Browsing by Author "Avery, Anne, advisor"
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Item Open Access CD4+ T-cell derived IL-10 mitigates malarial anemia(Colorado State University. Libraries, 2013) Kiser, Patti, author; Avery, Anne, advisor; Callan, Robert, committee member; Olver, Christine, committee member; Quackenbush, Sandra, committee memberIndividuals living in malaria endemic areas develop effective anti-parasite immunity over several years of repeated exposure, but become resistant to severe disease after just one or two infections. This observation suggests that the acquired immune system plays a role in both processes, but may involve different mechanisms. Using the mouse model of malaria caused by non-lethal Plasmodium yoelii, we test the hypothesis that CD4+ T-cell derived IL-10 contributes to disease resistance by mitigating severe anemia. Here we show that IL-10 deficient mice develop significant anemia despite a very low parasite burden. Anemia in this model is mediated primarily by increased erythrocyte destruction and not from suppressed erythropoiesis. Wild type mice that have recovered from P. yoelii infection have an expanded population of IL-10 producing CD4 T cells, with the majority of these cells co-expressing IFN-γ and display a Th1 phenotype. In the absence of IL-10, there is an increase in IFN-γ+ T cells. We demonstrate that IL-10 competent CD4+ T cells protect athymic nude mice from anemia when compared to CD4 T cells taken from recovered IL-10 deficient mice. Utilizing an ex vivo system that tests the function of APCs in activating CD4+ T-cells, we also determined that APCs exposed to P. yoelii in vivo induced a greater population of CD4+ T-cells that express IL-10 compared to naïve APCs. We also demonstrate that IFN-γ is required, with the possible involvement of IL-10 and IL-12, for efficient IL-10 expression in CD4+ T-cells. Our findings suggest that one mechanism by which the acquired immune system contributes to resistance to severe anemia may be the development of CD4 T cells that co-express IL-10 and IFN-γ, thereby self-regulating IFN-γ levels, which then inhibits pro-inflammatory mediated destruction of naïve red blood cells.Item Open Access Clinical and molecular characterization of canine small cell B-cell lymphocytosis disorders(Colorado State University. Libraries, 2020) Rout, Emily, author; Avery, Anne, advisor; Avery, Paul, committee member; Zabel, Mark, committee member; Weil, Michael, committee memberTo view the abstract, please see the full text of the document.Item Open Access Correlation of HAS2-associated gene duplications with biological aggressiveness of mast cell tumors in Chinese Shar-Pei dogs(Colorado State University. Libraries, 2013) Garner, Alana Pavuk, author; Avery, Anne, advisor; Thamm, Douglas, committee member; Basaraba, Randall, committee memberCutaneous mucinosis in Shar-Pei dogs is the result of excessive dermal hyaluronan, a protein associated with angiogenesis and tumor cell motility in multiple human and canine neoplasms. Cutaneous mucinosis in Shar-Pei has been associated with gene duplications upstream of the hyaluronic acid synthase 2 gene (HAS2). The objective of this study was to evaluate the relationship between HAS2, cutaneous mucinosis, and features of mast cell tumor (MCT) aggressiveness in Shar-Pei dogs. Biopsies of cutaneous MCTs from 149 Shar-Pei and 100 non-Shar-Pei were graded according to two schemes for canine cutaneous MCTs. Biopsies of the Shar-Pei MCTs were also evaluated for degree of cutaneous mucinosis, depth of invasion, and microvessel density (MVD). Shar-Pei and non-Shar-Pei MCTs were evaluated via qPCR for relative copy number of the gene duplication upstream of HAS2. The proportion of grade III tumors was significantly higher in Shar-Pei than the general canine population (p=1.044e-11), with no difference in average age at diagnosis. Shar-Pei biopsies had significantly higher HAS2-associated gene segment duplications than non-Shar-Pei (p=1.128e-11), and copy number was significantly associated with the development of grade III tumors (p=0.0077), mitotic index > or = 7 (p=0.022), and tumoral MVD (p<0.05). Relative copy number was not significantly associated with the degree of cutaneous mucinosis or depth of invasion. Our data suggest a relationship between HAS2 gene duplications and features of MCT aggressiveness in Shar-Pei dogs.Item Open Access Design and application of a droplet-digital PCR assay for detection of the STAT5BN642H mutation in feline T cell neoplasia(Colorado State University. Libraries, 2024) Bork, Sydney Bonnie, author; Avery, Anne, advisor; Olver, Christine, committee member; Webb, Craig, committee memberLymphoma is a commonly diagnosed hematopoietic neoplasm in cats. Small Cell T-cell Epitheliotropic Intestinal Lymphoma (SCL) is the most reported subtype of lymphoma in cats. Cats with SCL are presented with non-specific clinical signs such as chronic vomiting, diarrhea, and weight loss. Diagnostic work-up often includes collection of intestinal biopsies with histopathology for diagnosis. SCL is characterized by infiltration of neoplastic lymphocytes into the intestinal epithelium and lamina propria of the small intestines. Neoplastic cells are small to intermediate in size and of T-cell origin. Diagnosing SCL can be challenging for pathologists because cats also commonly develop a condition called inflammatory bowel disease (IBD), which has an almost identical clinical presentation and similar histopathologic patterns. However, in IBD, the lymphocytic infiltration is often heterogeneous (termed "lymphoplasmacytic enteritis"). When histopathology results are inconclusive, assessment of expression with immunohistochemistry markers can help further characterize the cell population. Additionally, advancements have been made with lymphocyte clonality testing by PARR (PCR for Antigen Receptor Rearrangement), a DNA-based assay that evaluates T-cell receptor (TCR) and Immunoglobulin (Ig) gene rearrangements. Cats diagnosed with SCL demonstrate a clonal TCR result, while cats with IBD demonstrate a polyclonal TCR result. Unfortunately, there are still cases where histopathology and PARR results are equivocal. Recent work in feline medicine has demonstrated that cats with SCL exhibit high expression of phosphorylated STAT5B with immunohistochemical staining on small intestinal biopsy samples compared to cats with IBD. Importantly, one group detected a STAT5BN642H mutation in cats diagnosed with SCL. In this study, 40% (17/42) of cats with intestinal lymphoma were classified as SCL by histopathology. A combination of Sanger sequencing and ARMS qPCR detected the STAT5BN642H mutation in 29.4% (5/17) of cats with SCL. This work correlates to a comparable disease entity in people, monomorphic epitheliotropic intestinal T-cell lymphoma (MEITL), which has reported the prevalence of the STAT5BN642H mutation to be 22-57%. Our group aimed to develop a droplet digital PCR (ddPCR) assay to detect wild-type and mutated STAT5B in cats. Our first aim was to design specific primers and locked-nucleic acid hydrolysis probes to detect and discriminate between wild-type and mutated STAT5B. The first step included analyzing data from control samples using wild-type DNA from cats without neoplasia and a positive control gene fragment block ("gBlock"). The second step included analyzing two cohorts of young cats (<6 years of age) without a diagnosis of lymphoid neoplasia to assess assay performance and determine if the mutated STAT5B could be considered a germ line polymorphism. The fractional abundance was calculated from the ddPCR data, estimating the percentage of mutated copies within a positive sample. The results of the first aim demonstrated that our ddPCR assay can distinguish between wild-type and mutated STAT5B with high sensitivity. Most young cats without a diagnosis of lymphoid neoplasia do not carry the STAT5BN642H mutation. Two cats with marked lymphoplasmacytic enteritis had detectable mutated STAT5BN642H. The second aim was to evaluate the prevalence of the STAT5BN642H mutation in cats with confirmed SCL. A sub-aim was to evaluate cats with CD4 T-cell leukemia to determine if this mutation could be found in other forms of T cell lymphoid neoplasia. The results from this aim demonstrate that cats with SCL frequently carry the STAT5B mutation (66.7%). We also discovered that this mutation is not exclusive to cats with SCL, as almost half of the cats with CD4 T-cell leukemia also carry this mutation (47.7%). These findings shed light on the prevalence of the STAT5BN642H mutation in cats with SCL and CD4 T-cell leukemia. This data suggests potential implications for ddPCR mutation detection to help further differentiate and diagnose cats with T cell neoplasia versus those with inflammatory conditions (such as SCL versus IBD), and investigate novel therapies (i.e., JAK/STAT inhibitors). Further research is warranted to investigate other JAK/STAT pathway mutations, particularly in cats where the STAT5BN642H mutation was not detected. Larger outcome studies should investigate the correlation of STAT5BN642H mutation status and the fractional abundance to evaluate disease risk, treatment response, and survival.Item Open Access Molecular characterization of canine peripheral T-cell lymphoma(Colorado State University. Libraries, 2020) Harris, Lauren, author; Avery, Anne, advisor; Avery, Paul, committee member; Basaraba, Randall, committee member; Bailey, Susan, committee memberTo view the abstract, please see the full text of the document.Item Open Access T zone lymphoma: cellular origin and function(Colorado State University. Libraries, 2018) Hughes, Kelly Lynn, author; Avery, Anne, advisor; Ehrhart, EJ, committee member; Rovank, Joel, committee member; Page, Rodney, committee member; Dow, Steve, committee memberThe lymphoid system is exceedingly complex with specialized subsets of lymphocytes involved in both the innate and adaptive immune response. Lymphocytes are subdivided into T cells and B cells with lymphoproliferative disorders comprising a heterogeneous group of diseases arising from various lymphocyte subsets. Dogs are a natural model for studying cancer in humans with overlapping subtypes of lymphoproliferative disease seen in both species. We were particularly interested in characterizing lymphoma subtypes in dogs as a model for studying human disease progression. We believe characterization of lymphoma subtypes in dogs will contribute to enhanced understanding of the pathogenic mechanisms leading to the development of malignancy in both humans and dogs. T zone lymphoma (TZL) is a subtype of peripheral T cell lymphoma (PTCL) occurring in both humans and dogs. Determination of the cell-of-origin of PTCLs in humans has pointed to a subset of the larger disease, but up to half of these lymphomas cannot be further classified and are referred to as PTCL – not otherwise specified (PTCL-NOS). TZL has unique phenotypic characteristics including absence of the pan-leukocyte transmembrane protein, CD45, allowing for reliable identification by flow cytometry. CD45 has been found to be critical for T cell signaling through the T cell receptor (TCR). We hypothesized TZL has been derived from an activated, mature T cell. We used gene expression in an attempt to classify the cell-of-origin in TZL. Then we used in vitro systems to identify proliferative mechanisms, cytokine production, and immunosuppression at play in this disease. We determined TZL cells express genes associated with T helper 2 (Th2) and T regulatory (Treg) cells. We further confirmed that TZL cells do not proliferate through the TCR, likely due to their lack of CD45. We identified IL-2 may be involved in TZL signaling mechanisms. There are reports of TZL occurring with an opportunistic follicular mite infection, suggesting a component of immunosuppression with this disease. We thus anticipated TZL cells would have an immunosuppressive phenotype and inhibit normal T cells. Using in vitro methods, we were unable to confirm immunosuppression through production of TGF-β, inhibition of proliferation and inhibition of IFN-γ production. Finally, we observed older Golden retrievers having a higher frequency of cells with the same aberrant phenotype as TZL, but without evidence or suspicion of lymphoproliferative disease. We believe this finding resembles clonopathies of unknown significance in older human individuals, also with rare progression to disease. We now believe TZL is derived from a Th2 or Treg-like mature T cell which may be responsible for the indolent nature of this disease due to the tolerant behavior of those T cell subsets. Furthermore, inducing proliferation in this disease was challenging, which is consistent with slowly progressive biologic behavior. Expression of immunosuppressive molecules may also contribute to the indolent nature of this disease. While we were not able to observe immunosuppression in vitro, it is reasonable to assume there may be in vivo mechanisms of immunosuppression resulting in clinical manifestations seen in TZL. Characterization of cell-of-origin of this disease, neoplastic cell function, along with recognition of a pre-neoplastic state, contributes to enhanced understanding of the pathogenesis of lymphoproliferative diseases in both humans and dogs.Item Open Access The acquisition of dendritic cell tolerance during malaria infection results in differential T-cell activation(Colorado State University. Libraries, 2008) Perry, James A., author; Avery, Anne, advisorMalaria is caused by intracellular protozoan parasites belonging to the genus Plasmodia. These single cell eukaryotes have a complex life cycle requiring both mammalian (and in certain Plasmodium species, avian) and mosquito hosts. Clinical malaria in humans and other animals is the result of red blood cell (RBC) infection. Although infection direcdy destroys erythrocytes, causing anemia, a significant degree of anemia and morbidity is the result of the host immune response. Inflammatory cytokines have been implicated in the pathogenesis of severe malaria anemia (SMA) and cerebral malaria (CM), two diseases that are responsible for most malaria-related morbidity. Therefore, understanding the regulation of host immunity and inflammatory cytokine production during malaria infection will improve our understanding of malaria related illness.Item Embargo Using gene expression and mutational profiling to characterize canine acute myeloid leukemia and assess their comparative features with human acute myeloid leukemia(Colorado State University. Libraries, 2023) Harris, Adam, author; Avery, Anne, advisor; Avery, Paul, committee member; Dow, Steven, committee member; Duval, Dawn, committee memberAcute myeloid leukemia (AML) is an aggressive heterogenous hematopoietic neoplasm that afflicts both dogs and people. Over 10,000 individuals (about the seating capacity of Cameron basketball stadium at Duke University) in the United States succumb to AML-related deaths every year. Treatment options for AML have made little progress in the past few decades and prognosis for both human and canine AML (cAML) remains dismal. However, there are large ongoing multi-institutional studies devoted to advancing medical management for human AML (hAML) by providing targeted therapeutics to patients based on their molecular characteristics. A preclinical model for testing novel therapies could accelerate the development of better treatments in people. We hypothesize that cAML will have similar underlying molecular features as human AML and dogs could be a translational model for developing therapeutics focused on treating AML. The goals of this thesis were to assess the gene expression programs and mutational profiles of cAML and compare our findings with available human AML data. First, we established diagnostic criteria for defining cAML using flow cytometry. Next, we globally assessed normal hematopoiesis in dogs using single cell transcriptomics to generate a hematopoietic tree for defining the cellular composition of cAML. Additionally, we investigated the mRNA expression and genetic variants in cAML to ultimately compare the molecular features with pediatric and adult AML subtypes. We hope this work advances our knowledge of cAML molecular characteristics and adds further credentials to the dog as spontaneous model for human AML.Item Open Access Using the dog as a model to investigate environmental and genetic risk factors for mature, antigen-driven lymphoproliferative disorders(Colorado State University. Libraries, 2017) Labadie, Julia, author; Magzamen, Sheryl, advisor; Avery, Anne, advisor; Anderson, Brooke, committee member; Feigelson, Heather, committee member; Morley, Paul, committee member; Page, Rodney, committee memberTo view the abstract, please see the full text of the document.