<p> Studies with the serine hydrolase inhibitor JZL184 have suggested that enhanced 2-arachidonoylglycerol signaling could be strategized to stimulate innate immune cells to combat invading pathogens and improve host defense by prompting the systemic release of proinflammatory cytokines. Although the neurochemical effects of JZL184 were found to cultivate within 30 min in mice, its immune-regulating effects have been studied much later and its effects on chickens have not been clear. To explore the modulations in the chickens&rsquo; immune responses, we studied the effects of intraperitoneal injections of JZL184 in APEC O78-infected chickens on pathogenicity, histopathology, and IL-1&beta; levels. The pathogenicity of the strain was assessed by isolating bacteria from livers, blood, air sacs, and hearts at 8, 28, and 56 h post-infection (p.i.). Air sacs, livers, and hearts were examined for histopathological changes at 8, 28, and 56 h p.i. Serum samples were collected at 8, 28, and 56 h p.i. and analyzed with a chicken IL-1&beta; ELISA kit. Liver and spleen samples were homogenized for detection of serine hydrolases and carboxylesterases. Our work showed that 10 mg/kg and 40 mg/kg of JZL184 did not reduce the severity and progression of lesions produced in chickens challenged with 108 CFU of E. coli O78. The JZL184 treatments made colibacillosis lesions in the E. coli O78-challenged chickens worse and we did not find evidence of the injections increasing the serum cytokine levels of IL-1&beta; at our sampling times.</p><p>

<p> Duchenne&rsquo;s Muscular Dystrophy is a debilitating disease that affects skeletal and cardiac muscle of 1 in 5000 male births. In the last thirty years, the gene responsible for the encoding of Dystrophin has been identified, sequenced and the variations of mutations described. There remains a void in the successful treatment of the disease although corticosteroid use has proven useful in delaying progression. Novel therapies are produced in the categories of virus-mediated gene delivery and stem cells, but evaluating their efficacy is hindered by an inability to contemporaneously assess the changes in muscle. The purpose of this pilot study was to characterize the changes in skeletal and cardiac muscle in a clinically advanced population of dogs affected with Duchenne Muscular Dystrophy. Using traditional sequences, delayed gadolinium enhancement, novel sequences and spectroscopy, changes in the investigated muscle were characterized. By establishing the differences between affected and unaffected dogs, the long-term goal of this body of work is to characterize these changes longitudinally and design a non-invasive method for tissue assessment as novel treatments are trialed.</p><p>

<p> This dissertation focuses on the use of whole genome sequencing (WGS) for the identification of disease causing variants in canine genomes. A brief review on the historical milestones of genetics, the creation and popularization of the fast throughput DNA sequencing technologies and their advantages and potential problems and biases, the importance of the study of canine genetics and the current state of the canine genome assembly is presented. Our lab sequenced ~100 dogs in the attempt to discover disease-causing variants. So far 20 such variants have been identified. This dissertation contains detailed accounts of the discovery variants likely to be responsible for four canine diseases. Those diseases are: Paroxysmal dyskinesia in Soft Coated Wheaten Terriers that is associated to the missense mutation <i>PIGN:c</i>.398C>T; two different forms of neuronal ceroid lipofuscinosis, one in Australian Cattle dogs caused by CLN5:c.619C>T, and one in the Cane Corso caused by the splice site mutation <i>PPT1c.124+1G>A</i>; and a Shiba Inu GM2 gangliosidosis caused by <i>HEXB.c:948_950delCCT</i>. Furthermore, examples of not so successful attempts, possible reasons for failures and suggestions to successfully conclude other ongoing investigations.</p><p>

Cryptosporidiosis, caused by the apicomplexan parasite Cryptosporidium parvum, has become a well recognized diarrheal disease of immunodeficient humans and other mammals throughout the world. Specific therapy and immunoprophylaxis are currently unavailable, but passive immunization with C. parvum-specific monoclonal antibodies (mAbs) has demonstrated efficacy in immunocompromised hosts. In particular, mAbs eliciting the circumsporozoite precipitate (CSP)-like reaction protected against C. parvum infection. The circumsporozoite-like antigen (CSL), an ∼1,300 kDa apical glycoprotein of sporozoites and merozoites, is the molecular species mechanistically bound by mAbs having the ability to elicit the CSP-like reaction. These findings indicated that CSL has a functional role in sporozoite infectivity. In the present study, a quantitative in vitro sporozoite infectivity assay was developed to evaluate neutralizing activity of mAbs. 3E2, a mAb which elicited the CSP-like reaction, demonstrated the greatest level of neutralizing activity against C. parvum infections. Here, I report that CSL has properties consistent with being a sporozoite ligand for epithelial cells. For these studies, CSL was isolated from sporozoites by isoelectric focusing (IEF). The 1,200-1,400 kDa Mᵣ region containing CSL in SDS-PAGE of sporozoites was comprised of 31 species when analyzed by two-dimensional electrophoresis. Eight species were present in IEF-isolated CSL. CSL specifically bound to Caco-2 cells in a dose-dependent, saturable, and self-displaceable manner with high affinity. CSL bound to Caco-2 cells inhibited the attachment and invasion of sporozoites in a dose-dependent manner. Characterization of the epitope recognized by 3E2 in a dot immunoblot indicated a β-glucose component. Sporozoites having undergone the CSP-like reaction after incubation with CSL-reactive 3E2, did not attach to or invade Caco-2 cells. These findings indicated that at least 1 of 8 CSL species isolated by IEF was a sporozoite ligand. The Caco-2 cell receptor for CSL was comprised of 16, 51, and 85 kDa molecules. Further, sporozoites incubated with isolated Caco-2 receptors failed to attach to and invade Caco-2 cells. Finally, CSL bound distinctly to receptors present on calf ileum. Based on these findings, I concluded that CSL ligand function is amenable to targeted disruption by CSL-reactive mAbs and that it is a rational target for immunization against cryptosporidiosis.

No description available.

<p> <i>PTEN</i> and <i>TP53</i> loss are common molecular alterations in aggressive prostate cancer that progresses to castrate resistant prostate cancer (CRPC). <i>PTEN/TP53</i> loss contributes to regulation of self-renewal and differentiation in prostate progenitor cells, the presumptive tumor and metastasis initiating cells for prostate cancer. <i>TP53</i> plays an important role in regulating normal cellular metabolism, and loss of function is responsible for metabolic alterations in tumor cells, including increased aerobic glycolysis. We use a novel model of <i>Pten/Tp53</i> deleted prostate cancer to investigate properties of tumor and metastasis initiating cells, and metabolic alterations that contribute to the evolution of CRPC. </p><p> We employed a genetically engineered mouse model of <i>Pten^-/-Tp53^-/-</i> prostate cancer to develop an orthotopic model derived from a clonal cell line from the parental heterogeneous prostate carcinoma. We used histopathology and immunohistochemistry to characterize the orthotopic primary tumors and metastases. We performed metabolomic screening followed by focused analysis of HK II enzyme levels, activity, and cellular distribution in androgen replete and androgen deprived tumors. We also compared HK II levels in primary and metastatic human prostate cancer. </p><p> Tumor heterogeneity was due to transformation of tumor and metastasis initiating cells with biphenotypic potential capable of basal and luminal differentiation. There was epithelial-to-mesenchymal transition (EMT) in cells of the luminal lineage. The model was capable of androgen independent growth, which influenced the differentiation of metastasis initiating cells. CRPC had increased reliance on glycolysis with increased cytoplasmic and mitochondrial-associated HK II. These metabolic adaptations afforded CRPC increased ability to withstand metabolic stress. HK II levels in human metastases were markedly increased compared to primary tumors. </p><p> <i>Pten/Tp53</i> loss in prostate cancer contributes to lineage plasticity in both tumor and metastasis initiating cells, contributing to heterogeneity observed in primary tumors and metastases. Increased glycolysis due to increased total and mitochondrial HK II is a metabolic adaptation that contributes to the evolution of aggressive disease, with progression to androgen independence, providing increased energy and carbon precursors for anabolic processes. Mitochondrial bound HK II blocks apoptosis and contributes to survival in the androgen deprived environment. Targeting this metabolic adaptation may provide improved treatment for this deadly disease.</p>

Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2008. / Source: Dissertation Abstracts International, Volume: 69-05, Section: B, page: 2838. Adviser: Peter D. Constable. Includes supplementary digital materials. Includes bibliographical references (leaves 215-240) Available on microfilm from Pro Quest Information and Learning.

Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2007. / Source: Dissertation Abstracts International, Volume: 68-07, Section: B, page: 4318. Adviser: Indrani C. Bagchi. Includes bibliographical references (leaves 86-93) Available on microfilm from Pro Quest Information and Learning.

Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 2008. / Source: Dissertation Abstracts International, Volume: 69-05, Section: B, page: 2839. Adviser: Ronald M. Weigel. Includes bibliographical references (leaves 120-128) Available on microfilm from Pro Quest Information and Learning.

<p> Allogeneic bone marrow-derived mesenchymal stem cells (MSCs) are a promising cell therapy for effective and efficient treatment of various inflammatory and immunemediated diseases. While the prevailing dogma has been that MSCs are immune privileged, very few studies have controlled for MHC haplotype or adequately measured MSC immunogenicity in vitro or in vivo. Controlled studies have found that major histocompatibility complex (MHC)-mismatched MSCs evoke both cell-mediated and humoral immune responses in vivo. Microcytotoxicity assays were used to show that horses injected with MHC-mismatched MSCs generate cytotoxic alloantibodies capable of killing MSCs as early as 7-days post-transplantation. Rejection of MSCs likely leads to reduced therapeutic efficacy and the development of strategies to avoid allorecognition and rejection are necessary to provide safe and efficacious allogeneic therapy.</p><p> Downregulation of MHC expression allows cells to avoid immune surveillance and may enhance the ability of MSCs to avoid allorecognition and rejection. Transforming growth factor-&beta;2 (TGF-&beta;2) has been shown to downregulate MHC surface expression in various cell types. In agreement with what has been demonstrated in other cell types, TGF-&beta;2 treatment significantly reduced constitutive MHC I and MHC II surface expression and partially blocked IFN-&gamma;-induced MHC expression on equine MSCs. TGF-&beta;2 treatment did not significantly affect the morphology, cell surface markers, viability, or secretion of TGF- &beta;1 and TGF-&beta;2, but did increase the cell yield from cultures. This data indicates that TGF-&beta;2 may reduce MSC immunogenicity without altering the immunomodulatory properties of the cells.</p><p> The immunomodulatory capabilities of TGF-&beta;2-treated MSCs were analyzed in modified one-way mixed leukocyte reactions and ELISAs. Naive and TGF-&beta;2-treated MSCs both significantly reduced T cell proliferation as measured by the relative division index and relative CFSE geometric mean fluorescent intensity attenuation. Similar amounts of PGE2 and TGF-&beta;2 were also measured in the supernatant of MLRs with naive and TGF-&beta;2-treated MSCs. This supports that TGF-&beta;2 treatment does not negatively affect the immunomodulatory properties of equine MSCs, which are critical for therapeutic function and evading immune responses in vivo.</p><p> In conclusion, although MHC-mismatched equine MSCs are immunogenic in vivo, MHC I and MHC II surface expression can be manipulated by treating cells with TGF-&beta;2 in vitro. Downregulate of MHC surface expression is a promising strategy for enhancing the ability of MSCs to evade immune responses allowing for allogenic use clinically without the risk of immune rejection. The ability of TGF-&beta;2-treated MSCs to avoid immune rejection should continue to be investigated in vitro and in vivo along with the mechanism by which TGF-&beta;2 downregulates MHC expression.</p><p>