scholarly journals Mechanisms subserving the trophic actions of insulin on ovarian cells. In vitro studies using swine granulosa cells

1983 ◽  
Vol 72 (3) ◽  
pp. 1046-1057 ◽  
Author(s):  
Johannes D. Veldhuis ◽  
Lisa A. Kolp ◽  
Michael E. Toaff ◽  
Jerome F. Strauss ◽  
Lawrence M. Demers
Keyword(s):  
Granulosa Cells ◽  
In Vitro Studies ◽  
Ovarian Cells

scholarly journals A rapid and robust method for the cryopreservation of human granulosa cells

2021 ◽  
Author(s):  
Sarah Beschta ◽  
Katja Eubler ◽  
Nancy Bohne ◽  
Ignasi Forne ◽  
Dieter Berg ◽  
...  
Keyword(s):  
Granulosa Cells ◽  
Large Scale ◽  
Ovarian Function ◽  
Oocyte Retrieval ◽  
Cell Contact ◽  
Cellular Model ◽  
Preovulatory Follicle ◽  
Human Granulosa Cells ◽  
Ovarian Cells

AbstractHuman primary granulosa cells (GCs) derived from women undergoing oocyte retrieval can be cultured and used as a cellular model for the study of human ovarian function. In vitro, they change rapidly, initially resembling cells of the preovulatory follicle and then cells of the corpus luteum. They are derived from individual patients, whose different medical history, lifestyle and age lead to heterogeneity. Thus, cells can rarely be ideally matched for cellular experiments or, if available, only in small quantities. We reasoned that cryopreservation of human GCs may be helpful to improve this situation. Previous studies indicated the feasibility of such an approach, but low survival of human GCs was reported, and effects on human GC functionality were only partially evaluated. We tested a slow freezing protocol (employing FCS and DMSO) for human GCs upon isolation from follicular fluid. We compared cryopreserved and subsequently thawed cells with fresh, non-cryopreserved cells from the same patients. About 80% of human GCs survived freezing/thawing. No differences were found in cell morphology, survival rate in culture, or transcript levels of mitochondrial (COX4, OPA1, TOMM20), steroidogenic (CYP11A1, CYP19A1) or cell–cell contact genes (GJA1) between the two groups in cells cultured for 1–5 days. A proteomic analysis revealed no statistically significant change in the abundance of a total of 5962 proteins. The two groups produced comparable basal levels of progesterone and responded similarly to hCG with elevation of progesterone. Taken together, our results show this to be a rapid and readily available method for the cryopreservation of human GCs. We anticipate that it will allow future large-scale experiments and may thereby improve cellular studies with human ovarian cells.

scholarly journals Serum deprivation affects secretory activity of cultured porcine ovarian follicles and granulosa cells and their response to hormones

2009 ◽  
Vol 54 (No. 10) ◽  
pp. 455-460
Author(s):  
A.V. Sirotkin
Keyword(s):  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Secretory Activity ◽  
Ovarian Follicles ◽  
Serum Deprivation ◽  
Ovarian Cells ◽  
Igf I ◽  
Granulose Cells ◽  
Vitro Model

The aim of the present study is to understand the hormonal mechanisms of the effect of malnutrition on ovarian follicle functions. For this purpose, we examined the effect of malnutrition/serum deprivation, addition of metabolic hormones and gonadotropin (IGF-I, leptin and FSH) and their combination on the release of progesterone (P<sub>4</sub>), testosterone (T), estradiol (E<sub>2</sub>) and insulin-like growth factor I (IGF-I) by cultured whole ovarian follicles and on P<sub>4</sub> and IGF-I output by cultured granulosa cells isolated from porcine ovaries. It was observed that in ovarian follicles cultured with nutrients/serum addition of IGF-I reduced release of P<sub>4</sub>, but not of T or E<sub>2</sub>. Exogenous leptin reduced output of E<sub>2</sub>, but not of P<sub>4</sub> or T, and increased IGF-I output. No significant effect of FSH on release of steroid hormones by isolated follicles was found. Serum deprivation did not affect release of P<sub>4</sub>, but reduced output of T and E<sub>2</sub>, and promoted IGF-I release by cultured ovarian follicles. Addition of hormones failed to prevent the effect of malnutrition on the secretory activity of cultured ovarian follicles. In cultured granulose cells, all the tested hormones promoted release of both P<sub>4</sub> and IGF-I. Food restriction/serum deprivation reduced both P<sub>4</sub> and IGF-I output. Additions of either IGF-I, leptin and FSH prevented the inhibitory action of malnutrition on both P<sub>4</sub> and IGF-I release. The present observations (1) confirm the involvement of the hormones IGF-I, leptin and FSH in the control of secretory activity of ovarian cells, (2) demonstrate, that both isolated ovarian granulosa cells and whole follicles cultured in the absence of serum nutrients could be an adequate in-vitro model for studying the effect of malnutrition on ovarian secretory functions, and (3) suggest, that malnutrition could affect ovarian functions through changes in the release of ovarian hormones.

The Role of Estradiol as a Biological Amplifier of the Actions of Follicle-Stimulating Hormone: In Vitro Studies in Swine Granulosa Cells*

Endocrinology ◽  
1982 ◽  
Vol 111 (1) ◽  
pp. 144-151 ◽  
Author(s):  
JOHANNES D. VELDHUIS ◽  
PATRICIA A. KLASE ◽  
JEROME F. STRAUSS ◽  
JAMES M. HAMMOND
Keyword(s):  
Granulosa Cells ◽  
Follicle Stimulating Hormone ◽  
In Vitro Studies ◽  
Stimulating Hormone

scholarly journals FSH, Oxytocin and IGF-I Regulate the Expression of Sirtuin 1 in Porcine Ovarian Granulosa Cells

2020 ◽  
pp. 461-466
Author(s):  
A SIROTKIN ◽  
P DEKANOVÁ ◽  
A HARRATH
Keyword(s):  
Granulosa Cells ◽  
Intracellular Signaling ◽  
Sirtuin 1 ◽  
Signaling System ◽  
Factor I ◽  
Ovarian Granulosa Cells ◽  
Ovarian Cells ◽  
Igf I ◽  
Hormonal Action

The involvement of the mTOR system/enzyme sirtuin 1 (SIRT1) intracellular signaling system in the control of ovarian functions and its role in mediating hormonal action on the ovary has been proposed, but this hypothesis should be supported by a demonstrated influence of hormones on mTOR/SIRT1. Therefore, the aim of our in vitro experiments was to examine the effect of the known hormonal regulators of ovarian functions, such as follicle-stimulating hormone (FSH), oxytocin (OT) and insulin-like growth factor I (IGF-I), on mTOR/SIRT1. The accumulation of SIRT1 in porcine ovarian granulosa cells cultured with and without these hormones (at doses of 1, 10 or 100 ng.ml-1) was evaluated using immunocytochemistry. It was observed that the addition of FSH (at 10 ng.ml-1 but not at 1 or 100 ng/ml) and OT (at all tested doses) increased the expression of SIRT1 in ovarian cells. In addition, 100 ng.ml-1, but not at 1 or 10 ng.ml-1, of IGF-I decreased SIRT1 accumulation. Our observations are the first demonstration that hormones can directly regulate the ovarian mTOR/SIRT1 system and that this system could mediate the action of hormonal regulators on the ovary.

scholarly journals KIRREL is differentially expressed in adipose tissue from ‘fertil+’ and ‘fertil−’ cows: in vitro role in ovary?

Reproduction ◽  
2018 ◽  
Vol 155 (2) ◽  
pp. 181-196
Author(s):  
S Coyral-Castel ◽  
C Ramé ◽  
J Cognié ◽  
J Lecardonnel ◽  
S Marthey ◽  
...  
Keyword(s):  
Adipose Tissue ◽  
Dairy Cows ◽  
Granulosa Cells ◽  
Tiling Array ◽  
Bovine Chromosome ◽  
Bovine Kidney ◽  
Ovarian Cells ◽  
Expression Of Genes ◽  
Trait Locus

We have previously shown that dairy cows carrying the ‘fertil−’ haplotype for one quantitative trait locus affecting female fertility located on the bovine chromosome three (QTL-F-Fert-BTA3) have a significantly lower conception rate and body weight after calving than cows carrying the ‘fertil+’ haplotype. Here, we compared by Tiling Array the expression of genes included in the QTL-F-Fert-BTA3 in ‘fertil+’ and ‘fertil−’ adipose tissue one week after calving when plasma non-esterified fatty acid concentrations were greater in ‘fertil−’ animals. We observed that thirty-one genes were overexpressed whereas twelve were under-expressed in ‘fertil+’ as compared to ‘fertil−’ cows (P < 0.05). By quantitative PCR and immunoblot we confirmed that adipose tissue KIRREL mRNA and protein were significantly greater expressed in ‘fertil+’ than in ‘fertil−’. KIRREL mRNA is abundant in bovine kidney, adipose tissue, pituitary, and ovary and detectable in hypothalamus and mammary gland. Its expression (mRNA and protein) is greater in kidney of ‘fertil+’ than ‘fertil−’ cows (P < 0.05). KIRREL (mRNA and protein) is also present in the different ovarian cells with a greater expression in granulosa cells of ‘fertil+’ than ‘fertil−’ cows. In cultured granulosa cells, recombinant KIRREL halved steroid secretion in basal state (P < 0.05). It also decreased cell proliferation (P < 0.05) and in vitro oocyte maturation (P < 0.05). These results were associated to a rapid increase in MAPK1/3 and MAPK14 phosphorylation in granulosa cells and to a decrease in MAPK1/3 phosphorylation in oocyte. Thus, KIRREL could be a potential metabolic messenger linking body composition and fertility.

scholarly journals Gamma Tocopherol Reduced Chemotherapeutic-Induced ROS in an Ovarian Granulosa Cell Line, But Not in Breast Cancer Cell Lines In Vitro

Antioxidants ◽  
2020 ◽  
Vol 9 (1) ◽  
pp. 51 ◽  
Author(s):  
Daniela Figueroa Gonzalez ◽  
Fiona Young
Keyword(s):  
Breast Cancer ◽  
Granulosa Cells ◽  
Breast Cancer Cell Lines ◽  
Human Breast ◽  
Gamma Tocopherol ◽  
Ovarian Cells ◽  
Viable Cells ◽  
Ovarian Granulosa Cell ◽  
Treat Breast Cancer

Doxorubicin and cyclophosphamide are used to treat breast cancer, but they also cause infertility through off-target cytotoxicity towards proliferating granulosa cells that surround eggs. Each chemotherapeutic generates reactive oxygen species (ROS) but the effects of the combination, or the antioxidants alpha (αToc) and gamma tocopherol (γToc) on ROS in breast cancer or ovarian cells are unknown. Human breast cancer (MCF7, T47D) and ovarian cancer (OVCAR, COV434) cells were loaded with DCDFA and exposed (1, 2, 3, 24 h) to the MCF7-derived EC25 values of individual agents, or to combinations of these. ROS were quantified and viable cells enumerated using crystal violet or DAPI. Each chemotherapeutic killed ~25% of MCF7, T47D and OVCAR cells, but 57 ± 2% (doxorubicin) and 66 ± 2% (cyclophosphamide) of the COV434 granulosa cells. The combined chemotherapeutics decreased COV434 cell viability to 34 ± 5% of control whereas doxorubicin + cyclophosphamide + γToc reduced ROS within 3 h (p < 0.01) and reduced cytotoxicity to 54 ± 4% (p < 0.05). αToc was not cytotoxic, whereas γToc killed ~25% of the breast cancer but none of the ovarian cells. Adding γToc to the combined chemotherapeutics did not change ROS or cytotoxicity in MCF7, T47D or OVCAR cells. The protection γToc afforded COV434 granulosa cells against chemotherapy-induced ROS and cytotoxicity suggests potential for fertility preservation.

scholarly journals Apelin (APLN) regulates progesterone secretion and oocyte maturation in bovine ovarian cells

Reproduction ◽  
2017 ◽  
Vol 153 (5) ◽  
pp. 589-603 ◽  
Author(s):  
J Roche ◽  
C Ramé ◽  
M Reverchon ◽  
N Mellouk ◽  
A Rak ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Germinal Vesicle ◽  
Meiotic Progression ◽  
Ovarian Cells ◽  
Progesterone Secretion ◽  
Maturation Medium ◽  
G Protein Coupled

APLN and its G-protein coupled receptor APLNR are expressed in the bovine ovary. However their role in granulosa cells and oocytes is unknown. Here, we studied their expression in bovine ovarian cells and investigated their regulation in cultured luteinizing granulosa cells in response to IGF1 and FSH. We determined the effect and the molecular mechanism of APLN (isoforms 17 and 13) on bovine granulosa cell progesterone secretion and on oocyte maturation. By RT-qPCR and immunoblot, we showed that the expression of both APLN and APLNR in granulosa and oocytes significantly increased with ovarian follicles size whereas it was similar in theca interstitial cells.In vitro, in unstimulated luteinizing bovine granulosa cells and in response to IGF1 (10−8 M) but not to FSH (10−8 M), we observed that APLN (-17 and -13) (10−9 M) increased progesterone production; this was abolished in response to the APLNR antagonist ML221. These latter effects were dependent on the MAPK ERK1/2 kinase. Furthermore, we showed that APLN (-17 and -13) (10−9 M) increased cell proliferation through AKT signaling. Conversely, the addition of APLN-13 and APLN-17 toin vitromaturation medium containing IGF1 (10−8 M) but not FSH (10−8 M) arrested most oocytes at the germinal vesicle stage, which was associated with a decrease in progesterone secretion, an inhibition in MAPK ERK1/2 phosphorylation and an increase in PRKA phosphorylation in oocytes. Thus, APLN can increase progesterone secretion and cell proliferation in bovine luteinizing granulosa cellsin vitro, while it blocks meiotic progression at the germinal vesicle stage during bovine oocytein vitromaturation.

Interaction between oocytes, cortical germ cells and granulosa cells of the mouse and bat, following the dissociation–re-aggregation of adult ovaries

Zygote ◽  
2020 ◽  
Vol 28 (3) ◽  
pp. 223-232
Author(s):  
Tania Janeth Porras-Gómez ◽  
Norma Moreno-Mendoza
Keyword(s):  
Germ Cells ◽  
Granulosa Cells ◽  
Fluorescent Protein ◽  
Primordial Germ Cells ◽  
Morphological Characteristics ◽  
Active Role ◽  
Cellular Interaction ◽  
Cortical Cells ◽  
Ovarian Cells

SummaryIt is widely accepted that the oocyte plays a very active role in promoting the growth of the follicle by directing the differentiation of granulosa cells and secreting paracrine growth factors. In turn, granulosa cells regulate the development of the oocytes, establishing close bidirectional communication between germ and somatic cells. The presence of cortical cells with morphological characteristics, similar to primordial germ cells that express specific germline markers, stem cells and cell proliferation, known as adult cortical germ cells (ACGC) have been reported in phyllostomid bats. Using magnetic cell separation techniques, dissociation–cellular re-aggregation and organ culture, the behaviour of oocytes and ACGC was analyzed by interacting in vitro with mouse ovarian cells. Bat ACGC was mixed with disaggregated ovaries from a transgenic mouse that expressed green fluorescent protein. The in vitro reconstruction of the re-aggregates was evaluated. We examined the viability, integration, cellular interaction and ovarian morphogenesis by detecting the expression of Vasa, pH3, Cx43 and Laminin. Our results showed that the interaction between ovarian cells is carried out in the adult ovary of two species, without them losing their capacity to form follicular structures, even after having been enzymatically dissociated.

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