Uterine kallikrein and arterial bradykinin activities and uterine arterial proliferation in response to acute estradiol-17ß exposure in ovariectomized ewes.

Estradiol-17ß (E2) increases kallikrein in rodent and human reproductive tissues. Kallikrein specific activity is increased in the porcine uterus when conceptus E2 is secreted at maternal recognition of pregnancy. When kallikrein acts on kininogen to liberate bradykinin, angiogenic and vasoactive factors are released. The uterus of ovariectomized ewes administered E2 undergoes rapid vascular changes via different patterns of angiogenic and vasoactive factors. Our hypothesis was that E2 would increase the specific activity and protein secretion of tissue kallikrein in endometrial explants culture media (ECM) and ewes exposed to E2 would have uterine arteries that would be more sensitive to the vasodilatory effects of bradykinin. Ovariectomized ewes received 100 mg of E2 implants for 0, 12, 24, or 48 h. After treatment, uterine weights were determined, and caruncles were processed for ECM. Uterine weights and uterine weight per ewe body weight were significantly greater in the 12 and 24 h ewes compared with the 0 h ewes, with the 48 h ewes being similar to the 24 h ewes. There were no statistically significant differences in caruncular tissue kallikrein protein secretion among the treatment groups. There was a tendency (P = 0.09) for duration of E2 exposure to influence tissue kallikrein specific activity where kallikrein activity was greater (P ≤ 0.05) in the 12 and 48 h ewes compared with the 0 h ewes, with 24 h ewes being intermediate (unprotected F test). Uterine arteries from ewes with E2 for 24 and 48 h had more sensitivity to bradykinin, via the bradykinin receptor 2, than uterine arteries from ewes with 0 or 12 h E2 exposure. We fail to reject our hypothesis as E2 did elicit a positive response in tissue kallikrein specific activity and bradykinin response. Further investigations are needed to determine how kallikrein and bradykinin may be involved in vascular remodeling of the ovine uterus.

Supplementation of metabolizable protein during late gestation and fetal number impact ewe organ mass, maternal serum hormone and metabolite concentrations, and conceptus measurements.

To examine the effects of maternal metabolizable protein (MP) supplementation during late gestation on serum hormone and metabolites and organ masses, multiparous ewes (n = 45) carrying singletons or twins were allotted randomly (within pregnancy group) to 1 of 3 treatments: 60% (MP60), 80% (MP80), or 100% (MP100) of MP requirements. Blood samples were drawn before the initiation of diets (day 100) and before slaughter (day 130) for chemistry panel analysis and weekly for hormone analysis including progesterone (P4) and estradiol-17ß (E2). At day 130, ewe organ masses were recorded. Despite being fed isocaloric diets, MP60 ewes gained less weight throughout pregnancy compared with MP80 and MP100 ewes which were similar. Although diet did not impact E2 or P4 concentrations, ewes carrying twins had greater (P < 0.05) concentrations of both as gestation advanced. Albumin, aspartate aminotransferase, and total protein were reduced (P < 0.05) in MP60 compared with MP100 ewes near term. There was a diet by fetal number interaction (P = 0.03) for lactate dehydrogenase. Twin-carrying MP80 ewes had greater lactate dehydrogenase compared with all other groups on day 130 of gestation. Ewes that were fed MP80 had greater body weight on day 130 of gestation compared with MP60 ewes. Kidney and heart weights were lighter in MP60 ewes compared with MP80 ewes. There was a maternal diet by fetal number interaction (P = 0.05) on fetal weight per unit empty ewe body weight. In ewes carrying singletons, MP60 ewes supported less fetal weight compared with MP100. In contrast, MP60 ewes supported more fetal mass compared with MP100 ewes when carrying twins. The level of protein, and not just total energy, in the diet appears to impact some aspects of the maternal system. Moreover, it appears some measurements of mobilizing maternal body resources are enhanced in ewes carrying twins.

Maternal metabolizable protein restriction during late gestation on uterine and umbilical blood flows and maternal and fetal amino acid concentrations near term in sheep.

To examine the effects of maternal metabolizable protein (MP) restriction during late gestation on uterine and umbilical blood flows, conceptus size, and amino acid concentrations in the uterine and umbilical vessels, 11 ewes with singleton pregnancies were assigned to one of three isocaloric diets formulated to provide 60% of MP (MP60), 80% of MP (MP80), or 100% of MP (MP100) requirements from days 100 to 130 of gestation. On day 130 of gestation, intraoperative uterine and umbilical blood flows were obtained as well as serum samples from the uterine artery, uterine vein, umbilical artery, and umbilical vein. Ewes on the MP60 diet had lighter (P=0.04) and smaller (P≤0.05) fetuses, but increased (P=0.02) uterine blood flow relative to fetal weight compared with MP100 ewes, with MP80 being intermediate. Umbilical blood flow was similar (P=0.70) across treatments. Glutamine, glycine, isoleucine, leucine, ornithine, serine, and valine concentrations were impacted (P≤0.02) by maternal treatment. While uterine flux of total serum nitrites was greater (P=0.03) in MP60 and MP80 ewes compared with MP100 ewes, fetal flux did not differ. Decreased maternal protein intake resulted in less (P<0.01) maternal cytochrome P450 1A enzyme activity. There were minimal impacts of maternal diet on steroid concentrations. Maternal dietary protein may alter fetal growth by impacting placental vasculature function and nutrient absorptive capabilities.

Effects of short-term oilseed supplementation on plasma fatty acid composition, progesterone and prostaglandin F metabolite in lactating beef cows.

Twenty-four 3-year-old Angus cows (512.2±21.6 kg) and six ruminally cannulated beef heifers (523.1±16.9 kg) were used to determine the impact of feeding oilseeds starting at the beginning of estrous synchronization until maternal recognition of pregnancy on plasma fatty acid composition. Starting ~60 days postpartum cows were synchronized with the Select Synch+controlled internal drug-release (CIDR) device and timed artificial insemination (AI) protocol. The day CIDR was inserted; cattle were randomly assigned to one of the three treatments being grazing only (CON) or a supplement containing whole soybeans (SOY); or whole flaxseed (FLX). Cattle continued to receive these diets for 28 days. Blood was collected every 3 days until 10 days after insemination and then every day until 18 days after insemination. All cattle grazed a common pasture and supplemented cattle were individually fed their respective supplements once daily. Ruminally cannulated heifers were used to evaluate the impact supplements had on forage intake, which was reduced (P=0.05) with oilseed supplementation. Feeding oilseeds increased total fatty acid intake (P<0.001) across treatments with SOY having greater (P<0.001) 18:2n-6 intake than either CON or FLX. Likewise, cattle fed FLX had greater (P<0.001) 18:3n-3 intake than either CON or SOY. There was a treatment×time interaction (P≤0.05) for all fatty acids identified except for 20:5n-3 (P=0.99). Within 3 days after the start of supplementation, plasma concentrations of 18:2n-6 increased (P<0.001) for cattle fed SOY compared with CON or FLX, whereas flax-fed cattle did not exhibit an increase (P=0.02) until day 15 of supplementation over that of CON. Plasma concentrations for 18:3n-3 was greater (P<0.013) for FLX than both CON and SOY by day 12. Feeding flaxseed tended to (P=0.07) increase and increased (P=0.01) plasma concentrations of 20:4n-6 by day 18 over CON and SOY, respectively. Overall, treatment did not affect serum concentration of progesterone (P=0.18) or prostaglandin F metabolite (P=0.89). However, day after breeding had an effect on serum progesterone (P=0.01) with day 16 after timed AI being lower compared with other days. Feeding oilseeds during the time of estrous synchronization will not only increase the energy density of the diet but will provide key fatty acids around the time of maternal recognition of pregnancy.

Supplementing metabolizable protein to ewes during late gestation: I. Effects on ewe performance and offspring performance from birth to weaning.

We hypothesized that a maternal diet greater in MP during the last third of gestation would yield improved offspring growth due to enhanced placental or mammary gland function. Our objective was to determine how MP intake during late gestation affected ewe and offspring performance from birth to weaning in ewes fed diets similar in total energy. In yr 1, maternal dietary treatments were applied at d 100 of gestation, were similar in total energy, and contained 60% of MP requirements (60MP1), 80% of MP requirements (80MP1), and 100% of the MP requirements (100MP1) on a DM basis during late gestation. In yr 2, maternal dietary treatments were similar in total energy and contained 60% of MP requirements (60MP2), 100% of the MP requirements (100MP2), and 140% of MP requirements (140MP2) on a DM basis during late gestation. In yr 1, there was no effect (P ≥ 0.30) of maternal MP restriction on gestation length, lamb birth weight, or milk production of the dam. At lambing, ewe BW (P = 0.02) and BCS (P = 0.01) increased linearly as MP in the diet increased. While there were no differences in lamb BW at birth, lamb weaning BW (P = 0.08) and ADG from birth to weaning (P = 0.10) tended to increase linearly as maternal MP intake increased. In yr 2, maternal BW increased with increased dietary MP. However, there were no significant effects (P ≥ 0.45) of maternal dietary treatment on maternal BCS or lamb birth weight. Moreover, lamb weaning weight and ADG were not influenced by maternal MP levels. These results suggest that ewes maintain and gain BW and BCS when consuming diets similar in total energy with increased MP during late gestation, but the increased MP had minimal effects on lamb performance through weaning.

Maternal selenium supplementation and timing of nutrient restriction in pregnant sheep: Impacts on nutrient availability to the fetus.

To determine the effects of maternal Se intake and plane of nutrition during mid or late gestation or both on AA concentrations and metabolite concentrations in the dam and fetus, pregnant ewe lambs (n = 64) were assigned to 1 of 8 treatments arranged in a 2 × 2 × 2 factorial array: Se level [initiated at breeding; adequate (ASe; 3.05 µg/kg of BW) or high (HSe; 70.4 µg/kg of BW)] and nutritional level [100% (control; CON) or 60% (restricted; RES) of NRC recommendations] fed at different times of gestation [d 50 to 90 (mid) or d 91 to 132 (late)]. A blood sample was obtained from each ewe and fetus on d 132 of gestation and used to measure circulating concentrations of glucose, NEFA, blood urea N, and AA. The late RES ewes and their fetuses had less (P ≤ 0.03) circulating glucose compared with late CON ewes and fetuses at d 132; however, no effect (P ≥ 0.14) of diet on the fetal:maternal glucose concentration ratio was observed. Late RES ewes had a smaller (P = 0.01) fetal:maternal NEFA ratio compared with late CON ewes. Ewes fed ASe had a greater (P = 0.01) fetal:maternal blood urea N ratio compared with HSe ewes. Fetal:maternal ratios of total circulating AA, total essential AA, and total nonessential AA were each affected (P ≤ 0.03) by the combination of Se treatment and late gestation nutritional level.

Maternal selenium supplementation and timing of nutrient restriction in pregnant sheep: effects on maternal endocrine status and placental characteristics.

To determine the effects of maternal Se intake and plane of nutrition during midgestation, late gestation, or both on hormone and metabolite concentrations in the dam and on placental characteristics, pregnant ewe lambs (n = 64) were assigned to 1 of 8 treatments arranged in a 2 x 2 x 2 factorial array: Se level [initiated at breeding; adequate (3.05 microg/kg of BW) or high (70.4 microg/kg of BW)] and nutritional level [100% (control) or 60% (restricted) of NRC recommendations] fed at different times of gestation [d 50 to 90 (midgestation) or d 91 to 130 (late gestation)]. The control ewes had a greater (P = 0.01) percentage change in BW from d 50 than restricted ewes during both mid- and late gestation. Although blood urea N was not affected by either Se or nutritional level, restricted ewes had greater (P = 0.01) concentrations of circulating Se on d 66, 78, 106, 120, and 130 of gestation compared with control ewes. Both Se and timing of the nutritional level affected circulating progesterone; however, only nutritional level affected thyroxine and triiodothyronine concentrations in the dam. Nutrient restriction during late gestation decreased (P or= 0.1) placental, caruncular, or cotyledonary weights, cotyledonary cellular proliferation was decreased (P < 0.05) in ewes receiving a high concentration of Se compared with those receiving adequate Se. In addition, either Se or nutritional level affected vascular endothelial growth factor (VEGFA), VEGFA-receptor 1, VEGFA-receptor 2, and NO synthase mRNA abundance in the cotyledonary tissue. In the caruncular tissue, either Se or nutritional level affected VEGFA-receptor 1, placental growth factor, and NO synthase mRNA abundance. Selenium supplementation and the duration or timing of nutrient restriction appear to influence the endocrine and metabolic status of the ewe, which may influence nutrient transport and placental function.

Cotyledonary responses to maternal selenium and dietary restriction may influence alterations in fetal weight and fetal liver glycogen in sheep.

To examine the effects of maternal supranutritional selenium (Se) and nutrient restriction during mid and late gestation on placental characteristics and fetal liver glycogen, ewes received either adequate Se (ASe) or high Se (HSe) prior to breeding. On d 64 of gestation, ASe and HSe ewes remained at 100% of requirements (controls; CON) or were restricted (RES; 60% of requirements). On d 135 of gestation, fetal weight (P< or =0.08) was greatest in both HSe and CON ewes. Placentome number, mass, and caruncular and cotyledonary weight were not different (P> or =0.17) among treatments. Fetal mass:placental mass ratio was less (P=0.06) in RES compared to CON ewes. Compared to ASe, HSe exhibited increased (P< or =0.08) cellular proliferation and DNA concentration and decreased (P=0.07) cellular size in cotyledonary tissue. Nutritional restriction decreased (P< or =0.08) cotyledonary protein concentration and cellular size. VEGF receptor 1 (Flt) mRNA in cotyledonary tissue was greater in HSe compared with ASe ewes (P=0.06) and in RES compared with CON ewes (P=0.08). There was no effect of diet on caruncular growth variables (P> or =0.13) or on placental vascularity (P> or =0.11). Progesterone was greater (P< or =0.08) in ASe-RES ewes compared to all groups at d 90 and ASe-CON and HSe-CON at d 104. Although fetal glucose and cortisol concentrations were not affected by diet, fetal liver glycogen was greater (P=0.04) in ASe-RES compared to ASe-CON and HSe-RES ewes with HSe-CON being intermediate. Both Se and nutritional plane may impact placental function and fetal growth, as fetal weight and liver glycogen are altered despite similar placental vascularity measurements.