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Introduction: Pathogenesis of cutaneous leishmaniases involves parasite growth, persistent inflammation, and likely participation of lipoproteins (LP). The cholesteryl ester transfer protein (CETP), involved in LP remodeling, has been shown to participate in the inflammatory response and the evolution of infectious conditions. Methods: We evaluated the impact of the presence of CETP on infection by Leishmania (L.) amazonensis in an experimental model of cutaneous leishmaniasis using C57BL6/J mice transgenic for human CETP (CETP), having as control their littermates that do not express the protein, wild-type (WT) mice. The progression of the lesion after infection in the footpad was monitored for 12 weeks. Two groups of animals were formed to collect the plantar pad in the 4th and 12th week post-infection. Results: The lesion increased from the 3rd week onwards, in both groups, with a gradual decrease from the 10th week onwards in the CETP group compared to the WT group, showing a reduction in parasitism and an improvement in the healing process, a reduction in CD68+ cells, and an increase in CD163+ and CD206, characterizing a population of M2 macrophages. A reduction in ARG1+ cells and an increase in INOS+ cells were observed. During infection, the LP profile showed an increase in triglycerides in the VLDL fraction in the CETP group at 12 weeks. Gene expression revealed a decrease in the CD36 receptor in the CETP group at 12 weeks, correlating with healing and parasite reduction. In vitro, macrophages derived from bone marrow cells from CETP mice showed lower parasite load at 48 h and, a reduction in arginase activity at 4 h accompanied by increased NO production at 4 and 24 h compared to WT macrophages, corroborating the in vivo findings. Discussion: The data indicate that the presence of CETP plays an important role in resolving Leishmania (L.) amazonensis infection, reducing parasitism, and modulating the inflammatory response in controlling infection and tissue repair.
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Proteínas de Transferencia de Ésteres de Colesterol , Leishmaniasis Cutánea , Macrófagos , Ratones Endogámicos C57BL , Ratones Transgénicos , Animales , Proteínas de Transferencia de Ésteres de Colesterol/genética , Proteínas de Transferencia de Ésteres de Colesterol/metabolismo , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Leishmaniasis Cutánea/metabolismo , Ratones , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/parasitología , Humanos , Progresión de la Enfermedad , Modelos Animales de EnfermedadRESUMEN
We studied some fibrotic aspects of chronic interstitial pneumonitis in the lungs of dogs infected with Leishmania infantum. The lungs of eleven naturally infected dogs, twelve experimentally infected with two distinct strains of L. infantum (BH401 and BH46), and six uninfected (controls) dogs, were analyzed by histological, parasitological, and immunohistochemical studies. Conventional histology (HE), collagen deposition (Gomori's silver staining for reticulin collagen fibers), and immunohistochemistry for myofibroblast characterization were carried out based on the cellular expression of alpha-smooth muscle actin, vimentin, cytokeratin, E-cadherin, snail antigen homologue 1 (SNAI1) (Snail), and the cytokine expression of transforming growth factor-beta (TGF-ß). Parasitological screening was carried out using conventional polymerase chain reaction (PCR) and the immunohistochemical reaction of streptavidin-peroxidase for visualizing Leishmania amastigotes. Dogs naturally infected with L. infantum and experimentally infected with L. infantum BH401 strains showed intense interstitial pneumonitis characterized by thickening of the alveolar septa as a consequence of an intense diffuse and focal (plaques) chronic exudate of mononuclear cells associated with fibrogenesis. The expression of alpha-actin, vimentin, and TGF-ß was higher in the lung interstitium of all infected dogs than in the other two groups (BH46 strain and controls). Moreover, in both the naturally and experimentally infected dog (BH401 strain) groups, the expression of Snail was moderate to intense in contrast to the other groups. Based on these immunohistochemical results, we concluded that mesenchymal cells are active in promoting changes in the extracellular matrix in the lungs of dogs naturally and experimentally infected with L. infantum, but it depends on the virulence of the parasite.
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BACKGROUND: Eosinophils are granulocytes that rapidly increase frequency in the bloodstream during helminthic infections and allergic responses. They are found in tissue infected by Leishmania during early disease, but their role during infection is not entirely understood. OBJECTIVES: We aim to compare the disease due to Leishmania amazonensis in BALB/c and Δdbl-GATA1 mice, which lack eosinophils. METHODS: BALB/c and Δdbl-GATA1 mice infected with L. amazonensis were observed for several weeks. The parasite load and dissemination pattern were assessed. FINDINGS: The Δdbl-GATA1 mice developed an anticipated dissemination of L. amazonensis and a worsening disease. No differences were found in the lesion development or the parasite load in the footpad among Δdbl-GATA1 mice and BALB/c eight weeks after infection. However, nine weeks after infection, massive growth of metastatic lesions appeared in several parts of the skin in Δdbl-GATA1 mice, weeks earlier than BALB/c. We observed increased parasites in the bloodstream, probably an essential dissemination route. Thirteen weeks after infection, metastatic lesions were found in all Δdbl-GATA1 mice. MAIN CONCLUSION: These results suggest a protective role of eosinophils in delaying the disease caused by L. amazonensis, although several limitations of this mice strain must be considered.
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Leishmania mexicana , Leishmania , Animales , Ratones , Eosinófilos , Carga de Parásitos , PielRESUMEN
BACKGROUND Eosinophils are granulocytes that rapidly increase frequency in the bloodstream during helminthic infections and allergic responses. They are found in tissue infected by Leishmania during early disease, but their role during infection is not entirely understood. OBJECTIVES We aim to compare the disease due to Leishmania amazonensis in BALB/c and Δdbl-GATA1 mice, which lack eosinophils. METHODS BALB/c and Δdbl-GATA1 mice infected with L. amazonensis were observed for several weeks. The parasite load and dissemination pattern were assessed. FINDINGS The Δdbl-GATA1 mice developed an anticipated dissemination of L. amazonensis and a worsening disease. No differences were found in the lesion development or the parasite load in the footpad among Δdbl-GATA1 mice and BALB/c eight weeks after infection. However, nine weeks after infection, massive growth of metastatic lesions appeared in several parts of the skin in Δdbl-GATA1 mice, weeks earlier than BALB/c. We observed increased parasites in the bloodstream, probably an essential dissemination route. Thirteen weeks after infection, metastatic lesions were found in all Δdbl-GATA1 mice. MAIN CONCLUSION These results suggest a protective role of eosinophils in delaying the disease caused by L. amazonensis, although several limitations of this mice strain must be considered.
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Schistosoma mansoni eggs retained in host tissues induce innate cytokine release, contributing to the induction of Type-2 immune responses and granuloma formation, important to restrain cytotoxic antigens, but leading to fibrosis. Interleukin(IL)-33 participates in experimental models of inflammation and chemically induced fibrosis, but its role in S. mansoni-induced fibrosis is still unknown. To explore the role of the IL-33/suppressor of the tumorigenicity 2 (ST2) pathway, serum and liver cytokine levels, liver histopathology, and collagen deposition were comparatively evaluated in S. mansoni-infected wild-type (WT) and IL-33-receptor knockout (ST2-/-) BALB/c mice. Our data show similar egg counts and hydroxyproline in the livers of infected WT and ST2-/- mice; however, the extracellular matrix in ST2-/- granulomas was loose and disorganised. Pro-fibrotic cytokines, such as IL-13 and IL-17, and the tissue-repairing IL-22 were significantly lower in ST2-/- mice, especially in chronic schistosomiasis. ST2-/- mice also showed decreased α-smooth muscle actin (α-SMA) expression in granuloma cells, in addition to reduced Col III and Col VI mRNA levels and reticular fibres. Therefore, IL-33/ST2 signalling is essential for tissue repairing and myofibroblast activation during S. mansoni infection. Its disruption results in inappropriate granuloma organisation, partly due to the reduced type III and VI collagen and reticular fibre formation.
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Schistosoma mansoni , Esquistosomiasis mansoni , Ratones , Animales , Proteína 1 Similar al Receptor de Interleucina-1/genética , Interleucina-33/genética , Cirrosis Hepática/patología , Hígado/metabolismo , Fibrosis , Citocinas , Ratones Endogámicos BALB C , Colágeno/metabolismo , Granuloma/patologíaRESUMEN
Fibrosis is a common pathophysiological response of many tissues and organs subjected to chronic injury. Despite the diverse aetiology of keloid, lacaziosis and localized scleroderma, the process of fibrosis is present in the pathogenesis of all of these three entities beyond other individual clinical and histological distinct characteristics. Fibrosis was studied in 20 samples each of these three chronic cutaneous inflammatory diseases. An immunohistochemical study was carried out to explore the presence of α-smooth muscle actin (α-SMA) and vimentin cytoskeleton antigens, CD31, CD34, Ki67, p16; CD105, CD163, CD206 and FOXP3 antigens; and the central fibrotic cytokine TGF-ß. Higher expression of vimentin in comparison to α-SMA in all three lesion types was found. CD31- and CD34-positive blood vessel endothelial cells were observed throughout the reticular dermis. Ki67 expression was low and almost absent in scleroderma. p16-positive levels were higher than ki67 and observed in reticular dermis of keloidal collagen in keloids, in collagen bundles in scleroderma and in the external layers of the granulomas in lacaziosis. The presence of α-actin positive cells and rarely CD34 positive cells, observed primarily in keloids, may be related to higher p16 antigen expression, a measure of cell senescence. Low FOXP3 expression was observed in all lesion types. CD105-positive cells were mainly found in perivascular tissue in close contact with the adventitia in keloids and scleroderma, while, in lacaziosis, these cells were chiefly observed in conjunction with collagen deposition in the external granuloma layer. We did not find high involvement of CD163 or CD206-positive cells in the fibrotic process. TGF-ß was notable only in keloid and lacaziosis lesions. In conclusion, we have suggested vimentin to be the main myofibroblast general marker of the fibrotic process in all three studied diseases, while endothelial-to-mesenchymal transition (EndoMT) and mesenchymal stem cells (MSCs) and M2 macrophages may not play an important role.
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Queloide , Lobomicosis , Esclerodermia Localizada , Piel , Humanos , Células Endoteliales/metabolismo , Células Endoteliales/patología , Fibroblastos/metabolismo , Fibrosis , Factores de Transcripción Forkhead/metabolismo , Queloide/metabolismo , Queloide/patología , Antígeno Ki-67/metabolismo , Lobomicosis/patología , Esclerodermia Localizada/metabolismo , Esclerodermia Localizada/patología , Piel/metabolismo , Piel/patología , Factor de Crecimiento Transformador beta/metabolismo , Vimentina/metabolismoRESUMEN
BACKGROUND: Leishmania (Mundinia) enriettii is a species commonly found in the guinea pig, Cavia porcellus. Although it is a dermotropic species, there is still an uncertainty regarding its ability to visceralise during Leishmania life cycle. OBJECTIVE: Here, we investigated the ability of L. enriettii (strain L88) to visceralise in lungs, trachea, spleen, and liver of C. porcellus, its natural vertebrate host. METHODS: Animals were infected sub-cutaneously in the nose and followed for 12 weeks using histological (hematoxilin-eosin) and molecular tools (polymerase chain reaction-restriction fragment length polymorphism - PCR-RFLP). To isolate parasite from C. porcellus, animals were experimentally infected for viscera removal and PCR typing targeting hsp70 gene. FINDINGS: Histological analysis revealed intense and diffuse inflammation with the presence of amastigotes in the trachea, lung, and spleen up to 12 weeks post-infection (PI). Molecular analysis of paraffin-embedded tissues detected parasite DNA in the trachea and spleen between the 4th and 8th weeks PI. At the 12th PI, no parasite DNA was detected in any of the organs. To confirm that the spleen could serve as a temporary site for L. enriettii, we performed additional in vivo experiments. During 6th week PI, the parasite was isolated from the spleen confirming previous histopathological and PCR observations. MAIN CONCLUSION: Leishmania enriettii (strain L88) was able to visceralise in the trachea, lung, and spleen of C. porcellus.
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Leishmania enriettii , Leishmania , Animales , Cobayas , Leishmania/genética , Pulmón , Bazo , TráqueaRESUMEN
Extracellular matrix (ECM) alterations in visceral leishmaniasis are related mainly to collagen deposition (fibropoiesis). In canine visceral leishmaniasis (CVL), an intense fibrosis associated to chronic inflammation in organs such as kidneys is described. However, renal fibropoiesis has not been described in natural or experimental infections with L. (L.) infantum. We aimed to characterize renal nephropathies by histology and confocal microscopy comparing renal lesions in dogs naturally and experimentally infected with L. (L.) infantum. Sixty-two mixed-breed symptomatic dogs naturally infected with L. (L.) infantum, sixteen beagles experimentally infected with two strains of L. infantum (eleven dogs with the BH400 strain and five dogs with the BH401 strain), and five uninfected beagles (controls) were used. Samples were stained with hematoxylin & eosin for routine histology. Congo red was used to visualize amyloid protein deposits, periodic acid-Schiff to identify glomerular basal membrane anomalies, Masson's trichrome for collagen deposits, and Jones' methenamine silver to reveal membranous glomerulonephropathy. Immunohistochemistry was used to identify Leishmania amastigotes, and confocal microscopy was used for macrophage characterization (L1/calprotectin and CD163 antigen receptors). The most common lesions were chronic glomerular and interstitial nephritis, which was found in all naturally infected dogs and dogs experimentally infected with L. infantum strain BH401 but not with the BH400 strain. Glomeruloesclerosis was the main lesion presented in all BH401 group. Morphometric analysis revealed positive correlation of renal glomeruli tufts with cellular expression of L1/calprotectin and CD163 antigens. Leishmania infantum strain BH401 shows pathogenicity that may be sufficient to induce classic chronic visceral renal leishmaniasis.
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Enfermedades de los Perros , Leishmania infantum , Leishmaniasis Visceral , Perros , Animales , Hematoxilina , Eosina Amarillenta-(YS) , Rojo Congo , Metenamina , Ácido Peryódico/metabolismo , Riñón/patología , Microscopía Confocal/veterinaria , Complejo de Antígeno L1 de Leucocito , Proteínas Amiloidogénicas/metabolismoRESUMEN
BACKGROUND Leishmania (Mundinia) enriettii is a species commonly found in the guinea pig, Cavia porcellus. Although it is a dermotropic species, there is still an uncertainty regarding its ability to visceralise during Leishmania life cycle. OBJECTIVE Here, we investigated the ability of L. enriettii (strain L88) to visceralise in lungs, trachea, spleen, and liver of C. porcellus, its natural vertebrate host. METHODS Animals were infected sub-cutaneously in the nose and followed for 12 weeks using histological (hematoxilin-eosin) and molecular tools (polymerase chain reaction-restriction fragment length polymorphism - PCR-RFLP). To isolate parasite from C. porcellus, animals were experimentally infected for viscera removal and PCR typing targeting hsp70 gene. FINDINGS Histological analysis revealed intense and diffuse inflammation with the presence of amastigotes in the trachea, lung, and spleen up to 12 weeks post-infection (PI). Molecular analysis of paraffin-embedded tissues detected parasite DNA in the trachea and spleen between the 4th and 8th weeks PI. At the 12th PI, no parasite DNA was detected in any of the organs. To confirm that the spleen could serve as a temporary site for L. enriettii, we performed additional in vivo experiments. During 6th week PI, the parasite was isolated from the spleen confirming previous histopathological and PCR observations. MAIN CONCLUSION Leishmania enriettii (strain L88) was able to visceralise in the trachea, lung, and spleen of C. porcellus.
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Lacaziosis is a cutaneous mycosis caused by the fungus Lacazia loboi, described in different countries of Latin America and prevalent in the Amazon region. The ineffective immune response against the agent seems to be related to a Th2 pattern of cytokines. There are few reports exploring elements of the humoral response in these lesions. Our aim was to investigate some elements focusing on B cells, plasma cells and local expression of IgG and IgM antibodies. Forty skin biopsies of lower limbs were selected. The diagnosis of lacaziosis was based on direct mycological examination and histological analysis. The visualization of fungal cells was improved by using Gridley's staining. An immunohistochemical protocol was performed to detect the expression of B cells, plasma cells, IgG and IgM. A double staining was performed to explore the presence of yeasts in the cytoplasm of keratinocytes, using an anti-AE1 AE3 antibody over Gridley's staining. The inflammatory infiltrate consisted of macrophages, multinucleated giant cells, lymphocytes, and fibrosis. Fungal cells were frequent in the stratum corneum and in both, the dermis and, in 50% of the specimens, also in the epidermis. Cells expressing IgG were more abundant when compared to cells expressing IgM. B cells and the presence of IgG might indicate that the humoral response promotes a Th2 immune response resulting in an anti-inflammatory phenotype. Our results lead us to suggest a possible role of B cells and immunoglobulins in the mechanisms of lacaziosis pathogenesis.
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Dermatomicosis , Lacazia/aislamiento & purificación , Lobomicosis/diagnóstico , Biopsia , Humanos , Inmunoquímica , PielRESUMEN
OBJECTIVES: To analyse the accuracy of American tegumentary leishmaniasis (ATL) diagnostic methods and evaluate the quality of the existing publications by means of a systematic review. METHODS: Diagnostic tests evaluated in at least two studies with common reference standards were included in the sensitivity and/or specificity meta-analyses. Quality and susceptibility to bias were analysed using the QUADAS-2 and STARD tools. RESULTS: The title and abstract of 3387 publications were evaluated after deduplication resulting from database searches. Thirty-eight studies were included in the review, and 26 of them had results inserted in meta-analyses. The diagnostic methods with the highest pooled sensitivity values were ELISA, polymerase chain reaction (PCR), indirect immunofluorescence reaction and Montenegro's intradermal reaction. Cytometry was assessed in only two studies and presented 100% sensitivity in both. Smear slide microscopy and histopathology showed low pooled values of sensitivity. For specificity, the highest pooled values were identified for PCR. High values were also identified for ELISA, except for studies in which the reference standard for defining negative participants included individuals with Chagas' disease or paracoccidioidomycosis, which also occurred for cytometry. IFR had lower specificities than ELISA. There was a predominance of case-control designs of phase 1 or 2 and only four studies were strongly recommended as evidence generators. Several reference standards were adopted, and different methods were assessed in a small number of studies. CONCLUSION: PCR showed the highest accuracy for the diagnosis of ATL, and its use should be encouraged in clinical practice. ELISA is recommended for the screening of suspected individuals, but the possibility of cross-reactions should be considered. New validation studies for the tests evaluated in few publications and studies of phase 3 with appropriate methods are needed.
OBJECTIFS: Analyser l'exactitude des méthodes de diagnostic de la leishmaniose tégumentaire américaine (LTA) et évaluer la qualité des publications existantes au moyen d'une analyse systématique. MÉTHODES: Les tests diagnostiques évalués dans au moins deux études avec des étalons de référence communs ont été inclus dans les méta-analyses de sensibilité et/ou de spécificité. La qualité et la sensibilité au biais ont été analysées à l'aide des outils QUADAS-2 et STARD. RÉSULTATS: Le titre et le résumé de 3387 publications ont été évalués après déduplication résultant de recherches dans la base de données. 38 études ont été incluses dans la revue et 26 d'entre elles ont eu des résultats inclus dans des méta-analyses. Les méthodes de diagnostic avec les valeurs de sensibilité poolées les plus élevées étaient ELISA, la réaction en chaîne par polymérase (PCR), la réaction d'immunofluorescence indirecte et la réaction intradermique du Monténégro. La cytométrie a été évaluée dans seulement deux études et présentait une sensibilité de 100% dans les deux. La microscopie et l'histopathologie sur lame de frottis ont montré de faibles valeurs poolées de sensibilité. Pour la spécificité, les valeurs poolées les plus élevées ont été identifiées pour la PCR. Des valeurs élevées ont également été identifiées pour l'ELISA, à l'exception des études dans lesquelles la norme de référence pour définir les participants négatifs incluait des individus atteints de la maladie de Chagas ou de paracoccidioïdomycose, qui s'est également produite pour la cytométrie. L'IFR avait des spécificités plus faibles que l'ELISA. Il y avait une prédominance des essais cas-témoins de phases 1 ou 2 et seules quatre études étaient fortement recommandées comme génératrices de preuves. Plusieurs normes de référence ont été adoptées et différentes méthodes ont été évaluées dans un petit nombre d'études. CONCLUSION: la PCR a montré la plus grande exactitude pour le diagnostic de l'ATL et son utilisation doit être encouragée dans la pratique clinique. L'ELISA est recommandé pour le dépistage des personnes suspectées, mais la possibilité de réactions croisées doit être considérée. De nouvelles études de validation des tests évalués dans quelques publications et des études de phase 3 avec des méthodes appropriées sont nécessaires.
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Leishmania/aislamiento & purificación , Leishmaniasis Cutánea/diagnóstico , Animales , Pruebas Diagnósticas de Rutina , Ensayo de Inmunoadsorción Enzimática , Humanos , Leishmania/genética , América del Norte , Reacción en Cadena de la Polimerasa , Sensibilidad y Especificidad , América del SurRESUMEN
To maintain homeostasis, macrophages must be capable of assuming either an inflammatory or an anti-inflammatory phenotype. To better understand the latter, we stimulated human macrophages in vitro with TLR ligands in the presence of high-density immune complexes (IC). This combination of stimuli resulted in a broad suppression of inflammatory mediators and an upregulation of molecules involved in tissue remodeling and angiogenesis. Transcriptomic analysis of TLR stimulation in the presence of IC predicted the downstream activation of AKT and the inhibition of GSK3. Consequently, we pretreated LPS-stimulated human macrophages with small molecule inhibitors of GSK3 to partially phenocopy the regulatory effects of stimulation in the presence of IC. The upregulation of DC-STAMP and matrix metalloproteases was observed on these cells and may represent potential biomarkers for this regulatory activation state. To demonstrate the presence of these anti-inflammatory, growth-promoting macrophages in a human infectious disease, biopsies from patients with leprosy (Hanseniasis) were analyzed. The lepromatous form of this disease is characterized by hypergammaglobulinemia and defective cell-mediated immunity. Lesions in lepromatous leprosy contained macrophages with a regulatory phenotype expressing higher levels of DC-STAMP and lower levels of IL-12, relative to macrophages in tuberculoid leprosy lesions. Therefore, we propose that increased signaling by FcγR cross-linking on TLR-stimulated macrophages can paradoxically promote the resolution of inflammation and initiate processes critical to tissue growth and repair. It can also contribute to infectious disease progression.
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Complejo Antígeno-Anticuerpo/metabolismo , Lepra Lepromatosa/inmunología , Lepra Tuberculoide/inmunología , Macrófagos/inmunología , Biopsia , Diferenciación Celular/inmunología , Línea Celular , Progresión de la Enfermedad , Glucógeno Sintasa Quinasa 3/antagonistas & inhibidores , Glucógeno Sintasa Quinasa 3/metabolismo , Humanos , Lepra Lepromatosa/patología , Lepra Tuberculoide/patología , Activación de Macrófagos , Macrófagos/metabolismo , Masculino , Persona de Mediana Edad , Neovascularización Fisiológica/inmunología , Proteínas Proto-Oncogénicas c-akt/metabolismo , RNA-Seq , Receptores de IgG/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunología , Piel/citología , Piel/inmunología , Piel/patología , Receptores Toll-Like/metabolismo , Adulto JovenRESUMEN
Feline leishmaniasis (FeL) is an emerging infectious disease of cats caused by Leishmania infantum with global distribution. This study investigated the cause of chronic progressive cutaneous lesions in two cats from Central-west Brazil by using cytological, histopathologic, and immunohistochemical (IHC) analyses. Clinically, both cats had ulcerative cutaneous lesions at the nasal planum and ear resulting in a tentative diagnosis of squamous cell carcinoma (SCC). Moreover, both cats had varying degrees of onychogryphosis. However, cytology revealed chronic inflammatory reactions associated with intralesional amastigotes; histopathology confirmed chronic ulcerative dermatitis associated with intralesional and intracytoplasmic parasitic organisms consistent with amastigotes of Leishmania spp. within histiocytes. The IHC assay demonstrated that the intralesional parasitic structures identified by cytology and histopathology were immunoreactive to antigens of Leishmania spp., confirming the participation of this infectious disease agent in the development of the cutaneous lesions of these cats. The observation of onychogryphosis must be highlighted, since this lesion is frequently observed in dogs with visceral leishmaniasis but is underreported in FeL. Collectively, the pathologic and IHC findings of the chronic cutaneous disease confirmed active infections due to Leishmania spp. in these cats. Additionally, FeL with associated lesions to the ear and nasal planum must be considered as differential diagnosis for SCC in cats.
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Enfermedades de los Gatos/parasitología , Leishmania infantum/aislamiento & purificación , Leishmaniasis Cutánea/veterinaria , Animales , Biopsia con Aguja Fina/veterinaria , Brasil/epidemiología , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/veterinaria , Enfermedades de los Gatos/diagnóstico , Enfermedades de los Gatos/epidemiología , Gatos , Ciudades , Diagnóstico Diferencial , Oído Externo/patología , Femenino , Miembro Anterior/patología , Miembro Posterior/patología , Inmunohistoquímica/veterinaria , Leishmaniasis Cutánea/diagnóstico , Leishmaniasis Cutánea/epidemiología , Leishmaniasis Cutánea/parasitología , Nariz/patología , Neoplasias Cutáneas/diagnóstico , Neoplasias Cutáneas/veterinariaRESUMEN
BACKGROUND: Despite the very low or absent parasitism in the lungs, the interstitial pneumonitis is a common lesion found in humans and dogs with visceral leishmaniasis. The lung is a neglected organ in the study of dogs and humans with visceral leishmaniasis, but interstitial pneumonitis represents an important lesion characterized by thickening of the alveolar septum due to fibrosis and inflammatory exudate, and its pathogenesis is still uncertain. In this study, the polymerase chain reaction (PCR) was used to detect Leishmania infantum in paraffin-embedded lung biopsies from naturally infected dogs from an endemic area in Minas Gerais State, Brazil; PCR was compared to histological and immunohistochemical techniques for detecting Leishmania. RESULTS: Eighteen dogs in which leishmaniasis had been diagnosed by serological tests - indirect immunofluorescence assay (IFAT), enzyme-linked immunosorbent assay (ELISA) and complement fixation tests (CFT) - were classified as asymptomatic, oligosymptomatic or symptomatic. Nine of the 18 dogs studied had a positive PCR (50%) but parasites were not detected by histopathological and immunocytochemistry methods. CONCLUSIONS: These data indicate that PCR on DNA extracted from paraffin-embedded tissue is a valuable method for detecting Leishmania infantum parasites in lungs of naturally infected dogs, despite the apparent absence of parasites from standard HE (hematoxylin and eosin) stained slides and of labeled parasites from immunocytochemical preparations.
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ADN Protozoario/análisis , Enfermedades de los Perros/diagnóstico , Leishmaniasis Visceral/veterinaria , Pulmón/parasitología , Animales , Brasil , Enfermedades de los Perros/parasitología , Enfermedades de los Perros/patología , Perros , Inmunohistoquímica/normas , Inmunohistoquímica/veterinaria , Leishmania infantum/genética , Leishmaniasis Visceral/diagnóstico , Leishmaniasis Visceral/parasitología , Leishmaniasis Visceral/patología , Pulmón/química , Reacción en Cadena de la Polimerasa/normas , Reacción en Cadena de la Polimerasa/veterinariaRESUMEN
Visceral Leishmaniasis is a chronic and lethal, parasitic disease. In the later infection stages, it is known that expressive hematological disorders can be observed, including changes in the frequency and phenotype of certain leukocytes. There is a lack of good prognostic indicators to characterize the on-goin clinical status of the patient. In this study, we have analyzed the frequency of monocyte subpopulations in mice infected with Leishmania major (L. major). Our results show a significant correlation between increased blood monocyte frequency and lesion development in both BALB/c and in the C57BL/6 mice infected with L. major. In BALB/c mice we observed a significant correlation between the frequency of GR1+ monocytes and lesion size. Furthermore, treatment of infected BALB/c mice with Anfotericin B, to resolve lesions, resulted in a lower frequency of GR1+ monocytes compared to untreated infected BALB/c mice. C57BL/6 infected mice, which normally resolve infections, show decreased numbers of monocytes during the healing phase of infection. The results indicate that disease severity can be predicted by analyzing monocyte frequency. Thus, we propose that the frequency of monocytes, can be used to define the severity of the disease as well as the success of the treatment in experimental leishmaniasis.
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Anfotericina B/farmacología , Leishmania major/aislamiento & purificación , Leishmaniasis Visceral/parasitología , Monocitos/parasitología , Animales , Antiprotozoarios/farmacología , Biomarcadores , Modelos Animales de Enfermedad , Femenino , Leishmania major/efectos de los fármacos , Leishmaniasis Visceral/tratamiento farmacológico , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Monocitos/metabolismo , Índice de Severidad de la EnfermedadRESUMEN
Naturally Leishmania infantum infected bitches were divided into oestrogenized (n = 11) and non-oestrogenized (n = 6) groups. Vaginal secretions were collected for polymerase chain reaction (PCR), and vulval, vaginal and uterine tissues were collected for the immunohistochemical (IHQ) identification of L. infantum. Parasite DNA was identified in vaginal secretions of non-oestrogenized (41.8%) and oestrogenized (18.2%) bitches (P<0.05; Fisher's Exact test). IHQ was positive in vulvar dermis (23.5%) and vaginal mucosa (17.7%) but negative in endometrium of all bitches. Poor association between positive vaginal secretion PCR and tissue IHQ (Kappa index) were observed. The results showed that genital secretions are a potential source for dog contamination.
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Líquidos Corporales/parasitología , Enfermedades de los Perros/parasitología , Estradiol/análogos & derivados , Leishmania infantum , Leishmaniasis Visceral/veterinaria , Animales , Anticonceptivos Femeninos/administración & dosificación , Anticonceptivos Femeninos/farmacología , ADN Protozoario/aislamiento & purificación , Transmisión de Enfermedad Infecciosa , Enfermedades de los Perros/transmisión , Perros , Estradiol/administración & dosificación , Estradiol/farmacología , Femenino , Enfermedades de los Genitales Femeninos , Leishmaniasis Visceral/parasitología , Leishmaniasis Visceral/transmisiónRESUMEN
Interferon gamma (IFN-γ) is a key factor in the protection of hosts against intracellular parasites. This cytokine induces parasite killing through nitric oxide and reactive oxygen species production by phagocytes. Surprisingly, during Leishmania amazonensis infection, IFN-γ plays controversial roles. During in vitro infections, IFN-γ induces the proliferation of the amastigote forms of L. amazonensis. However, this cytokine is not essential at the beginning of an in vivo infection. It is not clear why IFN-γ does not mediate protection during the early stages of infection. Thus, the aim of our study was to investigate the role of IFN-γ during L. amazonensis infection. We infected IFN-γ(-/-) mice in the footpad and followed the development of leishmaniasis in these mice compared with that in WT mice. CD4(+) T lymphocytes and macrophages migrated earlier to the site of infection in the WT mice, and the earlier migration of these 2 cell types was associated with lesion development and parasite growth, respectively. These differences in the infiltrate populations were explained by the increased expression of chemokines in the lesions of the WT mice. Thus, we propose that IFN-γ plays a dual role during L. amazonensis infection; it is an important inducer of effector mechanisms, particularly through inducible nitric oxide synthase expression, and conversely, it is a mediator of inflammation and pathogenesis through the induction of the expression of chemokines. Our data provided evidence for a pathogenic effect of IFN-γ production during leishmaniasis that was previously unknown.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Interferón gamma/metabolismo , Leishmania mexicana/inmunología , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Animales , Quimiocinas/metabolismo , Quimiotaxis de Leucocito/inmunología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Interacciones Huésped-Parásitos , Mediadores de Inflamación/metabolismo , Interferón gamma/genética , Leishmaniasis Cutánea/genética , Leishmaniasis Cutánea/parasitología , Masculino , Ratones , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismoRESUMEN
Knowledge of Leishmania virulence is essential for understanding how the contact between the pathogen and host cells can lead to pathogenesis. Virulence in two L. infantum strains was characterized using macrophages and hamsters. Next, we used difference gel electrophoresis (DIGE) and mass spectrometry to identify the differentially expressed proteins. A total of 63 spots were identified corresponding to 36 proteins; 20 were up-regulated, in which 16 had been previously associated with Leishmania virulence. Considering our results and what has been reported before, we suggest the hypothesis that L. infatum virulence could be a result of the increased expression of KMP-11 and metallopeptidase, associated with an improved parasite-host interacting efficiency and degradation of the protective host proteins and peptides, respectively. Other factors are tryparedoxin peroxidase and peroxidoxin, which protect the parasite against the stress response, and 14-3-3 protein-like, which can prolong infected host cell lifetime. Proteins as chaperones and endoribonuclease L-PSP can increase parasite survival. Enolase is able to perform versatile functions in the cell, acting as a chaperone or in the transcription process, or as a plasminogen receptor or in cell migration events. As expected in more invasive cells with high replication rates, energy consumption and protein synthesis are higher, with up-regulation of Rieske iron-sulfur protein precursor, EF-2, S-adenosylhomocysteine, and phosphomannomutase.
Asunto(s)
Leishmania infantum/metabolismo , Leishmania infantum/patogenicidad , Proteínas Protozoarias/análisis , Factores de Virulencia/análisis , Animales , Células Cultivadas , Cricetinae , Macrófagos , Ratones , Ratones Endogámicos BALB C , Proteoma/análisis , Proteómica/métodos , Proteínas Protozoarias/química , Proteínas Protozoarias/metabolismo , Factores de Virulencia/química , Factores de Virulencia/metabolismoRESUMEN
BACKGROUND: American tegumentary leishmaniasis (ATL) is endemic in Latin America, where Brazil has over 27 thousand cases per year. The aim of the present study was to develop an immunohistochemical method (IHC) for ATL diagnosis. For this purpose, we used serum from a dog naturally infected with Leishmania (L) infantum (canine hyperimmune serum) as the primary antibody, followed by a detection system with a secondary biotinylated antibody. METHODOLOGY: Skin samples were obtained from 73 patients in an endemic area of Caratinga, Minas Gerais (MG) State, Brazil all testing positive for ATL with the Montenegro skin test, microscopy, and PCR. Canine hyperimmune serum of a dog naturally infected with Leishmania (L.) infantum was employed as a primary antibody in an immunohistochemical diagnostic method using streptavidin-biotin peroxidase. To assess the specificity of this reaction, IHC assays employing two monoclonal antibodies were carried out. As the polymer-based technology is less time-consuming and labor intensive than the IHC labeled streptavidin-biotin peroxidase method, we compared the two methods for all samples. RESULTS: The IHC method detected ATL in 67 of the 73 cases (91.8%). Immunolabeled parasites were primarily detected inside macrophages either in the superficial or the deep dermis. Detection was facilitated by the high contrast staining of amastigotes (dark brown) against the light blue background. A lower detection rate (71.2%) was observed with the both of the monoclonal Leishmania antibodies compared to the canine hyperimmune serum. This may have been due to a non-specific background staining observed in all histological samples rendering positive detection more difficult. The higher efficacy of the canine hyperimmune serum in the IHC method was confirmed by the method using streptavidin-biotin peroxidase as well as that with the polymer-based technology (biotin-avidin-free system). CONCLUSIONS: The data are encouraging with regard to validating IHC as a standard alternative method for ATL diagnosis.
Asunto(s)
Leishmania/aislamiento & purificación , Leishmaniasis Cutánea/patología , Leishmaniasis Cutánea/parasitología , Piel/patología , Piel/parasitología , Adolescente , Adulto , Anciano , Animales , Brasil , Niño , Preescolar , Perros , Femenino , Humanos , Inmunohistoquímica , Lactante , Masculino , Persona de Mediana Edad , Parasitemia/parasitología , Parasitemia/patología , Adulto JovenRESUMEN
This case report describes an atypical clinical presentation of visceral leishmaniasis affecting the digestive tract and causing malabsorption syndrome in a patient without recognized immunosuppressive condition. After appropriate treatment for the classical visceral form of the disease, diarrhea persisted as the main symptom and massive infection by Leishmania was detected by histopathology analysis of the duodenal mucosa. Schistosoma mansoni coinfection was also confirmed and treated without impact on diarrhea. New course of amphotericin B finally led to complete improvement of diarrhea. Atypical visceral leishmaniasis involving the gastrointestinal tract is well recognized in HIV coinfection but very rare in immunocompetent patients. The factors determining the control or evolution of the Leishmania infection have not been completely identified. This case stresses the importance of atypical symptoms and the unusual location of visceral leishmaniasis, not only in immunodepressed patients, and raises the possible influence of chronic infection by S. mansoni reducing the immune response to Leishmania.