The biological cycles of milk production and reproduction determine dairying profitability thus making management decisions dynamic and time-dependent. Diseases also negatively impact on net earnings of a dairy enterprise. Transition cows in particular face the challenge of negative energy balance (NEB) and/or disproportional energy metabolism (fatty liver, ketosis, subacute, acute ruminal acidosis); disturbed mineral utilization (milk fever, sub-clinical hypocalcemia); and perturbed immune function (retained placenta, metritis, mastitis). Consequently NEB and reduced dry matter intake are aggravated.The combined effects of all these challenges are reduced fertility and milk production resulting in diminishing profits. Risk factors such as NEB, inflammation and impairment of the immune response are highly cause-and-effect related. Thus, managing cows during the transition period should be geared towards reducing NEB or feeding specially formulated diets to improve immunity. Given that all cows experience a reduced feed intake and body condition, infection and inflammation of the uterus after calving, there is a need for further research on the immunology of transition dairy cows. Integrative 2 approaches at the molecular, cellular and animal level may unravel the complex interactions between disturbed metabolism and immune function that predispose cows to periparturient diseases.
After calving, the bovine endometrium undergoes marked morphological and functional changes that are necessary for subsequent re-breeding. Regulation and integration of these key events are largely uncharacterised. Here, endometrial swabs and biopsies were taken at 15, 30 and 60 days postpartum (DPP) from 13 healthy primiparous cows, 10 of which subsequently conceived, with a view to characterising innate and inflammatory gene expression profiles. Endometrial biopsies exhibited severe inflammation (>75 leukocytes per high-power field) at 15 DPP, which had begun to resolve by 30 DPP and had completely resolved by 60 DPP. The severe inflammation at 15 DPP coincided with uterine infection in all cows and a significant increase (P < 0.05) in the expression of all of 16 genes investigated, including CD45, IL8, IL6, IL1, TNF, TAP, SAA3 and HP at 15 DPP, relative to 60 DPP. All of these parameters had begun to return to normal physiological levels at 30 DPP. Systemically, serum protein concentrations of IL-8 were elevated at 15 DPP compared with 60 DPP (78 pg mL–1 vs 48 pg mL–1; P = 0.02). These results indicate that endometrial inflammation, leukocyte infiltration and increased expression of pro-inflammatory, antimicrobial and acute-phase protein genes are expected features of the postpartum period, critical to bacterial clearance and uterine involution.
The anti-microbial peptides beta-defensins constitute a large family of innate immune effector molecules, conserved across a wide species range. In this paper, we describe a systematic search of the sequenced bovine genome to characterise this extensive gene family in Bos taurus, providing an insight into the pattern of conservation of beta-defensin genes between species. We have sequenced a sub-set of these newly discovered bovine beta-defensin genes and also report expression data for these genes across a range of tissues. We have synthesised the peptide product of one of these genes, bovine beta-defensin 123, and found it to be a potent inhibitor of several pathogenic microbes, particularly Escherichia coli and Listeria monocytogenes.
BackgroundThe regulation of endometrial inflammation has important consequences for the resumption of bovine fertility postpartum. All cows experience bacterial influx into the uterus after calving; however a significant proportion fail to clear infection leading to the development of cytological endometritis (CE) and compromised fertility. We hypothesised that early immunological changes could not only act as potential prognostic biomarkers for the subsequent development of disease but also shed light on the pathogenesis of endometritis in the postpartum dairy cow.MethodsEndometrial biopsy RNA was extracted from 15 cows at 7 and 21 days postpartum (DPP), using the Qiagen RNeasy® Plus Mini kit and quality determined using an Agilent 2100 bioanalyser. Disease status was determined by histpathology based on inflammatory cell infiltrate. RNA-seq of both mRNA and miRNA libraries were performed on an Illumina® HiSeq™ 2000. Paired reads were aligned to the bovine genome with Bowtie2 and differentially expressed genes were identified using EdgeR. Significantly over-represented Gene Ontology terms were identified using GO-seq, and pathway analysis was performed using KEGG. Quanititative real-time PCR was also performed for validation (ABI 7500 fast). Haematology was assessed using an automated ADVIA 2120 analyser. Serum proteins were evaluated by ELISA and metabolite analysis was performed using a Beckman Coulter AU 400 clinical analyser. Terminal-restriction fragment length polymorphism (T-RFLP) was used to obtain fingerprints of the microbial communities present.ResultsNext-generation sequencing from endometrial biopsies taken at 7 DPP identified significant induction of inflammatory gene expression in all cows. Despite the common inflammatory profile and enrichment of the Toll-like receptor and NFκB pathways, 73 genes and 31 miRNAs were significantly differentially expressed between healthy cows (HC, n = 9) and cows which subsequently developed CE at 7 DPP (n = 6, FDR < 0.1). While significant differential expression of 4197 genes in the transcriptome of healthy cows between 7 and 21 DPP showed the transition from a proinflammatory to tissue profliferation and repair, only 31 genes were differentially expressed in cows with CE (FDR < 0.1), indicating the arrest of such a transition. A link betwene the dysregulated inflammatory response and the composition of the uterine microbial communities was suggested by the presence of significant differences in uterine bacterial tRFLP profiles between HC and CE groups. Furthermore, inflammatory activity was not confined to the uterus; decreased circulating granulocytes and increased Acute Phase Protein (SAA and HP) expression levels were detected in plasma at 7 DPP in cows that developed CE.ConclusionOur data suggests that the IL1 and IL17 inflammatory cascade activated early postpartum is resolved thereby restoring homeostasis in healthy cows by 21 DPP, but this transition fails to occur in cows which develop CE. Despite a common early inflammatory profile, elevated and differe...
β-defensins are small cationic peptides, with potent immunoregulatory and antimicrobial activity which are produced constitutively and inducibly by eukaryotic cells. This study profiles the expression of a cluster of 19 novel defensin genes which spans 320 kb on chromosome 13 in Bos taurus. It also assesses the genetic variation in these genes between two divergently selected cattle breeds. Using quantitative real-time PCR (qRT-PCR), all 19 genes in this cluster were shown to be expressed in the male genital tract and 9 in the female genital tract, in a region-specific manner. These genes were sequenced in Norwegian Red (NR) and Holstein-Friesian (HF) cattle for population genetic analysis. Of the 17 novel single nucleotide polymorphisms (SNPs) identified, 7 were non-synonymous, 6 synonymous and 4 outside the protein coding region. Significant frequency differences in SNPs in bovine β-defensins (BBD) 115, 117, 121, and 122 were detected between the two breeds, which was also reflected at the haplotype level (P<0.05). There was clear segregation of the haplotypes into two blocks on chromosome 13 in both breeds, presumably due to historical recombination. This study documents genetic variation in this β-defensin gene cluster between Norwegian Red and Holstein-Friesian cattle which may result from divergent selection for production and fertility traits in these two breeds. Regional expression in the epididymis and fallopian tube suggests a potential reproductive-immunobiology role for these genes in cattle.
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