Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Gene expression in cultured bovine ovarian granulosa

ABSTRACT: The growth of the mammalian ovarian follicle requires the formation of a fluid filled antrum, and maturation and differentiation of the ovarian granulosa cells, largely under the control of Follicle Stimulating Hormone (FSH). Many follicles will regress and die by a process called atresia at this early antral stage. We therefore decided to analyse the gene expression profiles of granulosa cells cultured in the presence or absence of FSH and Tumour Necrosis Factor-alpha (TNF-alpha), an apoptotic factor, to simulate the key influences. Different concentratons of FSH and TNFa in granulosa culture were used to determine effective conditions via estradiol and progesterone production, and cell number. RNA for the array experiments and quantitative real time PCR was extracted from cells cultured with FSH added at 0.33 and TNF-alpha at 50 ng/ml. Four treatments of : (1) FSH alone, (2) TNF-alpha alone, (3) FSH + TNF-alpha and (4) control = neither drug, with replicates (n=4, except controls n=3 ) were used to generate RNA for the gene expression arrays (n=15)

ORGANISM(S): Bos taurus

SUBMITTER: Nicholas Hatzirodos

PROVIDER: E-GEOD-42535 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The global effect of follicle-stimulating hormone and tumour necrosis factor α on gene expression in cultured bovine ovarian granulosa cells.

Glister Claire C Hatzirodos Nicholas N Hummitzsch Katja K Knight Philip G PG Rodgers Raymond J RJ

BMC genomics 20140128

<h4>Background</h4>Oocytes mature in ovarian follicles surrounded by granulosa cells. During follicle growth, granulosa cells replicate and secrete hormones, particularly steroids close to ovulation. However, most follicles cease growing and undergo atresia or regression instead of ovulating. To investigate the effects of stimulatory (follicle-stimulating hormone; FSH) and inhibitory (tumour necrosis factor alpha; TNFα) factors on the granulosa cell transcriptome, bovine ovaries were obtained fr ...[more]

PMID: 24467805

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Project description:The forkhead box transcription factor FOXO1 is highly expressed in granulosa cells of growing follicles but is down-regulated by FSH in culture or by LH-induced luteinization in vivo. To analyze the function of FOXO1, we infected rat and mouse granulosa cells with adenoviral vectors expressing two FOXO1 mutants: a gain-of-function mutant FOXOA3 that has two serine residues and one threonine residue mutated to alanines rendering this protein constitutively active and nuclear, and a FOXOA3-mutant DNA-binding domain (mDBD) in which the DBD is mutated. The infected cells were then treated with vehicle or FSH for specific time intervals. Infection of the granulosa cells was highly efficient, caused only minimal apoptosis, and maintained FOXO1 protein at levels of the endogenous protein observed in cells before exposure to FSH. RNA was prepared from control and adenoviral infected cells exposed to vehicle or FSH for 12 and 24 h. Affymetrix microarray and database analyses identified, and real time RT-PCR verified, that genes within the lipid, sterol, and steroidogenic biosynthetic pathways (Hmgcs1, Hmgcr, Mvk, Sqle, Lss, Cyp51, Tm7sf2, Dhcr24 and Star, Cyp11a1, and Cyp19), including two key transcriptional regulators Srebf1 and Srebf2 of cholesterol biosynthesis and steroidogenesis (Nr5a1, Nr5a2), were major targets induced by FSH and suppressed by FOXOA3 and FOXOA3-mDBD in the cultured granulosa cells. By contrast, FOXOA3 and FOXOA3- mDBD induced expression of Cyp27a1 mRNA that encodes an enzyme involved in cholesterol catabolism to oxysterols. The genes up-regulated by FSH in cultured granulosa cells were also induced in granulosa cells of preovulatory follicles and corpora lutea collected from immature mice primed with FSH (equine choriogonadotropin) and LH (human choriogonadotropin), respectively. Conversely, Foxo1 and Cyp27a1 mRNAs were reduced by these same treatments. Collectively, these data provide novel evidence that FOXO1 may play a key role in granulosa cells to modulate lipid and sterol biosynthesis, thereby preventing elevated steroidogenesis during early stages of follicle development. Experiment Overall Design: Immature rat granulosa cells were affected with either control (GFP) or FOXO1 mutant adenovirus (FOXOA3 and FOXOA3-mDBD) for 4 hours. Then the cells were treated with either FSH or vehicle for 24 hours. The gene expression profiles for the four treatment samples (control, control +FSH, FOXOA3 + FSH, FOXOA3-mDBD + FSH) were compared using Rat Genome 230.2 Arrays with one array per sample.

Dataset Information

Gene expression in cultured bovine ovarian granulosa

Publications

The global effect of follicle-stimulating hormone and tumour necrosis factor α on gene expression in cultured bovine ovarian granulosa cells.

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