Low-dose lipopolysaccharide exposure during oocyte maturation disrupts early bovine embryonic development.

Gram-negative bacteria release of lipopolysaccharide (LPS) endotoxin elicits robust immune responses capable of disrupting normal ovarian function contributing to female infertility. However, effects of subclinical or non-detectable infections on oocyte competence and subsequent embryo development remain to be fully elucidated. The aim of this study was to investigate the effects of exposing bovine oocytes to low LPS doses on oocyte and embryo competence. Bovine oocytes were collected from slaughterhouse-derived ovaries and matured with vehicle-control or increasing doses of LPS (0.01, 0.1, and 1 µg/mL) for 21 h. Oocytes (n = 252) were evaluated for nuclear maturation. A set of embryos from LPS-matured oocytes (n = 300) were cultured for 8 d to evaluate day 3 cleavage rates and day 8 blastocyst rates along with blastocyst cell counts. A subset of oocytes (n = 153) was fertilized and cultured for time-lapse image capture and analysis of embryo development. Results demonstrate no significant treatment differences among treatment groups in percent of oocytes at germinal vesicle (GV; P = 0.90), germinal vesicle breakdown (GVBD; P = 0.13), meiosis I (MI; P = 0.26), or metaphase II (MII; P = 0.44). Likewise, treatment differences were not observed in cleavage rates (P = 0.97), or blastocyst rates (P = 0.88) evaluated via traditional microscopy. Treatment with LPS did not affect total blastocyst cell count (P = 0.68), as indicated by trophectoderm (P = 0.83), and inner cell mass (P = 0.21) cell counts. Time-lapse embryo evaluation demonstrated no differences among control or LPS matured oocytes in number of zygotes that did not cleave after fertilization (P = 0.84), or those that cleaved but arrested at the 2-cell stage (P = 0.50), 4-cell (P = 0.76), prior to morula (P = 0.76). However, embryos derived from oocytes challenged with 0.1 µg/mL LPS tended to have reduced development to the morula stage compared with vehicle-treated controls (P = 0.06). Additionally, the percentage of blastocysts derived from oocytes matured in 0.01 µg/mL LPS tended to decrease compared to vehicle-treated controls (11.38 and 25.45 %, respectively; P = 0.09). Similarly, the proportion of oocytes that developed to the blastocyst stage was greater in vehicle-treated controls (25.45 %) compared with embryos derived from oocytes matured in 0.1 and 1 µg/mL (5.92 and 6.55 %, respectively; P = 0.03) LPS. These data suggest LPS-matured oocytes that subsequently underwent in vitro fertilization, experienced decreased competence to develop to the blastocyst stage.

Recycling nutrients in the beef supply chain through circular manuresheds: Data to assess tradeoffs.

Nutrient circularity can help supply chain participants meet sustainability targets. Across the segmented beef supply chain, opportunity exists to reinforce and introduce nutrient circularity by recycling surplus manure nutrients from cattle feedlots to lands where cattle feed is produced. We describe four datasets developed to evaluate options in U.S. and Canadian beef systems. The datasets delineate three "circular manuresheds," each encompassing a hay-grazing landscape where beef cattle are raised on grazingland and supplemented with hay grown nearby, and the distant feedlots where those cattle produce manure nutrients for potential import back to the hayfields. We selected the hay-grazing landscapes of New Mexico, USA; Florida, USA; and western Canada (Manitoba, Saskatchewan, Alberta, British Columbia) because of their significant grazingland production and potential to substitute feedlot manure for commercial fertilizer on hayfields. In each circular manureshed, the manure nutrients from major feedlot destinations could supply a considerable proportion of the P used by hay for grazing cattle: 34% of the P requirements in New Mexico, 36% in Florida, and 6% in western Canada. The average distance to return the resource was 647 km for New Mexico, 1,884 km for Florida, and 1,587 km for western Canada. These magnitudes and distances suggest that the New Mexico circular manureshed may be the most economically viable in the current agri-food system, but this reflects only part of a greater, multi-factor assessment of tradeoffs. The circular manureshed concept provides a platform for simultaneous consideration of competing factors for sustainability via circularity.

Nutrient circularity can help animal supply chain participants meet sustainability goals. The manureshed concept can help willing consortia operationalize nutrient circularity. Data delineate three circular manuresheds where feedlot manure is recycled onto hayfields in landscapes where feedlot cattle originated. Available nutrients and transport distances for the three circular manuresheds are calculated. These findings can inform a comprehensive assessment of tradeoffs of circular manureshed management.

Effects of lipopolysaccharide on follicular estrogen production and developmental competence in bovine oocytes.

Reproductive efficiency and female fertility is essential for productive and sustainable beef cattle operations. Gram-negative bacterial infections cause release of the endotoxin lipopolysaccharide (LPS) which initiates immune responses shown to alter ovarian steroidogenesis and impair oocyte development. The current study was designed to investigate the impact of varying levels of naturally occurring infection and follicular LPS on estradiol (E2) production and oocyte maturation. Bovine ovary pairs were harvested from a slaughterhouse, and oocytes were aspirated from small follicles and matured in vitro. Meiotic events were evaluated on nuclear maturation and spindle morphology to classify oocytes as normal or abnormal. Follicular fluid LPS concentrations were measured and subsequently separated into Low or High LPS groups. A marked difference was detected between the percent of abnormal oocytes matured from Low LPS follicles, compared to the percent of abnormal oocytes matured from High LPS follicles (P = 0.1). Follicular E2 concentrations tended to be greater for high LPS follicles (P = 0.1), however, relative abundance of mRNA transcripts for aromatase (P = 0.93) and beta-catenin (P = 0.63) were similar between groups. No changes were detected in Toll-like Receptor 4 (P = 0.15), Myeloid Differentiation Factor-2 (P = 0.61), or cluster of differentiation 14 (P = 0.46) mRNA transcript abundance in follicles with high LPS, compared to low. Therefore, even Low levels of follicular LPS indicating a subacute infection is capable of impacting the ovarian milieu and may represent an unappreciated factor leading to reduced female fertility and decreased cow retention.

Effects of lipopolysaccharide on beta-catenin, aromatase, and estrogen production in bovine granulosa cells in vivo and in vitro.

Diseases resulting from Gram-negative bacterial infection can induce an immune response by releasing a lipopolysaccharide (LPS) endotoxin that may lead to impaired fertility in cows. To evaluate the effects of LPS on follicular dynamics in a subacute inflammatory disease state, 14 Angus heifers (BW = 413 kg±14) were blocked by weight and assigned to vehicle (n = 7) or LPS treated (n = 7) groups. Heifers received subcutaneous injections of saline (CON) or 2.0 µg/kg LPS on d 2, 5, and 8 of a select synch plus controlled internal drug release device (CIDR) follicular wave synchronization protocol. Fifty hours following CIDR withdrawal, ovaries were harvested, and follicular fluid was collected for hormone and LPS analysis. Daily blood samples were collected from d 0 to d 7. Beginning on d 8 blood samples were collected at 0, 16, 24, 32, 40, and 50 h following LPS challenge. Rectal temperatures were recorded prior to treatment and at regular intervals after each LPS challenge. Heifers treated with LPS exhibited mild (+0.5 °C) hyperthermia (P < 0.05) at 3, 4, and 8 h after the initial LPS challenge (d 2) when compared to vehicle-treated controls. Follicular fluid concentrations of estradiol (E2) increased (P = 0.04) in LPS-treated heifers compared to controls (1,595 ng/mL and 808 ng/mL±240, respectively), while follicular fluid progesterone (P4) concentrations did not differ (P = 0.27) between treatment groups. Additionally, LPS concentrations tended to be increased (P = 0.59) in dominant follicles of LPS-treated heifers, but no difference was detected (P = 0.81) in small developing follicles. To further delineate the impact of LPS on ovarian signaling pathways, a granulosa cell line (KGN) was incubated in the presence or absence of LPS (10 µg/mL) for 48 h. Cells were then collected for gene expression and protein analysis. Cells in both treatment groups expressed toll-like receptor 4, myeloid differentiation factor-2 receptor, and CD-14 complex genes required for LPS signaling. Cells treated with LPS exhibited decreased mRNA expression of aromatase (P = 0.03) and beta-catenin (P = 0.02). However, no change (P > 0.10) was detected in abundance of total beta-catenin protein or beta-catenin phosphorylated isoforms at serine 552 or 675. Based on results from this in vivo experiment, these investigators concluded that low doses of LPS can alter E2 concentrations and this effect may be modulated in part through beta-catenin regulation of aromatase transcription.

Special-Fed Veal: Separable components, proximate composition, and nutrient analysis of selected raw and cooked, wholesale and retail cuts.

Nutrition research continues to be important for consumers to make informed food purchasing decisions and is used in nutrition policy decisions. The objective of this study was to analyze the nutrient concentration of raw and cooked cuts from special-fed veal calves to update nutrient data in the USDA National Nutrient Database for Standard Reference (SR) Release 27. Packages of wholesale (whole loin roasts, center-cut hindshanks and ground veal) and retail veal cuts (osso buco foreshanks, loin chops, leg cutlets and shoulder blade chops) were randomly collected in original vacuum packaging from six U.S. suppliers. Packages were shipped to the Colorado State University Meat Laboratory for cut dissection, cooking, and nutrient analysis. Composites of lean, external fat and seam fat were formed for analysis of proximate, fatty acid, vitamin and mineral composition. Results from this study identified additional fatty acids, established choline concentration, and provided updated veal nutrient composition information for inclusion in USDA SR 27.

National Beef Quality Audit-2016: Phase 1, Face-to-face interviews.

The National Beef Quality Audit (NBQA) is conducted every 5 yr and was most recently again conducted in 2016. Face-to-face interviews gauged progress in quality associated with live cattle production using procedures first utilized in NBQA 2011. The 2016 NBQA was the first in which interviews concerning fed steers and heifers were combined with an audit of market cow and bull beef. Face-to-face interviews were designed to illicit definitions for beef quality, estimate willingness to pay (WTP) for quality attributes, establish relative importance rankings for important quality factors, and assess images, strengths, weaknesses, potential threats, and shifting trends in the beef industry since the 2011 audit. Individuals making purchasing decisions in 5 market sectors of the steer/heifer and cow/bull beef supply chain were interviewed, including packers (n = 36), retailers (including large and small supermarket companies and warehouse food sales companies; n = 35), food service operators (including quick-serve, full-service, and institutional establishments; n = 29), further processors (n = 64), and peripherally-related government and trade organizations (GTO; n = 30). Face-to-face interviews were conducted between January and November of 2016 using a designed dynamic routing system. Definitions (as described by interviewees) for 7 pre-determined quality factors, including: (1) How and where the cattle were raised, (2) Lean, fat, and bone, (3) Weight and size, (4) Visual characteristics, (5) Food safety, (6) Eating satisfaction, and (7) Cattle genetics were recorded verbatim and categorized into similar responses for analysis. Compared to NBQA-2011, a higher percentage of companies were willing to pay premiums for guaranteed quality attributes, but overall were willing to pay lower average premiums than the companies interviewed in 2011. Food safety had the highest share of preference among all interviewees, generating a double-digit advantage over any other quality factor. The 2 beef industries have an overall positive image among interviewees, and despite lingering weaknesses, product quality continued to be at the forefront of the strengths category for both steer and heifer beef and market cow and bull beef.

Effect of bovine respiratory disease during the receiving period on steer finishing performance, efficiency, carcass characteristics, and lung scores.

Bovine respiratory disease (BRD) is responsible for the majority of morbidity, mortality, and production losses occurring in feedlots. This experiment evaluated the effects of BRD incidence on subsequent finishing performance, efficiency, carcass characteristics, and lung scores of steers. Crossbred steers (n = 516) were monitored daily for clinical signs of BRD (BRD attributed morbidity and mortality were 66.5 and 13.2%, respectively). A subset of calves (n = 174) were grouped by the number of times treated for BRD (BRDX) and randomly allocated to finishing pens. The BRDX experimental groups included never treated for BRD (0X; 8 pens) and treated 1 time (1X; 8 pens), 2 times (2X; 8 pens), or 3 or 4 times (3/4X; 8 pens). Arrival BW was not different among BRDX groups (P = 0.17); however, BRDX during the receiving period decreased performance, resulting in BW of 324, 316, 285, and 260 kg for 0X, 1X, 2X, and 3/4X, respectively, at the start of finishing (P < 0.001). Ultrasound estimates, BW, and visual appraisal were used to target a common body composition (average days on feed = 182). With increasing BRDX, days on feed increased linearly (P = 0.002), whereas HCW, DP, rib eye area, QG, and unconsolidated lungs decreased linearly (P ≤ 0.03). These results suggest that with additional days on feed, calves treated multiple times for BRD are able to reach similar compositional endpoints as their untreated cohorts; however, it may not be possible for these calves to reach the same QG and carcass yield.

Effect of copper, manganese, and zinc supplementation on the performance, clinical signs, and mineral status of calves following exposure to bovine viral diarrhea virus type 1b and subsequent infection.

Research has indicated that trace mineral (TM) supplementation may alter immune function and reduce morbidity associated with bovine respiratory disease. The objective of this experiment was to determine the influence of dietary Cu, Mn, and Zn supplementation on the performance, clinical signs, and TM balance of calves following a bovine viral diarrhea virus (BVDV) and (MH) combination respiratory pathogen challenge. Steers ( = 16; 225 ± 20 kg BW) from a single ranch were processed, weaned, and randomly pairwise assigned to either the TM-supplemented (MIN) or the control (CON) experimental treatments. The MIN calves received an additional 150 mg of Cu, 130 mg of Mn, and 320 mg of Zn daily and the CON calves received the basal diet with no additional Cu, Mn, or Zn supplementation. The basal diet contained sufficient Mn and Zn but inadequate Cu based on published nutrient requirements. After 46 d on the experimental treatments, all calves were naturally exposed to a heifer persistently infected with BVDV type 1b for 4 d and then subsequently intratracheally challenged with MH. Data were analyzed using the GLIMMIX procedure of SAS with sampling time serving as a repeated measure and calf serving as the experimental unit. The respiratory challenge was validated via increased BVDV type 1b antibody concentrations, MH whole cell and leukotoxin antibody concentrations, rectal temperatures (TEMP), and subjective clinical severity scores (CS). Calf performance ( ≥ 0.48) was not affected by TM supplementation. Mineral supplementation also did not impact the CS or TEMP of calves ( ≥ 0.53). There was a treatment × time ( < 0.001) interaction observed for liver Cu concentrations. The concentrations of Cu, Mn, Zn, and Fe within the liver; Cu, Mn, and Zn within the muscle; and Cu, Zn, and Fe within the serum were all impacted by time ( ≤ 0.03). Calves receiving the MIN treatment had greater ( < 0.01) liver Cu and Mn concentrations compared with CON calves. In contrast, serum Cu and Fe concentrations were increased ( ≤ 0.05) in CON calves compared with MIN calves. Mineral supplementation did not impact TM concentrations within the muscle ( ≥ 0.38). The supplementation of Cu, Mn, and Zn can improve the Cu and Mn status within the liver and serum of calves in response to a BVDV and MH challenge. When Cu is supplemented to calves receiving a marginally Cu-deficient diet, Cu status within the body is significantly improved.

Protein kinase B is required for follicle-stimulating hormone mediated beta-catenin accumulation and estradiol production in granulosa cells of cattle.

Follicle-stimulating hormone regulation of ovarian estradiol (E2) production requires involvement of beta-catenin (CTNNB1), a transcriptional co-factor. In cultured granulosa cells (GC) of cattle, FSH treatment increased protein abundance of CTNNB1 as well as protein kinase B (AKT), a molecule known to regulate components of the CTNNB1 degradation complex. However, whether FSH induction of CTNNB1 is through direct modulation of AKT remains to be determined. To investigate specific contributions of AKT to CTNNB1 accumulation, GC were treated with insulin-like growth factor-I (IGF-I), a well-established AKT activator, in the presence or absence of FSH. Granulosa cells treated with FSH, IGF-I, and IGF-I plus FSH had increased CTNNB1 accumulation compared with controls (P ≤ 0.02; n=6). E2 medium concentrations were greater (P=0.09; n=4) in FSH treated cells compared to controls (166 and 100 ± 28 pg/mL, respectively). Treatment with IGF-I and IGF-I plus FSH increased (P<0.01) E2 to comparable concentrations. Subsequently, GC treated with lithium chloride (LiCl), a pharmacological activator of AKT, provided a response consistent with IGF-I treated cells, as LiCl, FSH, and FSH plus LiCl increased CTNNB1 accumulation compared with non-treated controls (P ≤ 0.03; n=3). In contrast, inhibition of AKT signaling with LY294002 suppressed the ability of FSH and IGF-I to regulate CTNNB1. Additionally, LY294002 treatment reduced FSH and IGF-I mediated E2 medium concentrations (P ≤ 0.004). These results demonstrate that activation of AKT is required for gonadotropin regulation of CTNNB1 accumulation and subsequent ovarian E2 production.

Cattle with increased severity of bovine respiratory disease complex exhibit decreased capacity to protect against histone cytotoxicity.

Bovine respiratory disease complex (BRDC) is the leading cause of morbidity and mortality in feedlot cattle. Significant inflammation and lesions are often observed in lungs of infected cattle. During acute inflammatory responses, histones contribute to mortality in rodents and humans and serum proteins can protect against histone-induced cytotoxicity. We hypothesized that cattle experiencing chronic or fatal cases of BRDC have reduced ability to protect against cytotoxic effects of histones. Serum samples were collected from 66 bull calves at the time of normal feedlot processing procedures. Animals were retrospectively assigned to groups consisting of calves never treated for BRDC (control [CONT]; n = 10), calves treated with antimicrobials once for BRDC (1T; n = 16), calves treated twice for BRDC (2T; n = 13), calves treated 3 times for BRDC (3T; n = 14), or calves treated 4 times for BRDC (4T; n = 13). Samples were also collected each time animals received antimicrobial treatment; animals within a group were further sorted by calves that recovered and calves that died to test histone cytotoxicity. Bovine kidney cells were cultured in duplicate in 96-well plates and exposed to 0 or 50 µg/mL of total histones for 18 h with 1% serum from each animal. Cell viability was assessed by the addition of resazurin for 6 h followed by fluorescent quantification. Fluorescent values from serum alone were subtracted from values obtained for histone treatment for each animal. Serum from CONT, 1T, and 2T at initial processing all exhibited a similar (P > 0.10) response to histone treatment with fluorescent values of -312 ± 557, -1,059 ± 441, and -975 ± 489, respectively. However, 3T and 4T demonstrated an impaired capacity (P < 0.05) to protect against histones (-2,778 ± 471 and -3,026 ± 489) at initial processing when compared to the other groups. When sorted by mortality within group, calves that were treated twice and recovered (-847 ± 331) demonstrated a greater (P < 0.05) protective capacity than calves that were treated twice and died (-2,264 ± 412), indicating that calves that contract BRDC and ultimately die might have reduced protective capacity against histone cytotoxicity. Results suggest that calves that require multiple treatments for BRDC have reduced ability to protect against cytotoxicity of histones. Understanding the primary mechanism responsible for protecting against histone cytotoxicity could lead to improved identification of animals susceptible to severe cases of BRDC, improved focus and use of available resources, or better treatments for severe cases of BRDC

Effect of anabolic implants on adrenal cortisol synthesis in feedlot beef cattle implanted early or late in the finishing phase.

Implantation of anabolic steroids to increase growth rate in beef cattle impacts adrenal glucocorticoid production. The mechanism by which combination androgen and estrogen implants reduce cortisol biosynthesis in heifers is not clear. The objective of this study was to identify whether pituitary or adrenal gene expression and liver enzyme activity may contribute to altered serum cortisol concentrations in heifers receiving a combination implant. On d 0 of a 122-d finishing phase, 187 predominantly Angus heifers (361 kg) approximately 14 months old were randomly assigned to one of three implant groups: (1) non-implanted control, (2) implanted at the beginning of the finishing phase (d 0; early implant) with a combination implant (200mg TBA+20mg E2; Revalor 200®), and (3) implanted during the late stage of the finishing phase (d 56; late implant) with Revalor 200®. At d 56, body weight (BW) was greater (P<0.0001) for the early implanted heifers (456 ± 1.9 kg) compared to 437 and 435 (± 1.8) kg for control and late implanted heifers, respectively. Final BW (d 122) was similar between both implanted groups and heavier than non-implanted controls (P<0.0001). Serum cortisol was similar among groups at d 0 (P=0.86) however, by d 28 heifers receiving the combination implant had reduced (P<0.05) serum cortisol concentrations (31.2 ng/mL) compared to controls (49.4 ng/mL) and late (48.2 ng/mL) groups. On d 84 cortisol was similar (P=0.75) among implanted heifers and was less (P<0.01) than non-implanted heifers. Expression of pituitary and adrenal genes involved in glucocorticoid synthesis was evaluated at d 28/29 or 84/85; however, despite decreased serum cortisol in implanted heifers, no change in mRNA expression was demonstrated. Liver CYP3A enzyme activity at d 28/29 was decreased 59% in early implanted heifers compared to control heifers (P=0.01). Additionally, at d 84/85 AKR1C activity was greatest (P=0.01) in control heifers compared to both implanted groups. Data suggest that components of hypothalamic-pituitary-adrenal axis are influenced by exposure to exogenous hormones and this should be recognized when considering cortisol levels as a marker for stress response.

Follicle-stimulating hormone regulation of estradiol production: possible involvement of WNT2 and ß-catenin in bovine granulosa cells.

Follicle-stimulating hormone regulation of estrogen biosynthesis in the adult rodent ovary requires ß-catenin (CTNNB1), but whether CTNNB1 is involved in FSH-induced estrogen production in cattle is unknown. To elucidate the effect of FSH in regulating specific wingless-type mouse mammary tumor virus integration site (WNT)/CTNNB1 pathway components in bovine folliculogenesis and steroidogenesis, granulosa cells and follicular fluid were collected from large antral follicles (8 to 22 mm) from ovaries containing stage-III corpora lutea (d 11 to 17 of an estrous cycle). Follicles were categorized as high estradiol (n = 3; ≥ 25 ng/mL) or low estradiol (n = 3; ≤ 14 ng/mL) based on intra-follicular estradiol concentrations. Protein fractions were collected from granulosa cells and CTNNB1 abundance was analyzed by Western blot. Follicles with increased estradiol concentrations had 6-fold greater (P < 0.001) abundances of CTNNB1 compared with those classified as low-estradiol follicles, indicating that the hormonal milieu responsible for increased estradiol content could result in CTNNB1 accumulation. To ascertain specific contributions of FSH to increases in CTNNB1 protein abundances, granulosa cells were isolated from small ovarian follicles (1 to 5 mm) and cultured in the presence or absence of 100 ng/mL FSH for 24 or 48 h. Real-time PCR quantification of aromatase (CYP19A1) and select WNT family members were evaluated in response to FSH treatment. Successful stimulation of granulosa cells with FSH was confirmed by induction of CYP19A1 mRNA and parallel temporal increases of medium estradiol concentrations. Additionally, protein kinase b (AKT), a known FSH target, increased 1.7-fold (P = 0.07). Of the WNT family members analyzed, only WNT2 mRNA was induced after 24 h of FSH treatment compared with controls (0.12-fold and 3.7-fold for control and FSH-treated, respectively; P < 0.05), and WNT2 expression tended (P = 0.11) to remain increased at 48 h in FSH-treated cells compared with controls (1.0- and 3.14-fold, respectively). Furthermore, FSH-treated granulosa cells had greater abundances of total CTNNB1 (P = 0.04) protein. These data demonstrate for the first time that FSH regulates CTNNB1 protein and WNT2 mRNA expressions in bovine granulosa cells, suggesting a potential role of canonical WNT signaling in ovarian steroidogenesis and follicular growth of cattle. Future studies are necessary to determine if FSH directly regulates CTNNB1 through modulation of AKT or indirectly by up regulating WNT2, which subsequently activates the canonical WNT pathway.

Growth and Development Symposium: Impacts of inflammation on cattle growth and carcass merit.

Inflammation caused by bovine respiratory disease (BRD) continues to be one of the greatest challenges facing beef cattle producers and feedlot managers. Inflammation decreases DMI, ADG, and G:F in feedlot calves, decreasing growth rate and increasing days on feed, which results in economic losses during the feeding period. During the past decade, marketing of feedlot animals has changed from selling cattle on a live basis to a grid-based marketing system. When cattle are marketed on a live basis, the economic effects of BRD stop at increased health cost and decreased feedlot performance, carcass weight, and death loss. However, when cattle are marketed in a grid-based system, inflammation has the potential to also affect carcass cutability and quality. The effects of inflammation on feedlot cattle in regards to performance are well understood; however, specific effects on cattle growth and ultimately carcass merit are not as well described. Recent studies in feedlot cattle have indicated that the incidence of BRD decreases both HCW and marbling; however, mechanisms are not understood. Research in other species has demonstrated that during the acute phase response, pro-inflammatory cytokines promote skeletal muscle catabolism to supply AA and energy substrates for immune tissues. Further, during this early immune response, the liver changes its metabolic priorities to the production of acute phase proteins for use in host defense. Together these dramatic shifts in systemic metabolism may explain the detrimental effects on performance and carcass traits commonly associated with BRD in feedlot calves. Moreover, recent studies relative to human health have revealed complex multilevel interactions between the metabolic and immune systems, and highlighted inflammation as being a significant contributor to major metabolic diseases. The objective of this paper is to review data to help explain the economical and physiological effects of inflammation on cattle growth and carcass merit.

Receptor transporter protein 4 (RTP4) in endometrium, ovary, and peripheral blood leukocytes of pregnant and cyclic ewes.

Interferon-tau (IFNT) is secreted by the conceptus trophoblast and signals pregnancy recognition in ruminants. IFNT regulates expression of genes in the endometrium, peripheral blood leukocytes (PBLs), and corpus luteum (CL). Microarray analysis identified that expression of (chemosensory) receptor transporter protein 4 (RTP4) increased in PBLs during early pregnancy in cows. In the present study, we cloned and characterized RTP4 transcription during early pregnancy in ewes. Endometrium, PBLs, and CL were collected on Days 11, 13, and 15 of the cycle and on Days 11, 13, 15, 17, and 19 of pregnancy. Northern blot analysis revealed an expected 1.6-kb mRNA and an unexpected 2.6-kb mRNA. In endometria, RTP4 mRNA levels in cyclic ewes remained low, whereas RTP4 mRNA increased from Day 11 to Day 17 in pregnant ewes. Levels of RTP4 mRNA also increased from Day 15 to Day 19 in CL and PBL samples from pregnant ewes only. The RTP4 mRNA was located in the glandular epithelium, stratum compactum, and caruncular stroma. Ovine glandular epithelial cells were treated with IFNT to determine if IFNT alone could induce RTP4. IFNT increased RTP4 more than 70-fold at 1.5 h after treatment, with maximal induction of nearly 300-fold above values observed in nontreated controls at 6 h after treatment. These results indicate that RTP4 mRNA levels are induced in the ovine endometrium, PBLs, and CL by IFNT during early pregnancy and in cell culture in response to IFNT. If RTP4 expression affects G protein-coupled receptor function, it may be important for establishment of pregnancy in domestic ruminants.

Results of laparoscopic reoperation for failed antireflux surgery: does the indication for redo surgery affect the outcome?

Short and medium term outcomes from laparoscopic antireflux surgery are generally excellent. A small number of patients suffer recurrent reflux or intolerable side-effects and may require reoperation. In this paper we describe our experience of 35 laparoscopic reoperations from a single center. Data on patients undergoing antireflux surgery in our unit has been prospectively collected and includes more than 600 primary laparoscopic antireflux operations since 1993. Laparoscopic reoperations have been performed between 1996 and 2005 for patients suffering recurrent reflux, dysphagia or severe gas bloat symptomatic despite medical treatment. All patients underwent preoperative barium studies and endoscopy with selective manometry and pH studies. Symptomatic outcomes were evaluated at 6 weeks and 12 months with Visick scores. Anatomical results were assessed with barium studies at between 6 and 12 months. Thirty-five laparoscopic reoperations were performed in 20 women and 13 men (median age 56 years). Primary surgery had been performed in our unit in 27 (77%) and elsewhere in eight (23%). Median time from primary surgery was 28.5 months (5-360). Two patients underwent a second reoperation. Indication was recurrent reflux in 28 (80%), dysphagia in five (14%) and gas bloat in two (6%). Thirty-two of the 35 reoperations (91.4%) were completed laparoscopically, median operating time was 120.5 min (65-210) and median hospital stay 2 days. There was no mortality and there were only five minor complications. Twelve-month follow-up was available for 32 reoperations (91%). Overall good symptomatic outcomes were obtained in 26 (74%) Visick I or II at 6 weeks and 24 of 32 (75%) at 12 months. In reoperations for dysphagia/gas bloat there was a relative risk of 4.26 of a poor symptomatic outcome (Visick III or IV) at 12 months compared to those for recurrent reflux (P < 0.05, Fisher's exact test). Laparoscopic reoperation is feasible with low conversion rates and minimal morbidity for patients who have undergone previous abdominal or thoracic hiatal repair. Symptomatic outcomes are generally good, particularly if the indication is recurrent reflux.

Serum thyroid hormones and performance of offspring in ewes receiving propylthiouracil with or without melatonin.

Two experiments were conducted during mid-gestation to examine effects in ewes of propylthiouracil (PTU) treatment alone or with melatonin on serum thyroid hormones, postpartum reproduction, and lamb performance. In the first experiment, beginning on day 0 (first day of treatment when all animals were 72.2+/-0.9 days of gestation), ewes received daily treatments (gavage) consisting of either 0mg (n=6) or 40 mg (n=6) PTU/kg body weight/day for 15 days. After 15 days, the 40 mg dosage was decreased to 20mg/kg body weight for an additional 20 days (35 days of PTU). Serum thyroxine (T4) did not differ (P>0.10) between groups through day 4; but on day 5, control females had a serum value of 67 ng/ml compared with 46 (+/-5)ng/ml for PTU-treated ewes (P=0.02). On the last day that 40 mg of PTU was administered, serum T4 averaged 67 and 7 (+/-5)ng/ml (P<0.001) in the two respective groups. Serum T4 remained low and was 80 and 1 ng/ml (P<0.001) in control and treated ewes on day 34. Serum T4 rose gradually after PTU but remained different from that observed in control ewes through day 48. Lambs from control and treated ewes had similar (P=0.46) T4 values at birth but lambs from PTU-treated ewes had lower (P=0.03) birth weights than did those from control ewes. Serum progesterone (P4) after parturition indicated a lack of cyclicity in all ewes. In the second experiment, beginning on day 0 (76.8+/-4.7 days of gestation), ewes received PTU as in Experiment 1. In addition, after 15 days of PTU, melatonin was given (i.m. injections at 5mg/day) for 30 days. Propylthiouracil decreased (P0.60) for lambs born to control and treated ewes. Female offspring of PTU+melatonin-treated dams reached puberty, became anestrus, and returned to cyclicity at similar (P>0.10) times to contemporary ewe lambs. Results indicate that 40/20mg PTU alone or with melatonin does not induce cyclicity after lambing in spring lambing ewes and has little effect on offspring performance.

Regulation of interferon-stimulated genes in peripheral blood leukocytes in pregnant and bred, nonpregnant dairy cows.

In ruminants, pregnancy results in up-regulation of a large number of IFN-stimulated genes (ISG) in the uterus. Recently, one of these genes was also shown to increase in peripheral blood leukocytes (PBL) during early pregnancy in sheep. Our working hypothesis is that conceptus signaling activates maternal gene expression in PBL in dairy cattle. The objectives of this study were to characterize ISG expression in PBL from pregnant (n = 20) and bred, nonpregnant (n = 30) dairy cows. Steady-state levels of mRNA for Mx1, Mx2, beta2-microglobulin, ISG-15, IFN regulatory factor-1, and IFN regulatory factor-2 were quantified. Holstein cows were synchronized to estrus and artificially inseminated (d 0). Blood samples were collected (coccygeal venipuncture) on d 0 and 16, 18, and 20 d after insemination for progesterone analysis and PBL isolation. Pregnancy was confirmed by transrectal ultrasonography at approximately 40 d after breeding. A status x day interaction was detected for Mx1, Mx2, and ISG-15 gene expression. When analyzed within day, levels of mRNA for ISG-15 and Mx1 were greater in pregnant compared with bred, nonpregnant cows on d 18 and 20, respectively. Expression of the Mx2 gene increased in the pregnant group compared with bred, nonpregnant cows on d 16, 18, and 20 after insemination. beta2-Microglobulin, IFN regulatory factor-1, and IFN regulatory factor-2 were not different between groups. The results clearly indicated that components of the innate immune response are activated in PBL during the period of pregnancy recognition and early embryo signaling. The physiological implications of these changes on maternal immune function are as yet unknown; however, they do provide a unique opportunity to identify bred, nonpregnant, cows 18 d after insemination in dairy cattle.

Enhancing patient safety through teamwork training.

The effective reduction of medical errors depends on an environment of safety for patients in both clinically-based and systems-oriented arenas. Formal teamwork training is proposed as a systems approach that will achieve these ends. In a study conducted by Dynamics Research Corporation, weaknesses and error patterns in Emergency Department teamwork were assessed, and a prospective evaluation of a formal teamwork training intervention was conducted. Improvements were obtained in five key teamwork measures, and most importantly, clinical errors were significantly reduced.

Evidence towards the role of arylamine N-acetyltransferase in Mycobacterium smegmatis and development of a specific antiserum against the homologous enzyme of Mycobacterium tuberculosis.

Arylamine N-acetyltransferase (NAT) in humans inactivates the anti-tubercular drug isoniazid (INH). Homologues of human NAT are present in Mycobacterium tuberculosis and Mycobacterium smegmatis, where they can acetylate, and hence inactivate, INH. The in vivo role of mycobacterial NAT is not known but heterologous expression of the M. tuberculosis gene increases the INH resistance. The 0.85 kb nat gene is part of a gene cluster in M. smegmatis. The gene is transcribed as a large, 7.5 kb mRNA as demonstrated by Northern analysis. A nat knockout strain of M. smegmatis was generated by targeted disruption. The new strain was confirmed to be devoid of NAT activity. The growth of the knockout strain is considerably delayed compared with the wild-type, due to an extended lag phase. The knockout mutant has an increased sensitivity to INH as would be predicted. The NATs from M. smegmatis and M. tuberculosis have a high degree of homology, except in the region of the C terminus. A specific polyclonal antiserum raised against recombinant NAT protein from M. tuberculosis is described that recognizes a stretch of about twenty residues within the C terminus of M. tuberculosis NAT. This highly specific antiserum will enable comparison of nat expression between isolates of M. tuberculosis.

Neural processing of gravito-inertial cues in humans. II. Influence of the semicircular canals during eccentric rotation.

All linear accelerometers, including the otolith organs, respond equivalently to gravity and linear acceleration. To investigate how the nervous system resolves this ambiguity, we measured perceived roll tilt and reflexive eye movements in humans in the dark using two different centrifugation motion paradigms (fixed radius and variable radius) combined with two different subject orientations (facing-motion and back-to-motion). In the fixed radius trials, the radius at which the subject was seated was held constant while the rotation speed was changed to yield changes in the centrifugal force. In variable radius trials, the rotation speed was held constant while the radius was varied to yield a centrifugal force that nearly duplicated that measured during the fixed radius condition. The total gravito-inertial force (GIF) measured by the otolith organs was nearly identical in the two paradigms; the primary difference was the presence (fixed radius) or absence (variable radius) of yaw rotational cues. We found that the yaw rotational cues had a large statistically significant effect on the time course of perceived tilt, demonstrating that yaw rotational cues contribute substantially to the neural processing of roll tilt. We also found that the orientation of the subject relative to the centripetal acceleration had a dramatic influence on the eye movements measured during fixed radius centrifugation. Specifically, the horizontal vestibuloocular reflex (VOR) measured in our human subjects was always greater when the subject faced the direction of motion than when the subjects had their backs toward the motion during fixed radius rotation. This difference was consistent with the presence of a horizontal translational VOR response induced by the centripetal acceleration. Most importantly, by comparing the perceptual tilt responses to the eye movement responses, we found that the translational VOR component decayed as the subjective tilt indication aligned with the tilt of the GIF. This was true for both the fixed radius and variable radius conditions even though the time course of the responses was significantly different for these two conditions. These findings are consistent with the hypothesis that the nervous system resolves the ambiguous measurements of GIF into neural estimates of gravity and linear acceleration. More generally, these findings are consistent with the hypothesis that the nervous system uses internal models to process and interpret sensory motor cues.