Low CXCL13 Expression, Splenic Lymphoid Tissue Atrophy and Germinal Center Disruption in Severe Canine Visceral Leishmaniasis

Visceral leishmaniasis is associated with atrophy and histological disorganization of splenic compartments. In this paper, we compared organized and disorganized splenic lymphoid tissue from dogs naturally infected with Leishmania infantum assessing the size of the white pulp compartments, the distribution of T, B and S100+ dendritic cells, using immunohistochemistry and morphometry and the expression of CCR7 and the cytokines, CXCL13, lymphotoxin (LT)-α, LT-β, CCL19, CCL21, TNF-α, IL-10, IFN-γ and TGF-β, using by real time RT-PCR. The lymphoid follicles and marginal zones were smaller (3.2 and 1.9 times, respectively; Mann-Whitney, P<0.02) in animals with disorganized splenic tissue in comparison to those with organized splenic lymphoid tissue. In spleens with disorganized lymphoid tissue, the numbers of T cells and S100+ dendritic cells were decreased in the follicles, and the numbers of B cells were reduced in both the follicles and marginal zones. CXCL13 mRNA expression was lower in animals with disorganized lymphoid tissue (0.5±0.4) compared to those with organized lymphoid tissue (2.7±2.9, both relative to 18S expression, P = 0.01). These changes in the spleen were associated with higher frequency of severe disease (7/12) in the animals with disorganized than in animals with organized (2/13, Chi-square, P = 0.01) splenic lymphoid tissue. The data presented herein suggest that natural infection with Leishmania infantum is associated with the impairment of follicular dendritic cells, CXCL13 expression, B cell migration and germinal center formation and associates these changes with severe clinical forms of visceral leishmaniasis. Furthermore the fact that this work uses dogs naturally infected with Leishmania infantum emphasizes the relevance of the data presented herein for the knowledge on the canine and human visceral leishmaniasis

Severe clinical presentation of visceral leishmaniasis in naturally infected dogs with disruption of the splenic white pulp.

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2014-05-23T13:30:54Z No. of bitstreams: 1 Lima IS Severe clinical....pdf: 446652 bytes, checksum: 95b0b58332d2883e5a7c188c1db02d5e (MD5)Made available in DSpace on 2014-05-23T13:30:54Z (GMT). No. of bitstreams: 1 Lima IS Severe clinical....pdf: 446652 bytes, checksum: 95b0b58332d2883e5a7c188c1db02d5e (MD5) Previous issue date: 2014Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilCentro de Referência em Doenças Endêmicas Pirajá da Silva (PIEJ). Jequié, BA, BrasilCentro de Referência em Doenças Endêmicas Pirajá da Silva (PIEJ). Jequié, BA, BrasilUniversidade Federal da Bahia. Escola de Medicina Veterinária. Salvador, BA, BrasilUniversidade Federal da Amazônia. Faculdade de Ciências Farmacêuticas. Manaus, AM, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Escola de Medicina Veterinária. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Salvador, BA, BrasilIn this work, we investigated the association between the disruption of splenic lymphoid tissue and the severity of visceral leishmaniasis in dogs. Clinical and laboratory data from 206 dogs were reviewed. Spleen sections collected during the euthanasia of these animals were analyzed, and the splenic lymphoid tissue samples were classified as well organized (spleen type 1), slightly disorganized (spleen type 2), or moderately to extensively disorganized (spleen type 3). Of 199 dogs with evidence of Leishmania infection, 54 (27%) had spleen type 1, 99 (50%) had spleen type 2, and 46 (23%) had spleen type 3. The number of clinical signs associated with visceral leishmaniasis was significantly higher in the animals with evidence of Leishmania infection and spleen type 2 or 3 than in the animals with spleen type 1. Alopecia, anemia, dehydration, dermatitis, lymphadenopathy, and onychogryphosis were all more frequent among animals with evidence of Leishmania infection and spleen type 3 than among the dogs with evidence of Leishmania infection and spleen type 1. The association between the severity of canine visceral leishmaniasis and the disorganization of the splenic lymphoid tissue was even more evident in the group of animals with positive spleen culture. Conjunctivitis and ulceration were also more common in the animals with spleen type 3 than in the animals with spleen type 1. The serum levels (median, interquartile range) of albumin (1.8, 1.4–2.3 g/dL) and creatinine (0.7, 0.4–0.8 mg/dL) were significantly lower and the serum levels of aspartate aminotransferase were significantly higher (57, 39–95 U) in animals with spleen type 3 than in animals with spleen type 1 (2.8, 2.4–3.4 g/dL; 0.9, 0.7–1.2 mg/dL and 23, 20–32 U, respectively). Our data confirm the hypothesis that disruption of the splenic lymphoid tissue is associated with a more severe clinical presentation of canine visceral leishmaniasis

Laboratory findings of included non-infected dogs, dogs without active infection with organized white pulp (TYPE1SC), infected dogs with organized white pulp (TYPE1SC+) and infected animals with disorganized white pulp (TYPE3SC+).

<p>Laboratory findings of included non-infected dogs, dogs without active infection with organized white pulp (TYPE1SC), infected dogs with organized white pulp (TYPE1SC+) and infected animals with disorganized white pulp (TYPE3SC+).</p

An assessment of the genetic diversity of Leishmania infantum isolates from infected dogs in Brazil.

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2014-05-30T14:14:34Z No. of bitstreams: 1 Batista LFS An Assessment of the....pdf: 276239 bytes, checksum: 4e9777b507a531756c6e08755b0b58e8 (MD5)Made available in DSpace on 2014-05-30T14:14:34Z (GMT). No. of bitstreams: 1 Batista LFS An Assessment of the....pdf: 276239 bytes, checksum: 4e9777b507a531756c6e08755b0b58e8 (MD5) Previous issue date: 2012Fundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilUniversidade Federal de Minas Gerais. Laboratório de Genética Bioquímica. Departamento de Bioquímica e Imunologia. ICB. Belo Horizonte, MG,BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilUniversidade Federal da Bahia. Hospital Universitário Professor Edgard Santos. Serviço de Imunologia. Salvador, BA, BrasilUniversidade Federal de Mato Grosso do Sul. Departamento de Morfofisiologia. Campo Grande, MS, BrasilPrefeitura Municipal de Campo Grande. Secretaria Municipal de Saúde Pública Sesau. Centro de Controle de Zoonoses. Campo Grande, MS, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilLaboratório de Genética Bioquímica. Departamento de Bioquímica e Imunologia. ICB. Universidade Federal de Minas Gerais. Belo Horizonte, MG,BrasilUniversidade Federal da Bahia. Hospital Universitário Professor Edgard Santos. Serviço de Imunologia. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisa Gonçalo Moniz. Laboratório de Patologia e Biointervenção. Salvador, BA, BrasilCorrelations between the genetic diversity of Leishmania infantum (syn. L. chagasi) isolates and their respective geographic origins support the theoretic assumption that visceral leishmaniasis probably originated in the Old World. Because dogs are widely considered to be the main reservoir of this disease, the present study aimed to investigate the degree of genetic divergence among 44 leishmanial canine isolates from two Brazilian cities, Jequie´ and Campo Grande, located approximately 2,028 km from each other. We hypothesized that a low degree of genetic divergence would be observed among these isolates. In fact, statistical analyses found no significant differences between the isolates using both random amplified polymorphic DNA and multilocus microsatellite typing genotyping techniques with three and seven markers, respectively. These findings provide support for the recent introduction of L. infantum into the New World

Splenic white pulp compartments defined for morphometric estimative.

<p><b>A</b> – Dog spleen section showing the red pulp and the white pulp compartments; <b>B</b> – white pulp compartments estimated by morphometry. RP = red pulp, MZ = marginal zone, FL = follicle, P = PALS.</p

Primer sequences and annealing temperatures used in real time PCR to detect cytokine and cytokine receptor expression in the spleens of dogs with visceral leishmaniasis.

<p>Primer sequences and annealing temperatures used in real time PCR to detect cytokine and cytokine receptor expression in the spleens of dogs with visceral leishmaniasis.</p

Gene expression of APRIL, BAFF, IL6, CXCL12 and of the CXCR4 chemokine receptor in the spleens of normal dogs, those with inactive (TYPE1SC-) or active (TYPE1SC+) <i>Leishmania</i> infection, or those with disorganized white pulp and active <i>Leishmania</i> infection (TYPE3SC+).

<p>(Kruskall-Wallis test).</p

B cell, T cell, dendritic and proliferating cells number in organized and disorganized spleens.

<p>Estimation of the average number of CD79α⁺ B and CD3⁺ T lymphocytes, S100⁺ dendritic cells and Ki-67⁺ proliferating cells in cross sections of the splenic compartments of dogs infected with <i>L. infantum</i> with and without disruption of splenic lymphoid tissue structure. Box and whisker plots represent median, 25% and 75% percentiles and ranges. Y-axis scale is in Log₁₀. The number of animals used in each estimative is shown on the top of the up range bar. P values (Mann-Whitney test) only shown for statistically significant differences.</p

Cytokine expression in organized and disorganized spleens.

<p>The gene expression of cytokines and CCR7 in the splenic compartments of stray dogs uninfected with organized splenic white pulp (ORG-N), infected with <i>L. infantum</i> with (DES-P) and without (ORG-P) disruption of splenic lymphoid tissue structure. Horizontal lines represent the median of the estimates from RT-PCR, and the values are relative to 18S gene expression. Each dot corresponds to a different animal with 12 organized (12) or disorganized (9) splenic lymphoid tissue. P values (Kruskal-Wallis test) only shown for statistically significant differences.</p

Estimation of the areas represented by the different compartments of the spleen in animals with organized or disorganized splenic histological architecture.

a<p>Organized/disorganized ratios;</p>b<p>Unpaired t test or Mann-Whitney test when recommended.</p>c<p>Proportion (%) of the splenic tissue represented by the specific compartment either red or white pulp in the histological sections;</p>d<p>Mean and sd of the compartment (PALS, follicle or marginal zone) area expressed in mµ².</p