Steroidogenic properties of a spontaneously established porcine granulosa cell line (PGC-2)

Kwan, I; Farookhi, Riaz; Murphy, Bruce D.; Turner, Jeffrey D.; Downey, B.R.

doi:10.1002/(sici)1098-2795(199611)45:3<299::aid-mrd6>3.0.co;2-n

Cited by 14 publications

(11 citation statements)

References 26 publications

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“…2D and data not shown). Similar results were obtained in an established porcine granulosa cell line, PGC-2 [34] (data not shown). DC3 cells transfected with a constitutively active form of CTNNB1 (S37A) or treated with lithium chloride displayed dose-dependent increases in pGL3-OT activity, indicating functional CTNNB1/TCF-dependent (pGL3-OT) transcriptional responsiveness in this cell line (Fig.…”

Section: Dc3 Cells Expressed Fzd4 But Not Wnt2 Mrnamentioning

confidence: 99%

Characterization of Wnt2 Overexpression in a Rat Granulosa Cell Line (DC3): Effects on CTNNB1 Activation1

Finnson

Kontogiannea

et al. 2012

Biology of Reproduction

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WNTs comprise a family of secreted glycoproteins that are essential for normal embryonic development of the female reproductive system. The functional role that WNTs play in the postnatal ovary is poorly defined. We have shown previously that Wnt2 and Fzd4 mRNAs are expressed in granulosa cells of the postnatal rat ovary. Here we examine the effects of Wnt2 overexpression in a rat granulosa cell line (DC3) that displays characteristics of granulosa cells at an early stage of follicular development. We show that DC3 cells express a 7.7-kb Fzd4 mRNA transcript similar in size to that detected in the rat and human ovary. Our results demonstrate that Wnt2 overexpression in DC3 promotes cytosolic and nuclear accumulation of beta-catenin (CTNNB1), but does not stimulate CTNNB1/TCF-dependent (pGL3-OT) transcriptional activity. We show that chibby (CBY1), a nuclear CTNNB1-associated antagonist of the WNT pathway, is expressed in DC3 cells and associates with CTNNB1 in the presence and absence of Wnt2 overexpression, suggesting that Cby1 contributes to suppression of CTNNB1/TCF-dependent transcription in these cells. Our results show that Wnt2 overexpression in DC3 cells increases follistatin (Fst) mRNA expression and promotes resistance to activin-induced cell deletion. Taken together, our results suggest that WNT2 opposes activin activity in granulosa cells by up-regulating expression of the activin antagonist Fst in a CTNNB1/TCF-independent manner, and that rat granulosa cells express factors, including Cby1, that suppress CTNNB1/TCF-dependent signal transduction in the presence of a WNT signal.

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Section: Dc3 Cells Expressed Fzd4 But Not Wnt2 Mrnamentioning

confidence: 99%

Characterization of Wnt2 Overexpression in a Rat Granulosa Cell Line (DC3): Effects on CTNNB1 Activation1

Finnson

Kontogiannea

et al. 2012

Biology of Reproduction

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“…The porcine granulosa cell line PGC-2 (20) was cultured in MEM with the additives described above and 10% fetal calf serum (Gibco BRL). Y-1 mouse adrenal tumor cells [American Type Culture Collection (ATCC), Manassas, VA] were maintained in Dulbecco's modified Eagle's medium/F12 (Gibco BRL) supplemented with 10% horse serum, 2.5% FBS, and antibiotics.…”

Section: Plasmid Constructionsmentioning

confidence: 99%

Cholesterol supply and SREBPs modulate transcription of the Niemann-Pick C-1 gene in steroidogenic tissues

Gévry

Schoonjans

Guay

et al. 2008

Journal of Lipid Research

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We tested whether sterol-regulatory element binding proteins (SREBPs) mediate sterol-regulated transactivation of the Niemann-Pick C-1 (NPC-1) gene. Loading granulosa cells with 22-or 25-hydroxycholesterol decreased NPC-1 mRNA, whereas culturing in cholesterol-depleted medium or inhibition of cholesterol biosynthesis increased NPC-1 promoter activity and NPC-1 mRNA abundance. Cotransfection of SREBP1a, SREBP1c, and SREBP2 and the NPC-1 promoter-luciferase reporter into granulosa cell lines increased the transcriptional activity of porcine, human, and mouse NPC-1 promoters. Deletion analysis of the 5 ¶ flanking region of the pig NPC-1 gene demonstrated significant promoter activity between fragments 2934 and 2636 bp upstream from the transcription initiation site. Sequence analysis revealed three sterol-regulatory elements (SREs) clustered between 2558 and 2650 bp. Each site, along with E-box sequences, bound recombinant SREBP in electromobility shift assays. Mutation of all three sites attenuated the SREBP induction of promoter activity. Chromatin immunoprecipitation (ChIP) assays revealed that cholesterol depletion enriched the association of both SREBP and acetylated histone H3 with the NPC-1 promoter fragment containing the three SREs. ChIP analysis confirmed that SREBP's association with SRE and the E-box was enriched in cells cultured in cholesterol-depleted medium. We conclude that NPC-1 is sterol-regulated, achieved by SREBP acting via SRE and the E-box sequences.-Gévry,

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“…Most of the granulosa cell lines described in the literature express only some of the functions known to be expressed in granulosa cells in vivo. In only three cases, the cell lines conserve the expression of aromatase [14,19,23]. The GK cell lines described here express granulosa cell-specific genes like AMH and the steroidogenic enzyme P450 scc , with AMH representing a gene specifically expressed in growing follicles [41] and P450 scc being up-regulated around the time of ovulation with high levels of expression in luteinized granulosa cells [5].…”

Section: Discussionmentioning

confidence: 99%

“…The majority of these cell lines represent luteinizing granulosa cells, demonstrating that the process of immortalization does not prevent the spontaneous luteinization of granulosa cells in culture. One cell line conserving FSH receptor expression and the stimulation of estradiol production by FSH has been described [14]; two other lines show induction of P450 aromatase (P450 arom ) expression by cAMP [19] or constitutive P450 arom expression [23]. Recently, one cell line has been described that responds to FSH by an irreversible stop of proliferation followed by cell differentiation [25].…”

Section: Introductionmentioning

confidence: 96%

“…Main indicators used are an increase in progesterone production and the downregulation of estradiol synthesis. In order to develop experimental systems better suited for the study of granulosa cell functions, numerous granulosa cell lines have been established from several species, mostly by immortalization of primary granulosa cells by viral oncogenes or stimulation of granulosa cell growth by mitogens [11][12][13][14][15][16][17][18][19][20][21][22][23][24][25]. The majority of these cell lines represent luteinizing granulosa cells, demonstrating that the process of immortalization does not prevent the spontaneous luteinization of granulosa cells in culture.…”

Section: Introductionmentioning

confidence: 99%

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Expression of Granulosa Cell-Specific Genes and Induction of Apoptosis in Conditionally Immortalized Granulosa Cell Lines Established from H-2Kb-tsA58 Transgenic Mice

Walther¹,

Einspanier

Jansen³

1999

Biology of Reproduction

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Granulosa cell lines have been established from H-2Kb-tsA58 transgenic mice. Using immunocytochemistry, significant amounts of insulin-like growth factor-I (IGF-I) were found in all cell lines investigated, whereas estrogen and progesterone receptor expression could be detected in only some of the lines. All cell lines showed low basal production of the gonadal steroids estradiol and progesterone. The genes for the ovarian paracrine regulators IGF-I and basic fibroblast growth factor were expressed, as well as the genes for anti-Müllerian hormone and for the P450 side-chain cleavage enzyme (P450scc). Expression of P450scc could be shown to be up-regulated in the cell lines under conditions mimicking the hormonal environment of the luteinizing granulosa cells in vivo. Inactivation of the temperature-sensitive SV40 T antigen by a shift of the cell lines to the nonpermissive temperature of 39.5 degrees C led to massive induction of apoptosis in several cell lines. These cell lines will allow a detailed study of the mechanisms regulating the expression of granulosa cell-specific functions, as well as the induction of granulosa cell apoptosis.

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Steroidogenic properties of a spontaneously established porcine granulosa cell line (PGC-2)

Cited by 14 publications

References 26 publications

Characterization of Wnt2 Overexpression in a Rat Granulosa Cell Line (DC3): Effects on CTNNB1 Activation1

Characterization of Wnt2 Overexpression in a Rat Granulosa Cell Line (DC3): Effects on CTNNB1 Activation1

Cholesterol supply and SREBPs modulate transcription of the Niemann-Pick C-1 gene in steroidogenic tissues

Expression of Granulosa Cell-Specific Genes and Induction of Apoptosis in Conditionally Immortalized Granulosa Cell Lines Established from H-2Kb-tsA58 Transgenic Mice

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