Browsing by Subject "immunity"

Now showing 1 - 5 of 5
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    Computational Methods for Sequencing and Interpreting Killer-cell Immunoglobulin-like Receptors (KIRs) at Multiple Resolutions
    (2021-01) Roe, David
    The era of personalized genetic medicine has arrived. It is now routine to sequence an individual’s DNA, either whole genome sequences (WGS) or targeted, as part of a patient’s medical plan. One of the exceptions is an ~200 kilobase region in chromosome 19 containing the killer-cell immunoglobulin-like receptors (KIR). These genes encode proteins that influence the actions of natural killer (NK) cells based on whether or not they bind with peptide-bound human leukocyte antigen receptors. This is evolutionary important to fight pathogens and mediate pregnancy. Modern medicine has correlated low-resolution KIR with many diseases and treatments, although the findings are often relatively vague and sometimes contradictory due to low resolution interpretation and/or small cohort sizes. Whether for personal medicine or population studies, the current best practices for KIR genotyping are to determine the presence/absence (PA) or copy number variation (CNV) of each gene using oligo- or primer-based polymerase chain reaction.The goal of our research is to advance DNA sequencing and interpretation of human KIR haplotypes. To that end, we have created algorithms and workflows to enhance interpretations of KIR at resolutions from PA genotyping via short-read WGS to full-haplotype assembly from long-read targeted or whole-genome sequences. First, we developed the first workflow to efficiently and accurately capture, sequence, assemble, and annotate full KIR haplotype sequences. As part of this workflow, we designed small sequences to capture the DNA fragments. Next, we use the alignment pattern of those short sequences across finished KIR haplotypes to define and annotate haplotype structures. The results show, for the first time, that the KIR region is composed of 9 genes in 14 loci. Next, we annotated all 68 reported human haplotypes, aligned them at the structural level, and then refined the alignment down the base level, providing the first KIR haplotype multiple sequence alignment. These efforts have led to this region being the best annotated and most diverse in the human genome reference. We next leveraged the MSA to discover PA markers and leveraged them in the first KIR WGS genotyping application. It was evaluated independently and reported to be at least 97% accurate. These discoveries and inventions are the culmination of several computational methods we have developed that interpret KIR under different typing resolutions. This multi-resolution aspect is crucial to overall understanding; it improves resolution at any given level by leveraging references and/or markers from other resolutions. From low-resolution genotyping from any kind of DNA sequence to the first efficient full-haplotype assembly method, these results advance interpretation of this important genetic region to the personal and population levels.
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    Effects of milk yield genotype on immune, endocrine and metabolite interactions in dairy cows
    (2018-08) Cousillas, Georgina
    Studies were designed to assess the impact of milk yield genotype on the innate immune response and its interactions with endocrine and metabolic components when growing heifers and lactating cows are challenged with lipopolysaccharide (LPS). A novel aspect of these studies was the comparison of unique Holsteins that have not been subjected to selection since 1964 (unselected Holsteins; UH) with contemporary Holsteins (CH). In addition, our animal models included heifers from contemporary Red-Black Angus cows (CA) and our experimental designs included the use of repeated LPS administration to assess the impact of genotype on the acute immune response and on the development of a refractory (endotoxin tolerance) state. Pro-inflammatory cytokine and glucose concentrations were greater and cortisol concentrations increased more rapidly in Holsteins than in Angus heifers which is consistent with results from our collaborators that indicate Holsteins have a more robust immune response than Angus cattle. Differences in plasma concentrations of pro-inflammatory cytokines, glucose and cortisol, and in expression of adhesion molecules and phagocytic activity of polymorphonuclear leukocytes (PMNL) after LPS administration indicates UH heifers and cows have a more robust immune response than CH heifers and cows. This was further supported by hepatic gene expression data, which indicated greater expression of genes in the TLR4 signaling pathway and of genes involved in the production of pro and anti-inflammatory mediators (IL6, TNF, IL1RN, TGFB1) in UH than in CH cows during the acute immune response. In addition, results during the second challenge indicated greater development of tolerance to immunotoxin stimulation in UH than CH cows. Overall, our results demonstrate that TLR4 signaling pathways have been altered by five decades of selective breeding and these changes contribute to a less robust and less controlled innate immune response in CH cows. Thus, immune activation and the ability to minimize negative effects of prolonged inflammation are reduced in the CH cow. Although we did not assess the impact of selection, we assessed functionality of several anti-ADAM17 antibodies and inhibitors and for, the first time, characterized the presence and activity of ADAM17 (a disintegrin and metalloproteinase-17) protein in bovine PMNL.
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    Effects of Peripartum Recombinant Bovine Somatotropin Treatment and Prepartum Stocking Density on Immune Responses, Metabolism, Health, and Performance of Dairy Cows
    (2016-10) Basso Silva, Paula Regina
    Excessive fatty acids release from the adipose tissue of periparturient cows may result in hyperketonemia and hepatic lipidosis, which are known to compromise liver and damage immune cells. Immunosuppression in transition cows is a result of shortage in energy, nutrients and calcium impairing immune cells’ metabolism. Thereafter, immunosuppressed periparturient cows are at higher risks for developing infectious and non-infectious health disorders. Strategies that improve metabolism and immune function of periparturient dairy cows may reduce incidence of diseases. Six experiments were performed to test two main strategies: the use of recombinant bovine somatotropin (rbST) during the peripartum period and the reduction of prepartum pens’ stocking density (SD) from 100 to 80% of headlocks. The specific objectives of these experiments were to evaluate the effects of treating peripartum dairy cows with rbST on immune, inflammatory, and metabolic responses, incidences of postpartum diseases, performance, and hepatic and leukocyte gene expression; and to evaluate the effects of two prepartum SD (80 vs. 100%) on milk yield, concentration of metabolites, health and reproductive parameters, innate and adaptive leukocyte responses, and serum and hair cortisol concentrations. Results demonstrated that treatment of dairy cows with 125 mg of rbST improved innate immune responses and IgG concentration, with limited effects on metabolism, decreased the incidence of uterine disorders in Holstein and Jersey cows and increased yield of energy corrected milk during the first 30 DIM in Holstein cows. Administration of rbST during the periparturient period may improve liver function and health by increasing hepatic capacity for gluconeogenesis and lipid transport and by reducing inflammation and oxidative stress. Treatment of dairy cows with 125 mg of rbST during the periparturient period may improve leukocyte functions by upregulating mRNA expression of genes involved in glycolysis, pathogen recognition, phagocytosis and oxidative burst, antimicrobial peptides, and antibody production. Finally, in herds with weekly or twice weekly movement of new cows to the prepartum pen and separate housing of nulliparous and parous animals, 100% SD of headlocks on the day of movement does not affect health, metabolic, reproductive, and productive parameters and 80% did not improve leukocyte responses compared with 100% target SD.
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    Innate Immunogenetics Of Keratinocyte Carcinomas
    (2014-12) Vineretsky, Karin
    Keratinocyte carcinomas (KC) which include basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) of the skin represent the most common malignancies in the world. UVR is the major risk factor associated with KC, and contributes to skin cancer carcinogenesis both as a mutagen and an immune suppressing agent. Exposures to arsenic, immunosupressive medications, beta-HPV, and individual susceptibility risk factors including genetic variability have also been implicated in KC development. Previous research was largely focused on the role of adaptive immunity in KC, while the role of innate immunity has not been fully investigated. Recent studies, including those on the action of Imiquimod, point to the centrality of innate immunity in KC pathogenesis. Natural killer (NK) cells of the innate immune system are the first line of defense against transformed and infected cells. NK cell killing capacity is determined by multiple inhibitory and activating cell surface receptors, including broad cellular-stress sensing natural killer group 2 member D (NKG2D) receptors, and highly diverse in gene content, copy number, and allelic polymorphisms killer-cell immunoglobulin-like receptors (KIR) that recognize self-human leukocyte antigen class I (HLA-I) ligands. Inter-individual variability of the KIR locus, and its interaction with cognate HLA-I, has been increasingly implicated in various disease settings including cancer. Genetic variability of NKG2D receptor activity has also been described in epithelial cancers. Given that NK cell function is influenced by these two classes of receptors, we hypothesized that genetic variability at these loci impacts innate immunity and BCC and SCC risk. Using a large population-based case-control study, a three part investigation was conducted. First investigation examined whether two genetic variants related to high natural cytotoxic activity of the NKG2D receptor decrease risk of KC and are modified by susceptibility and exposure factors including: sex, skin type, number of severe sunburns, glucocorticoids, and arsenic. This revealed differential effects of NKG2D genotype on KC risk by sex and skin type. The second study, determined whether KIR gene content alone and in combination with HLA-I ligands was associated with KC risk. The results suggested interactions of the activating KIR and HLA-I ligands may be implicated in control of BCC and to a lesser degree of SCC. The third study investigated the combined effects of KIR and NKG2D. NKG2D activity was assigned based on the number of alleles with high cytotoxic activity, and activating KIR were grouped based on the presence of 2DS1, 3DS1 and 2DS5 that were previously associated with reduced KC risk. The results showed reduced KC risk with the highest activating profile (containing 3 activating KIR and 2 or more NKG2D high activity alleles). Taken together, the results suggest a greater involvement of the innate immune response in the etiology of BCC, and to a lesser extent of SCC. In the context of SCC, these data suggest increased inflammation may deregulate the innate immune response.
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    Positive And Negative Regulation Of Defense Responses Against Pseudomonas Syringae In Arabidopsis
    (2014-03) Sreekanta, Suma
    Immune signaling in plants involves both positive and negative regulators. Maintaining a balance between growth and defense responses is important because there is a fitness cost to the plants if immune responses are left unchecked. Suppression of immune responses in the absence of pathogens as well as after the threat has passed is critical in maintaining such a balance between growth and defense responses. Upon pathogen perception, the positive regulators counter the immune repression to induce defense responses. We investigated the roles of two genes, CBP60a and PCRK1 in the regulation of defense responses against Pseudomonas syringae pathogen in the model system Arabidopsis thaliana . CBP60a is a negative regulator of immune responses. We showed that CBP60a is a CaM binding protein and that CaM binding is important for its function in transducing defense signals. Mutants of CBP60a were more resistant to Pseudomonas syringae infection suggesting that CBP60a was a negative regulator of defense responses. We found that CBP60a functions in repressing immune signaling under conditions where the plants are not challenged by a pathogen. We also investigated the role of a putative kinase, PCRK1, in immune signaling. We showed that pcrk1 mutants are more susceptible to Pseudomonas syringae than wild type plants suggesting that PCRK1 has a positive role in immune responses. We also showed that PCRK1 is important for immunity triggered by some of the conserved Microbe Associated Molecular Patterns (MAMP) as well endogenous signals generated as a result of pathogen activity called Damage Associated Molecular Patterns (DAMP).