Characterization of proteins in cryopreserved and non-cryopreserved seminal plasma of dairy bulls of dif-fering fertility

Seminal plasma is composed of secretions from accessory sex glands, which are mixed with sperm at ejaculation and contribute the majority of semen volume. Seminal plasma is considered a transport and support medium for sperm in the female reproductive tract. Because seminal plasma is not required for fertilization, the importance of its constituents to the establishment of normal pregnancy has been overlooked. Four seminal plasma proteins, Osteopontin, Sper-madhesin Z13, BSP 30 kDa and Phospholipase A2, have been identified as markers of fertility in dairy bulls (Cancel et al., 1997;Moura et al., 2006, 2007). The objective of the present study was to characterize the expression patterns of these proteins and other proteins found to be of interest in seminal plasma of cryopreserved and non-cryopreserved bull semen. Seminal plasma samples were obtained from 16 mature Hol-stein-Friesian bulls at Select Sires Inc. Samples were divided into two groups based on assigned fertility score expressed as the percentage point deviation (PD) of the bull’s non-return rate (NRR) from the average NRR of all bulls in the Select Sires Inc. reproductive management program. Group 1 (high fertility bulls, n = 8) 1.9 ≤ PD ≤ 2.7%, and group 2 (low fertility bulls, n = 8) -6.5 ≤ PD ≤ 1.8 %. Additionally, the samples were categorized as processed (cryopreserved) or unprocessed (non-cryopreserved) for protein analysis. Protein expression was analyzed by 2-D fluorescence difference gel electrophoresis (2D-DIGETM). Protein spots were picked from a reference gel, analyzed by mass spectrometry and, subsequently identified by MS/MS ion searches performed on the SwissProt database. Protein expression did not differ (P > 0.05) with fertility grouping but displayed two distinct patterns among the processing groups: majority of the functional proteins were highly expressed in seminal plasma of non-cryopreserved semen while the cryopreserved semen contained mainly structural/extender derived proteins. Functional proteins identified included Spermadhesin Z13, BSP A1/2, BSP 30 kDa, Nucleobindin-1 and metalloproteinase inhibitor 2. Some of these proteins have been identified as anti-fertility or fertility enhancing agents in males. Whether this alteration in protein expression after processing might affect semen fertility is worthy of further evaluation.

Feeding Different Omega-3 Polyunsaturated Fatty Acid Sources Influences Renal Fatty Acid Composition, Inflammation, and Occurrence of Nephrocalcinosis in Female Sprague-Dawley Rats

The general population is encouraged to increase omega-3 polyunsaturated fatty acid (n-3 PUFA) intake in order to optimize health for preventative health care. Consumers are typically unaware that different amounts, types, and structural forms of n-3 PUFA have different efficacy. Therefore, the objectives of this study were to characterize different sources of n-3 PUFAs and to determine whether consumption of these oils influences renal fatty acid composition and renal health. Lipid classes and fatty acid profile of corn (CO), flaxseed (FO), menhaden (MO), salmon (SO), tuna (TO) or krill (KO) oils were determined by thin-layer and gas chromatography. All dietary oils consisted of >65% triglyceride with the exception of KO. KO and FO also contained phospholipids. FO was rich in the n-3 PUFA, alpha-linolenic acid (18:3n-3) whereas, the marine oils were rich in the long-chain n-3 PUFAs (>18 carbons). Following characterization of the oil sources, female Sprague-Dawley rats (age 28 d) were randomly assigned (n = 10/group) to be fed a high fat 12% (wt) diet consisting of these different oil sources for 8 weeks. Rats fed MO, TO, and SO had significantly higher renal eicosapentaenoic acid (20:5n-3) and docosahexaenoic acid (22:6n-3) deposition and this in turn, modulated inflammatory responses. Feeding rats MO, SO and TO reduced urinary excretion of 13,14-dihydro-15-keto prostaglandin E2. Feeding rats TO and SO reduced (P ≤ 0.002) nuclear factor kappa B activity and circulating TNFα (P < 0.05). In contrast, rats consuming KO had heavier kidney weights (P < 0.001), total calcium content, and histological evidence of renal calcification and tubulo-interstitial injury. This was due to increased (P < 0.001) urinary phosphorus excretion associated with the phospholipids content of KO. The study results indicated that consumption of n-3 PUFAs influences renal health and the effects varied depending on the n-3 PUFA source consumed.

Evaluation of forage-based weaning systems in spring-born cross-bred beef calves

Preconditioned calves have greater marketvalue per unit weight than normal-weaned calves. Development of a low cost forage-based preconditioning system allows producers to add value to their calf-crop. This study evaluated calf performance in three forage-based weaning systems;early-weaned calves were backgrounded in legume/grass forage plots and supplemented with commercial preconditioning feed (Treatment 1) or an on-farm corn-mix (Treatment 2). Control (Treatment 3) calves suckled for an additional 45 days. Supplements provided2.17 kgTDN/calf/ day. Weights were collected on days -30, 0 and 45 with respect to early weaning, from135 inyear 1 and 150 calves in each of the two subsequent years. Effects of treatment, age of dam, sex of calf and their interactions on calf weight gain were analyzed by analysis of covariance using GLM procedures of SAS. Marginal effects of treatment and feed cost were used to evaluate economic feasibility. Sensitivity analyses were evaluated for anticipated market fluctuations in feed costs and calf premiums. Data are reported as least squares means. Calf weight gains differed (P < 0.001) among treatments and averaged 1.16, 1.03 and1.04 kg/calf/day for commercial supplement, corn-mix and controls, respectively. Calves from 2-year-old cows gained less (P < 0.001) weight compared to those from cows 3 - 4 and ≥5 years of age (44.8, 48.9 and51.5 kg, respectively). Steers calves gained more (P < 0.001) weight compared to heifer calves (51.2 vs.45.7 kg, respectively). Net returns for corn mix were greater than those for commercial feed ($1.48 vs. $1.35/kg weight gain, respectively). Sensitivity analyses indicated that selection of preconditioning treatment to a large degree was less sensitive to significant changes in market conditions due to the large gap in marginal costs between the two treatments. In conclusion, forage-based weaning systems can be utilized to precondition calves providing an economical means for calf weight gain and profit potential as long as feed costs are held within reasonable limits.

Effect of Lipopolysaccharide-Induced Immune Responses on Pregnancy Loss in Ewes

An acute phase response induced by Gram-negative bacteria can reduce pregnancy rate. Early pregnant ewes were used to monitor effects of lipopolysaccharide (LPS), an endotoxin in the outer cell membrane of Gram-negative bacteria, on acute phase/innate immunity response. In Exp. 1, mixed breed ewes were assigned to receive either LPS or LPS and flunixin meglumine, an inhibitor of prostaglandin synthetase, intravenously on day 5 after mating. In Exp. 2, mixed breed ewes were assigned to receive an intravenous injection on day 5 after mating of either saline, LPS, recombinant human tumor necrosis factor (TNF)-α or LPS after pretreatment with dexamethasone. Pregnancy was diagnosed ultrasonographically on d 25, and live births were recorded at parturition. Challenge with LPS induced acute phase responses (fever, mucosal responses, lethargy and increased serum TNF, haptoglobin and?serum amyloid A) and decreased pregnancy rates. Predictably, flunixin meglumine attenuated fever but did not increase pregnancy rate in LPS-treated ewes. Similarly, exogenous TNF alone induced mucosal and serum amyloid A responses but did not affect pregnancy. Pre-treatment with dexamethasone blocked fever and mucosal and lethargic responses and attenuated increases in TNF and haptoglobin but did not ameliorate LPS-induced pregnancy loss. In summary, acute challenge with LPS mimics bacterial-induced pregnancy losses in early pregnant ewes. Although pretreatment with dexamethasone decreased clinical signs and some innate immune responses, neither it nor flunixin meglumine prevented LPS-induced pregnancy loss. That exogenous TNF alone did not promote pregnancy loss indicates that other cytokines also contribute to LPS-induced embryonic loss.

Periconceptional growth hormone treatment alters early uterine environment

We have shown that an injection of sustained release growth hormone (GH), given just prior to breeding, results in lambs that are 25% heavier at birth, with an altered body composition as evidenced by an increased abdominal girth, but no difference in crown rump length. The mechanisms by which these differences occur from a single periconceptional injection are not yet known. Therefore, the objective of this experiment was to determine the effect of an injection of GH given prior to breeding on the composition of the uterine luminal environment and the embryo at the time of blastocyst. Ewes were synchronized with two injections of prostaglandin F2α given eight days apart. On the day of the second injection, ewes were randomly assigned to be given an injection of sustained release GH or remain as controls and penned with a ram. On day 6.5 following breeding embryos were collected. Prior to surgery a jugular blood sample was taken to determine plasma progesterone and urea concentration. The uterine content of urea, prostaglandin F2α, prostaglandin E2, transforming growth factor β-1 (TGF β), and nitric oxide metabolites were measured. Collected embryos were differentially stained to calculate a trophectoderm to inner cell mass ratio. The concentration of progesterone in maternal plasma was greater in the treated group compare to controls, but blood urea nitrogen was not different between groups. The uterine content of urea in the GH group was lower than that of the control group. There was a trend for TGF β to be increased in the GH treated group compared to control (P = 0.07). There was no difference (P > 0.05) in the total uterine content prostaglandin F2α, prostaglandin E2 or nitric oxide metabolites. The trophectoderm to inner cell mass ratio was not different between treatment groups. Thus, we suggest that the observed difference in fetal development following periconceptional growth hormone administration just prior to breeding may be the result of an alteration in the uterine luminal environment, which may alter the cellular program of the conceptus and later development beyond the embryonic stage.

Reactive oxygen and nitrogen species regulate porcine embryo development during pre-implantation period:A mini-review

Significant porcine embryonic loss occurs during conceptus morphological elongation and attachment from d 10 to 20 of pregnancy,which directly decreases the reproductive efficiency of sows.A successful establishment of pregnancy mainly depends on the endometrium receptivity,embryo quality,and utero-placental microenvironment,which requires complex cross-talk between the conceptus and uterus.The understanding of the molecular mechanism regulating the uterine-conceptus communication during porcine conceptus elongation and attachment has developed in the past decades.Reactive oxygen and nitrogen species,which are intracellular reactive metabolites that regulate cell fate decisions and alter their biological functions,have recently reportedly been involved in porcine conceptus elongation and attachment.This mini-review will mainly focus on the recent researches about the role of reactive ox-ygen and nitrogen species in regulating porcine embryo development during the pre-implantation period.