RESUMEN

Parasitic diseases have a significant impact on human and animal health, representing a major hazard to the public and causing economic and health damage worldwide. Extracellular vesicles (EVs) have long been recognized as diagnostic and therapeutic tools but are now also known to be implicated in the natural history of parasitic diseases and host immune response modulation. Studies have shown that EVs play a role in parasitic disease development by interacting with parasites and communicating with other types of cells. This review highlights the most recent research on EVs and their role in several aspects of parasite-host interactions in five key parasitic diseases: Chagas disease, malaria, toxoplasmosis, leishmaniasis and helminthiases. We also discuss the potential use of EVs as diagnostic tools or treatment options for these infectious diseases.

Asunto(s)

Vesículas Extracelulares , Interacciones Huésped-Parásitos , Enfermedades Parasitarias , Humanos , Vesículas Extracelulares/metabolismo , Animales , Enfermedades Parasitarias/terapia , Enfermedades Parasitarias/diagnóstico , Enfermedades Parasitarias/inmunología , Enfermedad de Chagas/terapia , Enfermedad de Chagas/diagnóstico , Enfermedad de Chagas/inmunología

RESUMEN

BACKGROUND: Leishmania (Viannia) braziliensis Thor strain exhibits a heterogeneous composition comprised of subpopulations with varying levels of infectivity. Clonal subpopulations were previously obtained from the strain Thor by sorting single-parasites and proceeding cultivation. The subpopulations used in this study are named Thor03, Thor 10 and Thor22. OBJECTIVES: Phenotypic characteristics of the parasite, specially focusing on virulence factors and resistance to the antimicrobial mechanisms of macrophages, were investigate in these subpopulations. METHODS: Cellular and molecular biology, as well as biochemistry approaches were applied to obtain the data analysed in this study. FINDINGS: Relative quantification of gene expression was measured for calpain, cysteine protease B (CPB), and subtilisin proteases but no significant differences in these genes' expression among subpopulations was observed. However, subtilisin and CPB proteins were assessed as more abundant in Thor03 by fluorescence-labelled flow cytometry technique. Western Blotting assays, as semi-quantitative analysis in gel, showed higher concentrations of subtilisin (110 to 50 kDa) and CPB (40 to 18 kDa) in extract of intracellular amastigotes from subpopulations Thor03 and Thor10 and calpain (60 to 25 kDa) showed no significant differences among subpopulations. Complementary, higher trypanothione reductase activity was observed in Thor10 intracellular amastigotes and assays of susceptibility to hydrogen peroxide-inducing agents and nitric oxide donors conducted with promastigotes revealed greater resistance to in vitro oxidative stress induction for Thor10, followed by Thor03. MAIN CONCLUSIONS: The data obtained for the virulence factors explored here suggest how multiple coexisting phenotypic-distinct subpopulations may contribute in adaptability of a single L. (V.) braziliensis strain during infection in the host cells.

Asunto(s)

Leishmania braziliensis , Leishmania braziliensis/enzimología , Leishmania braziliensis/genética , Leishmania braziliensis/efectos de los fármacos , Animales , Macrófagos/parasitología , Western Blotting , Citometría de Flujo , Factores de Virulencia , Péptido Hidrolasas/metabolismo , Fenotipo , NADH NADPH Oxidorreductasas

RESUMEN

There are no approved vaccines yet for human visceral leishmaniasis (VL), the most severe form of the leishmaniasis clinical manifestations that is fatal in over 95 % of untreated cases. It is well-accepted that immunological changes during aging have deleterious impact on the efficacy of vaccines and response to infections. In this work, we compared the response of young and aged mice to intranasal vaccination with killed Leishmania amazonensis promastigote antigens (LaAg) that were then challenged with L. infantum infection, a species that causes visceral leishmaniasis. Intranasal vaccination with LaAg induced a similar reduction in parasitism and hepatosplenomegaly in both young and aged mice compared to their unvaccinated counterparts. Following infection, there was also a less prominent inflammatory profile particularly in the vaccinated aged group, with lower production of TNF-α and nitrite compared to the respective unvaccinated group. Interestingly, the LaAg intranasal vaccination promoted increased production of IFN-γ that was observed in both young- and aged vaccinated groups. Additionally, CD4+ and CD8+T cells from both vaccinated groups presented decreased expression of the inhibitory receptors PD-1 and KLRG1 compared to their unvaccinated controls. Interestingly, a strong positive correlation was observed between the expression of both inhibitory receptors PD-1 and KLRG1 and parasitism, which was more conspicuous in the unvaccinated-aged mice than in the others. Overall, this study helps define new strategies to improve vaccine effectiveness and provides a perspective for prophylactic alternatives against leishmaniasis.

Asunto(s)

Leishmania infantum , Leishmania mexicana , Vacunas contra la Leishmaniasis , Leishmaniasis Visceral , Leishmaniasis , Vacunas Antiprotozoos , Humanos , Animales , Ratones , Anciano , Leishmaniasis Visceral/prevención & control , Receptor de Muerte Celular Programada 1 , Antígenos de Protozoos , Ratones Endogámicos BALB C , Citocinas

RESUMEN

BACKGROUND Leishmania (Viannia) braziliensis Thor strain exhibits a heterogeneous composition comprised of subpopulations with varying levels of infectivity. Clonal subpopulations were previously obtained from the strain Thor by sorting single-parasites and proceeding cultivation. The subpopulations used in this study are named Thor03, Thor 10 and Thor22. OBJECTIVES Phenotypic characteristics of the parasite, specially focusing on virulence factors and resistance to the antimicrobial mechanisms of macrophages, were investigate in these subpopulations. METHODS Cellular and molecular biology, as well as biochemistry approaches were applied to obtain the data analysed in this study. FINDINGS Relative quantification of gene expression was measured for calpain, cysteine protease B (CPB), and subtilisin proteases but no significant differences in these genes' expression among subpopulations was observed. However, subtilisin and CPB proteins were assessed as more abundant in Thor03 by fluorescence-labelled flow cytometry technique. Western Blotting assays, as semi-quantitative analysis in gel, showed higher concentrations of subtilisin (110 to 50 kDa) and CPB (40 to 18 kDa) in extract of intracellular amastigotes from subpopulations Thor03 and Thor10 and calpain (60 to 25 kDa) showed no significant differences among subpopulations. Complementary, higher trypanothione reductase activity was observed in Thor10 intracellular amastigotes and assays of susceptibility to hydrogen peroxide-inducing agents and nitric oxide donors conducted with promastigotes revealed greater resistance to in vitro oxidative stress induction for Thor10, followed by Thor03. MAIN CONCLUSIONS The data obtained for the virulence factors explored here suggest how multiple coexisting phenotypic-distinct subpopulations may contribute in adaptability of a single L. (V.) braziliensis strain during infection in the host cells.

RESUMEN

The severity of lesions that develop in patients infected by Leishmania braziliensis is mainly associated with a highly cytotoxic and inflammatory cutaneous environment. Recently, we demonstrated that senescent T and NK cells play a role in the establishment and maintenance of this tissue inflammation. Here, we extended those findings using transcriptomic analyses that demonstrate a strong co-induction of senescence and pro-inflammatory gene signatures in cutaneous leishmaniasis (CL) lesions. The senescence-associated signature was characterized by marked expression of key genes such as ATM, Sestrin 2, p16, p21 and p38. The cell type identification from deconvolution of bulk sequencing data showed that the senescence signature was linked with CD8+ effector memory and TEMRA subsets and also senescent NK cells. A key observation was that the senescence markers in the skin lesions are age-independent of patients and were correlated with lesion size. Moreover, a striking expression of the senescence-associated secretory phenotype (SASP), pro-inflammatory cytokine and chemokines genes was found within lesions that were most strongly associated with the senescent CD8 TEMRA subset. Collectively, our results confirm that there is a senescence transcriptomic signature in CL lesions and supports the hypothesis that lesional senescent cells have a major role in mediating immunopathology of the disease.

Asunto(s)

Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Inmunosenescencia/genética , Leishmania braziliensis/inmunología , Leishmaniasis Cutánea/etiología , Leishmaniasis Cutánea/patología , Transcriptoma , Biomarcadores , Biopsia , Biología Computacional/métodos , Citocinas/genética , Citocinas/metabolismo , Bases de Datos Genéticas , Susceptibilidad a Enfermedades/inmunología , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Mediadores de Inflamación/metabolismo , Leishmaniasis Cutánea/metabolismo , Carga de Parásitos , Piel/patología

RESUMEN

Cytotoxic activity mediated by CD8+ T cells is the main signature of the immunopathogenesis of cutaneous leishmaniasis (CL). Here, we performed a broad evaluation of natural killer (NK) cell phenotypic and functional features during cutaneous leishmaniasis. We demonstrate for the first time that CL patients present the accumulation of circulating NK cells with multiple features of replicative senescence including low proliferative capacity and shorter telomeres, elevated expression of CD57, KLRG1 but diminished CD27 stimulatory receptor expression. Moreover, they exhibited higher cytotoxic and inflammatory potential than age-matched controls. The accumulation of circulating senescent NK cells (CD56dim  CD57bright ) correlated positively with skin lesion size in the same patients, suggesting that they, like circulating senescent CD8+ T cells, may contribute to the immunopathology of CL. However, this senescent population had lower cutaneous lymphocyte antigen expression and so had diminished skin-homing potential compared with total or senescent CD8+ T cells. This was confirmed in CL skin lesions where we found a predominance of CD8+ T cells (both senescent and non-senescent) that correlated with the severity of the disease. Although there was also a correlation between the proportions of senescent NK cells (CD56+  CD57+ ) in the skin and lesion size, this was less evident. Collectively our results demonstrate first-hand that senescent cytotoxic cells may mediate skin pathology during human cutaneous leishmaniasis. However, as senescent cytotoxic CD8+ T cells predominate in the skin lesions, they may have a greater role than NK cells in mediating the non-specific skin damage in CL.

Asunto(s)

Citotoxicidad Inmunológica , Células Asesinas Naturales/patología , Leishmania braziliensis/patogenicidad , Leishmaniasis Cutánea/patología , Piel/patología , Linfocitos T Citotóxicos/patología , Antígeno CD56/genética , Antígeno CD56/inmunología , Antígenos CD57/genética , Antígenos CD57/inmunología , Estudios de Casos y Controles , Senescencia Celular/inmunología , Femenino , Regulación de la Expresión Génica , Interacciones Huésped-Parásitos/genética , Interacciones Huésped-Parásitos/inmunología , Humanos , Interferón gamma/genética , Interferón gamma/inmunología , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/parasitología , Lectinas Tipo C/genética , Lectinas Tipo C/inmunología , Leishmania braziliensis/inmunología , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Masculino , Oligosacáridos/genética , Oligosacáridos/inmunología , Receptores Inmunológicos/genética , Receptores Inmunológicos/inmunología , Índice de Severidad de la Enfermedad , Antígeno Sialil Lewis X/análogos & derivados , Antígeno Sialil Lewis X/genética , Antígeno Sialil Lewis X/inmunología , Transducción de Señal , Piel/inmunología , Piel/parasitología , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/parasitología

RESUMEN

The CAF01 adjuvant has previously been shown to be safe for human use and to be a potent adjuvant for several vaccine antigens. In the present work, we sought to optimize the Leishmania amazonensis antigens (LaAg) intranasal vaccine in an attempt to enhance the protective immune responses against Leishmania (infantum) chagasi by using the CAF01 association. LaAg/CAF01 vaccinated mice that were challenged 15 days after booster dose with L. (infantum) chagasi showed a significant reduction in their parasite burden in both the spleen and liver, which is associated with an increase in specific production of IFN-γ and nitrite, and a decrease in IL-4 production. In addition, LaAg/CAF01 intranasal delivery was able to increase lymphoproliferative immune responses after parasite antigen recall. These results suggest the feasibility of using the intranasal route for the delivery of crude antigens and of a human-compatible adjuvant against visceral leishmaniasis.

Asunto(s)

Antígenos de Protozoos/inmunología , Leishmania infantum/inmunología , Leishmania mexicana/inmunología , Leishmaniasis Visceral/prevención & control , Vacunas Antiprotozoos/administración & dosificación , Adyuvantes Inmunológicos/administración & dosificación , Administración Intranasal , Animales , Anticuerpos Antiprotozoarios/sangre , Citocinas/biosíntesis , Epítopos , Femenino , Inmunidad Celular , Inmunoglobulina G/sangre , Hígado/parasitología , Ratones , Ratones Endogámicos BALB C , Nitritos/metabolismo , Bazo/citología , Bazo/inmunología , Bazo/parasitología , Vacunas de Productos Inactivados/administración & dosificación

RESUMEN

The major diagnostic antigens of Histoplasma capsulatum var. capsulatum are the H and M antigens, pluripotent glycoproteins that elicit both humoral and T-cell-mediated immune responses. The gene encoding the M antigen has previously been sequenced, and its sequence has significant overall homology to those of the genes for fungal catalases. Regions of the M-antigen gene with little or no homology were used to design four oligonucleotide sequences for application in the PCR detection and identification of H. capsulatum var. capsulatum. The PCR correctly identified the 31 H. capsulatum var. capsulatum strains isolated from human, animal, and soil specimens and 1 H. capsulatum var. duboisii isolate. PCR products of 111 and 279 bp were amplified with primers Msp1F-Msp1R and Msp2F-Msp2R, respectively. No amplification product was obtained from DNA extracted from an H. capsulatum var. farciminosum isolate. The specificity of the PCR with the M-antigen-derived primers was confirmed by the total absence of amplification products when genomic DNA from Paracoccidioides brasiliensis, Candida spp., Sporothrix schenckii, Cryptococcus neoformans, Blastomyces dermatitidis, Coccidioides immitis, Aspergillus niger, and Aspergillus fumigatus were applied in the reaction. This rapid, sensitive, and specific assay provides a way to identify typical and atypical isolates of H. capsulatum var. capsulatum.

Asunto(s)

Antígenos Fúngicos/genética , Glicoproteínas/genética , Histoplasma/aislamiento & purificación , Reacción en Cadena de la Polimerasa/métodos , ADN de Hongos/análisis , Histoplasma/genética , Humanos