Establishment and Characterization of a Steroidogenic Human Granulosa-Like Tumor Cell Line, KGN, That Expresses Functional Follicle-Stimulating Hormone Receptor

Nishi, Yoshihiro

doi:10.1210/en.142.1.437

Cited by 146 publications

(129 citation statements)

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“…The following primers were used for real-time PCR: for VEGF, forward 5 0 -TGCAGATTA-TGCGGATCAAACC, reverse 5 0 -TGCATTCACATTTGT-TGTGCTGTAG; for VEGFR-2, forward 5 0 -GGAAGCTC-CTGAAGATCTGT, reverse 5 0 -GAGGATATTTCGTGCCGC; for GAPDH control, forward 5 0 -TCATTTCCTGGTATGA-CAACG, reverse 5 0 -TTACTCCTTGGAGGCCATGT. Standard curve method was applied using purified mRNA from an established human GCT cell line (KGN) (27) as standard. All analyses were performed in triplicate with an ABI PRISM 7700 sequence detection system (Applied Biosystems) according to the manufacturer's instructions.…”

Section: Quantitative Real-time Pcrmentioning

confidence: 99%

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 are highly expressed in ovarian granulosa cell tumors

Färkkilä¹,

Anttonen²,

Pociuviene³

et al. 2011

European Journal of Endocrinology

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Objective: Ovarian granulosa cell tumors (GCTs) are hormonally active sex cord stromal tumors accounting for 3-5% of all ovarian cancers. These tumors are generally diagnosed at an early stage but there is a high risk of recurrence, associated with high mortality. Treatment of recurrent GCTs is difficult, and biologically targeted treatment modalities are lacking. GCTs are highly vascularized, and angiogenic factors most probably play a role in their pathology. Vascular endothelial growth factor (VEGF) is a key regulator of tumor angiogenesis, but in GCTs, the role of VEGF and its receptors VEGFR-1 (FLT1) and VEGFR-2 (KDR) remains largely unknown. Our objective is to study the expression of VEGF and its receptors in human GCTs. Methods: We analyzed GCTs from 106 patients for the expressions of VEGF and its receptors utilizing tumor tissue microarray, tumor mRNA, and patient serum samples. Results: We found that VEGF and its main biologically active receptor VEGFR-2 were highly expressed in primary and recurrent GCTs, when compared with normal granulosa-lutein cells. The expression of VEGF correlated positively to tumor microvessel density and to VEGFR-2 expression at the protein (P!0.05) and mRNA (P!0.05) levels. In contrast to VEGFR-2, the expression of VEGFR-1 was weak. Tumor VEGF protein expression was not prognostic for recurrence, however, we found high levels of circulating VEGF in the serum of patients with primary GCT. Conclusions:The results suggest an important role of VEGF and VEGFR-2 in GCT pathology and support the possibility of applying novel VEGF-or VEGFR-2-targeted treatments to patients with GCT.

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Section: Quantitative Real-time Pcrmentioning

confidence: 99%

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 are highly expressed in ovarian granulosa cell tumors

Färkkilä¹,

Anttonen²,

Pociuviene³

et al. 2011

European Journal of Endocrinology

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“…These data suggested that FOXL2 may function as a suppressor of ovarian follicle progression in small and medium follicles by the prevention of premature differentiation and/or proliferation of granulosa cells. However, testing this hypothesis in a human system is limited by lack of access to human granulosa cells other than cell lines derived from granulosa cell tumors, such as the KGN cell line (Nishi et al 2001), which contains a C402G mutation in FOXL2 (Shah et al 2009). To determine whether mouse FOXL2 functions similarly to human FOXL2, and the utility of mouse granulosa cells as a model system, we cloned the mouse Foxl2 cDNA and determined its role in the regulation of the mouse Star, Cyp19a1, Cyp11a1, and Ccnd2 genes under the influence of FSH in mouse granulosa cells.…”

Section: Introductionmentioning

confidence: 99%

Mouse forkhead L2 maintains repression of FSH-dependent genes in the granulosa cell

Kuo¹,

Fan²,

Bentsi-Barnes³

et al. 2012

Reproduction

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The forkhead transcription factor forkhead box L2 (FOXL2) is expressed in granulosa cells of small and medium follicles in the mouse ovary. Foxl2 female knockout mice exhibit primordial follicle depletion and primary ovarian failure, but evidence from adult female conditional Foxl2 knockout mice suggests that FOXL2 may also play a significant role in maintenance of ovarian differentiation at stages beyond the primordial follicle and initial wave of folliculogenesis. We previously showed that human FOXL2 functions as a transcriptional repressor of several key genes involved in granulosa cell proliferation and differentiation, including steroidogenic acute regulatory protein (STAR), P450aromatase (CYP19A1 (CYP19)), P450scc (CYP11A1 (CYP11A)), and cyclin D2 (CCND2). To elucidate the role of mouse FOXL2, we determined its role in transcriptional regulation in Chinese hamster ovary (CHO) cells and then confirmed our findings in mouse granulosa cells. We found that mouse FOXL2 represses the activities of the mouse Star, Cyp19a1, Cyp11a1 promoters in CHO cells, but may not repress the Ccnd2 promoter, and identified the minimal mouse Star, Cyp19a1, and Cyp11a1 promoter regions responsive to FOXL2 regulation. We then knocked down Foxl2 in mouse granulosa cells using siRNA, which resulted in significantly increased expression levels of mouse Star, Cyp19a1, and Cyp11a1 but not Ccnd2. To increase Foxl2 expression levels, we generated a mouse Foxl2 lentiviral construct and used it to infect mouse granulosa cells. Following lentiviral infection, the expression levels of mouse Star, Cyp19a1, and Cyp11a1, but not Ccnd2, decreased significantly. These data confirm that mouse FOXL2 functions as a transcriptional repressor of key granulosa cell genes that influence ovarian development.

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“…Primary granulosa cells (hGCs), an immortalized human granulosa cell line, SVOG, and a granulosa tumor cell line, KGN, were used as cell models. The KGN cell line (stocked in the RIKEN CELL Bank) was derived from a human ovarian granulosa cell tumor, which expresses the functional FSH receptor and maintains the functions of steroidogenesis and Fas-mediated apoptosis [26]. The SVOG cell line was produced by transfecting granulosa cells with SV40 Tag as described previously [27].…”

Section: Introductionmentioning

confidence: 99%

Homeobox A7 increases cell proliferation by up-regulation of epidermal growth factor receptor expression in human granulosa cells

Huang

Cheng

Nishi³

et al. 2010

Reprod Biol Endocrinol

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BackgroundHomeobox (HOX) genes encode transcription factors, which regulate cell proliferation, differentiation, adhesion, and migration. The deregulation of HOX genes is frequently associated with human reproductive system disorders. However, knowledge regarding the role of HOX genes in human granulosa cells is limited.MethodsTo determine the role of HOXA7 in the regulation and associated mechanisms of cell proliferation in human granulosa cells, HOXA7 and epidermal growth factor receptor (EGFR) expressions were examined in primary granulosa cells (hGCs), an immortalized human granulosa cell line, SVOG, and a granulosa tumor cell line, KGN, by real-time PCR and Western blotting. To manipulate the expression of HOXA7, the HOXA7 specific siRNA was used to knockdown HOXA7 in KGN. Conversely, HOXA7 was overexpressed in SVOG by transfection with the pcDNA3.1-HOAX7 vector. Cell proliferation was measured by the MTT assay.ResultsOur results show that HOXA7 and EGFR were overexpressed in KGN cells compared to hGCs and SVOG cells. Knockdown of HOXA7 in KGN cells significantly decreased cell proliferation and EGFR expression. Overexpression of HOXA7 in SVOG cells significantly promoted cell growth and EGFR expression. Moreover, the EGF-induced KGN proliferation was abrogated, and the activation of downstream signaling was diminished when HOXA7 was knocked down. Overexpression of HOXA7 in SVOG cells had an opposite effect.ConclusionsOur present study reveals a novel mechanistic role for HOXA7 in modulating granulosa cell proliferation via the regulation of EGFR. This finding contributes to the knowledge of the pro-proliferation effect of HOXA7 in granulosa cell growth and differentiation.

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Establishment and Characterization of a Steroidogenic Human Granulosa-Like Tumor Cell Line, KGN, That Expresses Functional Follicle-Stimulating Hormone Receptor

Cited by 146 publications

References 0 publications

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 are highly expressed in ovarian granulosa cell tumors

Vascular endothelial growth factor (VEGF) and its receptor VEGFR-2 are highly expressed in ovarian granulosa cell tumors

Mouse forkhead L2 maintains repression of FSH-dependent genes in the granulosa cell

Homeobox A7 increases cell proliferation by up-regulation of epidermal growth factor receptor expression in human granulosa cells

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