The effect of parity number on the metabolism, inflammation, and oxidative status of dairy sheep during the transition period.

The objective of this study was to evaluate whether dairy sheep during the transition period are affected by their parity numbers with regard to (1) body weight (BW), body condition score (BCS), and production performance (milk yield and composition) and (2) metabolic, inflammation, and stress biomarkers. For this purpose, 30 Sarda dairy ewes [15 primiparous (PRP) and 15 multiparous (MUP) ewes] were recruited on d 90 of gestation. Each group was homogeneous according to age, BW, and BCS. Sampling was carried out at -60, -30, -7, 0, +30, and +60 d from lambing. The MUP ewes showed a higher BW (46.32 vs. 38.71 kg) and larger litter size (1.45 vs. 1.06 kg) but a lower BCS (2.47 vs. 2.70) than the PRP ewes. Furthermore, the MUP ewes had lower concentrations of glucose (3.49 vs. 4.27 mol/L), cholesterol (1.63 vs. 1.81 mmol/L), free fatty acids (0.47 vs. 0.62 mmol/L), and triglycerides (0.22 vs. 0.25 mmol/L) compared with PRP ewes. With regard to inflammation and oxidative stress parameters, the PRP group had higher haptoglobin (0.48 vs. 0.18 g/L) and paraoxonase (187.90 vs. 152.11 U/L) activity than the MUP group. Overall, the MUP ewes were characterized by greater milk production performance and greater feed intake, resulting in a better energy balance, than the PRP ewes. Interestingly, these findings highlighted a different metabolic and inflammatory response over the transition period between PRP and MUP ewes, with the latter displaying lower concentrations of inflammatory-related biomarkers

Detection of buffalo milk adulteration with cow milk by capillary electrophoresis analysis.

The addition of cow milk during the production of buffalo mozzarella is a common fraud in dairy industries because of the lower price and greater availability of cow milk throughout the year. The aim of this study was to develop a new, rapid, and robust capillary electrophoresis method for detecting and quantifying cow milk in buffalo milk by exploiting cow α-lactalbumin as a marker of adulteration. In particular, a linear calibration curve was generated, using a training set of calibrators consisting of 7 series of 17 buffalo/bovine whey mixtures, obtained after casein precipitation, with increasing percentages of cow whey. The capillary electrophoresis method showed high linearity (R2 = 0.968), repeatability [relative standard deviation (RSD) = 2.11, 3.02, 4.38, and 1.18%, respectively for 5, 10, 20, and 50% of buffalo/bovine whey mixtures], and intermediate precision (RSD = 2.18, 2.49, 5.09, and 3.19%, respectively, for 5, 10, 20, and 50% buffalo/bovine whey mixtures). Moreover, the minimum amount of detectable fraudulent cow milk was 1%, and the limit of quantification was 3.1%

Methyl donor supply to heat stress-challenged polymorphonuclear leukocytes from lactating Holstein cows enhances 1-carbon metabolism, immune response, and cytoprotective gene network abundance

[EN] Mechanisms controlling immune function of dairy cows are dysregulated during heat stress (HS). Methyl donor supply-methionine (Met) and choline (Chop-positively modulates innate immune function, particularly antioxidant systems of polymorphonuclear leukocytes (PMN). The objective of this study was to investigate the effect of Met and Chol supply in vitro on mRNA abundance of genes related to 1-carbon metabolism, inflammation, and immune function in short-term cultures of PMN isolated from mid-lactating Holstein cows in response to heat challenge. Blood PMN were isolated from 5 Holstein cows (153 +/- 5 d postpartum, 34.63 +/- 2.73 kg/d of milk production; mean +/- SD). The PMN were incubated for 2 h at thermal-neutral (37 degrees C; TN) or heat stress (42 degrees C; HS) temperatures with 3 levels of Chol (0, 400, or 800 mu g/mL) or 3 ratios of Lys:Met (Met; 3.6:1, 2.9:1, or 2.4:1). Supernatant concentrations of IL-1 beta, IL-6, and tumor necrosis factor-alpha were measured via bovine-specific ELISA. Fold-changes in mRNA abundance were calculated separately for Chol and Met treatments to obtain the fold-change response at 42 degrees C (HS) relative to 37 degrees C (TN). Data were subjected to ANOVA using PROC MIXED in SAS (SAS Institute Inc., Cary, NC). Orthogonal contrasts were used to determine the linear or quadratic effect of Met and Chol for mRNA fold-change and supernatant cytokine concentrations. Compared with PMN receiving 0 mu g of Chol/mL, heat-stressed PMN supplemented with Chol at 400 or 800 mu g/mL had greater fold-change in abundance of CBS, CSAD, GSS, GSR, and GPX1. Among genes associated with inflammation and immune function, fold-change in abundance of TLR2, TLR4, IRAK1, IL1B, and IL10 increased with 400 and 800 mu g of Chol/mL compared with PMN receiving 0 mu g of Chol/mL. Fold-change in abundance of SAHH decreased linearly at increasing levels of Met supply. A linear effect was detected for MPO, NFKB1, and SOD1 due to greater fold-change in abundance when Met was increased to reach Lys: Met ratios of 2.9:1 and 2.4:1. Although increasing Chol supply upregulated BAX, BCL2, and HSP70, increased Met supply only upregulated BAX. Under HS conditions, enhancing PMN supply of Chol to 400 mu g/mL effectively increased fold-change in abundance of genes involved in antioxidant production (conferring cellular processes protection from free radicals and reactive oxygen species), inflammatory signaling, and innate immunity. Although similar outcomes were obtained with Met supply at Lys:Met ratios of 2.9:1 and 2.4:1, the response was less pronounced. Both Chol and Met supply enhanced the cytoprotective characteristics of PMN through upregulation of heat shock proteins. Overall, the modulatory effects detected in the present experiment highlight an opportunity to use Met and particularly Chol supplementation during thermal stress.M. Vailati-Riboni was supported in part by Hatch funds under project ILLU-538-914, National Institute of Food and Agriculture (Washington, DC). The authors declare no conflict of interest.Lopreiato, V.; Vailati-Riboni, M.; Parys, C.; Fernández Martínez, CJ.; Minuti, A.; Loor, J. (2020). Methyl donor supply to heat stress-challenged polymorphonuclear leukocytes from lactating Holstein cows enhances 1-carbon metabolism, immune response, and cytoprotective gene network abundance. Journal of Dairy Science. 103(11):10477-10493. https://doi.org/10.3168/jds.2020-18638S104771049310311Abdelmegeid, M. K., Vailati-Riboni, M., Alharthi, A., Batistel, F., & Loor, J. J. (2017). Supplemental methionine, choline, or taurine alter in vitro gene network expression of polymorphonuclear leukocytes from neonatal Holstein calves. Journal of Dairy Science, 100(4), 3155-3165. doi:10.3168/jds.2016-12025Armentano, L. E., Bertics, S. J., & Ducharme, G. A. (1997). Response of Lactating Cows to Methionine or Methionine Plus Lysine Added to High Protein Diets Based on Alfalfa and Heated Soybeans. Journal of Dairy Science, 80(6), 1194-1199. doi:10.3168/jds.s0022-0302(97)76047-8Banerjee, R., Evande, R., Kabil, Ö., Ojha, S., & Taoka, S. (2003). Reaction mechanism and regulation of cystathionine β-synthase. Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, 1647(1-2), 30-35. doi:10.1016/s1570-9639(03)00044-xBatistel, F., Arroyo, J. M., Bellingeri, A., Wang, L., Saremi, B., Parys, C., … Loor, J. J. (2017). Ethyl-cellulose rumen-protected methionine enhances performance during the periparturient period and early lactation in Holstein dairy cows. Journal of Dairy Science, 100(9), 7455-7467. doi:10.3168/jds.2017-12689Baumgard, L. H., & Rhoads, R. P. (2013). Effects of Heat Stress on Postabsorptive Metabolism and Energetics. Annual Review of Animal Biosciences, 1(1), 311-337. doi:10.1146/annurev-animal-031412-103644Bernabucci, U., Biffani, S., Buggiotti, L., Vitali, A., Lacetera, N., & Nardone, A. (2014). The effects of heat stress in Italian Holstein dairy cattle. Journal of Dairy Science, 97(1), 471-486. doi:10.3168/jds.2013-6611Bernabucci, U., Lacetera, N., Baumgard, L. H., Rhoads, R. P., Ronchi, B., & Nardone, A. (2010). Metabolic and hormonal acclimation to heat stress in domesticated ruminants. Animal, 4(7), 1167-1183. doi:10.1017/s175173111000090xBoldyrev, A., Bryushkova, E., Mashkina, A., & Vladychenskaya, E. (2013). Why Is Homocysteine Toxic for the Nervous and Immune Systems? Current Aging Science, 6(1), 29-36. doi:10.2174/18746098112059990007Catozzi, C., Ávila, G., Zamarian, V., Pravettoni, D., Sala, G., Ceciliani, F., … Lecchi, C. (2020). In-vitro effect of heat stress on bovine monocytes lifespan and polarization. Immunobiology, 225(2), 151888. doi:10.1016/j.imbio.2019.11.023Chinenov, Y., Gupte, R., & Rogatsky, I. (2013). Nuclear receptors in inflammation control: Repression by GR and beyond. Molecular and Cellular Endocrinology, 380(1-2), 55-64. doi:10.1016/j.mce.2013.04.006Chorąży, M., Kontny, E., Marcinkiewicz, J., & Maśliński, W. (2002). Amino Acids, 23(4), 407-413. doi:10.1007/s00726-002-0204-0Coleman, D. N., Lopreiato, V., Alharthi, A., & Loor, J. J. (2020). Amino acids and the regulation of oxidative stress and immune function in dairy cattle. Journal of Animal Science, 98(Supplement_1), S175-S193. doi:10.1093/jas/skaa138Collier, R. J., Stiening, C. M., Pollard, B. C., VanBaale, M. J., Baumgard, L. H., Gentry, P. C., & Coussens, P. M. (2006). Use of gene expression microarrays for evaluating environmental stress tolerance at the cellular level in cattle1. Journal of Animal Science, 84(suppl_13), E1-E13. doi:10.2527/2006.8413_supple1xCouper, K. N., Blount, D. G., & Riley, E. M. (2008). IL-10: The Master Regulator of Immunity to Infection. The Journal of Immunology, 180(9), 5771-5777. doi:10.4049/jimmunol.180.9.5771Del Vesco, A. P., Gasparino, E., Grieser, D. de O., Zancanela, V., Soares, M. A. M., & de Oliveira Neto, A. R. (2015). Effects of methionine supplementation on the expression of oxidative stress-related genes in acute heat stress-exposed broilers. British Journal of Nutrition, 113(4), 549-559. doi:10.1017/s0007114514003535Ekremoğlu, M., Türközkan, N., Erdamar, H., Kurt, Y., & Yaman, H. (2006). Protective effect of taurine on respiratory burst activity of polymorphonuclear leukocytes in endotoxemia. Amino Acids, 32(3), 413-417. doi:10.1007/s00726-006-0382-2El-Benna, J., Hurtado-Nedelec, M., Marzaioli, V., Marie, J.-C., Gougerot-Pocidalo, M.-A., & Dang, P. M.-C. (2016). Priming of the neutrophil respiratory burst: role in host defense and inflammation. Immunological Reviews, 273(1), 180-193. doi:10.1111/imr.12447Esposito, G., Irons, P. C., Webb, E. C., & Chapwanya, A. (2014). Interactions between negative energy balance, metabolic diseases, uterine health and immune response in transition dairy cows. Animal Reproduction Science, 144(3-4), 60-71. doi:10.1016/j.anireprosci.2013.11.007Fear, J. M., & Hansen, P. J. (2011). Developmental Changes in Expression of Genes Involved in Regulation of Apoptosis in the Bovine Preimplantation Embryo1. Biology of Reproduction, 84(1), 43-51. doi:10.1095/biolreprod.110.086249Gao, S. T., Guo, J., Quan, S. Y., Nan, X. M., Fernandez, M. V. S., Baumgard, L. H., & Bu, D. P. (2017). The effects of heat stress on protein metabolism in lactating Holstein cows. Journal of Dairy Science, 100(6), 5040-5049. doi:10.3168/jds.2016-11913Han, Z.-Y., Mu, T., & Yang, Z. (2014). Methionine protects against hyperthermia-induced cell injury in cultured bovine mammary epithelial cells. Cell Stress and Chaperones, 20(1), 109-120. doi:10.1007/s12192-014-0530-7Heiser, A., LeBlanc, S. J., & McDougall, S. (2018). Pegbovigrastim treatment affects gene expression in neutrophils of pasture-fed, periparturient cows. Journal of Dairy Science, 101(9), 8194-8207. doi:10.3168/jds.2017-14129Horowitz, M. (2001). Heat acclimation: phenotypic plasticity and cues to the underlying molecular mechanisms. Journal of Thermal Biology, 26(4-5), 357-363. doi:10.1016/s0306-4565(01)00044-4Hunter-Lavin, C., Davies, E. L., Bacelar, M. M. F. V. G., Marshall, M. J., Andrew, S. M., & Williams, J. H. H. (2004). Hsp70 release from peripheral blood mononuclear cells. Biochemical and Biophysical Research Communications, 324(2), 511-517. doi:10.1016/j.bbrc.2004.09.075Ingvartsen, K. L., & Moyes, K. (2013). Nutrition, immune function and health of dairy cattle. Animal, 7, 112-122. doi:10.1017/s175173111200170xJoshi, B. C., Joshi, H. B., McDowell, R. E., & Sadhu, D. P. (1968). Composition of Skin Secretions from Three Indian Breeds of Cattle Under Thermal Stress. Journal of Dairy Science, 51(6), 917-920. doi:10.3168/jds.s0022-0302(68)87105-xKobayashi, S. D., & DeLeo, F. R. (2009). Role of neutrophils in innate immunity: a systems biology‐level approach. Wiley Interdisciplinary Reviews: Systems Biology and Medicine, 1(3), 309-333. doi:10.1002/wsbm.32Kumar, H., Kawai, T., & Akira, S. (2011). Pathogen Recognition by the Innate Immune System. International Reviews of Immunology, 30(1), 16-34. doi:10.3109/08830185.2010.529976Lacetera, N., Bernabucci, U., Basiricò, L., Morera, P., & Nardone, A. (2009). Heat shock impairs DNA synthesis and down-regulates gene expression for leptin and Ob-Rb receptor in concanavalin A-stimulated bovine peripheral blood mononuclear cells. Veterinary Immunology and Immunopathology, 127(1-2), 190-194. doi:10.1016/j.vetimm.2008.09.020Lacetera, N., Bernabucci, U., Scalia, D., Basiricò, L., Morera, P., & Nardone, A. (2006). Heat Stress Elicits Different Responses in Peripheral Blood Mononuclear Cells from Brown Swiss and Holstein Cows. Journal of Dairy Science, 89(12), 4606-4612. doi:10.3168/jds.s0022-0302(06)72510-3Lecchi, C., Rota, N., Vitali, A., Ceciliani, F., & Lacetera, N. (2016). In vitro assessment of the effects of temperature on phagocytosis, reactive oxygen species production and apoptosis in bovine polymorphonuclear cells. Veterinary Immunology and Immunopathology, 182, 89-94. doi:10.1016/j.vetimm.2016.10.007Loos, H., Roos, D., Weening, R., & Houwerzijl, J. (1976). Familial deficiency of glutathione reductase in human blood cells. Blood, 48(1), 53-62. doi:10.1182/blood.v48.1.53.53Lopreiato, V., Vailati-Riboni, M., Bellingeri, A., Khan, I., Farina, G., Parys, C., & Loor, J. J. (2019). Inflammation and oxidative stress transcription profiles due to in vitro supply of methionine with or without choline in unstimulated blood polymorphonuclear leukocytes from lactating Holstein cows. Journal of Dairy Science, 102(11), 10395-10410. doi:10.3168/jds.2019-16413Lubos, E., Loscalzo, J., & Handy, D. E. (2011). Glutathione Peroxidase-1 in Health and Disease: From Molecular Mechanisms to Therapeutic Opportunities. Antioxidants & Redox Signaling, 15(7), 1957-1997. doi:10.1089/ars.2010.3586Lushchak, V. I. (2012). Glutathione Homeostasis and Functions: Potential Targets for Medical Interventions. Journal of Amino Acids, 2012, 1-26. doi:10.1155/2012/736837McGuire, M. A., Beede, D. K., DeLorenzo, M. A., Wilcox, C. J., Huntington, G. B., Reynolds, C. K., & Collier, R. J. (1989). Effects of Thermal Stress and Level of Feed Intake on Portal Plasma Flow and Net Fluxes of Metabolites in Lactating Holstein Cows2,3. Journal of Animal Science, 67(4), 1050-1060. doi:10.2527/jas1989.6741050xMin, L., Zheng, N., Zhao, S., Cheng, J., Yang, Y., Zhang, Y., … Wang, J. (2016). Long-term heat stress induces the inflammatory response in dairy cows revealed by plasma proteome analysis. Biochemical and Biophysical Research Communications, 471(2), 296-302. doi:10.1016/j.bbrc.2016.01.185Moyes, K. M., Drackley, J. K., Morin, D. E., & Loor, J. J. (2010). Greater expression of TLR2, TLR4, and IL6 due to negative energy balance is associated with lower expression of HLA-DRA and HLA-A in bovine blood neutrophils after intramammary mastitis challenge with Streptococcus uberis. Functional & Integrative Genomics, 10(1), 53-61. doi:10.1007/s10142-009-0154-7Moyes, K. M., Graugnard, D. E., Khan, M. J., Mukesh, M., & Loor, J. J. (2014). Postpartal immunometabolic gene network expression and function in blood neutrophils are altered in response to prepartal energy intake and postpartal intramammary inflammatory challenge. Journal of Dairy Science, 97(4), 2165-2177. doi:10.3168/jds.2013-7433Nakamura, M. (2000). Preconditioning decreases Bax expression, PMN accumulation and apoptosis in reperfused rat heart. Cardiovascular Research, 45(3), 661-670. doi:10.1016/s0008-6363(99)00393-4Oeckinghaus, A., & Ghosh, S. (2009). The NF- B Family of Transcription Factors and Its Regulation. Cold Spring Harbor Perspectives in Biology, 1(4), a000034-a000034. doi:10.1101/cshperspect.a000034Osorio, J. S., Ji, P., Drackley, J. K., Luchini, D., & Loor, J. J. (2014). Smartamine M and MetaSmart supplementation during the peripartal period alter hepatic expression of gene networks in 1-carbon metabolism, inflammation, oxidative stress, and the growth hormone–insulin-like growth factor 1 axis pathways. Journal of Dairy Science, 97(12), 7451-7464. doi:10.3168/jds.2014-8680Salama, A. A. K., Duque, M., Wang, L., Shahzad, K., Olivera, M., & Loor, J. J. (2019). Enhanced supply of methionine or arginine alters mechanistic target of rapamycin signaling proteins, messenger RNA, and microRNA abundance in heat-stressed bovine mammary epithelial cells in vitro. Journal of Dairy Science, 102(3), 2469-2480. doi:10.3168/jds.2018-15219Schell, M. T., Spitzer, A. L., Johnson, J. A., Lee, D., & Harris, H. W. (2005). Heat Shock Inhibits NF-kB Activation in a Dose- and Time-Dependent Manner. Journal of Surgical Research, 129(1), 90-93. doi:10.1016/j.jss.2005.05.025Silanikove, N. (2000). Effects of heat stress on the welfare of extensively managed domestic ruminants. Livestock Production Science, 67(1-2), 1-18. doi:10.1016/s0301-6226(00)00162-7Stankiewicz, A. R., Lachapelle, G., Foo, C. P. Z., Radicioni, S. M., & Mosser, D. D. (2005). Hsp70 Inhibits Heat-induced Apoptosis Upstream of Mitochondria by Preventing Bax Translocation. Journal of Biological Chemistry, 280(46), 38729-38739. doi:10.1074/jbc.m509497200Steel, G. J., Fullerton, D. M., Tyson, J. R., & Stirling, C. J. (2004). Coordinated Activation of Hsp70 Chaperones. Science, 303(5654), 98-101. doi:10.1126/science.1092287Sun, D., Chen, D., Du, B., & Pan, J. (2005). Heat Shock Response Inhibits NF-κB Activation and Cytokine Production in Murine Kupffer Cells. Journal of Surgical Research, 129(1), 114-121. doi:10.1016/j.jss.2005.05.028Taraktsoglou, M., Szalabska, U., Magee, D. A., Browne, J. A., Sweeney, T., Gormley, E., & MacHugh, D. E. (2011). Transcriptional profiling of immune genes in bovine monocyte-derived macrophages exposed to bacterial antigens. Veterinary Immunology and Immunopathology, 140(1-2), 130-139. doi:10.1016/j.vetimm.2010.12.002Trevisi, E., Jahan, N., Bertoni, G., Ferrari, A., & Minuti, A. (2015). Pro-Inflammatory Cytokine Profile in Dairy Cows: Consequences for New Lactation. Italian Journal of Animal Science, 14(3), 3862. doi:10.4081/ijas.2015.3862Tsan, M.-F., & Gao, B. (2004). Cytokine function of heat shock proteins. American Journal of Physiology-Cell Physiology, 286(4), C739-C744. doi:10.1152/ajpcell.00364.2003Vailati-Riboni, M., Zhou, Z., Jacometo, C. B., Minuti, A., Trevisi, E., Luchini, D. N., & Loor, J. J. (2017). Supplementation with rumen-protected methionine or choline during the transition period influences whole-blood immune response in periparturient dairy cows. Journal of Dairy Science, 100(5), 3958-3968. doi:10.3168/jds.2016-11812Yan, J., Meng, X., Wancket, L. M., Lintner, K., Nelin, L. D., Chen, B., … Liu, Y. (2012). Glutathione Reductase Facilitates Host Defense by Sustaining Phagocytic Oxidative Burst and Promoting the Development of Neutrophil Extracellular Traps. The Journal of Immunology, 188(5), 2316-2327. doi:10.4049/jimmunol.1102683Zhou, Z., Bulgari, O., Vailati-Riboni, M., Trevisi, E., Ballou, M. A., Cardoso, F. C., … Loor, J. J. (2016). Rumen-protected methionine compared with rumen-protected choline improves immunometabolic status in dairy cows during the peripartal period. Journal of Dairy Science, 99(11), 8956-8969. doi:10.3168/jds.2016-10986Zhou, Z., Ferdous, F., Montagner, P., Luchini, D. N., Corrêa, M. N., & Loor, J. J. (2018). Methionine and choline supply during the peripartal period alter polymorphonuclear leukocyte immune response and immunometabolic gene expression in Holstein cows. Journal of Dairy Science, 101(11), 10374-10382. doi:10.3168/jds.2018-14972Zhou, Z., Vailati-Riboni, M., Trevisi, E., Drackley, J. K., Luchini, D. N., & Loor, J. J. (2016). Better postpartal performance in dairy cows supplemented with rumen-protected methionine compared with choline during the peripartal period. Journal of Dairy Science, 99(11), 8716-8732. doi:10.3168/jds.2015-1052

Evaluation of the capillary electrophoresis method for measurement of immunoglobulin concentration in ewe colostrum

ABSTRACT Capillary electrophoresis (CE) is a technique routinely used in clinical laboratories that allows the separation and quantification of blood serum proteins in a rapid, precise, accurate, and inexpensive manner. Recently, CE has been proposed to separate and measure colostral proteins, but an evaluation of the agreement between CE and radial immunodiffusion (RID) method, currently used to quantify IgG in colostrum, is still lacking. The purpose of this study was to test the ability of a CE instrument, normally used in blood serum protein analysis, to realize the correct quantification of total Ig concentration in ewe colostrum, using RID assay as reference. Colostrum samples (n = 68) were collected from 35 multiparous Sarda ewes at first milking (n = 33) and at 24 h postpartum (n = 35). The mean ± standard deviation of IgG concentration measured by RID and whey colostrum total Ig concentration measured by CE were 54.76 ± 41.82 g/L and 54.70 ± 41.43 g/L, respectively. Lin's concordance correlation coefficient (r = 0.993; 95% confidence interval=0.989 to 0.996) and linear regression analysis results (RID = 1.0022CE − 0.063; R 2 = 0.986) showed an excellent agreement between these 2 methods. Bland-Altman analysis confirmed that CE method can be a suitable alternative to RID: the mean of the differences between CE and RID was −0.055 ± 4.95 g/L (95% confidence interval=−1.25 to 1.14 g/L) and the agreement limits were −9.75 to 9.60 g/L (low limit 95% confidence interval=−11.82 to −7.68 g/L; high limit 95% confidence interval=7.57 to 11.72 g/L). In conclusion, the current study indicates that CE method may be a reliable tool for the quantification of the total Ig concentration in ewe colostrum

Effect of litter size on prepartum metabolic and amino acidic profile in rabbit does.

The use of modern prolific lines of rabbit does in intensive production systems leads to an increase in productivity but also causes a rise in several problems related to the does' health status. Hence, the aim of this study was to investigate the effect of the litter size on the metabolic, inflammatory and plasma amino acid profile in rabbit does. The blood of 30 pregnant does was sampled on the 27th day of pregnancy. The does were retrospectively grouped according to the number of offspring into a high litter size group (HI, does with ≥ 12 kits; n = 16) and a low litter size group (LO, does with ≤ 11 kits; n = 14). Data were subjected to Pearson's correlation analysis. Further, data were analysed in agreement to a completely randomized design in which the main tested effect was litter size. The linear or quadratic trends of litter size on parameters of interests were post hoc compared by using orthogonal contrasts. In addition, compared with the LO group, the HI group had lower levels of glucose (-5%; P < 0.01), zinc (-19%; P < 0.05), albumin (-6%; P < 0.05) and total cholesterol (-13%; P < 0.07), but the total bilirubin level was higher in the HI group (+14%; P < 0.05). Regarding the plasma amino acids, the HI group had lower concentrations of threonine (-15%), glycine (-16%), lysine (-16%) and tryptophan (-26%) and a higher level of glutamic acid (+43%; P < 0.05) compared with the LO group. The exclusively ketogenic amount of amino acids was lower (P < 0.06) in the HI (55.8 mg/100 ml) does compared with the LO does (56.8 mg/100 ml). These results show that a few days before delivery, rabbit does that gave birth to a higher number of offspring had a metabolic profile and an inflammatory status that was less favourable with respect to does who gave birth to a lower number of offspring. Moreover, the plasma amino acid profile points out that there was an enhanced catabolic condition in the rabbit does with a high number of gestated foetuses; it was likely related to the greater energy demand needed to support the pregnancy and an early inflammatory response

Hyaluronic acid reduces bacterial fouling and promotes fibroblasts’ adhesion onto chitosan 2D-wound dressings

Wound healing is a dynamic process that can be seriously delayed by many factors including infectious complications. The development of dressings with intrinsic wound healing activity and/or releasing bioactive compounds may help with addressing such an issue. In this study, hyaluronic acid (HA) at different percentages (1–35%) was used to modify chitosan (CS) biological and physico-chemical properties in order to obtain 2D-matrices able to promote healing and protect from infection. HA incorporation in the CS matrix decreased film transparency and homogeneity, but improved film water uptake and surface wettability. The water vapor transmission rate (WVTR) increased up to a 5% HA content, where it reached the highest value (672 g/m2 day), and decreased for higher HA contents. At all of the tested HA concentrations, HA affected mechanical properties providing matrices more flexible than pure CS with benefit for wound care. Pure CS films permitted S. epidermidis adhesion and biofilm formation. That was not true for CS/HA matrices, where HA at concentrations equal to or greater than 5% was able to avoid S. epidermidis adhesion. Fibroblasts adhesion also took benefit from the HA presence in the film, especially at 5% content, where the best adhesion and proliferation was found

Inflammation and oxidative stress transcription profiles due to in vitro supply of methionine with or without choline in unstimulated blood polymorphonuclear leukocytes from lactating Holstein cows.

Neutrophils are the most important polymorphonuclear leukocytes (PMNL), representing the front-line defense involved in pathogen clearance upon invasion. As such, they play a pivotal role in immune and inflammatory responses. Isolated PMNL from 5 mid-lactating Holstein dairy cows were used to evaluate the in vitro effect of methionine (Met) and choline (Chol) supplementation on mRNA expression of genes related to the Met cycle and innate immunity. The target genes are associated with the Met cycle, cell signaling, inflammation, antimicrobial and killing mechanisms, and pathogen recognition. Treatments were allocated in a 3 × 3 factorial arrangement, including 3 Lys-to-Met ratios (L:M, 3.6:1, 2.9:1, or 2.4:1) and 3 levels of supplemental Chol (0, 400, or 800 μg/mL). Three replicates per treatment group were incubated for 2 h at 37°C and 5% atmospheric CO2. Both betaine-homocysteine S-methyltransferase and choline dehydrogenase were undetectable, indicating that PMNL (at least in vitro) cannot generate Met from Chol through the betaine pathway. The PMNL incubated without Chol experienced a specific state of inflammatory mediation [greater interleukin-1β (IL1B), myeloperoxidase (MPO), IL10, and IL6] and oxidative stress [greater cysteine sulfinic acid decarboxylase (CSAD), cystathionine gamma-lyase (CTH), glutathione reductase (GSR), and glutathione synthase (GSS)]. However, data from the interaction L:M × Chol indicated that this negative state could be overcome by supplementing additional Met. This was reflected in the upregulation of methionine synthase (MTR) and toll-like receptor 2 (TLR2); that is, pathogen detection ability. At the lowest level of supplemental Chol, Met downregulated GSS, GSR, IL1B, and IL6, suggesting it could reduce cellular inflammation and enhance antioxidant status. At 400 µg/mL Chol, supplemental Met upregulated PMNL recognition capacity [higher TLR4 and L-selectin (SELL)]. Overall, enhancing the supply of methyl donors to isolated unstimulated PMNL from mid-lactating dairy cows leads to a low level of PMNL activation and upregulates a cytoprotective mechanism against oxidative stress. Enhancing the supply of Met coupled with adequate Chol levels enhances the gene expression of PMNL pathogen-recognition mechanism. These data suggest that Chol supply to PMNL exposed to low levels of Met effectively downregulated the entire repertoire of innate inflammatory-responsive genes. Thus, Met availability in PMNL during an inflammatory challenge may be sufficient for mounting an appropriate biologic response

Dietary energy level affects adipose depot mass but does not impair in vitro subcutaneous adipose tissue response to short-term insulin and tumor necrosis factor-α challenge in nonlactating, nonpregnant Holstein cows.

We assessed effects of overfeeding energy to nonlactating and nonpregnant Holstein cows during a length of time similar to a typical dry period on body lipid storage and the abundance of genes related to insulin signaling, inflammation, and ubiquitination in subcutaneous adipose tissue (SAT) in vitro challenged with insulin and recombinant bovine tumor necrosis factor-α. Fourteen cows were randomly assigned to either a high-energy (OVE; net energy for lactation = 1.60 Mcal/kg of dry matter; n = 7) or control (CON; net energy for lactation = 1.30 Mcal/kg of dry matter; n = 7) diet for 6 wk. Immediately after slaughter, liver, kidneys, and mammary gland were separated and weighed. The adipose tissue mass in the omental, mesenteric, and perirenal depots was dissected and weighed. Subcutaneous adipose tissue was collected from the tail-head region and was used as follows: control, bovine insulin (INS) at 1 µmol/L, tumor necrosis factor-α at 5 ng/mL (TNF), and their combination. Despite a lack of difference in final body condition score, OVE cows had greater energy intake and were heavier than CON cows. Furthermore, overfeeding led to greater mass of mesenteric and perirenal adipose, liver, and mammary gland. Overall, SAT incubated with INS had an upregulation of insulin receptor (INSR), interleukin-10 (IL10), small ubiquitin-like modifier 3 (SUMO3), and ubiquitin conjugating enzyme E2I (UBC9), whereas TNF upregulated peroxisome proliferator-activated receptor gamma (PPARG), diacylglycerol O-acyltransferase 2 (DGAT2), interleukin-6 (IL6), nuclear factor kappa B subunit 1 (NFKB1), small ubiquitin-like modifier 2 (SUMO2), and UBC9. Regardless of in vitro treatment, feeding OVE upregulated PPARG, fatty acid synthase (FASN), and insulin induced gene 1 (INSIG1). Abundance of PPARG was greater in SAT of OVE cows cultured individually with INS and TNF. The interaction between diet and in vitro treatment revealed that sterol regulatory element binding transcription factor 1 (SREBF1) had greater abundance in SAT from the CON group in response to culture with INS, whereas SAT from OVE cows had greater SREBF1 abundance in response to culture with TNF. The mRNA abundance of IL6 and NFKB1 was greater in response to TNF treatment and overall in CON cows. Furthermore, SAT from these cows had greater IL10 abundance when cultured with INS and TNF. Overall, data highlighted that overfeeding energy increases adipose tissue mass in part by stimulating transcription of key genes associated with insulin signaling, adipogenesis, and lipogenesis. Because SAT thickness or mass was not measured, the lack of effect of overfeeding on body condition score limits its use to predict overall body lipid storage. An overt inflammatory response in SAT after a 6-wk period of over-consumption of energy could not be discerned

The Role of Innate Immune Response and Microbiome in Resilience of Dairy Cattle to Disease: The Mastitis Model

Animal health is affected by many factors such as metabolic stress, the immune system, and epidemiological features that interconnect. The immune system has evolved along with the phylogenetic evolution as a highly refined sensing and response system, poised to react against diverse infectious and non-infectious stressors for better survival and adaptation. It is now known that high genetic merit for milk yield is correlated with a defective control of the inflammatory response, underlying the occurrence of several production diseases. This is evident in the mastitis model where high-yielding dairy cows show high disease prevalence of the mammary gland with reduced effectiveness of the innate immune system and poor control over the inflammatory response to microbial agents. There is growing evidence of epigenetic effects on innate immunity genes underlying the response to common microbial agents. The aforementioned agents, along with other non-infectious stressors, can give rise to abnormal activation of the innate immune system, underlying serious disease conditions, and affecting milk yield. Furthermore, the microbiome also plays a role in shaping immune functions and disease resistance as a whole. Accordingly, proper modulation of the microbiome can be pivotal to successful disease control strategies. These strategies can benefit from a fundamental re-appraisal of native cattle breeds as models of disease resistance based on successful coping of both infectious and non-infectious stressors

Technical note: Capillary electrophoresis as a rapid test for the quantification of immunoglobulin G in serum of newborn lambs

ABSTRACT Finding a rapid and simple method of serum IgG determination in lambs is essential for monitoring failure of passive transfer of immunity. The aim of this study was to assess the ability of capillary electrophoresis (CE), an instrument mainly used in blood serum protein analysis, to estimate IgG content in serum of newborn lambs through determination of only total Ig percentage by comparing the results with those obtained with radial immunodiffusion (RID), the reference method for serum IgG quantification. Serum samples were collected at 24 h after birth from 40 Sarda lambs. The IgG concentration measured by RID and serum total Ig concentration measured by CE were (mean ± standard deviation) 29.8 ± 16.1 g/L and 37.8 ± 15.0%, respectively. Data provided by RID and CE analysis showed a polynomial trend (RID = 0.02CE2 − 0.04CE + 4.13; coefficient of determination, R2 = 0.96), displaying a strong relationship between these 2 methods. Applying the polynomial equation, the IgG values were predicted. Predicted IgG values were highly correlated (r = 0.98) and related (R2 = 0.96) to IgG values obtained by RID assay. These data were subjected to Bland–Altman analysis, revealing a high level of agreement between CE and RID methods with a bias that was not different from 0 (−0.04 g/L) and agreement limits of −6.38 g/L (low) and +6.30 g/L (high). In addition, the linear regression analysis between differences (dependent variable) and average of IgG concentration by CE and RID (independent variable) did not show proportional bias (R2 = 0.01). In conclusion, CE is a reliable instrument for a lamb health monitoring program, where Bland–Altman analysis also confirmed that the CE method can be a suitable alternative to RID