Virtually all multicellular organisms rely on a highly conserved innate immune system for defense against foreign microorganisms. Innate immunity consists primarily of a humeral response that culminates in the expression of antimicrobial peptides. In contrast to adaptive immunity seen in high order organisms, the innate immune response is not specific to the invader. In aging organisms, some of the most dramatic transcriptional changes take place within the innate immune system. In aging mammals, innate immune reorganization coincides with declining immune function, which often manifests itself as chronic inflammation. Similar to this state of chronic inflammation in mammals, Drosophila exhibit a marked upregulation of many innate immunity related genes. However, it remains unclear if this upregulation results in a similar decrease in immune function to that seen in mammals. If Drosophila is to be considered as a model organism in which to study the relationship between immunity and aging, it must first be determined whether it too undergoes declining immune function with age. By examining the response to quantifiable injections of bacteria, we were able to deduce that adult Drosophila do indeed undergo immune senescence. Elderly wildtype flies infected with various doses of bacteria showed a decreased ability to survive infection. Moreover, because the ability to clear the infection remains intact despite decreased survival following infection, it is believed that a bacterially produced factor is responsible for immune senescence in adult Drosophila. / Thesis (Master, Biology) -- Queen's University, 2007-10-02 09:37:18.314

The neonatal period represents a critical time period in the development of the immune system. Adaptive human immune responses are generally viewed as immature at birth. However little is known about innate immune capacity at birth. The TLR system plays an integral role as pattern recognition receptors in the innate immune response. It is also critical in initiating and regulating the adaptive immune response. Preterm birth is associated with increased risk of developing infections in early life and has been associated with increased risk of development of other chronic disorders in later life; however the underlying mechanisms are not at all well understood. Recently, late preterm neonates (34-36 weeks gestation vs full term, 37+ weeks) have been identified as having significantly greater risks of morbidity and mortality in the perinatal period than their full term counterparts. Hence, we focus on examination of TLR responses in late preterm and full term neonates to better understand immune potential and function in these populations. We examined cord blood cytokine and chemokine responses following stimulation with a broad range of TLR agonists. Our results show for the first time that late preterm neonates have reduced capacity to produce both pro- and anti-inflammatory cytokines following stimulation with a panel of TLR agonists. This reduced responsiveness was not due to a reduction in the number of responding cells, but instead appears to be mediated by a reduction in the intrinsic levels of expression of TLRs and associated adaptor proteins. Because little is known about how the innate immune system develops throughout life, we next compared TLR responses in full term neonates to 7 children, adolescents and adults. We found that neonates had selective impairments in TLR responses, most notably in anti-inflammatory cytokine production and anti-viral immune responses compared to the other age groups. Epigenetic modifications, such as the addition or removal of acetyl groups to histone proteins by histone acetyl transferase (HAT) and histone deactylase (HDAC) respectively, are able to modify the expression of genes. Hence, environmental stimuli have been shown to influence gene expression in part by modifying the level or activity of these epigenetic regulators. Currently there are no studies which have examined how epigenetic modifications may influence neonatal innate immune responses. Hence, we sought to determine how modulation of endogenous HDAC activity would affect neonatal innate immune responses. We found that inhibition of HDAC had both inhibitory and enhancing effects on cytokine expression depending on the TLR pathway activated, indicating that the endogenous HDAC expression does not have a global inhibitory impact on all TLR-dependent responses. In summary, this body of work demonstrates that neonatal innate immune responses vary depending on gestational age, indicating that the final few weeks of gestation are crucial for maturation of responses to both bacteria and viruses. Neonates respond differently to TLR stimuli than do older individuals, further highlighting a maturation process of the innate immune system which continues throughout life. Finally, we have shown that environmental exposures may have powerful effects on immune responses in early life.

No description available.

The neonatal period represents a critical time period in the development of the immune system. Adaptive human immune responses are generally viewed as immature at birth. However little is known about innate immune capacity at birth. The TLR system plays an integral role as pattern recognition receptors in the innate immune response. It is also critical in initiating and regulating the adaptive immune response. Preterm birth is associated with increased risk of developing infections in early life and has been associated with increased risk of development of other chronic disorders in later life; however the underlying mechanisms are not at all well understood. Recently, late preterm neonates (34-36 weeks gestation vs full term, 37+ weeks) have been identified as having significantly greater risks of morbidity and mortality in the perinatal period than their full term counterparts. Hence, we focus on examination of TLR responses in late preterm and full term neonates to better understand immune potential and function in these populations. We examined cord blood cytokine and chemokine responses following stimulation with a broad range of TLR agonists. Our results show for the first time that late preterm neonates have reduced capacity to produce both pro- and anti-inflammatory cytokines following stimulation with a panel of TLR agonists. This reduced responsiveness was not due to a reduction in the number of responding cells, but instead appears to be mediated by a reduction in the intrinsic levels of expression of TLRs and associated adaptor proteins. Because little is known about how the innate immune system develops throughout life, we next compared TLR responses in full term neonates to 7 children, adolescents and adults. We found that neonates had selective impairments in TLR responses, most notably in anti-inflammatory cytokine production and anti-viral immune responses compared to the other age groups. Epigenetic modifications, such as the addition or removal of acetyl groups to histone proteins by histone acetyl transferase (HAT) and histone deactylase (HDAC) respectively, are able to modify the expression of genes. Hence, environmental stimuli have been shown to influence gene expression in part by modifying the level or activity of these epigenetic regulators. Currently there are no studies which have examined how epigenetic modifications may influence neonatal innate immune responses. Hence, we sought to determine how modulation of endogenous HDAC activity would affect neonatal innate immune responses. We found that inhibition of HDAC had both inhibitory and enhancing effects on cytokine expression depending on the TLR pathway activated, indicating that the endogenous HDAC expression does not have a global inhibitory impact on all TLR-dependent responses. In summary, this body of work demonstrates that neonatal innate immune responses vary depending on gestational age, indicating that the final few weeks of gestation are crucial for maturation of responses to both bacteria and viruses. Neonates respond differently to TLR stimuli than do older individuals, further highlighting a maturation process of the innate immune system which continues throughout life. Finally, we have shown that environmental exposures may have powerful effects on immune responses in early life.

No description available.

No description available.

This study investigated the immunological aspects of Entamoeba histolytica infection in Syrian hamsters. Immunization of hamsters by an intradermal injection with live axenic amoebae, conferred complete protection against amoebic liver abscess. Protection was achieved with homologous or heterologous strains of E. histolytica and was dose-dependent. Combination of thymectomy and anti T-cell serum treatment significantly increased the size of liver abscess and metastatic dissemination of the parasite. It was shown that a cell-mediated immune response controls the healing of skin ulcers in vaccinated animals and thus confers on them resistance to intrahepatic amoebic challenge infection. Resistance to hepatic infection with E. histolytica can be passively transferred to normal recipients with sensitized cells, but not with immune serum. Sensitized cells from vaccinated, protected or infected animal kill E. histolytica trophozoites in vitro. Splenectomy reduces the resistance of hamsters to amoebic infection. It is suggested that macrophages are required in the host defence against the establishment of amoebic abscess in the liver and dissemination of amoebae to other organs.

The incidences of endogenous viral genes (ev-genes) and myb-gene polymorphisms were determined in substrains of strains R, M and G which had been divergently selected or susceptibility to tumour formation induced by Rous Sarcoma virus (RSV) of type A and B. Frequencies of myb gene polymorphisms were also determined in two replicates of strains selected for high and low multiple immune response to challenge with Pasteurella multocida and Mycoplasma gallisepticum. Strains R, M and G were found to contain different sets of ev-genes reflecting their distinct genetic origins. Among eleven ev-genes identified, two showed a significantly increased frequency in the susceptible substrains. One was ev-6, which expresses the viral envelope protein of the endogenous avian leucosis, while the other was a new endogenous viral gene New-E, whose phenotype is unknown. A significant increased incidence of ev-4, reported to be a silent ev-gene, was observed in resistant rather than susceptible substrain. Myb gene polymorphisms were assessed using a cDNA probe and a genomic probe yielding 2 and 3 restriction fragment length polymorphisms (RFLPs) respectively. In strains M and G, only one polymorphism (PM5$ sp+$) observed at an Msp I site located downstream of the last of the c-myb exons was found to be significantly co-selected for susceptibility. Analysis of RFLPs of myb-gene in strains selected for high or low multiple immune response did not reveal any significant response to selection. Rather, polymorphisms seemed to reflect a founder effect as revealed by opposite frequencies obtained in the two replicates. DNA methylation, a possible epigenetic mechanism regulating gene expression, was also investigated in the myb-gene. DNA from semen, blood, spleen, liver and thymus was extracted from organs obtained from chickens at different ages.

Four experiments evaluated the impact of probiotic administration on the immune response of broilers vaccinated with a live coccidiosis vaccine. Experiment one showed that probiotic administration increased heterophil and monocyte oxidative burst, and lymphocyte proliferation at multiple time points. In experiment two, probiotic + vaccine increased heterophil and monocyte oxidative burst on d 15 when compared with the negative controls. Overall, vaccine administration alone showed the highest response when compared to all other treatments. In the second trial, all birds were exposed to Eimeria oocysts in the litter and oral gavaged. The results showed that probiotic + vaccine resulted in greater heterophil and monocyte oxidative burst levels on d 14 and 28 when compared to the negative controls. Increases in lymphocyte proliferation were also seen in the probiotic + vaccine and probiotic alone broilers on d 14 among other treatments. In experiment three, heterophil oxidative burst was increased (p <= 0.05) in the vaccine alone group, vaccine with probiotic group, and the ionophore with probiotic group, when compared to the negative control. Monocyte oxidative burst was increased (p <= 0.05) in the vaccine with probiotic group on d 36 and 43, compared to the negative control. Lymphocyte proliferation was greater (p <= 0.05) on d 22 and 36 in the ionophore with probiotic group, when compared to the negative control. Experiment four showed that liver AVBD 2 gene expression elevated (p <= 0.05) in the probiotic + vaccine group relative to the probiotic alone group. Ileum AVBD 2 gene expression was not affected among any of the treatments was evaluated. Liver AVBD 9 was demonstrated to have higher (p <= 0.05) gene expression in the vaccine group when compared to controls. When AVBD 9 gene expression was evaluated in the ileum, a decrease (p <= 0.05) was observed in all treatments compared to the control group. These data suggest that simultaneous administration of probiotics during coccidiosis vaccination or ionophore treatment has the ability to modulate the immune response at varying time points.

The hepatitis C virus (HCV) currently infects 3% of the world???s population, with chronic infection in 50-80% of exposed individuals. A small subset of individuals who are exposed to HCV do not develop anti-HCV antibodies, persistent viraemia or chronic hepatitis despite generating HCV-specific CD4+ and CD8+ T cells. These individuals are believed to develop an immune response which rapidly clears viraemia prior to the induction of an antibody response. Circumstantial evidence supports the likelihood that some of these individuals may generate these same responses and outcomes on repeated occasions of HCV infection. HCV-specific cellular immune responses in seronegative subjects have been the subject of only limited prior study, in part due to the lack of appropriate recombinant antigens and assay systems. Therefore, this thesis described the development and validation of an interferon-? (IFN-?) ELISPOT assay using overlapping peptides (n=441). Using this assay, HCV-specific cellular immune responses were detected in 5/10 (50%) of chronically infected subjects. Responses were identified more frequently, and were directed against more regions of the HCV genome, than with traditional assay systems. This IFN-? ELISPOT assay, a comparable interleukin (IL)-2 ELISPOT assay, and a multiplex in vitro cytokine production assay were then used to evaluate HCV-specific cellular immune responses in three cohorts of seronegative subjects at high-risk of exposure to HCV ??? babies born to infected mothers, multiply-transfused subjects with thalassaemia, and high risk injecting drug users. Cellular immune responses were evaluated in 23 infants born to HCV-antibody positive women. Responses were not detected in infants born to HCV-PCR negative mothers. IFN-? production was detected in 1/11 infants born to viraemic mothers using the ELISPOT assay, with cytokine production observed in an additional 3/5 infants studied using the in vitro cytokine production assay. HCV-specific cellular immune responses were assessed in a cohort of multiply transfused subjects with thalassaemia using assays for cytotoxic T lymphocyte activity, IFN-? and IL-2 ELISPOT, as well as lymphocyte proliferation and in vitro cytokine production. Responses were detected in 6/13 chronically infected subjects (46%), 4/7 subjects who had cleared infection (71%), and 14/17 seronegative subjects (82%). The seronegative subjects had responses which were broader and higher in magnitude than those with chronic HCV infection, although lower and narrower than in subjects who had cleared prior HCV infection. IFN-? and IL-2 ELISPOT assays, in additional to in vitro cytokine production assays, were performed on 41 injecting drug users (IDUs), with responses detected in 6 (15%). Seronegative IDUs with HCV-specific cellular immune responses had been injecting for a mean of 7.7 years, and reported multiple risk factors for exposure to HCV. The combined data from these three cohorts indicate that the HCV-specific cellular immune responses detected in seronegative subjects were generally broad in specificity. Cytokine production was generally Th1-biased, a pattern which has previously been associated with an increased likelihood of clearance in primary infection. The findings also suggest that responses can be maintained for decades after exposure, and may provide protection against repeated exposures. In summary, cellular immunity against HCV is evident in some seronegative high risk subjects, suggesting that the cellular immune responses may efficiently facilitate viral clearance. Understanding the mechanisms of this immune response pattern will allow better understanding of the host response to HCV and may provide key insights into vaccine design.