|Cushman, Robert - Bob|
Submitted to: Society for the Study of Reproduction Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 3/28/2013
Publication Date: 7/1/2013
Citation: McFee, R.M., Kurz, S.G., Summers, A.F., Pohlmeier, W.E., Cushman, R.A., Wood, J.R., Cupp, A.S. 2013. Granulosa cells from bovine follicles activate different signal transduction pathways dependent on follicle health status and ability to convert androstenedione to estrogen [abstract]. Biology of Reproduction Supplement (46th Annual Meeting of the Society for the Study of Reproduction). p. 265 (Abstract #577). Interpretive Summary:
Technical Abstract: Since steroidogenesis is a critical component in the development of competent preovulatory follicles we hypothesized that granulosa cells from follicles of cows treated with normal levels of progesterone (CIDR) or with melengestrol acetate (MGA), which results in the development of persistent follicles, exhibit differences in the activation of signal transduction pathways that impact steroidogenesis. The CIDR cows underwent a modified Co-Synch protocol with GnRH on day 0, a CIDR in place from days 0 to 7, and prostaglandin F2alpha (PGF2alpha) on day 7. The MGA cows received MGA in the diet for 14 days and PGF2alpha on days 0, 7, and 14. Follicles were aspirated 36-60 hours after the final PGF2alpha injection. Follicular fluid was assayed to determine concentrations of estrogen (E2), progesterone (P4), and androstenedione (A4). Follicles with a follicular fluid E2:P4 ratio greater than 1 were classified as E2-active (EA) while those with an E2:P4 ratio less than 1 were classified as E2-inactive (EI). Granulosa cells were processed for Western blot analysis of MAPK1/3 (mitogen-activated protein kinase 1 and 3; also known as ERK2 and 1) of the Ras-MEK-ERK-signaling cascade, AKT1 (v-akt murine thymoma viral oncogene homolog 1) of the PI3K/AKT1 signaling pathway, and FOXO1 (forkhead box O1), a downstream substrate of AKT1. The abundance of total (P=0.0164) and phosphorylated (P=0.0483) MAPK1/3 was greater in granulosa cells from EA follicles (n=20) compared to EI follicles (n=10) from CIDR cows but no differences were detected between CIDR and MGA follicles (n=14). In addition, the abundance of total and phosphorylated AKT1 and FOXO1 did not differ between EA or EI follicles or treatment group. Within our herd, we have identified subpopulations of cows that have either high (greater than 40 ng/ml) or low (less than 40 ng/ml) A4 follicular fluid concentrations as well as differing abilities to convert available A4 to E2 (low = E2:A4 ratio less than 100 with high A4 levels; high = E2:A4 ratio greater than 100 with low A4 levels). These classifications are maintained over repeated estrous cycles and years. In addition, we noted that cows treated with MGA also have high follicular fluid A4 concentrations with reduced ability to convert A4 to E2 similar to Low E2:A4 cows. Consequently, we further classified the EA follicles from the CIDR treated cows into High E2:A4 (n=10) and Low E2:A4 (n=10) groups. Thus, our second hypothesis was that the reduced ability to convert A4 to E2 in mural granulosa cells is the result of altered signaling downstream of gonadotropin receptors. Follicular fluid concentrations of E2 were not different between CIDR High and Low E2:A4 follicles; however, A4 levels were 24.6-fold greater (P=0.0015) in Low E2:A4 follicles. Furthermore, the abundance of both phosphorylated AKT1 (P=0.05) and phosphorylated FOXO1 (P=0.0473) was greater in granulosa cells from Low E2:A4 follicles compared to High E2:A4 follicles. These results demonstrate that activation of MAPK1/3 and AKT1 associated signaling pathways differs in EA versus EI and Low E2:A4 versus High E2:A4 follicles, respectively. Therefore, these differences in signal transduction activity may contribute to the altered steroid hormone profiles identified in the dominant bovine follicles. USDA is an equal opportunity provider and employer.