RESUMO
Capsaicin, a pungent compound in chili peppers, is described as having potent anti-inflammatory, antioxidant, and antimicrobial properties. It is also described as a potential modulator of the immune system and intestinal microbiota. Oral or rectal administration of capsaicin has been studied to treat or prevent colitis. However, those vias are often not well accepted due to the burning sensation that capsaicin can cause. Our objective was to evaluate whether the application of capsaicin skin creams (0.075%) would be effective in improving inflammation and epithelial barrier function as well as the composition of the gut microbiota in a model of mild colitis induced by dextran sulfate sodium (1.5%). The results showed that the cutaneous application of capsaicin reversed weight loss and decreased colon shortening and diarrhea, all typical signs of colitis. There was also an improvement in the intestinal epithelial barrier, preserving proteins from tight junctions. We also evaluated the biodistribution of 99mtechnetium-radiolabeled capsaicin (99mTc-CAPS) applied to the back skin of the animals. We found significant concentrations of 99 mTc-Cap in the colon and small intestine after 2 and 4 h of administration. In addition, there was an increased expression of capsaicin receptor TRPV1 in the colon. Moreover, animals with colitis receiving cutaneous capsaicin presented a better short-chain fatty acid profile and increased levels of SIgA, suggesting increased microbiota diversity. In conclusion, our work opens avenues for further studies to better understand capsaicin's potential benefits and mechanisms in addressing colitis through cutaneous application.
RESUMO
The discovery of new therapeutic alternatives for cancer treatment is essential for improving efficacy and specificity, overcoming resistance, and enabling a more personalized approach for each patient. We investigated the antitumor activity of the crude ethanolic extract of the fungus Trichoderma asperelloides (ExtTa) and its interaction with chemotherapeutic drugs. It was observed, by MTT cytotoxicity assay, that ExtTa significantly reduced cell viability in breast adenocarcinoma, glioblastoma, lung carcinoma, melanoma, colorectal carcinoma, and sarcomas cell lines. The highest efficacy and selectivity of ExtTa were found against glioblastoma T98G and colorectal HCT116 cell lines. ExtTa is approximately four times more cytotoxic to those tumor cells than to non-cancer cell lines. A synergistic effect between ExtTa and doxorubicin was found in the treatment of osteosarcoma Saos-2 cells, as well as with 5-fluorouracil in the treatment of HCT116 colorectal carcinoma cells using CompuSyn software. Our data unravel the presence of bioactive compounds with cytotoxic effects against cancer cells present in T. asperelloides ethanolic crude extract, with the potential for developing novel anticancer agents.
RESUMO
Vaccination against visceral leishmaniasis (VL) should be considered as a safe and effective measure to disease control; however, few vaccines are available against canine VL and there is no an approved human vaccine. In this context, in the present study, we evaluated the endonuclease III (ENDO) protein, which was recently showed to be antigenic for human disease, as a vaccine candidate against Leishmania infantum infection. The recombinant protein (rENDO) was administered in BALB/c mice alone or associated with saponin (rENDO/Sap) or micelles (rENDO/Mic) as adjuvants. Controls received saline, saponin or empty micelles. Results showed that both rENDO/Sap and rENDO/Mic compositions induced higher levels of IFN-γ, IL-12, TNF-α, and GM-CSF cytokines, besides nitrite and IgG2a isotype antibodies, before and after challenge infection, which were related to both CD4+ and CD8+ T cell subtypes. The immunological results contributed to significant reductions in the parasite load found in the spleens, livers, bone marrows and draining lymph nodes of the vaccinated animals. In general, mice immunized with rENDO/Mic presented a slightly higher Th1-type cellular and humoral immune response, as compared to those receiving rENDO/Sap. In addition, saponin caused a slight to moderate inflammatory edema in their vaccinated footpads, which was not observed when micelles were used with rENDO. In addition, a preliminary analysis showed that the recombinant protein was immunogenic to human cells cultures, since PBMCs from treated VL patients and healthy subjects showed higher lymphoproliferation and IFN-γ production in the culture supernatants. In conclusion, data suggest that rENDO could be considered as a candidate to be evaluated in future studies as vaccine to protect against VL.
Assuntos
Leishmania infantum , Vacinas contra Leishmaniose , Leishmaniose Visceral , Leishmaniose , Saponinas , Humanos , Animais , Cães , Camundongos , Micelas , Proteínas Recombinantes , Leishmaniose/prevenção & controle , Adjuvantes Imunológicos , Camundongos Endogâmicos BALB C , Antígenos de ProtozoáriosRESUMO
Abstract Dengue fever (DF) is increasingly recognized as one of the worlds major mosquito borne diseases and causes significant morbidity and mortality in tropical and subtropical countries. Dengue fever is endemic in most part of Pakistan and continues to be a public health concern. Knowledge, attitude and practices can play an important role in management of the disease. Current study was aimed to determine the level of knowledge, attitude and practices regarding dengue fever among health practitioners, to study the level of knowledge and attitude with preventive practices for dengue fever. A cross sectional study was carried out in medical practitioners of the four districts of Malakand region during October to November 2019. A pre-structured questionnaire was used to collect data from medical practitioners. Data was analyzed using Graph Pad version 5. Significant value was considered when less than 0.05 (at 95% confidence of interval). The results revealed that most of participants have seen dengue vector (62%), the media being the most quoted source of information. Nearly 81.2% participants were aware from transmission of dengue fever is by mosquito bite. Practices based upon preventive measures were found to be predominantly focused towards prevention of mosquito bites rather than elimination of breeding places. Although the knowledge regarding DF and mosquito control measure was quite high among the medical practitioners but this knowledge was not put into practice. Further studies are required to aware the people about dengue and its vector in order to get prevention and control.
Resumo A dengue (DF) é cada vez mais reconhecida como uma das principais doenças transmitidas por mosquitos do mundo e causa significativa morbidade e mortalidade em países tropicais e subtropicais. A dengue é endêmica na maior parte do Paquistão e continua a ser um problema de saúde pública. Conhecimento, atitude e práticas podem desempenhar papel importante no manejo da doença. O presente estudo teve como objetivo determinar o nível de conhecimento, atitude e práticas em relação à dengue entre os profissionais de saúde, para estudar o nível de conhecimento e atitude com as práticas preventivas da dengue. Um estudo transversal foi realizado com médicos dos quatro distritos da região de Malakand de outubro a novembro de 2019. Um questionário pré-estruturado foi usado para coletar dados de médicos. Os dados foram analisados no Graph Pad versão 5. Valor significativo foi considerado quando menor que 0,05 (com intervalo de confiança de 95%). Os resultados revelaram que a maioria dos participantes já viu o vetor da dengue (62%), sendo a mídia a fonte de informação mais citada. Quase 81,2% dos participantes sabiam que a transmissão da dengue é por picada de mosquito. Constatou-se que as práticas baseadas em medidas preventivas se concentravam predominantemente na prevenção de picadas de mosquitos, e não na eliminação de criadouros. Embora o conhecimento sobre DF e medidas de controle de mosquitos fosse bastante elevado entre os médicos, esse conhecimento não foi colocado em prática. Mais estudos são necessários para conscientizar a população sobre a dengue e seu vetor, a fim de se obter prevenção e controle.
RESUMO
Dengue fever (DF) is increasingly recognized as one of the world's major mosquito borne diseases and causes significant morbidity and mortality in tropical and subtropical countries. Dengue fever is endemic in most part of Pakistan and continues to be a public health concern. Knowledge, attitude and practices can play an important role in management of the disease. Current study was aimed to determine the level of knowledge, attitude and practices regarding dengue fever among health practitioners, to study the level of knowledge and attitude with preventive practices for dengue fever. A cross sectional study was carried out in medical practitioners of the four districts of Malakand region during October to November 2019. A pre-structured questionnaire was used to collect data from medical practitioners. Data was analyzed using Graph Pad version 5. Significant value was considered when less than 0.05 (at 95% confidence of interval). The results revealed that most of participants have seen dengue vector (62%), the media being the most quoted source of information. Nearly 81.2% participants were aware from transmission of dengue fever is by mosquito bite. Practices based upon preventive measures were found to be predominantly focused towards prevention of mosquito bites rather than elimination of breeding places. Although the knowledge regarding DF and mosquito control measure was quite high among the medical practitioners but this knowledge was not put into practice. Further studies are required to aware the people about dengue and its vector in order to get prevention and control.
A dengue (DF) é cada vez mais reconhecida como uma das principais doenças transmitidas por mosquitos do mundo e causa significativa morbidade e mortalidade em países tropicais e subtropicais. A dengue é endêmica na maior parte do Paquistão e continua a ser um problema de saúde pública. Conhecimento, atitude e práticas podem desempenhar papel importante no manejo da doença. O presente estudo teve como objetivo determinar o nível de conhecimento, atitude e práticas em relação à dengue entre os profissionais de saúde, para estudar o nível de conhecimento e atitude com as práticas preventivas da dengue. Um estudo transversal foi realizado com médicos dos quatro distritos da região de Malakand de outubro a novembro de 2019. Um questionário pré-estruturado foi usado para coletar dados de médicos. Os dados foram analisados no Graph Pad versão 5. Valor significativo foi considerado quando menor que 0,05 (com intervalo de confiança de 95%). Os resultados revelaram que a maioria dos participantes já viu o vetor da dengue (62%), sendo a mídia a fonte de informação mais citada. Quase 81,2% dos participantes sabiam que a transmissão da dengue é por picada de mosquito. Constatou-se que as práticas baseadas em medidas preventivas se concentravam predominantemente na prevenção de picadas de mosquitos, e não na eliminação de criadouros. Embora o conhecimento sobre DF e medidas de controle de mosquitos fosse bastante elevado entre os médicos, esse conhecimento não foi colocado em prática. Mais estudos são necessários para conscientizar a população sobre a dengue e seu vetor, a fim de se obter prevenção e controle.
Assuntos
Conscientização , Dengue/epidemiologia , Doenças Transmitidas por Vetores , PaquistãoRESUMO
Leishmania virulence proteins should be considered as vaccine candidates against disease, since they are involved in developing infection in mammalian hosts. In a previous study, a Leishmania guanosine-5'-triphosphate (GTP)-binding protein was identified as a potential parasite virulence factor. In the present work, the gene encoding GTP was cloned and the recombinant protein (rGTP) was evaluated as a vaccine candidate against Leishmania infantum infection. The protein was associated with saponin (rGTP/Sap) or Poloxamer 407-based micelles (rGTP/Mic) as adjuvants, and protective efficacy was investigated in BALB/c mice after parasite challenge. Both rGTP/Sap and rGTP/Mic compositions induced a Th1-type immune response in vaccinated animals, with significantly higher levels of IFN-γ, IL-12, IL-2, TNF-α, GM-CSF, nitrite, specific IgG2a isotype antibody and positive lymphoproliferation, when compared to the control groups. This response was accompanied by significantly lower parasite load in the spleens, livers, bone marrows and draining lymph nodes of the animals. Immunological and parasitological evaluations indicated that rGTP/Mic induced a more polarized Th1-type response and higher reduction in the organ parasitism, and with lower hepatotoxicity, when compared to the use of rGTP/Sap. In conclusion, our preliminary data suggest that rGTP could be considered for further development as a vaccine candidate to protect against VL.
Assuntos
Leishmania infantum , Leishmaniose Visceral , Leishmaniose , Animais , Antígenos de Protozoários , Proteínas de Transporte , Guanosina , Guanosina Trifosfato , Mamíferos , Camundongos , Camundongos Endogâmicos BALB C , Micelas , Poloxâmero , Polifosfatos , Proteínas RecombinantesRESUMO
Vaccination against visceral leishmaniasis (VL) should be considered as a control measure to protect against disease, and amastigote-specific proteins could help to develop such vaccines, since this parasite form is in contact with the host immune system during the active disease. In this study, a Leishmania amastigote-specific protein, LiHyG, was evaluated as recombinant protein (rLiHyG) as vaccine candidate against Leishmania infantum infection in BALB/c mice. The protein was associated with saponin (rLiHyG/Sap) or Poloxamer 407-based polymeric micelles (rLiHyG/Mic) as adjuvants, and animals receiving saline, saponin or micelle as controls. Immunological and parasitological analyses were performed before (n = 8 per group; as primary endpoint) and after (n = 8 per group; as secondary endpoint) infection. Results showed that, in both endpoints, rLiHyG/Sap and rLiHyG/Mic induced higher levels of IFN-γ, IL-12 and GM-CSF in spleen cell cultures from vaccinated animals, besides elevated presence of IgG2a isotype antibodies. Decreased hepatotoxicity and 'positive lymphoproliferative response were also found after challenge. Such findings reflected in significantly lower levels of parasite load found in their spleens, livers, bone marrows and draining lymph nodes. In conclusion, rLiHyG associated with Th1-type adjuvant could be considered for future studies as vaccine candidate to protect against VL.
Assuntos
Leishmania infantum , Vacinas contra Leishmaniose , Leishmaniose Visceral , Leishmaniose , Saponinas , Adjuvantes Imunológicos , Animais , Antígenos de Protozoários/genética , Leishmaniose/prevenção & controle , Leishmaniose Visceral/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/genéticaRESUMO
Dengue fever (DF) is increasingly recognized as one of the world's major mosquito borne diseases and causes significant morbidity and mortality in tropical and subtropical countries. Dengue fever is endemic in most part of Pakistan and continues to be a public health concern. Knowledge, attitude and practices can play an important role in management of the disease. Current study was aimed to determine the level of knowledge, attitude and practices regarding dengue fever among health practitioners, to study the level of knowledge and attitude with preventive practices for dengue fever. A cross sectional study was carried out in medical practitioners of the four districts of Malakand region during October to November 2019. A pre-structured questionnaire was used to collect data from medical practitioners. Data was analyzed using Graph Pad version 5. Significant value was considered when less than 0.05 (at 95% confidence of interval). The results revealed that most of participants have seen dengue vector (62%), the media being the most quoted source of information. Nearly 81.2% participants were aware from transmission of dengue fever is by mosquito bite. Practices based upon preventive measures were found to be predominantly focused towards prevention of mosquito bites rather than elimination of breeding places. Although the knowledge regarding DF and mosquito control measure was quite high among the medical practitioners but this knowledge was not put into practice. Further studies are required to aware the people about dengue and its vector in order to get prevention and control.
Assuntos
Dengue , Conhecimentos, Atitudes e Prática em Saúde , Animais , Estudos Transversais , Dengue/epidemiologia , Dengue/prevenção & controle , Humanos , Paquistão/epidemiologia , Inquéritos e QuestionáriosRESUMO
Visceral leishmaniasis (VL) is an important public health problem in the world, and control measures are insufficient to avoid the spread of this neglected disease. Dogs are important domestic reservoirs of Leishmania parasites in countries where VL is a zoonosis, representing a major source of infection between sand fly vectors and humans. In this context, a precise diagnosis of canine leishmaniasis (CanL) could help to reduce the number of human cases. Distinct approaches for the diagnosis of CanL have used recombinant proteins in serological assays. However, variable results of the antigens have been found, mainly to diagnosis asymptomatic cases. The present study used bioinformatics to select specific B-cell epitopes of four Leishmania infantum proteins, which had previously been proven to be antigenic in VL, aiming to produce a novel chimeric protein and to evaluate it for the diagnosis of CanL. Seven B-cell epitopes were identified and used to construct the chimera, which was analyzed in a recombinant format through an ELISA assay against a canine serological panel. A soluble Leishmania antigenic extract (SLA) was used as an antigen control. Results showed 100 % sensitivity and specificity for chimera, while when using SLA the values were 26.0 % and 96.4 %, respectively. The performance of chimera was compared with a commercial kit using asymptomatic and symptomatic dog sera, and the data showed that no false-negative result was found when the recombinant protein was used. However, when using the commercial kit, 40.0 % and 16.0 % of the false-negative results were found, respectively. In conclusion, the recombinant chimera showed an antigenic potential to be evaluated in new studies against a larger serological panel for the diagnosis of CanL.
Assuntos
Doenças do Cão , Leishmania infantum , Leishmaniose Visceral , Animais , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Doenças do Cão/diagnóstico , Cães , Ensaio de Imunoadsorção Enzimática/veterinária , Epitopos de Linfócito B/genética , Epitopos de Linfócito B/imunologia , Leishmania infantum/genética , Leishmania infantum/imunologia , Leishmaniose Visceral/diagnóstico , Leishmaniose Visceral/veterinária , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Sensibilidade e Especificidade , Testes Sorológicos/veterináriaRESUMO
BACKGROUND: Ventricular pre-excitation is characterized by the presence of atrioventricular accessory pathways, predisposing to arrhythmias. Although it is well established that risk stratification in symptomatic patients should be invasive, there is a lack of evidence of the benefit in asymptomatic. OBJECTIVE: Evaluate ventricular pre-excitation in the electrocardiogram (ECG) as a risk factor for overall mortality in patients of Telehealth Network of Minas Gerais (TNMG), Brazil. METHODS: This observational study was developed with the database of digital ECGs (2010-2017) from TNMG. The electronic cohort was obtained by linking data from ECG exams and those from the national mortality information system. Only the first ECG was considered. Clinical data were self-reported, and ECGs were interpreted manually by cardiologists and automatically by the Glasgow University Interpreter software. Hazard ratio (HR) for mortality was estimated using weighted Cox regression. RESULTS: Nearly 1 665 667 patients were included (median age: 50 [Q1: 34; Q3: 63] years; 41.4% were male). In a mean follow-up of 3.7 years, the overall mortality rate was 3.1%. The prevalence of ventricular pre-excitation was 0.07%. In multivariate analysis, adjusting for sex and age, ventricular pre-excitation was not associated with an increased risk of mortality (HR: 1.41; 95% confidence interval [CI]: 0.56-3.57; p = .47) when compared to the whole sample or to patients with normal ECG (HR: 1.41; 95% CI: 0.53-4.36; p = .43). In a subanalysis on accessory pathway location, there was no evidence of a higher risk of death related to any location. CONCLUSION: Ventricular pre-excitation was not associated with an increased risk of mortality in a primary care cohort.
Assuntos
Feixe Acessório Atrioventricular , Síndromes de Pré-Excitação , Adulto , Arritmias Cardíacas , Eletrocardiografia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Síndromes de Pré-Excitação/diagnóstico , Atenção Primária à SaúdeRESUMO
Treatment for visceral leishmaniasis (VL) is hampered mainly by the toxicity and/or high cost of antileishmanial drugs. What is more, variability on sensitivity and/or specificity of diagnostic tests hinders effective disease management. In this context, prophylactic vaccination should be considered as a strategy to prevent disease. In the present study, immunogenicity of the Leishmania eukaryotic Elongation Factor-1 beta (EF1b) protein, classified as a Leishmania virulence factor, was evaluated in vitro and in vivo and tested, for the first time, as a vaccine candidate against Leishmania infantum infection. The antigen was administered as DNA vaccine or as recombinant protein (rEF1b) delivered in saponin. BALB/c mice immunization with a DNA plasmid and recombinant protein plus saponin induced development of specific Th1-type immunity, characterized by high levels of IFN-γ, IL-12, GM-CSF, both T cell subtypes and antileishmanial IgG2a isotype antibodies, before and after infection. This immunological response to the vaccines was corroborated further by parasitological analysis of the vaccinated and then challenged mice, which showed significant reductions in the parasite load in their liver, spleen, bone marrow and draining lymph nodes, when compared to the controls. Vaccination using rEF1b/saponin induced a more robust Th1 response and parasitological protection when compared to the DNA vaccine. Furthermore, in vitro analysis of lymphoproliferation, IFN-γ and IL-10 levels in human PBMC cultures showed as well development of a specific Th1-type response. In conclusion, data suggest that EF1b could be a promising vaccine candidate to protect against L. infantum infection.
Assuntos
Leishmania infantum , Vacinas contra Leishmaniose , Animais , Antígenos de Protozoários/genética , Leucócitos Mononucleares , Camundongos , Camundongos Endogâmicos BALB C , Fatores de Alongamento de PeptídeosRESUMO
Aims: Atrial fibrillation (AF) is a public health problem and its prevalence is increasing worldwide. Electronic cohorts, with large electrocardiogram (ECG) databases linked to mortality data, can be useful in determining prognostic value of ECG abnormalities. Our aim is to evaluate the risk of mortality in patients with AF from Brazil. Methods: This observational retrospective study of primary care patients was developed with the digital ECG database from the Telehealth Network of Minas Gerais, Brazil. ECGs performed from 2010 to 2017 were interpreted by cardiologists and the University of Glasgow automated analysis software. An electronic cohort was obtained linking data from ECG exams and those from a national mortality information system, using standard probabilistic linkage methods. We considered only the first ECG of each patient. Patients under 16 years were excluded. Hazard ratios (HR) for mortality were adjusted for demographic and self-reported clinical factors and estimated with Cox regression. Results: From a dataset of 1,773,689 patients, 1,558,421 were included, mean age 51.6 years; 40.2% male. There were 3.34% deaths from all causes in 3.68 years of median follow up. The prevalence of AF was 1.33%. AF was an independent risk factor for all-cause mortality (HR 2.10, 95%CI 2.03-2.17) and cardiovascular mortality (HR 2.06, 95%CI 1.86-2.29). Females with AF had a higher risk of overall and cardiovascular mortality compared with males (p < 0.001). Conclusions: AF was a strong predictor of cardiovascular and all-cause mortality in a primary care population, with increased risk in women. Condensed abstract: To assess risk of mortality in AF patients, an electronic cohort was obtained linking data from ECG exams of Brazilian primary care patients and a national mortality information system. From 1,558,421 patients, AF (prevalence 1.33%) carried a higher risk of overall and cardiovascular mortality, with increased risk in women. What's New: This is the first study with a large Brazilian electronic cohort to evaluate the risk of mortality linked to AF in primary care patients.AF patients from a Brazilian primary care population had a higher risk of death for all causes (HR 2.10, 95%CI 2.03-2.17) and cardiovascular mortality (HR 2.06, 95%CI 1.86-2.29).Female patients with AF had an increased risk of overall and cardiovascular mortality compared with male patients (p < 0.001).
Assuntos
Fibrilação Atrial/mortalidade , Eletrocardiografia/métodos , Frequência Cardíaca/fisiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Fibrilação Atrial/fisiopatologia , Brasil/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Taxa de Sobrevida/tendências , Adulto JovemRESUMO
Most studies evaluating vaccine candidates against visceral leishmaniasis (VL) have used parasite promastigote-expressed antigens; however, Leishmania proteins expressed in the amastigote forms should be considered, since few hours after infection this stage comes into contact with the host immune system and is responsible for the development of the disease. In this context, in the present study, a Leishmania amastigote-specific hypothetical protein, called LiHyJ, was evaluated as a recombinant protein plus saponin as an adjuvant or DNA vaccine to protect against VL. The vaccine effect was evaluated by means of the evaluation of immunological and parasitological analyses performed in BALB/c mice against Leishmania infantum infection. Results showed that rLiHyJ/saponin and DNA LiHyJ induced significantly higher levels of anti-protein and anti-parasite IFN-γ, IL-12, GM-CSF, and IgG2a isotype antibodies, which were associated with a low presence of IL-4 and IL-10. DNA vaccination induced higher IFN-γ production, mainly by CD8+ T cells, while rLiHyJ/saponin stimulated the production of this cytokine, mainly by CD4+ T cells. The parasite load evaluated in distinct organs showed that both immunization schedules significantly reduced organic parasitism, when compared to the controls. Similar results were found in the immunological and parasitological assays when using the recombinant protein or DNA, although the vaccination with rLiHyJ plus saponin induced a slightly higher Th1 response and lower parasite load, when compared to the use of DNA plasmid. The protein also proved to be immunogenic when peripheral blood mononuclear cells of treated VL patients and healthy subjects were in vitro stimulated, since higher IFN-γ and lower IL-4 and IL-10 levels were found in the culture supernatants. In conclusion, LiHyJ should be considered in future studies as a vaccine candidate to protect against VL.
Assuntos
Leishmania/imunologia , Vacinas contra Leishmaniose/imunologia , Vacinas de DNA/imunologia , Adjuvantes Imunológicos/administração & dosagem , Adulto , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Linfócitos T CD8-Positivos/imunologia , DNA/imunologia , Feminino , Humanos , Leishmania/patogenicidade , Leishmania infantum/imunologia , Leishmaniose Visceral/imunologia , Leucócitos Mononucleares/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pessoa de Meia-Idade , Proteínas de Protozoários/imunologia , Proteínas Recombinantes/imunologiaRESUMO
The treatment against visceral leishmaniasis (VL) presents problems, mainly related to the toxicity and/or high cost of the drugs. In this context, a prophylactic vaccination is urgently required. In the present study, a Leishmania protein called LiHyE, which was suggested recently as an antigenic marker for canine and human VL, was evaluated regarding its immunogenicity and protective efficacy in BALB/c mice against Leishmania infantum infection. In addition, the protein was used to stimulate peripheral blood mononuclear cells (PBMCs) from VL patients before and after treatment, as well as from healthy subjects. Vaccination results showed that the recombinant (rLiHyE) protein associated with liposome or saponin induced effective protection in the mice, since significant reductions in the parasite load in spleen, liver, draining lymph nodes, and bone marrow were found. The parasitological protection was associated with Th1-type cell response, since high IFN-γ, IL-12, and GM-CSF levels, in addition to low IL-4 and IL-10 production, were found. Liposome induced a better parasitological and immunological protection than did saponin. Experiments using PBMCs showed rLiHyE-stimulated lymphoproliferation in treated patients' and healthy subjects' cells, as well as high IFN-γ levels in the cell supernatant. In conclusion, rLiHyE could be considered for future studies as a vaccine candidate against VL.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Antígenos de Protozoários/administração & dosagem , Leishmania infantum/imunologia , Leishmaniose Visceral/prevenção & controle , Animais , Antígenos de Protozoários/imunologia , Feminino , Humanos , Imunogenicidade da Vacina , Leishmaniose Visceral/imunologia , Leishmaniose Visceral/parasitologia , Leucócitos Mononucleares/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Carga Parasitária , Células Th1/imunologia , VacinaçãoRESUMO
Leishmania infantum pyridoxal kinase (PK) protein was characterized after an immunoproteomics screening performed with the sera from patients suffering visceral leishmaniasis (VL). Since it was recognized by sera of mammalian hosts infected by a viscerotropic Leishmania species, PK could emerge as a new vaccine candidate against disease, due to its antigenicity and immunogenicity. In this context, in the present study, the effects of the immunization using PK were evaluated when administered as a DNA plasmid (pDNAA3/PK) or recombinant protein (rPK) plus saponin. The immune response elicited by both vaccination regimens reduced in significant levels the parasite load in spleen, liver, draining lymph nodes and bone marrow, being associated with the development of Th1-type immune response, which was characterized by high levels of IFN-γ, IL-12, GM-CSF, and specific IgG2a antibody, besides low production of IL-4, IL-10, and protein and parasite-specific IgG1 antibodies. CD8+ T cells were more important in the IFN-γ production in the pDNAA3/PK group, while CD4+ T cells contributed more significantly to production of this cytokine in the rPK/Saponin group. In addition, increased IFN-γ secretion, along with low levels of IL-10, were found when PBMCs from VL patients after treatment and healthy individuals were stimulated with the protein. In conclusion, when administered either as a DNA plasmid or recombinant protein plus adjuvant, PK can direct the immune response towards a Th1-type immune profile, protecting mice against L. infantum challenge; therefore, it can be seen as a promising immunogen against human VL.
Assuntos
Antígenos de Protozoários/imunologia , Vacinas contra Leishmaniose/imunologia , Leishmaniose Visceral/imunologia , Piridoxal Quinase/imunologia , Animais , Anticorpos Antiprotozoários/imunologia , Humanos , Leishmania infantum/imunologia , Camundongos , Proteínas Recombinantes/imunologia , Vacinas de DNA/imunologiaRESUMO
Background: Leishmaniases are neglected diseases caused by infection with Leishmania parasites and there are no human vaccines in use routinely. The purpose of this study was to examine the immunogenicity of ChimeraT, a novel synthetic recombinant vaccine against visceral leishmaniasis (VL), incorporated into a human-compatible liposome formulation. Methods: BALB/c mice were immunized subcutaneously with ChimeraT/liposome vaccine, ChimeraT/saponin adjuvant, or ChimeraT/saline and immune responses examined in vitro and in vivo. Results: Immunization with the ChimeraT/liposome formulation induced a polarized Th1-type response and significant protection against L. infantum infection. ChimeraT/liposome vaccine stimulated significantly high levels of interferon (IFN)-γ, interleukin (IL)-12, and granulocyte macrophage-colony stimulating factor (GM-CSF) cytokines by both CD4 and CD8 T-cells, with correspondingly lower levels of IL-4 and IL-10 cytokines. Induced antibodies were predominantly IgG2a isotype, and homologous antigen-stimulated spleen cells produced significant nitrite as a proxy for nitric oxide (NO). Furthermore, we examined a small number of treated VL patients and found higher levels of circulating anti-ChimeraT protein IgG2 antibodies, compared to IgG1 levels. Conclusions: Overall, the liposomal formulation of ChimeraT induced a protective Th1-type immune response and thus could be considered in future studies as a vaccine candidate against human VL.
RESUMO
The treatment against visceral leishmaniasis (VL) presents problems, mainly related to the toxicity and/or high cost of the drugs. In this context, a rapid and precise diagnosis of the disease should be performed, mainly to treat patients as soon as possible, aiming to reduce the treatment time and the toxicity of the therapeutics. In the present study, the diagnostic role of an amastigote-specific Leishmania protein was evaluated in the canine and human VL. Results showed that the recombinant protein (called rLiHyJ) and one specific B cell epitope (called PeptJ) predicted from protein sequence presented high sensitivity and specificity values to diagnose canine and human disease, showing also a low reactivity against cross-reactive samples. The rA2 protein and a parasite antigenic extract showed variable sensitivity and/or specificity values in the ELISA experiments. A prognostic evaluation of protein and peptide in the human VL indicated that specific IgG antibodies significantly decreased after treatment, when compared to be values obtained before therapy. The in vitro immunogenicity using rLiHyJ in peripheral blood mononuclear cell (PBMC) cultures collected of such patients and healthy subjects suggested that the protein induced lymphoproliferation and high IFN-γ production in the stimulated cells. In conclusion, although preliminary, results suggest that rLiHyJ and PeptJ could present distinct biotechnological applications in the canine and human VL.
Assuntos
Doenças do Cão , Leishmania infantum , Leishmaniose Visceral , Animais , Antígenos de Protozoários , Doenças do Cão/diagnóstico , Cães , Epitopos de Linfócito B , Humanos , Leishmaniose Visceral/diagnóstico , Leishmaniose Visceral/veterinária , Leucócitos MononuclearesRESUMO
The control measures against visceral leishmaniasis (VL) include a precise diagnosis of disease, the treatment of human cases, and reservoir and vector controls. However, these are insufficient to avoid the spread of the disease in specific countries worldwide. As a consequence, prophylactic vaccination could be interesting, although no effective candidate against human disease is available. In the present study, the Leishmania infantum amastin protein was evaluated regarding its immunogenicity and protective efficacy against experimental VL. BALB/c mice immunized with subcutaneous injections of the recombinant protein with or without liposome/saponin (Lip/Sap) as an adjuvant. After immunization, half of the animals per group were euthanized and immunological evaluations were performed, while the others were challenged with L. infantum promastigotes. Forty-five days after infection, the animals were euthanized and parasitological and immunological evaluations were performed. Results showed the development of a Th1-type immune response in rAmastin-Lip and rAmastin-Sap/vaccinated mice, before and after infection, which was based on the production of protein and parasite-specific IFN-γ, IL-12, GM-CSF, and nitrite, as well as the IgG2a isotype antibody. CD4+ T cells were mainly responsible for IFN-γ production in vaccinated mice, which also presented significant reductions in parasitism in their liver, spleen, draining lymph nodes, and bone marrow. In addition, PBMC cultures of treated VL patients and healthy subjects stimulated with rAmastin showed lymphoproliferation and higher IFN-γ production. In conclusion, the present study shows the first case of an L. infantum amastin protein associated with distinct delivery systems inducing protection against L. infantum infection and demonstrates an immunogenic effect of this protein in human cells.
Assuntos
Leishmania infantum/imunologia , Leishmaniose Visceral/imunologia , Proteínas de Protozoários/imunologia , Adjuvantes Imunológicos/farmacologia , Sequência de Aminoácidos , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/parasitologia , Células Cultivadas , Feminino , Humanos , Imunidade/imunologia , Interferon gama/imunologia , Leishmaniose Visceral/parasitologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/parasitologia , Linfonodos/imunologia , Linfonodos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/imunologia , Células Th1/imunologia , Células Th1/parasitologiaRESUMO
Visceral leishmaniasis (VL) is a highly neglected disease that is present in several countries worldwide. Present-day treatments against this disease are unsuitable, mainly due to the toxicity and/or high cost of drugs. In addition, the development of vaccines is still insufficient. In this scenario, a prompt VL diagnosis was deemed necessary, although sensitivity and/or specificity values of the tests have been. In this context, new antigenic candidates should be identified to be employed in a more precise diagnosis of canine and human VL. In this light, the present study evaluated the diagnostic efficacy of the Leishmania infantum pyridoxal kinase (PK) protein, applied in its recombinant version (rPK). In addition, one specific B-cell epitope derived of the PK sequence was predicted, synthetized, and evaluated as diagnostic marker. Results in ELISA tests showed that the antigens were highly sensitive to VL identification in dogs and human sera, presenting a low reactivity with VL-related disease samples. The recombinant A2 (rA2) protein and L. infantum antigenic preparation (SLA), used as controls, also proved to be highly sensitive in detecting symptomatic cases, although a low sensitivity was found when asymptomatic sera were analyzed. High cross-reactivity was also found when these antigens were evaluated against VL-related disease samples. The post-therapeutic serological follow-up showed that anti-rPK and anti-peptide IgG antibody levels decreased in significant levels after treatment. By contrast, the presence of high levels of the anti-rA2 and anti-SLA antibodies was still detected after therapy. In conclusion, rPK and its specific B-cell epitope should be considered for future studies as a diagnostic marker for canine and human VL.