Differential gene expression of granulosa cells after ovarian superstimulation in beef cattle

Abstract

Microarray analysis was used to compare the gene expression of granulosa cells from dominant follicles with that of those after superstimulatory treatment. Cows were allocated randomly to two groups (superstimulation and control, nZ6/group). A new follicular wave was induced by ablation of follicles R5 mm in diameter, and a progesterone-releasing device controlled internal drug release (CIDR) was placed in the vagina. The superstimulation group was given eight doses of 25 mg FSH at 12-h intervals starting from the day of wave emergence (day 0), whereas the control group was not given FSH treatment. Both groups were given prostaglandin F 2a twice, 12 h apart, on day 3 and the CIDR was removed at the second injection; 25 mg porcine luteinizing hormone (pLH) was given 24 h after CIDR removal, and cows were ovariectomized 24 h later. Granulosa cells were collected for RNA extraction, amplification, and microarray hybridization. A total of 190 genes were downregulated and 280 genes were upregulated. To validate the microarray results, five genes were selected for real-time PCR (NTS, FOS, THBS1, FN1, and IGF2). Expression of four genes increased significantly in the three different animals tested (NTS, FOS, THBS1, and FN1). The upregulated genes are related to matrix remodeling (i.e. tissue proliferation), disturbance of angiogenesis, apoptosis, and oxidative stress response. We conclude that superstimulation treatment i) results in granulosa cells that lag behind in maturation and differentiation (most of the upregulated genes are markers of the follicular growth stage), ii) activates genes involved with the NFE2L2 oxidative stress response and endoplasmic reticulum stress response, and iii) disturbs angiogenesis.

Granulosa cell function and oocyte competence: Super-follicles, super-moms and super-stimulation in cattle

“…To determine if and to what extend superstimulation affect follicles and oocytes, microarray technology was used to compare granulosa cell gene expression in superstimulated follicles versus dominant follicles from a natural cycle (Dias et al, 2013c). The superstimulation protocol used in this study included exogenous administration of FSH over 4 days after follicular wave synchronization and LH 24 h after progesterone-device removal.…”

Lengthened superstimulatory treatment in cattle: Evidence for rescue of follicles within a wave rather than continuous recruitment of new follicles

“…By providing continuous exogenous FSH support, regression of subordinate follicles can be prevented so that they assume qualities of a dominant follicle. Recently, a molecular study of granulosa cells indicated that gene expression in follicles undergoing superstimulation was altered compared to a single, naturally occurring dominant follicle [27]. Twenty-four hours after LH administration, genes related to growth and oxidative stress response were upregulated in granulosa cells of follicles undergoing a 4-day FSH superstimulation protocol as compared to the single dominant follicle of an unstimulated follicular wave.…”

“…Recently, the evaluation of the expression of LHR messenger RNA in granulosa cells of superstimulated and single, naturally occurring dominant follicles 12 hours after progesterone device removal revealed that expression of LHR was decreased in superstimulated follicles [29]. As a result, follicles following the conventional 4-day FSH superstimulation protocol appeared to be less mature and less responsive to LH when compared to a naturally occurring, single dominant follicle [27]. In this regard, studies evaluating the expression of these genes in granulosa cells of follicles undergoing extended FSH support (analogous to present study) may provide valuable insight into the mechanisms underlying the observed improvement in superovulatory response [19,26,27].…”

Expression and regulative function of tissue inhibitor of metalloproteinase 3 in the goat ovary and its role in cultured granulosa cells

“…Adam et al (2012) furthermore indicated that the EGFR is one of the growth factor receptors directly targeted by DCN in human GCs and DCN is a local regulator of folliculogenesis, ovulation and the functional lifespan of the CL in primates. Recent studies demonstrated that the expression of FN was elevated in the superstimulation group and its expression was reported to be inversely related to follicle maturation (Berkholtz et al, 2006;Dias et al, 2013;Yasuda et al, 2005). TIMP3 was first discovered as an ECM bound protein and participated in changing the ECM (Li and Curry, 2009).…”