, advanced age, residual disease after surgery, and need for adjuvant treatment were associated with disease-related mortality and advanced stage disease and absence of initial staging surgery were associated with disease recurrence. However, in multivariate analysis, only initial stage was found to be a significant prognostic factor. CONCLUSION: Initial stage seems to be the single most important prognostic factor in ovarian granulosa cell tumors. Therefore, a comprehensive staging surgery should be attempted to document the real extent of disease and to estimate the oncologic outcome more accurately.
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Adulto , Feminino , Humanos , Seguimentos , Tumor de Células da Granulosa , Células da Granulosa , Análise Multivariada , Neoplasias Ovarianas , Prognóstico , Recidiva , Estudos Retrospectivos , Hemorragia UterinaRESUMO
Objective To study the effect of serum contained Erzhi Tiangui Granule on the secretion function of ovarian granulosa cells from primary senile mice. Methods The serum with decoction from primary senile mice was added to cultured granulosa cells and incubated for 48 h. Then the concentration of estradiol (E2), progesterone (P) and inhibin B (INHB) in cultured medium were measured by chemiluminescence immunoassay (CLIA) and Sandwich ELISA respectively. Results The serum contained Erzhi Tiangui Granule could increase the secretion of E2, P and INHB. Conclusion Erzhi Tiangui Granule can improve the ovarian function of primary senile mice by regulating the secretion function of granulosa cells.
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OBJECTIVE: In the ovary apoptosis eliminates granulosa cells (GC) during follicle atresia. While we identified many cell death regulatory molecules not yet characterized in the ovary, of particular interest were members of the Bcl-2 family which contain only the Bcl-2 homology (BH)-3 domain. The objectives of this study was to investigate and compare the expression patterns of BH3-only Bcl-2 family members in various organs and evaluate their function in ovarian granulosa cells. METHODS: Total RNA was extracted from GC, ovaries, uteri, hearts (low rate of cell turnover) and livers (high rate of cell turnover) of prepubertal female mice. BH3-only Bcl-2 family members were cloned to make riboprobe. The expression patterns in the tissues evaluated by Northern blot analysis. Nix mRNA expression in ovarian granulosa cells after gonadotropin treatment also compared by Northern blot analysis. RESULTS: Bad, Bid, Bim, Bmf, Map-1 and Nix were expressed in granulosa cells. Nix was most abundantly expressed in GC. In contrast, Blk was expressed in the ovary, liver, heart and uterus, but not in GC. Bmf, a sensor or microfilament disassembly, was expressed in GC, ovary and uterus, with limited to no expression in non-reproductive tissues. Nix mRNA expression was not regulated by gonadotropin after 42 hour. CONCLUSION: These studies will help to complete a molecular blueprint of the regulatory network that controls GC death during follicular atresia. In addition, these data, which show a tissue/cell-selective profile of BH3-only expression, may also explain the known variation in the in vivo apoptotic response of different tissues/cells to generic stimuli that should be globally lethal.
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Animais , Feminino , Humanos , Camundongos , Citoesqueleto de Actina , Apoptose , Northern Blotting , Morte Celular , Células Clonais , Atresia Folicular , Gonadotropinas , Células da Granulosa , Coração , Fígado , Ovário , RNA , RNA Mensageiro , ÚteroRESUMO
Objective:To explore the gene differential expression pattern of polycystic ovary syn-drome.Methods:We carried out microarray analysis to define the gene networks by the PCOS granulosacells in order to identify differentially expressed genes in PCOS patients.These granulosa cells of fivePCOS cases and five control cases which were derived during oocyte retrieval from women undergoingIVF.Results:As compared with control human ovarian granulosa cells,46 genes were screened out,25genes were up-regulated,and 21genes were down-regulated in PCOS.These differentially expressedgenes were involved in various biologic functions,such as regulation of fatty acid metabolism,cell-cellsignal transduction,immune and inflammatory response,reflecting the complexity of clinical manifesta-tions of PCOS.Conclusion:Microarray analysis technology is an effective mothod to identify novel PCOSassociated candidate genes.
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McCune-Albright syndrome, which consists of cafe-au lait spots, multiple fibrous dysplasia, sexual precocity and endocrinal dysfunction such as pituitary, thyroid and adrenal aberrations, has been increasingly recognized recently. The etiology is now suggested that activating mutations on the gene that encodes the alpha subunit of stimulatory G protein cause McCune-Albright syndrome. The child presented with sexual precocity and elevated estrogen level was still persisted after left oophorectomy and right partial oophorectomy due to misdiagnosed as ovarian granulosa-theca cell tumor. Whole body bone scan revealed abnormal radiotracer uptake on left facial bone and left proximal femur. Facial bone MRI showed marked thickening of facial bone from fibrous dysplasia. And the patient had cafe-au-lait pigmentations on neck and buttock area. Other endocrine function was normal. Pathologic finding of ovary was finally confirmed with cystic follicles. Skull biopsy revealed fibrous dysplasia. And she was finally diagnosed McCune-Albright syndrome.