LUTEAL AND ENDOMETRIAL LH RECEPTOR FUNCTION MODULATED BY PGE1, PGE2, LH, PGF2A, AND ESTROGEN

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: COMPLETE
• Funding Source: SPECIAL GRANT
• Reporting Frequency: Annual
• Accession No.: 0207623
• Grant No.: 2006-34135-17281
• Proposal No.: 2006-04752
• Project Start Date: Aug 15, 2006
• Project End Date: Aug 14, 2009
• Grant Year: 2006
• Program Code: [AH]- (N/A)

Recipient Organization
UNIV OF HAWAII
3190 MAILE WAY
HONOLULU,HI 96822

Performing Department
HUMAN NUTRITION, FOOD & ANIMAL SCIENCES

Non Technical Summary
Thirty to forty percent of pregnancies are lost during the first third of gestation resulting in multi-millions of dollars in losses to livestock producers. Data collected from these experiments may provide data for decision making and development of protocols to reduce or eliminate pregnancy loss during the first third of gestation.

Animal Health Component

(N/A)

Research Effort Categories

Basic

100%

Applied

(N/A)

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
301	3610	1020	100%

Knowledge Area
301 - Reproductive Performance of Animals;

Subject Of Investigation
3610 - Sheep, live animal;

Field Of Science
1020 - Physiology;

Keywords

ewe

corpus luteum

caruncle

progesterone

pge

pgf2a

estrogen

lh

ep receptors

Goals / Objectives
1) Determine the effects of vehicle, PGE1 or PGE2 infused every 4 hours intrauterine adjacent to the CL-containing ovary during the ovine estrous cycle on unoccupied and occupied caruncular endometrial and luteal membrane LH receptors, luteal and caruncular endometrial m-RNA for LH receptors, luteal weights, and uterine secretion of PGF2a, PGE, and jugular progesterone. 2) Determine the effects of Vehicle or specific agonists for PGE receptor subtypes EP1, EP2, EP3, or EP4 given into the ovarian vascular pedicle of the CL-containing ovary during the estrous cycle on CL weight, jugular venous progesterone, and luteal unoccupied and unoccupied receptors for LH, and luteal m-RNA for LH receptors. 3) Determine the effects of Vehicle, estradiol-17B, LH, or LH+ estradiol-17B infused every 6 hr intrauterine adjacent to the CL-containing ovary during the estrous cycle on concentrations of progesterone in jugular venous blood, CL weight, uterine secretion of PGF2 and PGE2, unoccupied and occupied receptors for LH in luteal and caruncular endometrial tissue, m-RNA for LH receptors in luteal and caruncular endometrium, and endocannabinoid secretion. In addition, caruncular endometrial tissue of each ewe in each treatment group will be incubated in vitro with Vehicle, LH, estradiol-17B, and LH+ estradiol-17B to assess caruncular endometrial secretion of PGE and PGF2a and endocannabinoids. 4) Determine whether caruncular endometrium in vitro treated with Vehicle, dibutryl-cAMP, LH, interferon-tau, or specific agonists for each of the PGE receptor subtypes (EP1, EP2, EP3, EP4), or combinations of these treatments affect secretion of PGE or PGF2a, caruncular endometrial m-RNA for LH receptors, and unoccupied and occupied receptors for LH, and endocannabinoid secretion.

Project Methods
1) Vehicle, PGE1, or PGE2 will be given intrauterine every 4 hr from days 10-16 postestrus of the ovine estrous cycle adjacent to the CL-containing ovary. Jugular venous and inferior vena cava plasma will be collected for analysis for progesterone, PGF2a and PGE, respectively, by RIA from days 10-16. On day 16, endometrial and luteal tissue will be collected for analysis for unoccupied and occupied caruncular and luteal membrane LH receptors, luteal weights, luteal and caruncular endometrial m-RNA for LH receptors. Additional caruncles from day 10 and each ewe from each in vivo treatment group on day 16 will be incubated for 4 and 8 hours with vehicle, PGE1, PGE2, LH, LH+ PGE1, or LH+PGE2 and tissue or media analyzed for unbound and bound LH receptors, m-RNA for LH receptors, or secretion of PGE, PGF2a, and endocannabinoids. 2) Vehicle or 333 micrograms of specific agonists for PGE receptor subtypes EP1, EP2, EP3, and EP4 will be given adjacent to the ovine CL-containing ovary on day 9 postestrus. Jugular venous plasma w CL-containing ovary will be collected every 6 hr from 0 hr through 48 hr for analysis for progesterone by RIA. CL will be collected at 48 hr, split in half, weighed, frozen and stored in liquid nitrogen until analysis of the two CL halves for m-RNA for LH receptors or unoccupied and occupied receptors for LH in luteal tissue. 3) Vehicle, estradiol-17B (10 pg/ml), ovine LH (10 ng/ml), or estradiol-17B+LH will be infused intrauterine to ewes adjacent to the luteal-containing ovary every 6 hours from days 6-18 postestrus. Jugular venous plasma will be collected daily for analysis for progesterone by RIA. Inferior vena collected daily and analyzed for PGE and PGF2a by RIA. CL will be collected at 1800 hr on day 18, split in half, weighed, and frozen in liquid nitrogen for analysis for m-RNA in one half, and occupied and unoccupied receptors for LH. Six caruncles in the infused uterine horn adjacent to the CL-containing ovary will be collected at 1800 hr on day 18 postestrus, weighed, frozen and stored in liquid nitrogen for analysis for unbound and bound LH receptors and another six caruncles for quantification of m-RNA for LH receptors. 4) On days 11 and 16 of the ovine estrous cycle, caruncular endometrium will be collected for incubation in vitro with Vehicle, LH, dibutryl-cAMP, interferon-tau, and specific agonists of the PGE receptor subtypes (EP1, EP2, EP3, and EP4) or in the presence of LH, dibutryl cAMP, PGF2a, and interferon-tau. Caruncular endometrial m-RNA receptors for LH, unoccupied and occupied receptors for LH, and PGE, PGF2a, and endocannabinoids in media will be analyzed.

Progress 08/15/06 to 08/14/09

Outputs
OUTPUTS: PGF2a is the luteolysis. . However, PGF2a is not decreased during early pregnancy, but ewes become resistant to PGF2a. PGE1 and PGE2 increased two fold in endometrium of day 13 pregnant ewes. PGE1 or PGE2 prevented luteolysis only when infused chronically into the uterine horn lumen adjacent to the luteal (CL)-containing ovary. Experiment 1. The objective of this experiment was to elucidate the effects of chronic intrauterine infusion of PGE1 or PGE2 on mRNA for LH receptors and unbound and bound LH receptors on CL and endometrial tissue and CL function in ewes. Vehicle or 500 μg of PGE1 or PGE2 were infused into the uterine lumen every 4 hr from day 10 through 1800 hr on day 16 postestrus. The CL and caruncular endometrium were collected at 160 hr and analyzed for mRNA for LH receptors and unbound and bound LH receptors. Samples were collected from day-10 ewes for the same analyses. Progesterone profiles differed (P<0.05) among treatment groups: PGE1>PGE2>Vehicle-treated ewes. PGE1 increased (P<0.05) the mRNA for LH receptors and unbound and bound LH receptors on CL and endometrial tissue when compared to day-10 control ewes or day-16 Vehicle or PGE2 treatment groups. PGE1 or PGE2 did not inhibit (P>0.05) endometrial PGF2a secretion. PGE1 is antiluteolytic by maintaining CL mRNA for LH receptors and unbound and bound LH receptors, while PGE2 is antiluteolytic by preventing the loss of mRNA and unbound and bound LH receptors during luteolysis. PGE1 may upregulate the mRNA for LH receptors on CL and the endometrium for implantation. Experiment 2. Four PGE receptor subtypes (EP1, EP2, EP3, and EP4) and one PGF2a (FP) receptor have been identified. The objective of this experiment was to elucidate the effects of EP1, EP2, EP3, and FP receptor agonists on CL function. On day-9, ewes received a single treatment into the vascular interstitium adjacent to the CL-containing ovary of Vehicle, PGF2a (100 μg) an FP receptor agonist, or 500 μg of the EP receptor agonists 17-Phenyl-Tri-Nor-PGE2 (EP1 and EP3). Butaprost (EP2), 19-(R)-OH-PGE2 (EP2), or Sulprostone (EP3). Jugular plasma was collected every 6 hours up to 48 hours for analysis for P4 and CL collected at 48 hr for mRNA for LH receptors and unbound and bound LH receptors. PGF2a or Sulprostone reduced (P<0.05) CL weight, circulating P4, CL mRNA for LH receptors, and unbound and bound LH receptors. 17-Phenyl-Tri-Nor-PGE2 did not affect (P>0.05) any parameter analyzed. Butaprost and 19-(R)-OH-PGE2 increased (P<0.05) circulating P4, CL mRNA for LH receptors, and CL unbound and bound LH receptors. Luteal mRNA for LH receptors and unbound and bound receptors for LH may be increased via the EP2 receptor, while the EP3 receptor may decrease CL mRNA for LH receptors and unbound and bound receptors for LH similar to PGF2a to contribute to luteolysis. PARTICIPANTS: Collaborators Q. X. Li for analyses; Y. Weems for surgeries and tissue analyses; D. Johnson-Graduate student training and professional development; Undergraduate student training and professional development-Tracie Uchima, E. Lennon, A. Raney, Allison Ong, Georgia Bowers, Tara Harbert, Lehua Pahia, and Kylee Goto for animal handling, treatment administration, and surgery. Partnerships Torrance Nett, Tracy Davis, and Ron D. Randel for experimental design and mRNA for LH receptor analyses TARGET AUDIENCES: Scientists working in the area PROJECT MODIFICATIONS: Not relevant to this project.

Impacts
Data from the two experiments support previous data that PGE1 or PGE2 prevent luteolysis in ewes when infused chronically in ewes. Although both PGE1 and PGE2 are doubled in the endometrium during early pregnancy, they share some mechanisms to prevent luteolysis. PGE2 prevents decreases in luteal mRNA for LH receptors, occupied and unoccupied LH receptors, and circulating progesterone, while PGE1 increases luteal and endometrial mRNA for LH receptors and occupied and unoccupied receptors for LH and increases circulating progesterone to prevent luteolysis. This indicates that PGE1 is also involved in implantation in the uterus as well as preventing luteolysis. In addition, these antiluteolytic actions of PGE1 or PGE2 appear tp be mediated via EP2 receptors, while luteolysis is not only mediated via FP receptors, but also through EP3 receptors, since either an FP or an EP3 receptor agonist decreased similarly circulating progesterone and mRHA and occupied and unoccupied receptors for LH.

Publications

• Weems, C., Y. Weems, T. Nett, T. Davis, T. Uchima, D. Johnson, R. Randel. 2008. Effects of Eand F prostaglandin receptor agonists on luteal function in vivo in ewes. Intl. Cong. Anim. Reprod. ABSTR. # P379
• MANUSCRIPTS IN PRESS -Y. S. Weems, T. M. Nett , T. L. Davis, D. L. Johnson, T. Uchima; A. Raney; E. Lennon, J. Pang, T. Harbert, G. Bowers, K. Goto, A. Ong, N. Tsutahara, R. D. Randel, and C. W. Weems. 2010. Prostaglandin E1 (PGE1), But Not Prostaglandin E2 (PGE2), Alters Luteal and Endometrial Luteinizing Hormone (LH) Occupied and Unoccupied Receptors and mRNA LH Receptor:β Actin Ratio in Ovine Luteal Tissue to Prevent Luteolysis. Prostaglandins and Other Lipid Mediators. IN PRESS.
• Y. S. Weems, T. M. Nett, T. L. Davis, D. L. Johnson, T. Uchima, A. Raney, E. Lennon, T. Harbert, G. Bowers, R. D. Randel, C. W. Weems. 2010. Effects of Prostaglandin E and F Receptor Agonists In Vivo on Luteal Function in Ewes. Prostaglandins and Other Lipid Mediators. IN PRESS.
• ABSTRACTS-Weems, Charles, Yoshie Weems, Torrance Nett, Tracy Davis, Tracie Uchima, Aaron Raney, Esther Lennon, Drew Johnson, Allison Ong, Georgia Bowers, Tara Harbert, Lehua Pahia, Kylee Goto, and Ron Randel. 2008. Effects of chronic intrauterine infusion of prostaglandins E1 or E2 (PGE1; PGE2) on mRNA:actin ratio for LH receptors and unbound and bound LH receptors on corpus luteum (CL) and endometrial plasma membranes, and luteal function in ewes. Society for the Study of Reproduction (SSR) ABSTR. # 112

Progress 08/15/06 to 08/14/07

Outputs
Ewes received either Vehicle, PGE1, or PGE2 chronically intrauterine adjacent the luteal-containing ovary from days 10-16 of the estrous cycle. Collection of all samples for experiment one has been completed. Profiles of progesterone differed in all three treatment groups with progesterone decreasing in the vehicle, not decreasing in the PGE2, and increasing in the PGE1 treatment groups over the sampling period. On day-16, mRNA for LH receptors on luteal tissue was greater in the PGE1-treated ewes than in vehicle-treated ewes on day-16, samples from day-10 control ewes (peak luteal function), or PGE2-treated ewes on day-16. mRNA for LH receptors on luteal tissue was greater on day-16 in the PGE2-treated ewes than on day-16 in vehicle-treated ewes, but was similar to mRNA for LH receptors on luteal tissue of day-10 control ewes. In addition, only PGE1 increased mRNA for LH receptors on caruncular endometrium. On day-16, unbound and bound LH receptors on luteal tissue were greater in the PGE1-treated ewes than in vehicle-treated ewes on day-16, samples from day-10 control ewes (peak luteal function), or PGE2-treated ewes on day-16. Unbound and bound LH receptors on luteal tissue was greater on day-16 in the PGE2-treated ewes than on day-16 in vehicle-treated ewes, but was similar to LH receptors on luteal tissue of day-10 control ewes. In addition, only PGE1 increased unbound and bound LH receptors on caruncular endometrium. In summary, PGE1 or PGE2 prevents luteolysis, but only PGE1, not PGE2, increases mRNA on luteal or caruncular endometrial as antiluteolytic signals to prevent luteolysis. tissue

Impacts
We believe these data could lead to development of stable analogues as treatments to reduce early pregnancy loss during the first trimester of pregnancy.

Publications

• No publications reported this period