Source: UNIV OF WISCONSIN submitted to
PIVOTAL PERIODS OF PREGNANCY LOSS AND MAINTENANCE AFTER EMBRYO TRANSFER
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1019374
Grant No.
2019-67015-29704
Cumulative Award Amt.
$500,000.00
Proposal No.
2018-06481
Multistate No.
(N/A)
Project Start Date
Jun 1, 2019
Project End Date
May 31, 2023
Grant Year
2019
Program Code
[A1211]- Animal Health and Production and Animal Products: Animal Reproduction
Recipient Organization
UNIV OF WISCONSIN
21 N PARK ST STE 6401
MADISON,WI 53715-1218
Performing Department
DAIRY SCIENCE-GEN
Non Technical Summary
Pregnancy loss represents a substantial economic cost for livestock producers. Transfer of in vitro produced (IVP) embryos has increased dramatically in the US and is critical for genetic progress in both dairy and beef cattle; however, recipients of IVP embryos experience high rates (10-20%) of pregnancy loss. Transfer of cloned embryos is an interesting model for pregnancy loss in which pregnancies per embryo transferat 32 days is similar to IVP embryos (~50%), however, half of these pregnancies are lost during the second month of pregnancy. To reduce pregnancy loss, more basic knowledge is needed on the mechanisms that maintain the pregnancy and on the mechanisms that lead to loss of pregnancy. This research will investigate two important potential mechanisms that are involved in this process.The first mechanism is the increase in uterine blood flow that occurs during pregnancy. We will mainpulate uterine blood flow by either increasing or decreasing uterine blood and evaluate the effect on maintenance of pregnancy or pregnancy loss.The second key mechanism is the hormone, progesterone. This hormone circulating in the blood and is responsible for maintaining the pregnancy in all mammals, including cattle. Our experiments will increase circulating progesterone concentrations to see if we can improve fertility in cows after embryo transfer by reducing pregnancy loss.These experiments are important for determining how we can reduce pregnancy loss in cattle and thus improve the efficiency of reproduction in the US cattle industry.
Animal Health Component
25%
Research Effort Categories
Basic
75%
Applied
25%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
30134101020100%
Knowledge Area
301 - Reproductive Performance of Animals;

Subject Of Investigation
3410 - Dairy cattle, live animal;

Field Of Science
1020 - Physiology;
Goals / Objectives
The long-term goals of this proposal are two-fold: 1) We seek to understand the factors/signals that underlie maintenance of the corpus luteum (CL) during the second month of pregnancy and, thereby, elucidate some of the potential causes of pregnancy loss; and 2) We propose to develop and test physiologically rational strategies to reduce pregnancy loss after embryo transfer.Objective 1 investigates whether increasing blood flow to the uterus blocks the process of regression of the corpus luteum. During pregnancy, regression of the corpus luteum is blocked and we will test if increasing uterine blood flow will produce a similar effect. This will be evidence that the corpus luteum is maintained during pregnancy by an increase in uterine blood flow. Experiment 1 will use specific treatments to increase uterine blood flow to see if this blocks the normal process of regression of the corpus luteum. Experiment 2 will block the increase in uterine blood flow that accompanies early pregnancy to see if this blocks the effect of pregnancy on maintenance of the corpus luteum.Thus, two different types of evidence will evaluate the role of uterine blood flow in maintenance of the corpus luteum during pregnancy.Objective 2 evaluates two methods to reduce pregnancy loss of cloned embryos. Cloned embryos have extremely high loss of pregnancies between 30 and 60 days of pregnancy. Experiment 3 will evaluate if increasing uterine blood flow will reduce these pregnancy losses. This will be done by giving a specific stimulatory of uterine blood flow, Sildenafil. Experiment 4 will evaluate whether increasing circulating progesterone concentrations will reduce pregnancy losses in recipients of cloned embryos or in vitro produced embryos. Progesterone is the main hormone that maintains pregnancy in cattle and other mammals and this experiment evaluates whether increased progesterone will improve maintenance of pregnancy in cattle after embryo transfer.Objective 3 will evaluate the causes of pregnancy loss. From a broad perspective, loss of pregnancy occurs either because the embryo dies or the corpus luteum dies causing subsequent loss of the embryo. In a large experiment using 4,000 animals, we will determine if the primary cause of pregnancy loss is due to death of the embryo or due to death of the corpus luteum which is followed by loss of the embryo. This study will also evaluate a more practical way of increasing circulating progesterone concentrations in an attempt to increase maintenance of pregnancy.
Project Methods
The experimental approach will primarily evaluate heifers that have received in vitro produced embryos or cloned embryos. These embryo recipients will receive various strategies to either increase or decrease uterine blood flow and increase or decrease circulating progesterone concentrations. Evaluations of changes in uterine blood flow and pregnancy will be evaluated using ultrasound. Circulating hormones will also be evaluated to determine effects of treatments on hormone concentrations. Experiment 1- Determine whether increasing uterine blood flow using different vasodilators will maintain the CL during administration of intrauterine PGF2α pulses.Cycling non-lactating cows (N= 40) will be randomized on Day 8 of the estrous cycle into 5 treatments (four replicates;10 cows/replicate). Serial blood samples will be collected via jugular catheters, and intrauterine treatments will be performed by passing an AI rod with treatment contained in an empty AI straw, as we have described elsewhere.Effects of treatment will be analyzed with SAS using Proc MIXEDfor continuous variables over time (P4, blood flow, etc.), Proc ANOVA for continuous variable (gene expression) and Proc GLIMMIX for binomial variables (occurrence of luteolysis).Experiment 2-Determine whether inhibition of pregnancy-induced uterine vasodilation will alter maintenance of the CL during the second month of pregnancy.All experiments will be done on Day 39 of pregnancy in non-lactating cows that have received IVP embryos. Preliminary experiments will be performed to determine the appropriate dose of L-NAME required to decrease circulating NO metabolites and uterine blood flow in pregnant cows (doses of 12.5, 25, and 50 mg/kg; based on research in ewes). The timing of decrease in uterine blood flow relative to treatment and any return to normal flow will be used to determine the timing of treatments in the main experiment (every 6 h, 12 h). Heifers on day 39 of pregnancy (n= 32) will be randomized into 4treatmentsusing a 2 x2 factorial design. Treatments will be L-NAME at an appropriate dose and timing such as 25 mg/kg every 12 hours or saline. After 24 hours of L-NAME treatment, oxytocin pulses (50 IU/pulse) will be administered i.m. every 12 h for 2 d (4 total oxytocin pulses). Hourly blood samples will be collected via jugular catheters from 4 h before L-NAME treatment until 24 h after the last oxytocin treatment. Ultrasound/Doppler evaluations will be done every 12 h for CL volume, CL blood flow and uterine blood flow. Luteal biopsies will be collected before first oxytocin treatment (24 h of L-NAME treatment) and 1 h after the third oxytocin treatment and analyzed by RT-PCR for genes evaluated in Expt. 1 to determine whether PGF2αis reaching or acting on the CL.Effects of treatment will be analyzed with SAS using Proc MIXEDfor continuous variables over time (P4, blood flow, etc.), Proc ANOVA for continuous variable (gene expression) and Proc GLIMMIX for binomial variables (occurrence of luteolysis).Experiment 3- Determine whether uterine vasodilation by treatment with Sildenafil can block pregnancy loss in recipients of cloned embryos.The estrous cycles of heifers will be synchronized to receive transfer of a cloned embryo on Day 7 of the cycle, and pregnancy status will be determined on Day 29of pregnancy using ultrasound. A total of 50 heifers with a confirmed pregnancy from a cloned embryo will be randomly assigned to a 2 x 2 factorial arrangements of treatments to assess the main effects of embryo type (clones vs. IVP) and treatment.The effect of treatment will be analyzed with SAS using Proc MIXED for continuous variables over time (P4, PAGs, PGFM, blood flow, embryo size), Proc ANOVA for comparison between treatment groups (IVP vs clones), and Proc GLIMMIX for binomial variables (pregnancy loss, P/ET). Using 25 embryos per treatment we will be able to detect a decrease from 50% to 25% in pregnancy loss (P = 0.03 with one-tailed Fischer's exact test). Comparisons will be made of hormonal and physiological measurements between clones with or without pregnancy loss and compared to pregnancy from IVP embryos.Experiment 4- Determine the effect of increasing circulating P4 concentrations on pregnancy loss in recipients of IVP and cloned embryos.In this experiment, we propose to increase circulating P4by using2 intravaginal P4 inserts. In heifers without a CL, treatment with 2 CIDR insertsincreasesP4 to more than 3 ng/mL (unpublished results). To increase P4 release from the CIDR inserts, we will autoclave the inserts before use to further increase P4 release by ~60%. Our goal is to maintain circulating P4 at about 5ng/mL, even if the CL regresses.Experiment 5- Increasing progesterone using hCG to evaluate pregnancy loss in recipients of IVP embryos.Experiment 5will be conducted in collaboration with Sexing Technologies (ST; recipients housed Fond du Lac, WI). Nonlactating Holstein dairy heifers (n = 4,000) will be submitted to a 5-d CIDR-synch protocol (Day 0, CIDR in; Day 5, CIDR out + PGF; Day 8, GnRH) and will receive direct transfer of afresh Day 7 IVP embryo7 d after the final GnRH treatment. Heifers will be randomized to one of two treatments on Day of ET (i.e., 7 dafter GnRH treatment): 1) Saline (control, C, n = 2,000) or Treatment with 3,000 IU hCG on day of ET (hCG6; n = 2,000). On Day 33, heifers diagnosed pregnant will be randomly assigned to: Saline (Sal33) or Treatment with 3,000 IU hCG (hCG33).Procedure GLIMMIX of SAS will be used to analyze effect of treatment, parity, and the two-way interaction on P/ET and pregnancy loss using a backward selection procedure with treatment and parity (a blocking factor) retained as fixed factors in each of the models as described. Based on power calculations, inclusion of 2,000heifers per treatment allows detection of 3%differencesin P/ET (50 to 53%) and pregnancy loss (15% to 12%; assuming 1000 pregnancies/treatment; 95% confidence; 80% power; one-sided test).

Progress 06/01/19 to 05/31/23

Outputs
Target Audience:Scientists working in similar areas of reproductive physiology and reproductive management. Veterinarians, producers, and industry professionals working in beef and dairy cattle. Undergraduate and graduate students in animal science and veterinary medicine. Changes/Problems:Some of the experiments in Objective 1 were delayed and disrupted by the pandemic. The college insisted on moving our cows even though they were in the midst of an experiment. Thus, some experiments could not be completed and results were lost. What opportunities for training and professional development has the project provided?• Through laboratory meetings and individual meetings postdoctoral fellows and graduate students received training in the design, implementation, and analysis of reproductive biology research. They gave presentations in these meetings as well as in the Graduate Seminar of the Department of Dairy Science. These scientists also served as teaching assistants and instructors in part of the Animal Physiology course and Reproductive Management of Dairy Cattle course. • Graduate students attended scientific meetings in 2021 and 2022 (Society for the Study of Reproduction, International Ruminant Reproduction Symposium, American Society for Animal Science, and American Dairy Science Association). How have the results been disseminated to communities of interest?• Manuscripts were published in peer-reviewed journals • Abstracts and oral presentations at national and international scientific meetings. • Presentations were delivered by PI to stakeholder meetings (producers and veterinarians). What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Objective 1: We have completed three different projects under this specific objective using different vasodilators to increase uterine blood flow. It does not appear that any of these vasodilators delayed luteolysis but we are still analyzing the results. These experiments were done in a intermittent fashion during 2020, due to movement of cattle out of the on-campus dairy cattle center during pandemic restrictions. Two scientific manuscripts have now been published related to this research (Domingues et al., 2023; domingues et al., 2022). Objective 2: These studies were completed. Most of the results have now been published although one publication on the cloned embryos has been prepared for publication and will be submitted this year. Objective 3: We completed this project but in lactating dairy cows (Monteiro et al., 2021a; Monteiro et al., 2021b). This research has provided a method for decreasing pregnancy loss in lactating dairy cows by inducing an accessory corpus luteum that is ipsilateral to the pregnancy. We also continued the research in this area, based on our initial results, and have now completed a study that is published (Domingues et al., 2023) and one that has been provisionally accepted for publication (Domingues et al, unpublished).

Publications

  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Wiltbank MC, Monteiro PLJ, Domingues RR, Andrade JPN, Mezera MA. Review: Maintenance of the ruminant corpus luteum during pregnancy: interferon-tau and beyond. Animal 2023; 17. ARTN 100827 10.1016/j.animal.2023.100827
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Domingues RR, Andrade JPN, Cunha TO, Madureira G, Moallem U, Gomez-Leon V, Martins JPN, Wiltbank MC. Is pregnancy loss initiated by embryonic death or luteal regression? Profiles of pregnancy-associated glycoproteins during elevated progesterone and pregnancy loss. JDS Commun 2023; 4:149-154. 10.3168/jdsc.2022-0282
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Domingues RR, Ginther OJ, Gomez-Leon V, da Silva PN, Castro T, Hoppmann A, Wiltbank MC. Processes involved in prostaglandin F2alpha autoamplification in heifers. Reproduction 2023; 165:93-101. 10.1530/Rep-22-0242
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Consentini CEC, Alves RLOR, Silva MA, Galindez JPA, Madureira G, Lima LG, Gon�alves JRS, Wiltbank MC, Sartori R. What are the factors associated with pregnancy loss after timed-artificial insemination in cattle? Theriogenology 2023; 196:264-269. 10.1016/j.theriogenology.2022.10.037
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Toledo MZ, Stangaferro ML, Oliveira RC, Monteiro PLJ, Gennari RS, Luchini D, Shaver RD, Giordano JO, Wiltbank MC. Effects of feeding rumen-protected methionine pre- and postpartum in multiparous Holstein cows: Health disorders and interactions with production and reproduction. Journal of Dairy Science 2023; 106:2137-2152. 10.3168/jds.2022-21950
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Cunha TO, Statz LR, Domingues RR, Andrade JPN, Wiltbank MC, Martins JPN. Accessory corpus luteum induced by human chorionic gonadotropin on day 7 or days 7 and 13 of the estrous cycle affected follicular and luteal dynamics and luteolysis in lactating Holstein cows. Journal of Dairy Science 2022; 105:2631-2650. 10.3168/jds.2021-20619
  • Type: Journal Articles Status: Published Year Published: 2023 Citation: Monteiro PLJ, Consentini CEC, Andrade JPN, Beard AD, Garcia-Guerra A, Sartori R, Wiltbank MC. Research on timed AI in beef cattle: Past, present and future, a 27-year perspective. Theriogenology 2023; 211:161-171. 10.1016/j.theriogenology.2023.07.037
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Fricke PM, Wiltbank MC. : The implications of spontaneous versus synchronized ovulations on the reproductive performance of lactating dairy cows. Journal of Dairy Science 2022; 105:4679-4689. 10.3168/jds.2021-21431
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Drum JN, Madureira G, Rosa CO, Seneda MM, Wiltbank MC, Sartori R, Ortega MS. Male Embryos Produced Deviate From Their Counterparts in Placental Gene Expression on Day 32 of Pregnancy. Front Anim Sci 2022; 3. ARTN 807217 10.3389/fanim.2022.807217
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Domingues RR, Wiltbank MC, Hernandez LL. Pregnancy Complications and Neonatal Mortality in a Serotonin Transporter Null Mouse Model: Insight Into the Use of Selective Serotonin Reuptake Inhibitor During Pregnancy. Front Med-Lausanne 2022; 9. ARTN 848581 10.3389/fmed.2022.848581
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Domingues RR, Ginther OJ, Gomez-Leon V, Castro T, Wiltbank MC. Endometrial and luteal responses to a prostaglandin F2alpha pulse: a comparison between heifers and mares. Biology of Reproduction 2022; 106:979-991. 10.1093/biolre/ioac025
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Domingues RR, Fricke HP, Sheftel CM, Bell AM, Sartori LC, Manuel RSJ, Krajco CJ, Wiltbank MC, Hernandez LL. Effect of Low and High Doses of Two Selective Serotonin Reuptake Inhibitors on Pregnancy Outcomes and Neonatal Mortality. Toxics 2022; 10. ARTN 11 10.3390/toxics10010011
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Domingues RR, Beard AD, Connelly MK, Wiltbank MC, Hernandez LL. Fluoxetine-induced perinatal morbidity in a sheep model. Front Med-Lausanne 2022; 9. ARTN 955560 10.3389/fmed.2022.955560
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Hughes CHK, Mezera MA, Wiltbank MC, Pate JL. Insights from two independent transcriptomic studies of the bovine corpus luteum during pregnancy. Journal of Animal Science 2022; 100. ARTN skac115 10.1093/jas/skac115
  • Type: Journal Articles Status: Published Year Published: 2022 Citation: Bishop CV, Selvaraj V, Townson DH, Pate JL, Wiltbank MC. History, insights, and future perspectives on studies into luteal function in cattle. Journal of Animal Science 2022; 100. ARTN skac143 10.1093/jas/skac143
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Mezera MA, Li WL, Wiltbank MC. Pregnancy-induced changes in the transcriptome of the bovine corpus luteum during and after embryonic interferon-tau secretion. Biology of Reproduction 2021; 105:148-163. 10.1093/biolre/ioab034
  • Type: Journal Articles Status: Published Year Published: 2021 Citation: Mezera MA, Li WL, Liu LH, Meidan R, Pe�agaricano F, Wiltbank MC. Effect of natural pre-luteolytic prostaglandin F pulses on the bovine luteal transcriptome during spontaneous luteal regression. Biology of Reproduction 2021; 105:1016-1029. 10.1093/biolre/ioab123


Progress 06/01/21 to 05/31/22

Outputs
Target Audience:The target audience includes scientists that are working in reproductive physiology and reproductive management. It also includes veterinarians, producers, and industry professionals working in beef and dairy cattle. Students are also part of the target audience including graduate students, undergraduates, and veterinary students. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?Through laboratory meetings and individual meetings postdoctoral fellows and graduate students received training in the design, implementation, and analysis of reproductive biology research. They gave presentations in these meetings as well as in the Graduate Seminar of the Department of Dairy Science. These scientists also served as teaching assistants and instructors in part of the Animal Physiology course and Reproductive Management of Dairy Cattle course. • Graduate students attended scientific meetings (Society for the Study of Reproduction, International Ruminant Reproduction Symposium, American Society for Animal Science, and American Dairy Science Association). How have the results been disseminated to communities of interest?• Manuscripts were published in peer-reviewed journals • Abstracts and oral presentations at national and international scientific meetings. • Presentations were delivered by PI to stakeholder meetings (producers and veterinarians). What do you plan to do during the next reporting period to accomplish the goals?The publications and results from this period are included in the final report.

Impacts
What was accomplished under these goals? Objective 1: We have completed three different projects under this specific objective using different vasodilators to increase uterine blood flow. It does not appear that any of these vasodilators delayed luteolysis but we are still analyzing the results. These experiments were done in a intermittent fashion during 2020, due to movement of cattle out of the on-campus dairy cattle center during pandemic restrictions. Objective 2: Experiment 3 and 4 were completed and is being prepared to publication. We have completed multiple studies that have now been published reporting these findings and our new understanding of the process of luteolysis. Objective 3: We completed this project but in lactating dairy cows (Monteiro et al., 2021a; Monteiro et al., 2021b). This research has provided a method for decreasing pregnancy loss in lactating dairy cows by inducing an accessory corpus luteum that is ipsilateral to the pregnancy.

Publications


    Progress 06/01/20 to 05/31/21

    Outputs
    Target Audience:The target audience includes scientists that are working in reproductive physiology and reproductive management. It also includes veterinarians, producers, and industry professionals working in beef and dairy cattle. Students are also part of the target audience including graduate students, undergraduates, and veterinary students. Changes/Problems:We developed more accurate methods for determination of early pregnancies and redid many of the projects using these more accurate methods. The publications from this research will be published in 2023. What opportunities for training and professional development has the project provided?• Through laboratory meetings and individual meetings postdoctoral fellows and graduate students received training in the design, implementation, and analysis of reproductive biology research. They gave presentations in these meetings as well as in the Graduate Seminar of the Department of Dairy Science. These scientists also served as teaching assistants and instructors in part of the Animal Physiology course and Reproductive Management of Dairy Cattle course. • Graduate students attended scientific meetings in 2021 and 2022 (Society for the Study of Reproduction, International Ruminant Reproduction Symposium, American Society for Animal Science, and American Dairy Science Association). How have the results been disseminated to communities of interest?• Manuscripts were published in peer-reviewed journals • Abstracts and oral presentations at national and international scientific meetings. • Presentations were delivered by PI to stakeholder meetings (producers and veterinarians). What do you plan to do during the next reporting period to accomplish the goals?We are continuing to complete and submit scientific manuscripts for publication reporting the results of this research. We have now developed methods that allow us to accurately evaluate pregnancy loss before day 22 and publications from this project have been submitted for publication.

    Impacts
    What was accomplished under these goals? Objective 1: We have completed three different projects under this specific objective using different vasodilators to increase uterine blood flow. It does not appear that any of these vasodilators delayed luteolysis but we are still analyzing the results. These experiments were done in a intermittent fashion during 2020, due to movement of cattle out of the on-campus dairy cattle center during pandemic restrictions. Objective 2: Experiment 3 and 4 were completed and is being prepared to publication. We have completed multiple studies that have now been published reporting these findings and our new understanding of the process of luteolysis. Objective 3: We completed this project but in lactating dairy cows (Monteiro et al., 2021a; Monteiro et al., 2021b). This research has provided a method for decreasing pregnancy loss in lactating dairy cows by inducing an accessory corpus luteum that is ipsilateral to the pregnancy.

    Publications

    • Type: Journal Articles Status: Published Year Published: 2021 Citation: Monteiro PLJ, Sartori R, Canavessi AMO, Melo LF, Motta JCL, Consentini CEC, Wiltbank MC. Accessory corpus luteum regression during pregnancy I: timing, physiology, and P4 profiles. Reproduction 2021; 162:473-482.
    • Type: Journal Articles Status: Published Year Published: 2021 Citation: Monteiro PLJ, Gamarra CA, Genari RS, Prata AB, Barletta RV, Duran PG, Canavessi AMO, Sartori R, Wiltbank MC. Accessory corpus luteum regression during pregnancy II: reproductive outcomes. Reproduction 2021; 162:483-495.
    • Type: Journal Articles Status: Published Year Published: 2022 Citation: Atli MO, Mehta V, Vezina CM, Wiltbank MC. Expression patterns of chemokine (C-C motif) ligand 2, prostaglandin F2A receptor and immediate early genes at mRNA level in the bovine corpus luteum after intrauterine treatment with a low dose of prostaglandin F2A. Theriogenology 2022; 189:70-76.
    • Type: Journal Articles Status: Published Year Published: 2021 Citation: Stangaferro, M.L., Toledo, M.Z., Gennari, R.S., Perez, M.M., Gamarra, C.A., Sitko, E.M., Monteiro, P.L.J., Jr., Masello, M., Prata, A.B., Granados, G.E., Van Amburgh, M.E., Luchini, D., Shaver, R.D., Wiltbank, M.C., Giordano, J.O., 2021. Effects of feeding rumen-protected methionine pre- and postpartum on reproductive outcomes of multiparous Holstein cows. J Dairy Sci 104, 11210-11225.
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Mezera MA, Li W, Edwards AJ, Koch DJ, Beard AD, Wiltbank MC. Identification of stable genes in the corpus luteum of lactating Holstein cows in pregnancy and luteolysis: Implications for selection of reverse-transcription quantitative PCR reference genes. J Dairy Sci 2020; 103:4846-4857.
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Madureira, G., Gomez-Leon, V., Grillo, G.F., Nascimento Andrade, J.P., Lett, B., Moghbeli, S.M., Wiltbank, M.C., Kirkpatrick, B.W., 2020. Practical application of an impractical bovine genotype: creating bilateral twin pregnancies in Trio allele carriers. J Anim Sci 98:1-8
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Garcia-Guerra A, Sala RV, Carrenho-Sala L, Baez GM, Motta JCL, Fosado M, Moreno JF, Wiltbank MC. Postovulatory treatment with GnRH on day 5 reduces pregnancy loss in recipients receiving an in vitro produced expanded blastocyst. Theriogenology 2020; 141:202-210.
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Drum JN, Wiltbank MC, Monteiro PLJ, Prata AB, Gennari RS, Gamarra CA, Canavessi AMO, Sartori R. Oxytocin-induced prostaglandin F2-alpha release is low in early bovine pregnancy but increases during the second month of pregnancy. Biol Reprod 2020; 102:412-423. Theriogenology 2020; 141:202-210.


    Progress 06/01/19 to 05/31/20

    Outputs
    Target Audience:Target Audiences: Scientists working in similar areas of reproductive physiology and reproductive management; veterinarians, producers, and industry professionals working in beef and dairy cattle; undergraduate and graduate students in animal science and veterinary medicine. Changes/Problems:During 2020, we had many setbacks on our experiments during the pandemic. We could not go to commercial operations to do our large fertility experiments, due to strict restrictions by the College of Agriculture and Life Sciences at University of Wisconsin-Madison. We also had many problems with completing our intensive experiments. The on-campus Dairy Cattle Center was closed in April 2020 while we were in the midst of an experiment. We moved this experiment to Arlington but were not able to do the same intensive measurements. The cows then returned at the end of August but there were promptly many problems due to the undergraduate student labor being quarantined. These pandemic-related problems have delayed completion of a number of our experiments. They have, however, provided us with greater time to analyze our previous results and prepare publications on a more rapid timetable. What opportunities for training and professional development has the project provided? Through laboratory meetings and individual meetings postdoctoral fellows and graduate students received training in the design, implementation, and analysis of reproductive biology research. They gave presentations in these meetings as well as in the Graduate Seminar of the Department of Dairy Science. These scientists also served as teaching assistants and instructors in part of the Animal Physiology course and Reproductive Management of Dairy Cattle course. Graduate students attended scientific meetings in 2019 and virtual meetings in 2020 (Society for the Study of Reproduction, International Ruminant Reproduction Symposium, American Society for Animal Science, and American Dairy Science Association). How have the results been disseminated to communities of interest? Manuscripts were published in peer-reviewed journals Abstracts and oral presentations at national and international scientific meetings. Presentations were delivered by PI to stakeholder meetings (producers and veterinarians). What do you plan to do during the next reporting period to accomplish the goals?During 2021, we are completing manuscripts that report our results from experiments on this research project completed in 2019 and 2020. The major experiments in 2021 will focus on determining the effect of increasing and decreasing uterine blood flow on pregnancy loss. We found that increasing circulating P4 had a small but significant effect on decreasing pregnancy loss in recipients of IVF embryos, however the losses were still near 50% during the period from 30 to 60 days of pregnancy. Therefore, we are now focused on changing the cloned embryos to more closely match the IVF embryos in order to improve placental development and decrease pregnancy loss. Our studies are focused on altering DNA and histone methylation patterns to normalize mRNA expression in 8-celll embryos. The optimized treatment will then be evaluated for pregnancy losses in recipients of cloned vs modified cloned embryos.

    Impacts
    What was accomplished under these goals? Objective 1: We have completed three different projects under this specific objective using different vasodilators to increase uterine blood flow. It does not appear that any of these vasodilators delayed luteolysis but we are still analyzing the results. These experiments wer done in a intermittent fashion during 2020, due to movement of cattle out of the on-campus dairy cattle center during pandemic restrictions. Objective 2: Experiment 4 was completed and is being prepared to publication. We have completed two studies that were recently published reporting our findings on understanding the process of luteolysis (Domingues et al. 2020a and 2020b). Objective 3: Due to cahnges at the ST Genetics, we performed this experiment in lactating cows rather than recipients of IVF embryos. The results are intriguing and are in the final stages of preparation as scientific manuscripts. In the course of these experiments and based on some of the observed results, we began working on a procedure for rapid reinsemination, that we have termed ReBred21. We have now published a preliminary report on this procedure (Andrade et al., 2020). We have also completed a study to evaluate fertility during use of this procedure and evaluating whether ReBreed21 can be used to improve fertility in embryo transfer programs.

    Publications

    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Domingues RR, Ginther OJ, Gomez-Leon VE, Wiltbank MC. Up-regulation of endometrial oxytocin receptor is associated with the timing of luteolysis in heifers with two and three follicular waves. Biol Reprod 2020; 102:316-326. 10.1093/biolre/ioz165
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Domingues RR, Ginther OJ, Toledo MZ, Wiltbank MC. Increased dietary energy alters follicle dynamics and wave patterns in heifers. Reproduction 2020; 160:943-953. 10.1530/Rep-20-0362
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Andrade JPN, Gomez-Leon VE, Andrade FS, Carvalho BP, Lacouth KL, Garcia FZ, Jacob JCF, Sales JNS, Wiltbank MC, Mello MRB. Development of a novel 21-day reinsemination program, ReBreed21, in Bos indicus heifers. Theriogenology 2020; 155:125-131. 10.1016/j.theriogenology.2020.04.021
    • Type: Journal Articles Status: Published Year Published: 2020 Citation: Motta JCL, Madureira G, Silva LO, Alves R, Silvestri M, Drum JN, Consentini CEC, Prata AB, Pohler KG, Wiltbank MC, Sartori R. Interactions of circulating estradiol and progesterone on changes in endometrial area and pituitary responsiveness to GnRH. Biol Reprod 2020; 103:643-653. 10.1093/biolre/ioaa065