Regulation of Angiogenesis-Related Prostaglandin F2alpha-Induced Genes in the Bovine Corpus Luteum1

In order to characterize the transition of the corpora lutea (CL) from acquisition of luteolytic sensitivity to rescue of luteal function: i) the expression of 38 factors associated with steroids, prostanoids, and angiogenic systems and ii) concentrations of the main hormones responsible for maintenance of CL function in cyclic and pregnant pigs were examined. Additionally, the effect of prostaglandin (PG) E 2 and F 2a on luteal function during the estrous cycle and pregnancy was evaluated in vitro. Significantly up-regulated gene expression was revealed in CL collected on day 14 of the estrous cycle (CYP19A1, ESR2, PTGS2, HIF1A, and EDN1) and on days 12-14 of pregnancy (SCARB1, PGRMC1, STAR, HSD3B1, NR5A1, PTGFR, PTGER4, and VEGFA). Elevated concentrations of estradiol-17b and PGE 2 occurred in CL on days 12 and 14 of pregnancy respectively, while an increased intraluteal PGF 2a content was noted on day 14 of the estrous cycle. Both PGs increased the synthesis of progesterone by cultured luteal slices obtained on day 14 of pregnancy, in contrast to the action of PGF 2a on the corresponding day of the estrous cycle. PGE 2 stimulated cAMP production via PTGER2 and PTGER4, while PGF 2a elevated the content of CREB in cultured luteal slices from CL of pregnant pigs. In silico analysis showed that infiltration of lymphocytes and apoptosis of microvascular endothelium were activated in CL on day 12 of the estrous cycle vs pregnancy. Summarizing, an abundance of E 2 and PGE 2 during pregnancy regulates specific pathways responsible for steroidogenesis, the prostanoid signaling system and angiogenesis during rescue from luteolysis in porcine CL.

Section: Rescue Of Luteal Function During Early Pregnancymentioning

confidence: 52%

Steroid hormones, prostanoids, and angiogenic systems during rescue of the corpus luteum in pigs

Przygrodzka¹,

Kaczmarek²,

Kaczyński³

et al. 2016

“…Real-time PCRs were performed using the Mx3000P quantitative PCR system (Stratagene, Garden Grove, CA, USA), using Platinum SYBR Green qPCR SuperMix-UDG (Invitrogen), as described previously (Kisliouk et al 2005, Zalman et al 2012. ACTB was used as the reference gene.…”

Section: Rna Isolation and Real-time Pcrmentioning

confidence: 99%

HIF1A-dependent increase in endothelin 2 levels in granulosa cells: role of hypoxia, LH/cAMP, and reactive oxygen species

Yalu¹,

Oyesiji²,

Eisenberg³

et al. 2015

Self Cite

Hypoxia-inducible factor 1 alpha (HIF1A) and endothelin 2 (EDN2) are transiently expressed during the same time window in the developing corpus luteum (CL). In this study, we sought to investigate the involvement of LH/cAMP, reactive oxygen species (ROS), and a hypoxia-mimetic compound (CoCl 2 ) on HIF1A expression and how it affected EDN2 levels, using transformed human granulosa cells (thGCs) and primary bovine granulosa cells (GCs). CoCl 2 elevated HIF1A protein levels in thGCs in a dose-dependent manner. Forskolin alone had no significant effect; however, forskolin and CoCl 2 together further induced HIF1A protein and EDN2 mRNA expression in thGCs. Similarly, in primary GCs, LH with CoCl 2 synergistically augmented HIF1A protein levels, which resulted in higher expression of EDN2 and another well-known hypoxia-inducible gene, VEGF (VEGFA). Importantly, LH alone elevated HIF1A mRNA but not its protein. The successful knockdown of HIF1A in thGCs using siRNA abolished hypoxia-induced EDN2 and also the additive effect of forskolin and CoCl 2 . We then examined the roles of ROS in thGCs: hydrogen peroxide (20 and 50 mM) elevated HIF1A protein as well as the expression of EDN2, implying that induction of HIF1A protein levels is sufficient to stimulate the expression of EDN2 (and VEGF) in normoxia. A broad-range ROS scavenger, butylated hydroxyanisole, inhibited CoCl 2 -induced HIF1A protein with a concomitant reduction in the mRNA expression of EDN2 and VEGF in thGCs. The results obtained in this study suggest that HIF1A, induced by various stimuli, is an essential mediator of EDN2 mRNA expression. The results may also explain the rise in the levels of HIF1A-dependent genes (EDN2 and VEGF) in the developing CL.

“…Thrombospondin mainly acts as anti-angiogenic factor; however, it also promotes atresia of rat granulosa cells in vitro (Garside et al 2010a(Garside et al , 2010b. THBS1 is highly expressed in small follicles from bovine ovaries (Greenaway et al 2005) as well as in small and atretic follicles of many other species (Thomas et al 2008, Garside et al 2010b, Zalman et al 2012. Igf1 inhibits Thbs1 transcription in cultured rat granulosa cells and its expression is further lowered when FSH is added in culture (Dreyfus et al 1992, McGray et al 2011.…”

Section: The Effect Of Superstimulation On Folliclesmentioning

confidence: 99%

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

Dias¹,

Khan²,

Sirard³

et al. 2013

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.