Functional characterization of cultured human granulosa cells in serum free culture system

The aim of this study was to explore the direct action of noradrenaline and dopamine on progesterone and estradiol secretion of human granulosa cells cultured in serum-free medium. Progesterone and estradiol production was measured in the presence and absence of noradrenaline, dopamine or propranolol using radioimmunoassays; statistical analysis was performed by analysis of variance and Newman-Keul's multiple range test. Twenty-six women aged 31±3 years undergoing in vitro fertilization and embryo transfer for infertility treatment at University Women's Hospital, University of Tübingen, Germany, took part in this study. Noradrenaline significantly inhibited progesterone production by human granulosa cells in a dose-related manner at a concentration of 10−4–10−6 mol/l. Dopamine significantly stimulated estradiol secretion by granulosa cells in an inverse dose-related manner. Both effects were blocked by propranolol. The results suggest that catecholaminergic actions switch over the steroid production of human granulosa cells cultured in serum-free medium from progesterone to estradiol.

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Utilising FGF2, IGF2 and FSH in serum-free protocol for long-term in vitro cultivation of primary human granulosa cells

Molecular and Cellular Endocrinology ◽

10.1016/j.mce.2020.110816 ◽

2020 ◽

Vol 510 ◽

pp. 110816

Author(s):

Kati Hensen ◽

Martin Pook ◽

Anu Sikut ◽

Tõnis Org ◽

Toivo Maimets ◽

...

Keyword(s):

Granulosa Cells ◽

In Vitro Cultivation ◽

Serum Free ◽

Human Granulosa Cells

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Characterization of a serum-free culture system comparing growth factor requirements of transformed and untransformed cells

Experimental Cell Research ◽

10.1016/0014-4827(90)90167-9 ◽

1990 ◽

Vol 188 (2) ◽

pp. 254-261 ◽

Cited By ~ 4

Author(s):

Kathleen M. Mulder ◽

Karla E. Childress-Fields

Keyword(s):

Growth Factor ◽

Culture System ◽

Serum Free

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Relationship between the production of prostaglandins and progesterone by luteinizing human granulosa cells

Journal of Endocrinology ◽

10.1677/joe.0.1710455 ◽

2001 ◽

Vol 171 (3) ◽

pp. 455-462 ◽

Cited By ~ 8

Author(s):

RC Fowkes ◽

C Chandras ◽

EC Chin ◽

S Okolo ◽

DR Abayasekara ◽

...

Keyword(s):

Granulosa Cells ◽

Serum Free Medium ◽

Free Medium ◽

Progressive Decline ◽

Serum Free ◽

Prostaglandin Production ◽

Human Granulosa Cells ◽

Meclofenamic Acid ◽

High Concentrations ◽

The Relationship

Luteinizing granulosa cells synthesize high concentrations of progesterone, prostaglandin (PG) E(2) and PGF(2 alpha). The objective of this study was to explore the relationship between prostaglandin and progesterone output from human granulosa cells as they undergo functional luteinization in culture. Granulosa cells were partially purified from ovarian follicular aspirates and cultured at a density of 10(5) cells/ml in serum-supplemented DMEM:Ham's F(12) medium for 0, 1 or 2 days. Cells were then switched to serum-free medium for 24 h before measuring hormone concentrations in this spent medium by specific radioimmunoassays. Over the first 3 days in culture, PGF(2 alpha) and PGE(2) production declined progressively by up to 82+/-3% coincident with a 55+/-11% increase in progesterone output. In subsequent experiments, cells were treated for 24 h on the second day of culture with either 0.01 to 10 microM meclofenamic acid or with 10 microM and 100 microM aminoglutethimide. Meclofenamic acid inhibited synthesis of PGF(2 alpha) and PGE(2) by up to 70+/-9% and 64+/-7% respectively without affecting progesterone output. Likewise, 100 microM aminoglutethimide inhibited progesterone production by 62+/-6% without affecting concentrations of either PGF(2 alpha) or PGE(2). We have concluded that the progressive decline in prostaglandin production and the rise in progesterone output from luteinizing human granulosa cells occur independently of each other.

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Dynamic transcriptome and chromatin architecture in granulosa cells during chicken folliculogenesis

Nature Communications ◽

10.1038/s41467-021-27800-9 ◽

2022 ◽

Vol 13 (1) ◽

Author(s):

Diyan Li ◽

Chunyou Ning ◽

Jiaman Zhang ◽

Yujie Wang ◽

Qianzi Tang ◽

...

Keyword(s):

Granulosa Cells ◽

Functional Characterization ◽

Three Dimensional ◽

Range Interaction ◽

Ample Evidence ◽

Chromatin Architecture ◽

Complex Biological Process ◽

Preovulatory Follicles ◽

Long Range Interaction

AbstractFolliculogenesis is a complex biological process involving a central oocyte and its surrounding somatic cells. Three-dimensional chromatin architecture is an important transcription regulator; however, little is known about its dynamics and role in transcriptional regulation of granulosa cells during chicken folliculogenesis. We investigate the transcriptomic dynamics of chicken granulosa cells over ten follicular stages and assess the chromatin architecture dynamics and how it influences gene expression in granulosa cells at three key stages: the prehierarchical small white follicles, the first largest preovulatory follicles, and the postovulatory follicles. Our results demonstrate the consistency between the global reprogramming of chromatin architecture and the transcriptomic divergence during folliculogenesis, providing ample evidence for compartmentalization rearrangement, variable organization of topologically associating domains, and rewiring of the long-range interaction between promoter and enhancers. These results provide key insights into avian reproductive biology and provide a foundational dataset for the future in-depth functional characterization of granulosa cells.

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OXYTOCIN BIOSYNTHESIS IN SERUM-FREE CULTURES OF HUMAN GRANULOSA CELLS

Journal of Endocrinology ◽

10.1677/joe.0.124r005 ◽

1990 ◽

Vol 124 (2) ◽

pp. R5-R8 ◽

Cited By ~ 8

Author(s):

I. plevrakis ◽

C. Clamagirand ◽

G. Pontonnier

Keyword(s):

Granulosa Cells ◽

High Performance ◽

Serum Free Medium ◽

Vitro Fertilization ◽

Serum Free ◽

Human Granulosa Cells ◽

Cell Extracts ◽

Preovulatory Follicles ◽

Follicular Puncture

ABSTRACT Human granulosa cells were collected from preovulatory follicles during follicular puncture for in-vitro fertilization. They were cultured in serum-free medium supplemented with ascorbic acid. Using a combination of high-performance liquid chromatography and radioimmunoassay, the oxytocin material present in the cell extracts and secreted into the medium was identified. When cells were deprived of ascorbate, intermediary forms resulting of the post-translational processing of pro-oxytocin/neurophysin were detected. These data demonstrate that oxytocin biosynthesis occurs in human granulosa cells.

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Differential regulation of pig theca cell steroidogenesis by LH, insulin-like growth factor I and granulosa cells in serum-free culture

Reproduction ◽

10.1530/reprod/118.2.211 ◽

2000 ◽

pp. 211-219 ◽

Cited By ~ 11

Author(s):

EM Shores ◽

HM Picton ◽

MG Hunter

Keyword(s):

Granulosa Cells ◽

Culture System ◽

Theca Cell ◽

Serum Free ◽

Theca Cells ◽

Plating Density ◽

Factor I ◽

Viable Cells ◽

Igf I ◽

Oestradiol Production

The regulation of pig theca cell steroidogenesis was studied by the development of a physiological serum-free culture system, which was subsequently extended to investigate potential theca-granulosa cell interactions. Theca cells were isolated from antral follicles 6-9 mm in diameter and the effects of plating density (50-150x10(3) viable cells per well), LH (0.01-1.0 ng ml(-1)), Long R3 insulin-like growth factor I (IGF-I) (10, 100 ng ml(-1)) and insulin (1, 10 ng ml(-1)) on the number of cells and steroidogenesis were examined. The purity of the theca cell preparation was verified biochemically and histologically. Co-cultures contained 50x10(3) viable cells per well in granulosa to theca cell ratio of 4:1. Wells containing granulosa cells only were supplemented with 'physiological' doses of androstenedione or 100 ng ml(-1). Oestradiol production by co-cultures was compared with the sum of the oestradiol synthesized by granulosa and theca cells cultured separately. Oestradiol and androstenedione production continued throughout culture. High plating density decreased steroid production (P < 0.01). LH increased androstenedione (P < 0.001) and oestradiol (P < 0.05) synthesis and the sensitivity of the cells increased with time in culture. Oestradiol production was increased by 10 ng IGF-I ml(-1) (P < 0.001) but androstenedione required 100 ng ml(-1) (P < 0.001). Co-cultures produced more oestradiol than the sum of oestradiol synthesized by theca and granulosa cells cultured separately (P < 0. 001), irrespective of the androstenedione dose. This serum-free culture system for pig theca cells maintained in vivo steroidogenesis and gonadotrophin responsiveness. Thecal androstenedione and oestradiol production were differentially regulated and were primarily stimulated by LH and IGF-I, respectively. Theca-granulosa cell interactions stimulated oestradiol synthesis and this interaction was mediated by factors additional to the provision of thecal androgen substrate to granulosa cells.

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