Crookenden M A, Heiser A, Murray A, Dukkipati V S R, Kay J K, Loor J J, Meier S, Mitchell M D, Moyes K M, Walker C G, Roche J R
DairyNZ, c/o University of Auckland, Auckland 1010, New Zealand; Institute of Vet, Animal and Biomedical Sciences, Massey University, Palmerston North 4442, New Zealand.
Institute of Vet, Animal and Biomedical Sciences, Massey University, Palmerston North 4442, New Zealand; AgResearch, Hopkirk Research Institute, Palmerston North 4442, New Zealand.
J Dairy Sci. 2016 Aug;99(8):6470-6483. doi: 10.3168/jds.2015-10877. Epub 2016 May 11.
Extensive metabolic and physiologic changes occur during the peripartum, concurrent with a high incidence of infectious disease. Immune dysfunction is a likely contributor to the increased risk of disease at this time. Studies using high-yielding, total mixed ration-fed cows have indicated that neutrophil function is perturbed over the transition period; however, this reported dysfunction has yet to be investigated in moderate-yielding, grazing dairy cows. Therefore, we investigated changes in the expression of genes involved in neutrophil function. Blood was collected from cows at 5 time points over the transition period: precalving (-1wk; n=46), day of calving (d 0; n=46), and postcalving at wk 1 (n=46), wk 2 (n=45), and wk 4 (n=43). Neutrophils were isolated by differential centrifugation and gene expression was investigated. Quantitative reverse transcriptase PCR with custom-designed primer pairs and Roche Universal Probe Library (Roche, Basel, Switzerland) chemistry, combined with microfluidics integrated fluidic circuit chips (96.96 Dynamic Array, San Francisco, CA) were used to investigate the expression of 78 genes involved in neutrophil function and 18 endogenous control genes. Statistical significance between time points was determined using a repeated measures ANOVA. Genes that were differentially expressed over the transition period included those involved in neutrophil adhesion (SELL, ITGB2, and ITGBX), mediation of the immune response (TLR4, HLA-DRA, and CXCR2), maturation, cell cycle progression, apoptosis (MCL1, BCL2, FASLG, and RIPK1), and control of gene expression (PPARG, PPARD, and STAT3). We noted reduced gene expression of proinflammatory cytokines (IFNG, TNF, IL12, and CCL2) on the day of calving, whereas anti-inflammatory cytokine gene expression (IL10) was upregulated. Increased gene expression of antimicrobial peptides (BNBD4, DEFB10, and DEFB1) occurred on the day of calving. Collectively, transcription profiles are indicative of functional changes in neutrophils of grazing dairy cows over the transition period and align with studies in cows of conventional total mixed ration systems. This altered function may predispose cows to disease over the transition period and is likely to be a natural change in function due to parturition.
围产期会发生广泛的代谢和生理变化，同时传染病发病率很高。免疫功能障碍可能是此时疾病风险增加的一个因素。对采用高产全混合日粮喂养奶牛的研究表明，过渡期中性粒细胞功能受到干扰；然而，这种报道的功能障碍尚未在中等产量的放牧奶牛中进行研究。因此，我们研究了参与中性粒细胞功能的基因表达变化。在过渡期的5个时间点采集奶牛血液：产犊前（-1周；n = 46）、产犊日（第0天；n = 46）以及产后第1周（n = 46）、第2周（n = 45）和第4周（n = 43）。通过差速离心分离中性粒细胞并研究基因表达。使用定制设计的引物对和罗氏通用探针库（罗氏，巴塞尔，瑞士）化学方法，结合微流控集成流体电路芯片（96.96动态阵列，旧金山，加利福尼亚州），研究78个参与中性粒细胞功能的基因和18个内参基因的表达。使用重复测量方差分析确定时间点之间的统计学显著性。在过渡期差异表达的基因包括参与中性粒细胞黏附（SELL、ITGB2和ITGBX）、免疫反应介导（TLR4、HLA - DRA和CXCR2）、成熟、细胞周期进程、凋亡（MCL1、BCL2、FASLG和RIPK1）以及基因表达调控（PPARG、PPARD和STAT3）的基因。我们注意到产犊日促炎细胞因子（IFNG、TNF、IL12和CCL2）的基因表达降低，而抗炎细胞因子基因表达（IL10）上调。产犊日抗菌肽（BNBD4、DEFB10和DEFB1）的基因表达增加。总体而言，转录谱表明放牧奶牛在过渡期中性粒细胞的功能发生了变化，并且与传统全混合日粮系统奶牛的研究结果一致。这种功能改变可能使奶牛在过渡期易患疾病，并且很可能是由于分娩导致的功能自然变化。
