Thesis (Ph. D.)--University of Missouri--Columbia, 1998. / Typescript. Vita. Includes bibliographical references (leaves: 138-170). Also available on the Internet.

Thesis (Ph.D.)--Memorial University of Newfoundland, Faculty of Medicine, 1999. / Includes bibliographical references.

Orientador: Rosangela Zacarias Machado / Banca: Solange Maria Gennari / Banca: Aramis Augusto Pinto / Banca: Deise Aparecida de Oliveira Silva / Banca: Ana Patricia Yatsuda Natsui / Resumo: Neospora caninum é um protozoário do Filo Apicomplexa, que foi primeiramente descrito como causa de encefalomielite em filhotes caninos sorologicamente negativos para Toxoplasma gondii. Estudos anteriores neste importante hospedeiro da cadeia epidemiológica de N. caninum demonstram que as respostas de anticorpos IgG são tardiamente detectadas e que a infecção clínica é de difícil indução. Desta forma, este trabalho objetivou o estudo da imunidade de cães frente à infecção oral por N. caninum. Os resultados obtidos a partir de análises de diversos animais experimentalmente infectados indicam que os cães apresentam uma prolongada fase aguda da infecção, com eliminação de oocistos associado à queda nos níveis de linfócitos T CD4+ e CD8+ e diminuição de MHC de classe II por células apresentadoras de antígeno. Adicionalmente, os animais apresentam soroconversão instável durante o mesmo período, sendo que somente IgG1 e IgG3 foram detectados em adultos e filhotes, respectivamente, entre o 2o e 3o mês de infecção. De forma concomitante, observa-se uma predominância da expressão de citocinas imunomoduladoras como TGF 1, IL-4 e IL-10. Após dois meses de infecção, o perfil da resposta se inverte, sendo observado picos de produção dos marcadores CD4 e CD8 de linfócitos T e citocinas próinflamatórias como IFN , IL-6 e IL-12, além do aumento nos títulos de anticorpos, principalmente IgG1 e IgG4 nos cães jovens. Com base nestes resultados, conclui-se que os cães apresentam uma relação de equilíbrio com N. caninum, a qual induz nesta espécie uma modulação da resposta imunológica durante a fase de merogonia. / Abstract: Neospora caninum is an Apicomplexan parasite firstly described as the cause of encephalomyelitis in puppies serologically negative to Toxoplasma gondii. Previous reports on the parasite’s definitive host indicate a late IgG antibody response and that clinical disease is difficult to be induced. The aim of this study was to investigate canine immunity during N. caninum oral infection. The results obtained from the analysis of infected animal’s samples indicate that dogs present a protracted acute phase, with oocyst shedding correlated to a drop in CD4+ and CD8+ T cell levels, and low MHC class II expression by antigen presenting cells. Additionally, the dogs presented an unstable seroconversion pattern in the same period, with only IgG1 and IgG3 being detected in adult dogs and puppies, respectively, between the second and third months of infection. Concomitantly, dominant Th2 cytokine expression was observed, with peak expression levels of TGF 1, IL-4 and IL-10. After 2 months of infection, the immunity profile shifts towards a Th1 response, with high levels of CD4 and CD8 lymphocytary marker production and pro-inflammatory cytokine expression (IFN , IL-6 and IL-12), besides of the raise in antibody levels, especially IgG1 and IgG4 in puppies. Based in the results presented herein, we may conclude that dogs present a balanced host-parasite relationship, modulating the host immune response during N. caninum merogony. / Doutor

Magister Scientiae - MSc / The aim of this study was to investigate the effects of Sutherlandia frutescens on the inflammatory response and T cell differentiation in vitro using cytokines as biomarkers. Whole blood cells containing various concentrations of Sutherlandia frutescens were stimulated in vitro with either Lipopolysaccharide (LPS) or Phytohaemagglutinin (PHA). Results show that Sutherlandia frutescens is not toxic at any of the concentrations tested. The addition of Sutherlandia frutescens at high concentrations to the stimulated whole blood cell cultures reflects a significant down regulation of Interleukin(IL) 6 and IL-10 compared to the control (P<0.05) hence suppressed the inflammatory and humoral immune response. Results obtained for Inteferon-gamma (IFN ) shows that Sutherlandia frutescens is donor specific as it reflects both up and down regulation in the release of IFN at the concentrations tested. The in vitro data generated by this study supports the use of Sutherlandia frutescens in the management of inflammatory conditions and allergies such as asthma. However the effects of Sutherlandia frutescens on cell mediated immunity was found to be donor specific. Further investigation of Sutherlandia frutescens on cellular immunity is advised. / South Africa

The interleukin (IL)-12 family of cytokines plays a key role in the orchestration of cellular immune responses, bridging innate and adaptive immunity. The founding member, IL-12, was discovered in the late 1980s as the first heterodimeric cytokine, composed of a 40 kDa (p40) and 35 kDa (p35) subunit. Years of basic and clinical research on this prototypical T helper type (Th)1 cytokine revealed its importance in immunity to intracellular non-viral infections, as well as in cancer and autoimmune diseases. Since the discovery of IL-23 as another cytokine composed of the p40 subunit of IL-12 in the year 2000, IL-23, rather than IL-12, could be shown to be the key player in rodent models of autoimmune diseases such as multiple sclerosis and rheumatoid arthritis. With accumulating evidence revealing IL-23 as the crucial regulator of a non-classical pathway of cellular immunity which is hallmarked by IL-17 producing T cells it is intriguing to gain understanding of the importance of such findings in immunity to infections. The present work describes a series of in vivo studies investigating the role of endogenous as well as exogenous IL-23 in a murine model of chronic fungal infection, cryptococcosis. To address the role of endogenous IL-23, wild-type (WT), IL-12- (IL-12p35-/-), IL-23- (IL-23p19-/-) deficient, as well as IL-12- and IL-23- double deficient (p40-deficient) mice on a C57BL/6 background were infected with Cryptococcus neoformans (C. neoformans). Following infection, p40-deficient mice demonstrated higher mortality than IL-12p35-/- mice. Reconstitution of p40-deficient mice with recombinant murine IL-23 prolonged their survival to levels similar to IL-12p35-/- mice. IL-23p19-/- mice showed a moderately reduced survival time and delayed fungal clearance in the liver. While interferon (IFN)-γ production was similar in WT and IL-23p19-/- mice, production of IL-17 was strongly impaired in the latter. IL-23p19-/- mice produced fewer hepatic granulomata relative to organ burden and showed defective recruitment of mononuclear cells to the brain. Moreover, activation of microglia cells and expression of IL-1β, IL-6, and MCP-1 in the brain was impaired. SUMMARY - 80 - The second part of the present work explores the mechanisms underlying the IL-23 effects by characterizing the role of exogenous IL-23. C. neoformans-infected C57BL/6 WT mice treated with recombinant murine IL-23 showed significantly prolonged survival time as compared to mock-treated control mice. However, complete survival throughout the observation period (100 days) was only achieved following IL-12 treatment. At day 21 post infection (p.i.) the IL-23-treated mice as well as the IL-12 group had a significantly lower fungal burden in the brain than the control mice. However, while IL-12 treatment was associated with elevated serum levels of the proinflammatory mediators IFN-γ, tumor necrosis factor (TNF)-α and nitric oxide, IL-23-treated animals, although more resistant, developed a Th2 response similar to the control group as measured by serum IgE levels. Further experiments to assess the mechanism of action were based on the finding of reduced fungal burden at the site of infection, the peritoneal cavity, at day 8 p.i. following IL-23 treatment. This microbicidal effect was also seen in p40-deficient as well as in T and B cell deficient (RAG-deficient) mice. Administration of IL-23 led to enhanced recruitment of inflammatory cells, not only of T cells but also cells of the innate immune system such as DCs, natural killer cells and granulocytes to the infected site. Although numbers of macrophages were not altered following IL-23 treatment, co-stimulatory molecules were markedly up-regulated on such cells. The chemokine/cytokine pattern induced by IL-23 treatment was hallmarked by proinflammatory mediators such as MCP-1, IL-1β, IL-6, TNF-α and IL-17, but also the Th2 associated cytokine IL-5. From these results it can be concluded that a non-classical immune pathway induced by IL-23 complements the more dominant role of IL-12 in protection against C. neoformans. This novel immune response is characterized by an enhancement of the inflammatory cell response and the production of a proinflammatory cytokine pattern hallmarked by IL-1β, IL-6, TNF-α and IL-17.

Arsenic is a universal environmental toxicant associated mostly with skin related diseases in people exposed to low doses over a long term. Low dose arsenic trioxide (ATO) with long exposure will lead to chronic exposure. Experiments were performed to provide new knowledge on the incompletely understood mechanisms of action of chronic low dose inorganic arsenic in keratinocytes. Cytotoxicity patterns of ATO on long-term cultures of HaCaT cells on collagen IV was studied over a time course of 14 days. DNA damage was also assessed. The percentages of viable cells after exposure were measured on Day 2, Day 5, Day 8, and Day 14. Statistical and visual analytics approaches were used for data analysis. In the result, a biphasic toxicity response was observed at a 5 μg/ml dose with cell viability peaking on Day 8 in both chronic and acute exposures. Furthermore, a low dose of 1 μg/ml ATO enhanced HaCaT keratinocyte proliferation but also caused DNA damage. Global gene expression study using microarray technique demonstrated differential expressions of genes in HaCaT cell exposed to 0.5 μg/ml dose of ATO up to 22 passages. Four of the up-regulated and 1 down-regulated genes were selected and confirmed with qRT-PCR technique. These include; Aldo-Keto Reductase family 1, member C3 (AKR1C3), Insulin Growth Factor-Like family member 1 (IGFL1), Interleukin 1 Receptor, type 2 (IL1R2) and Tumour Necrosis Factor [ligand] Super-Family, member 18 (TNFSF18), and down-regulated Regulator of G-protein Signalling 2 (RGS2). The decline in growth inhibiting gene (RGS2) and increase in AKR1C3 may be the contributory path to chronic inflammation leading to metaplasia. This pathway is proposed to be a mechanism leading to carcinogenesis in skin keratinocytes. The observed over expression of IGFL1 may be a means of triggering carcinogenesis in HaCaT keratinocytes. In conclusion, it was established that at very low doses, arsenic is genotoxic and induces aberrations in gene expression though it may appear to enhance cell proliferation. The expression of two genes encoding membrane proteins IL1R2 and TNFSF18 may serve as possible biomarkers of skin keratinocytes intoxication due to arsenic exposure. This research provides insights into previously unknown gene markers that may explain the mechanisms of arsenic-induced dermal disorders including skin cancer / Environmental Sciences / D. Phil. (Environmental science)

Doctor of Philosophy / Department of Anatomy and Physiology / Deryl L. Troyer / Many cell types were known to have migratory properties towards tumors and different research groups have shown reliable results regarding cells as delivery vehicles of therapeutics for targeted cancer treatment. Present report discusses proof of concept for 1. Cell mediated delivery of Magnetic nanoparticles (MNPs) and targeted Magnetic hyperthermia (MHT) as a cancer treatment by using in vivo mouse cancer models, 2. Cells surface engineering with chimeric proteins for targeted cancer treatment by using in vitro models. 1. Tumor homing cells can carry MNPs specifically to the tumor site and tumor burden will decrease after alternating magnetic field (AMF) exposure. To test this hypothesis, first we loaded Fe/Fe3O4 bi-magnetic NPs into neural progenitor cells (NPCs), which were previously shown to migrate towards melanoma tumors. We observed that NPCs loaded with MNPs travel to subcutaneous melanoma tumors. After alternating magnetic field (AMF) exposure, the targeted delivery of MNPs by the NPCs resulted in a mild decrease in tumor size (Chapter-2). Monocytes/macrophages (Mo/Ma) are known to infiltrate tumor sites, and also have phagocytic activity which can increase their uptake of MNPs. To test Mo/Ma-mediated MHT we transplanted Mo/Ma loaded with MNPs into a mouse model of pancreatic peritoneal carcinomatosis. We observed that MNP-loaded Mo/Ma infiltrated pancreatic tumors and, after AMF treatment, significantly prolonged the lives of mice bearing disseminated intraperitoneal pancreatic tumors (Chapter-3). 2. Targeted cancer treatment could be achieved by engineering tumor homing cell surfaces with tumor proteases cleavable, cancer cell specific recombinant therapeutic proteins. To test this, Urokinase and Calpain (tumor specific proteases) cleavable; prostate cancer cell (CaP) specific (CaP1 targeting peptide); apoptosis inducible (Caspase3 V266ED3)- rCasp3V266ED3 chimeric protein was designed in silico. Hypothesized membrane anchored chimeric protein (rCasp3V266ED3, rMcherry red) plasmids were constructed. Membrane anchoring and activity of designed proteins were analyzed in RAW264.7 Mo/Ma and HEK293 cells in vitro. Further, Urokinase (uPA) mediated cleavage and release of rCasp3V266ED3 from engineered cells was tested (Chapter-4). Animal models for cancer therapy are invaluable for preclinical testing of potential cancer treatments. Final chapter of present report shows evidence for immune-deficient line of pigs as a model for human cancers (Chapter-5)

A infecção de cães pela Leishmania (Leishmania) infantum resulta em um espectro de manifestações imunopatológicas que dependem da interação parasito hospedeiro e são definidas por fatores ambientais e pela genética do hospedeiro. Apesar disso, a imunogenética da leishmaniose visceral canina (LVC) permanece inexplorada. Nós realizamos diagnóstico laboratorial, clínico, ensaio de linfoproliferação (LPA), quantificação de citocinas, teste de hipersensibilidade tardia à leishmanina (LST), quantificação de IgA, IgE, IgG, IgM anti L. (L. ) infantum, IgG anti saliva de flebotomíneo e genotipagem ampla afim de identificar polimorfismos de nucleotídeo único (SNPs) associados aos diferentes perfis de imunidades celular, humoral, resposta clínica e status de infecção em cães de área endêmica, utilizando modelo de componente de variância (EMMAX). O efeito de estrutura da amostra foi controlado em todas as análises. A presença ou ausência de infecção pela L. (L. ) infantum foi associado a regiões contendo os genes PRGR_CANFA, RAB38, NOX4, PRKCI e SMAD7, IL1RA, IL12A_CANFA relacionados à ativação de fagócitos, mecanismos microbicidas, sobrevivência intracelular de patógenos e resposta pró inflamatória; a resposta clínica foi associada a regiões contendo os genes CATA_CANFA, LIAS, IL17A e IL17F relacionados à proteção contra danos do estresse oxidativo e indução de resposta pró inflamatória; o resultado LST+ foi associado à resposta Th1, controle da infecção mas não preveniu a manifestação de sinais clínicos, enquanto o LST- foi associado à resposta Treg e aumento do parasitismo. O resultado do LST foi associado a regiões contendo os genes MEP1B, PTPRM, TLN1, TGFBR1, ITGA9, EPCAM e CALM1 envolvidos com maturação de fagócitos, migração e adesão de leucócitos, estabilidade da sinapse imunológica, diferenciação e proliferação de linfócitos. A linfoproliferação foi dependente da carga parasitária e associada a regiões contendo os genes FOCAD, PIAS2, SMAD2 e IL6R envolvidos com supressão tumoral e diferenciação de linfócitos Th17 ou Treg; o aumento dos níveis de IgA, IgE e IgG anti L. (L. ) infantum foi associado à LVC sintomática enquanto o de IgG anti saliva de Lutzomyia longipalpis foi associado à exposição e infecção assintomática. Quanto à resposta de IgM, foram identificados SNPs nos genes NXN e SH3BP5 relacionados com inibição do crescimento, diferenciação e ativação de linfócitos B e vias de sinalização de TLR4 e TLR9; para IgG anti - L. (L. ) infantum foram identificadas regiões contendo os genes IL17RB, SH2B3 e replicação do loci de susceptibilidade NOX4, RAB38, CTSC envolvidos com linfopoiese, citocinese, resposta pró inflamatória, mecanismos microbicidas, sobrevivência intracelular de Leishmania; os níveis de IgA foram associados a regiões contendo os genes LIN28A e MAFB implicados na predisposição à nefropatia glomerular, já os níveis de IgG anti saliva de Lu. longipalpis foram associados a regiões contendo os genes ERBB2IP, CD180, RAB7A_CANFA, FOXP1, RUNX1, SOD1, Q3HTU8_CANFA, IFNAR1, IFNAR2 e IFNGR2 envolvidos com supressão celular, produção de imunoglobulina via TLR4 e sobrevivência intracelular de Leishmania. Esses resultados apontam regiões cromossômicas úteis para a elucidação da resposta à infecção por L. (L. ) infantum em cães e alvos potenciais para estudos funcionais, estratégias profiláticas e terapêuticas / Leishmania (Leishmania) infantum infection in dogs leads to a range of immunopathological responses, which depend on a parasite - host interaction and are defined by environmental factors and genetic of host. Neverthless, immunogenetic of the canine leishmaniasis (CanL) remains unexplored. We performed diagnosis and clinical evaluation, lymphoproliferation assay (LPA), leishmanin skin test (LST), quantification of cytokine, anti-L. (L. ) infantum IgA, IgE, IgG, IgM, anti- sandfly saliva IgG levels and genome wide association scan of 110.165 SNPs (GWAS) in order to indentify loci associated to clinical, antibody, cell-mediated responses and status of infection in 189 dogs, employng a expedited efficient mix model of association (EMMAX). Control of stratification effects due to sample structure was evideced by the low inflation factors. Status of infection was associated to SNPs in linkage desequilibrium (LD) with PRGR_CANFA, RAB38, NOX4, PRKCI and in the neighborhood of SMAD7, IL1RA, IL12A_CANFA genes related to phagocyte maturation, killing of pathogens and proinflamatory response; clinical outcome was associated to CATA_CANFA, LIAS, IL17A, IL17F loci involved in prevention of oxidative burst mediated injury and proinflamatory response; LST+ was associated to Th1 response although it has not prevented symptoms, whereas LST- was associated to Treg response and enhanced parasite load. Overall, LST response was associated to MEP1B, PTPRM, TLN1, TGFBR1, ITGA9, EPCAM e CALM1 loci committed to phagocyte maturation, leukocyte adhesion and migration, stability in immunological synapse, lymphocyte diferenciation and proliferation. Lymphocyte proliferation was rely on parasit burden and associated to FOCAD, PIAS2 loci in the neighborhood of SMAD2 e IL6R genes, wich are implicated in tumor supression and Treg/Th17 decision; Increased levels of anti-L. (L. ) infantum IgA, IgE, IgG were observed in severity of CanL. In contrast, anti- sandfly saliva IgG was enhenced in asymptomatic dogs. IgM response was associated to NXN e SH3BP5 loci related to fate, growing and activation of B cells; anti-L. (L. ) infantum IgG levels was associated to region containing IL17RB, SH2B3 and replication of NOX4, RAB38, CTSC suscptibility loci involved in proinflamatory response, microbicidal activity, lymphopoiesis and cytokinesis; IgA levels were associated to SNPs on LIN28A and MAFB, wich are implicated glomerular nephropathy, whereas anti-sandfly saliva IgG levels were associated to ERBB2IP, CD180, RAB7A_CANFA, FOXP1, RUNX1, SOD1, Q3HTU8_CANFA, IFNAR1, IFNAR2 e IFNGR2 loci, wich are related to supression of inflamation, antibody response through the TLR4 pathway and survival of intracellular pathogens. These findins provide insights for responses to L. (L. ) infantum infection and point to potential targets for functional investigations, therapeutic and prophylactic strategies

A infecção de cães pela Leishmania (Leishmania) infantum resulta em um espectro de manifestações imunopatológicas que dependem da interação parasito hospedeiro e são definidas por fatores ambientais e pela genética do hospedeiro. Apesar disso, a imunogenética da leishmaniose visceral canina (LVC) permanece inexplorada. Nós realizamos diagnóstico laboratorial, clínico, ensaio de linfoproliferação (LPA), quantificação de citocinas, teste de hipersensibilidade tardia à leishmanina (LST), quantificação de IgA, IgE, IgG, IgM anti L. (L. ) infantum, IgG anti saliva de flebotomíneo e genotipagem ampla afim de identificar polimorfismos de nucleotídeo único (SNPs) associados aos diferentes perfis de imunidades celular, humoral, resposta clínica e status de infecção em cães de área endêmica, utilizando modelo de componente de variância (EMMAX). O efeito de estrutura da amostra foi controlado em todas as análises. A presença ou ausência de infecção pela L. (L. ) infantum foi associado a regiões contendo os genes PRGR_CANFA, RAB38, NOX4, PRKCI e SMAD7, IL1RA, IL12A_CANFA relacionados à ativação de fagócitos, mecanismos microbicidas, sobrevivência intracelular de patógenos e resposta pró inflamatória; a resposta clínica foi associada a regiões contendo os genes CATA_CANFA, LIAS, IL17A e IL17F relacionados à proteção contra danos do estresse oxidativo e indução de resposta pró inflamatória; o resultado LST+ foi associado à resposta Th1, controle da infecção mas não preveniu a manifestação de sinais clínicos, enquanto o LST- foi associado à resposta Treg e aumento do parasitismo. O resultado do LST foi associado a regiões contendo os genes MEP1B, PTPRM, TLN1, TGFBR1, ITGA9, EPCAM e CALM1 envolvidos com maturação de fagócitos, migração e adesão de leucócitos, estabilidade da sinapse imunológica, diferenciação e proliferação de linfócitos. A linfoproliferação foi dependente da carga parasitária e associada a regiões contendo os genes FOCAD, PIAS2, SMAD2 e IL6R envolvidos com supressão tumoral e diferenciação de linfócitos Th17 ou Treg; o aumento dos níveis de IgA, IgE e IgG anti L. (L. ) infantum foi associado à LVC sintomática enquanto o de IgG anti saliva de Lutzomyia longipalpis foi associado à exposição e infecção assintomática. Quanto à resposta de IgM, foram identificados SNPs nos genes NXN e SH3BP5 relacionados com inibição do crescimento, diferenciação e ativação de linfócitos B e vias de sinalização de TLR4 e TLR9; para IgG anti - L. (L. ) infantum foram identificadas regiões contendo os genes IL17RB, SH2B3 e replicação do loci de susceptibilidade NOX4, RAB38, CTSC envolvidos com linfopoiese, citocinese, resposta pró inflamatória, mecanismos microbicidas, sobrevivência intracelular de Leishmania; os níveis de IgA foram associados a regiões contendo os genes LIN28A e MAFB implicados na predisposição à nefropatia glomerular, já os níveis de IgG anti saliva de Lu. longipalpis foram associados a regiões contendo os genes ERBB2IP, CD180, RAB7A_CANFA, FOXP1, RUNX1, SOD1, Q3HTU8_CANFA, IFNAR1, IFNAR2 e IFNGR2 envolvidos com supressão celular, produção de imunoglobulina via TLR4 e sobrevivência intracelular de Leishmania. Esses resultados apontam regiões cromossômicas úteis para a elucidação da resposta à infecção por L. (L. ) infantum em cães e alvos potenciais para estudos funcionais, estratégias profiláticas e terapêuticas / Leishmania (Leishmania) infantum infection in dogs leads to a range of immunopathological responses, which depend on a parasite - host interaction and are defined by environmental factors and genetic of host. Neverthless, immunogenetic of the canine leishmaniasis (CanL) remains unexplored. We performed diagnosis and clinical evaluation, lymphoproliferation assay (LPA), leishmanin skin test (LST), quantification of cytokine, anti-L. (L. ) infantum IgA, IgE, IgG, IgM, anti- sandfly saliva IgG levels and genome wide association scan of 110.165 SNPs (GWAS) in order to indentify loci associated to clinical, antibody, cell-mediated responses and status of infection in 189 dogs, employng a expedited efficient mix model of association (EMMAX). Control of stratification effects due to sample structure was evideced by the low inflation factors. Status of infection was associated to SNPs in linkage desequilibrium (LD) with PRGR_CANFA, RAB38, NOX4, PRKCI and in the neighborhood of SMAD7, IL1RA, IL12A_CANFA genes related to phagocyte maturation, killing of pathogens and proinflamatory response; clinical outcome was associated to CATA_CANFA, LIAS, IL17A, IL17F loci involved in prevention of oxidative burst mediated injury and proinflamatory response; LST+ was associated to Th1 response although it has not prevented symptoms, whereas LST- was associated to Treg response and enhanced parasite load. Overall, LST response was associated to MEP1B, PTPRM, TLN1, TGFBR1, ITGA9, EPCAM e CALM1 loci committed to phagocyte maturation, leukocyte adhesion and migration, stability in immunological synapse, lymphocyte diferenciation and proliferation. Lymphocyte proliferation was rely on parasit burden and associated to FOCAD, PIAS2 loci in the neighborhood of SMAD2 e IL6R genes, wich are implicated in tumor supression and Treg/Th17 decision; Increased levels of anti-L. (L. ) infantum IgA, IgE, IgG were observed in severity of CanL. In contrast, anti- sandfly saliva IgG was enhenced in asymptomatic dogs. IgM response was associated to NXN e SH3BP5 loci related to fate, growing and activation of B cells; anti-L. (L. ) infantum IgG levels was associated to region containing IL17RB, SH2B3 and replication of NOX4, RAB38, CTSC suscptibility loci involved in proinflamatory response, microbicidal activity, lymphopoiesis and cytokinesis; IgA levels were associated to SNPs on LIN28A and MAFB, wich are implicated glomerular nephropathy, whereas anti-sandfly saliva IgG levels were associated to ERBB2IP, CD180, RAB7A_CANFA, FOXP1, RUNX1, SOD1, Q3HTU8_CANFA, IFNAR1, IFNAR2 e IFNGR2 loci, wich are related to supression of inflamation, antibody response through the TLR4 pathway and survival of intracellular pathogens. These findins provide insights for responses to L. (L. ) infantum infection and point to potential targets for functional investigations, therapeutic and prophylactic strategies

<p>This study shows that herbs can be effectively screened for potiential bio-activity using in vitro methods. Further studies will be needed to better explore Artemisia afra&rsquo / s effect on immunoregulation, particularly long term effects of the herb on the immune system and its effect on other disease states.</p>