Aim: To conduct a systematic review and meta-analysis of all published studies studying the risk of vitamin D deficiency in tuberculosis patients and in healthy controls. Additionally, subgroup meta-analysis was performed based on the level of vitamin D to test the risk in TB groups and in healthy groups. Methods: Pubmed was searched for observational studies in human and English that discussed the association between risk of low serum vitamin D and TB. Meta-analysis was performed on all relevant studies combined and for subgroups of each vitamin D level. Results: 22 studies were selected and pooled in the analysis. The results were consistent with previous studies examining the same risk. The overall log risk ratio (log RR) of low vitamin D was significantly higher in TB patients 1.68 times than healthy controls. In 4 subgroup meta- analyses based on vitamin D level below (20 nmol/L, 30 nmol/L, 50 nmol/L, and 75 nmol/L), the risk of having low vitamin D in TB patients was (1.82, 2.89, 1.38, 1.32) respectively. That subgroup analysis showed more clearly the higher RR were below 20 and 30 nmol/L. The smallest RR was at 75 nmol/L level. Conclusion: This study verified the association between risk of low vitamin D level and TB development. It also clarified that, the risk increased by decreasing vitamin D level.

ABSTRACT Alzheimer's disease (AD) is the most common progressive age related dementia and the fourth major cause of mortality in the elderly in the United States. AD is pathologically characterized by deposition of amyloid beta (Aβ) plaques in the brain parenchyma and neurofibrillary tangles (NFTs) within the neuronal soma. While pharmacological targets have been discovered, current strategies for the symptomatic or disease-modifying treatment of AD do not significantly slow or halt the underlying pathological progression of the disease. Consequently, more effective treatment is needed. One possibility for amelioration is using human umbilical cord blood cell (HUCBC) therapy. HUCBCs comprise a population of hematopoietic stem and progenitor cells. During recent years, functional recovery has been observed from the use of HUCBCs in pre-clinical animal models of brain and spinal cord injuries. Thus, modulation by cell therapy, specifically HUCBCs, may be a suitable treatment for AD and other models because of the observed cognitive and behavioral improvements. The studies presented in this dissertation centers on the suitability of using HUBCs as a potential treatment for AD. Expanding on this, the aims of the study sought to: (I) Investigate bio-distribution of HUCBC transplantation in PSAPP mice, (II) Characterize efficacy and determine therapeutic outcome of HUCBC following short and long term multi injections at early and late disease stages in PSAPP mice and (III) Determine AD-like pathological and cognitive changes associated with multiple HUCBC-derived monocyte (CD14) injections in PSAPP mice. Thus the findings of this work evolved from experiments that characterized the effects of low-dose infusions of HUCBC and HUCBC-derived monocytes into 6 month old Presenilin 1/Amyloid Precursor Protein (PSAPP) plaque-developing transgenic AD mice. Treated mice were studied using standard behavioral tests to determine the effects of infusion on the multiple cognitive domains affected by AD, followed by biochemical and histological analyses that included Aβ load and amyloid precursor protein (APP) processing. Specifically, PSAPP mice and their wild-type (WT) littermates were treated monthly with a peripheral HUCBC infusion over a period of 6 and 10 months, followed by cognitive and motor evaluation. Additionally, based on reports that tumor cells can originate from stem cells present in HUCB, we further examined whether monocytes purified from HUCBCs would have a similar significant effect on the reduction of AD-like pathology in PSAPP mice. HUCB cells homed into tissues including the brain. The principal finding was significant reduction in Aβ levels and β–amyloid plaques following low-dose infusions of both HUCBC– derived mononuclear cells as well as HUCBC-derived monocytes, with the monocytes providing a stronger effect. Results further demonstrated that HUCBC and HUCBC– derived monocyte infusion could improve memory function and locomotor ability in treated PSAPP mice. A possible reason for behavioral improvements in these animals may be the significant reduction in both Aβ levels and plaque load. This study also identified significant reduction in microglial activation and astrocytosis, both of which can contribute to AD pathology. In conclusion, our data suggest that it might be the HUCBC–derived monocytic population rather than stem cells that are responsible for the reduction in AD pathology.

Ross River virus (RRV) is a mosquito-borne alphavirus and the aetiological agent of epidemic polyarthritis (EPA). Arthropod borne-Alphaviruses that are related to RRV, such as Chikungunya virus, Sindbis virus and Barmah Forest virus, are usually associated with epidemics of infectious arthritides in different parts of the world. In humans, RRV-induced disease symptoms include fever, rash, myalgia and pain and stiffness of the joints. Muscle and joint pain are the most debilitating symptoms in RRV patients and the best treatment available is non-steroidal anti-inflammatory drugs (NSAID). Previous studies in mice have demonstrated that RRV infection results in inflammation of skeletal muscle and joints and that macrophages play a primary role in disease. The present study was carried out to further elucidate the underlying mechanisms mediating RRV-induced muscle and joint pathology. Previous studies have reported that encephalitic alphaviruses trigger apoptosis of brain cells in mice and that blocking apoptosis reduces mortality rates. In the present study, the ability of RRV to induce muscle apoptosis was investigated in vitro, using a murine myoblast cell line (C1C12), and in vivo, using a mouse model of RRV disease. RRV-infected C1C12 myofibres displayed an array of morphological and biochemical makers of apoptosis. Apoptosis was also observed in the skeletal muscle of RRV-infected C57BL/6J mice. Blocking apoptosis by general caspase inhibition resulted in milder disease symptoms, reduced myofibre damage and decreased inflammation of muscle and joint tissues. The total number of cell infiltrates as well as the number of macrophages infiltrating muscle was significantly reduced by the treatment with a caspase inhibitor. The effects of RRV infection on the skeletal system were also investigated. Primary human osteoblast cells were infected with RRV and monitored for viral-induced cytopathic effect. Osteoblasts supported rapid virus growth and, by 48 hours after infection, succumbed to viral-induced necrosis. In addition, histological examination of bone tissue from RRV-infected C57BL/6J mice showed clear evidence of bone resorption. Tibias from infected mice showed an increased number of activated osteoclasts, a reduction in bone density and thinning of cortical bone. The expression of host factors involved in inflammatory responses and bone remodelling was studied in RRV-infected myofibres and osteoblast cell cultures and in the muscle and joint tissues from infected mice. RRV-infected muscle cells and tissue showed elevated mRNA levels for the chemokines CCL-2, CCL3, CCL5 and CXCL1, all of which are known to mediate the migration of monocytic cells. With the exception of CXCL1, these chemokines were also found to be up-regulated in RRV-infected osteoblast cultures and in joint tissues from infected mice. Muscle and joint tissue from infected mice also showed elevated mRNA levels for type I and type II interferons, TNF- and NOS2. In addition, joint tissues from infected animals contained high levels of IL-6 and IL-1, two cytokines known to mediate bone remodelling. Finally, the therapeutic potential of the drug bindarit was investigated using the mouse model of RRV disease. Bindarit is a known inhibitor of CCL-2 and TNF- and has been found to prevent protein denaturation. Treatment with bindarit resulted in mice developing milder disease symptoms, reduced muscle damage and decreased inflammation of muscle and joint tissues. In particular, bindarit significantly reduced macrophage infiltration into skeletal muscle tissue. This thesis has contributed to the understanding of RRV pathogenesis. It has identified novel mechanisms of RRV-induced muscle and bone pathology and provided further evidence that associate pro-inflammatory host factors to RRV disease. This work has also demonstrated that bindarit should be considered as a candidate for treating RRV disease in humans.

Several systems were examined for the production and delivery of recombinant vaccines for fish. C. crescentus was employed to produce a fragment of the IHNV glycoprotein. When administered by injection to 0.5 gram rainbow trout (Oncorhynchus mykiss), one of the fusion proteins (184 amino acids of the IHNV glycoprotein fused to 242 amino acids of the C-terminus of the Caulobacter crescentus) protected the fish against lethal challenge with IHNV. Attenuated strains of Yersinia ruckeri were generated using allelic exchange mutagenesis. These strains were characterized in terms of in vitro growth characteristics and invasiveness. Attenuated E. coli and Y. ruckeri were exploited to deliver plasmid DNA to fish cells in vitro; attenuated Y. ruckeri bacteria were examined in vivo as bivalent vaccine delivery vehicles, either through the expression of a fragment of the IHNV glycoprotein or by carrying a plasmid DNA vaccine encoding the complete IHNV glycoprotein. A cell wall deficient strain (11.29��dap) protected rainbow trout against lethal challenge with pathogenic Y. ruckeri. Gene transfer to fish was not detected by luciferase reporter gene assays. No clear protection from IHNV challenge was observed. / Graduation date: 2002

Thesis (M.S.)--University of Delaware, 2007. / Principal faculty advisor: Jack Gelb, Dept. of Animal & Food Sciences. Includes bibliographical references.

Blood was obtained from 5 adult horses and 16 juvenile horses (foals) at the time of birth and subsequently at 2-, 4-, and 8-weeks of age. Neutrophils from adult horses were purified and incubated for 2 h and 4 h with media, avirulent R. equi, virulent R. equi, or recombinant-human granulocyte-macrophage colony stimulating factor (rhGM-CSF). Neutrophils from foals were purified and incubated for 2 h and 4 h with media or virulent R. equi. Total RNA was extracted from both adult and foal neutrophils immediately after purification to measure baseline expression levels (0 h), and immediately after each of the prescribed incubation times. For each sample, 1 µg of total RNA was reverse-transcribed and analyzed for differential gene expression using real-time PCR. After 2 h and 4 h incubation with virulent or avirulent R. equi, neutrophils from adult horses expressed significantly (P< 0.05) greater TNFα, IL-12p40, IL-6, IL-8, and IL-23p19 mRNA relative to expression by unstimulated neutrophils, but not IFNγ or IL-12p35 mRNA. Furthermore, virulent R. equi induced significantly greater IL-23p19 mRNA expression than avirulent R. equi. Stimulation with rhGM-CSF of adult equine neutrophils failed to induce significant changes in cytokine expression. In foal neutrophils, stimulation with virulent R. equi induced significantly greater expression of IFNγ, TNFα, IL-6, IL-8, IL-12p40, and IL-12p35 mRNA relative to expression by unstimulated neutrophils. Furthermore, there were significant effects of age on expression of IL-6, IL-8 and IL-12p40 mRNA. Neutrophil mRNA expression of IL-6 and IL-8 in newborn foals was significantly greater than expression at 2-, 4-, and 8-weeks of age. There was no significant difference between unstimulated and R. equi-stimulated neutrophils from newborn and 2-week-old foals in expression of IL-12p40; however, expression of IL-12p40 by R. equi-stimulated neutrophils from 4- and 8-week-old foals was significantly greater than expression by unstimulated neutrophils. These results demonstrate that R. equi-stimulated neutrophils are a source of many pro-inflammatory cytokines, and that the magnitude of this expression with respect to IL-6, IL-8, and IL-12p40 mRNA expression was influenced by age. Collectively, the data presented indicate a non-phagocytic role for neutrophils that may influence the type of adaptive immune response to R. equi.

Public health planning for influenza is based on morbidity and mortality estimates derived from statistical models. Lower than anticipated 2009 H1N1 pandemic death estimates have raised questions about the method. Examining the statistical method is important for future policy and program development. We compared the main methods of estimating influenza burden through a systematic literature review and by comparing statistical estimates of influenza-attributable burden at the Ottawa Hospital (TOH) to clinical estimates validated through chart review. We identified heterogeneity in methods used to estimate influenza-attributable mortality in the literature which resulted in within-season estimate variation by study. We found statistical estimates of influenza burden at TOH to be 4-8 times greater than clinically validated data. We also found no significant association between the outcomes examined and epidemic periods at TOH. The findings of this study suggest discordance between model estimates by model approach and between model estimates and validated findings. Examining reasons for these discordances should be pursued.

Severe malaria infections cause almost 1 million deaths annually, mostly among non-immune African children. The pathogenesis of severe malaria is poorly understood. It is increasingly appreciated that while host innate immune responses such as inflammation and phagocytosis are critical for control of parasite replication, they can become dysregulated and contribute to severe disease. The goals of this work were: (1) to characterize inflammatory responses to malaria by defining their relationship to phagocytosis and identifying novel molecular mediators, and (2) to evaluate the utility of biomarkers of inflammation and other host responses for predicting outcome in severe malaria infection. Using an in vitro model of the malaria-macrophage interaction, inflammatory and phagocytic responses to Plasmodium falciparum were found to be partially coupled. Activation of Toll-like receptors (TLRs) by purified parasite components increased internalization of parasitized erythrocytes, but uptake of parasitized erythrocytes did not require TLRs, nor did it trigger cytokine production via TLRs or other receptors. Two candidate molecules – Triggering receptor expressed on myeloid cells-1 (TREM-1) and Chitinase-3 like-1 (CHI3L1) – did not appear to critically modulate inflammation to malaria in vitro or in murine models. However, exogenous TREM-1 activation enhanced the pro- inflammatory nature of the response to P. falciparum, with potential implications for malarial-bacterial co-infection. CHI3L1-deficient mice showed a trend towards earlier death in experimental cerebral malaria, suggesting that CHI3L1 may protect against severe malaria; however, further investigation in more informative models is required. Admission levels of plasma TREM-1, CHI3L1, and other biomarkers of inflammation and endothelial activation were increased in Ugandan children with severe malaria. Simple combinations of these biomarkers predicted mortality among severe malaria patients with high accuracy, warranting larger validation studies. Taken together, these findings identify host responses as putative targets for adjunctive therapies, and suggest the utility of host biomarker combinations as prognostic tests for severe malaria.

Severe malaria infections cause almost 1 million deaths annually, mostly among non-immune African children. The pathogenesis of severe malaria is poorly understood. It is increasingly appreciated that while host innate immune responses such as inflammation and phagocytosis are critical for control of parasite replication, they can become dysregulated and contribute to severe disease. The goals of this work were: (1) to characterize inflammatory responses to malaria by defining their relationship to phagocytosis and identifying novel molecular mediators, and (2) to evaluate the utility of biomarkers of inflammation and other host responses for predicting outcome in severe malaria infection. Using an in vitro model of the malaria-macrophage interaction, inflammatory and phagocytic responses to Plasmodium falciparum were found to be partially coupled. Activation of Toll-like receptors (TLRs) by purified parasite components increased internalization of parasitized erythrocytes, but uptake of parasitized erythrocytes did not require TLRs, nor did it trigger cytokine production via TLRs or other receptors. Two candidate molecules – Triggering receptor expressed on myeloid cells-1 (TREM-1) and Chitinase-3 like-1 (CHI3L1) – did not appear to critically modulate inflammation to malaria in vitro or in murine models. However, exogenous TREM-1 activation enhanced the pro- inflammatory nature of the response to P. falciparum, with potential implications for malarial-bacterial co-infection. CHI3L1-deficient mice showed a trend towards earlier death in experimental cerebral malaria, suggesting that CHI3L1 may protect against severe malaria; however, further investigation in more informative models is required. Admission levels of plasma TREM-1, CHI3L1, and other biomarkers of inflammation and endothelial activation were increased in Ugandan children with severe malaria. Simple combinations of these biomarkers predicted mortality among severe malaria patients with high accuracy, warranting larger validation studies. Taken together, these findings identify host responses as putative targets for adjunctive therapies, and suggest the utility of host biomarker combinations as prognostic tests for severe malaria.

Blood was obtained from 5 adult horses and 16 juvenile horses (foals) at the time of birth and subsequently at 2-, 4-, and 8-weeks of age. Neutrophils from adult horses were purified and incubated for 2 h and 4 h with media, avirulent R. equi, virulent R. equi, or recombinant-human granulocyte-macrophage colony stimulating factor (rhGM-CSF). Neutrophils from foals were purified and incubated for 2 h and 4 h with media or virulent R. equi. Total RNA was extracted from both adult and foal neutrophils immediately after purification to measure baseline expression levels (0 h), and immediately after each of the prescribed incubation times. For each sample, 1 µg of total RNA was reverse-transcribed and analyzed for differential gene expression using real-time PCR. After 2 h and 4 h incubation with virulent or avirulent R. equi, neutrophils from adult horses expressed significantly (P< 0.05) greater TNFα, IL-12p40, IL-6, IL-8, and IL-23p19 mRNA relative to expression by unstimulated neutrophils, but not IFNγ or IL-12p35 mRNA. Furthermore, virulent R. equi induced significantly greater IL-23p19 mRNA expression than avirulent R. equi. Stimulation with rhGM-CSF of adult equine neutrophils failed to induce significant changes in cytokine expression. In foal neutrophils, stimulation with virulent R. equi induced significantly greater expression of IFNγ, TNFα, IL-6, IL-8, IL-12p40, and IL-12p35 mRNA relative to expression by unstimulated neutrophils. Furthermore, there were significant effects of age on expression of IL-6, IL-8 and IL-12p40 mRNA. Neutrophil mRNA expression of IL-6 and IL-8 in newborn foals was significantly greater than expression at 2-, 4-, and 8-weeks of age. There was no significant difference between unstimulated and R. equi-stimulated neutrophils from newborn and 2-week-old foals in expression of IL-12p40; however, expression of IL-12p40 by R. equi-stimulated neutrophils from 4- and 8-week-old foals was significantly greater than expression by unstimulated neutrophils. These results demonstrate that R. equi-stimulated neutrophils are a source of many pro-inflammatory cytokines, and that the magnitude of this expression with respect to IL-6, IL-8, and IL-12p40 mRNA expression was influenced by age. Collectively, the data presented indicate a non-phagocytic role for neutrophils that may influence the type of adaptive immune response to R. equi.