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1.
PLoS Negl Trop Dis ; 14(2): e0007991, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32023254

RESUMO

BACKGROUND: During infections involving intracellular pathogens, iron performs a double-edged function by providing the pathogen with nutrients, but also boosts the host's antimicrobial arsenal. Although the role of iron has been described in visceral leishmaniasis, information regarding its status in the dermal sequel, Post Kala-azar Dermal Leishmaniasis (PKDL) remains limited. Accordingly, this study aimed to establish the status of iron within monocytes/macrophages of PKDL cases. METHODOLOGY/PRINCIPAL FINDINGS: The intramonocytic labile iron pool (LIP), status of CD163 (hemoglobin-haptoglobin scavenging receptor) and CD71 (transferrin receptor, Tfr) were evaluated within CD14+ monocytes by flow cytometry, and soluble CD163 by ELISA. At the lesional sites, Fe3+ status was evaluated by Prussian blue staining, parasite load by qPCR, while the mRNA expression of Tfr (TfR1/CD71), CD163, divalent metal transporter-1 (DMT-1), Lipocalin-2 (Lcn-2), Heme-oxygenase-1 (HO-1), Ferritin, Natural resistance-associated macrophage protein (NRAMP-1) and Ferroportin (Fpn-1) was evaluated by droplet digital PCR. Circulating monocytes demonstrated elevated levels of CD71, CD163 and soluble CD163, which corroborated with an enhanced lesional mRNA expression of TfR, CD163, DMT1 and Lcn-2. Additionally, the LIP was raised along with an elevated mRNA expression of ferritin and HO-1, as also iron exporters NRAMP-1 and Fpn-1. CONCLUSIONS/SIGNIFICANCE: In monocytes/macrophages of PKDL cases, enhancement of the iron influx gateways (TfR, CD163, DMT-1 and Lcn-2) possibly accounted for the enhanced LIP. However, enhancement of the iron exporters (NRAMP-1 and Fpn-1) defied the classical Ferritinlow/Ferroportinhigh phenotype of alternatively activated macrophages. The creation of such a pro-parasitic environment suggests incorporation of chemotherapeutic strategies wherein the availability of iron to the parasite can be restricted.


Assuntos
Ferro/metabolismo , Leishmaniose Cutânea/metabolismo , Adolescente , Adulto , Antígenos CD/genética , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/genética , Antígenos de Diferenciação Mielomonocítica/metabolismo , Proteínas de Transporte de Cátions/genética , Proteínas de Transporte de Cátions/metabolismo , Feminino , Humanos , Índia , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/fisiologia , Leishmaniose Cutânea/parasitologia , Lipocalina-2/genética , Lipocalina-2/metabolismo , Macrófagos/metabolismo , Masculino , Monócitos/metabolismo , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores da Transferrina/genética , Receptores da Transferrina/metabolismo , Adulto Jovem
2.
PLoS Negl Trop Dis ; 14(2): e0008029, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32023240

RESUMO

Interleukin-32 is a novel inflammatory mediator that has been described to be important in the immunopathogenesis and control of infections caused by Leishmania parasites. By performing experiments with primary human cells in vitro, we demonstrate that the expression of IL-32 isoforms is dependent on the time exposed to L. amazonensis and L. braziliensis antigens. Moreover, for the first time we show the functional consequences of three different genetic variations in the IL32 (rs4786370, rs4349147, rs1555001) modulating IL-32γ expression, influencing innate and adaptive cytokine production after Leishmania exposure. Using a Brazilian cohort of 107 American Tegumentary Leishmaniasis patients and a control cohort of 245 healthy individuals, the IL32 rs4786370 genetic variant was associated with protection against ATL, whereas the IL32 rs4349147 was associated with susceptibility to the development of localized cutaneous and mucosal leishmaniasis. These novel insights may help improve therapeutic strategies and lead to benefits for patients suffering from Leishmania infections.


Assuntos
Predisposição Genética para Doença , Variação Genética , Interleucinas/genética , Leishmania/classificação , Leishmaniose Cutânea/genética , Adulto , Idoso , Brasil/epidemiologia , Citocinas/genética , Citocinas/metabolismo , Feminino , Regulação da Expressão Gênica/imunologia , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Leishmaniose Cutânea/epidemiologia , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Masculino , Pessoa de Meia-Idade , Proteína Adaptadora de Sinalização NOD2/genética , Proteína Adaptadora de Sinalização NOD2/metabolismo , Isoformas de Proteínas , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo
3.
BMC Res Notes ; 12(1): 803, 2019 Dec 12.
Artigo em Inglês | MEDLINE | ID: mdl-31831065

RESUMO

OBJECTIVE: The resistance to antimony-containing glucantime is a major obstacle to successful treatment, especially in endemic areas. Looking the molecular mechanisms involved in this drug resistance will help in choosing the best treatment. The aim of this study was to evaluate the expression of multidrug-resistance 1 (MDR1) and multidrug-resistance protein A (MRPA) genes in acute, chronic non-lupoid, and chronic lupoid forms of dry type cutaneous leishmaniasis (DTCL). RESULTS: MDR1 gene was over-expressed as 14.4- and 1.56-folds in the chronic lupoid and acute forms compared with the chronic non-lupoid form, respectively. Results comparison showed P < 0.05 between the chronic non-lupoid and acute groups, P < 0.01 between acute and chronic lupoid groups, and P < 0.001 between the chronic non-lupoid and chronic lupoid groups. MRPA gene was over-expressed as 266 and 17.7-fold in the chronic lupoid and chronic non-lupoid forms compared with the acute form, respectively. Statistical analysis showed P < 0.01 between the chronic non-lupoid and chronic lupoid groups, P < 0.05 between acute and chronic non-lupoid groups, and P < 0.001 between the acute and chronic lupoid groups.


Assuntos
Leishmaniose Cutânea/metabolismo , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Doença Aguda , Doença Crônica , Expressão Gênica , Humanos , Leishmaniose Cutânea/genética , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Pele/metabolismo , Regulação para Cima/genética
4.
Rev Soc Bras Med Trop ; 52: e20190361, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31800923

RESUMO

INTRODUCTION: Cutaneous leishmaniasis is caused by protozoa of the genus Leishmania and transmission occurs through the bite of sandflies. It is an infectious disease, which affects skin and mucosa. The aim was to quantify the macrophages M1 and M2 and the annexin A1 expression in the skin lesions of patients with cutaneous leishmaniasis. METHODS: Skin biopsies from patients (n = 50) were analyzed and classified according to the lesion type as: exudative cellular reaction, exudative granulomatous reaction, exudative necrotic reaction, exudative necrotic-granulomatous reaction. Using the immunofluorescence technique, macrophages were identified by CD163 marker, differentiated by anti-MHCII and anti-CD206 antibodies, and annexin A1 expression was determined by arbitrary unit (a.u.) densitometry. RESULTS: In M1 macrophages, a greater expression of this protein was observed in the exudative cellular reaction type lesions (136.3 ± 2.6 a.u., assuming mean and standard derivation) when compared to the expression in the lesions of exudative granulomatous reaction, exudative necrotic reaction and exudative necrotic-granulomatous reaction patients (108.0 ± 2.3, 121.6 ± 3.2 and 124.7 ± 2.4 a.u., respectively). Regarding M2 macrophages, it was observed that patients with exudative cellular reaction lesion also had a higher expression of this protein (128.8 ± 2.6 a.u.), when compared to the expression in the lesions of exudative granulomatous reaction, exudative necrotic reaction and exudative necrotic-granulomatous reaction patients (105.6 ± 2, 113.9 ± 2.8, 114.3 ± 2.1 a.u., respectively). CONCLUSIONS: These data suggest that annexin A1 is assisting macrophages in the phagocytosis process of patients with exudative cellular reaction lesion type.


Assuntos
Anexina A1/metabolismo , Leishmaniose Cutânea/metabolismo , Macrófagos/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Anexina A1/análise , Biópsia , Feminino , Imunofluorescência , Humanos , Leishmaniose Cutânea/patologia , Macrófagos/parasitologia , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição , Adulto Jovem
5.
Sensors (Basel) ; 19(21)2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31661834

RESUMO

Cutaneous leishmaniasis (CL) is a neglected tropical disease that requires novel tools for its understanding, diagnosis, and treatment follow-up. In the cases of other cutaneous pathologies, such as cancer or cutaneous ulcers due to diabetes, optical diffuse reflectance-based tools and methods are widely used for the investigation of those illnesses. These types of tools and methods offer the possibility to develop portable diagnosis and treatment follow-up systems. In this article, we propose the use of a three-layer diffuse reflectance model for the study of the formation of cutaneous ulcers caused by CL. The proposed model together with an inverse-modeling procedure were used in the evaluation of diffuse-reflectance spectral signatures acquired from cutaneous ulcers formed in the dorsal area of 21 golden hamsters inoculated with Leishmanisis braziliensis. As result, the quantification of the model's variables related to the main biological parameters of skin were obtained, such as: diameter and volumetric fraction of keratinocytes, collagen; volumetric fraction of hemoglobin, and oxygen saturation. Those parameters show statistically significant differences among the different stages of the CL ulcer formation. We found that these differences are coherent with histopathological manifestations reported in the literature for the main phases of CL formation.


Assuntos
Leishmaniose Cutânea/patologia , Úlcera Cutânea/patologia , Pele/química , Espectrofotometria/métodos , Animais , Colágeno/fisiologia , Cricetinae , Modelos Animais de Doenças , Processamento Eletrônico de Dados , Feminino , Hemoglobinas/química , Leishmaniose Cutânea/metabolismo , Masculino , Mesocricetus , Oxigênio/química , Pele/patologia , Úlcera Cutânea/metabolismo , Úlcera Cutânea/parasitologia
6.
Microb Pathog ; 137: 103738, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31513893

RESUMO

INTRODUCTION: Cutaneous leishmaniasis is a dermal disease caused by several species of the genus Leishmania. It is an endemic disease with 1.2 million new cases occurring annually and mostly in developing countries. Oxidative stress is a condition of an imbalance in oxidant/antioxidant which may play a role in many different pathologic conditions. For the first time in this study, we introduced isoprostane as a reliable index for oxidative stress in patients suffering from leishmaniasis. We also investigated the possible relation between quantitative CRP and this disease. METHOD AND MATERIAL: We collected 5 ml blood of 30 patients in addition to the same sample of the control healthy group. After applying appropriate methods, the plasma and serum specimens were extracted in order to conduct oxidant-antioxidant balance and CRP tests in serum as well as measuring isoprostane factor in plasma. STATISTICAL ANALYSIS: We used T-student, ANOVA as well as linear regression to analyze the gathered data with a 0.05 confidence interval in SPSS environment. RESULTS: The results showed a significant difference between the two groups in terms of the oxidant-antioxidant balance. Also, isoprostane and quantitative CRP levels were substantially higher in patients. There was no significant relationship between the mentioned factors and wound size and number. CONCLUSION: Leishmania Amastigotes plays an important role in disturbing the oxidant-antioxidant balance resulting in inflammation and stress in patients. Furthermore, isoprostane was confirmed as a reliable index for evaluating oxidative stress in patients.


Assuntos
Antioxidantes/análise , Proteína C-Reativa/análise , Isoprostanos/sangue , Leishmaniose Cutânea/sangue , Leishmaniose Cutânea/metabolismo , Oxidantes/sangue , Estudos de Casos e Controles , Feminino , Humanos , Irã (Geográfico) , Masculino , Estresse Oxidativo
7.
Infect Immun ; 87(11)2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31451620

RESUMO

Cutaneous leishmaniasis is characterized by vascular remodeling. Following infection with Leishmania parasites, the vascular endothelial growth factor A (VEGF-A)/VEGF receptor 2 (VEGFR-2) signaling pathway mediates lymphangiogenesis, which is critical for lesion resolution. Therefore, we investigated the cellular and molecular mediators involved in VEGF-A/VEGFR-2 signaling using a murine model of infection. We found that macrophages are the predominant cell type expressing VEGF-A during Leishmania major infection. Given that Leishmania parasites activate hypoxia-inducible factor 1α (HIF-1α) and this transcription factor can drive VEGF-A expression, we analyzed the expression of HIF-1α during infection. We showed that macrophages were also the major cell type expressing HIF-1α during infection and that infection-induced VEGF-A production is mediated by ARNT/HIF activation. Furthermore, mice deficient in myeloid ARNT/HIF signaling exhibited larger lesions without differences in parasite numbers. These data show that L. major infection induces macrophage VEGF-A production in an ARNT/HIF-dependent manner and suggest that ARNT/HIF signaling may limit inflammation by promoting VEGF-A production and, thus, lymphangiogenesis during infection.


Assuntos
Translocador Nuclear Receptor Aril Hidrocarboneto/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Leishmaniose Cutânea/metabolismo , Macrófagos/metabolismo , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Translocador Nuclear Receptor Aril Hidrocarboneto/genética , Células Cultivadas , Regulação da Expressão Gênica , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Leishmania major , Leishmaniose Cutânea/parasitologia , Leishmaniose Cutânea/patologia , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais , Pele/metabolismo , Pele/parasitologia , Fator A de Crescimento do Endotélio Vascular/genética
8.
Photobiomodul Photomed Laser Surg ; 37(5): 298-304, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31084559

RESUMO

Objective: In this study, we evaluated the effectiveness of photodynamic therapy (PDT) for the treatment of experimental cutaneous leishmaniasis (CL) and the profile of macrophages activation markers. Background: Leishmaniasis is an infectious disease caused by parasites of the genus Leishmania. CL is caused by Leishmania major in the old world and by Leishmania braziliensis in the Americas. Considering the targeted organs, PDT may constitute a valuable therapeutic intervention. Macrophages are the host cells of Leishmania in mammals and may be classified into type M1 or M2 depending on the pattern of activation. Methods: BALB/c mice were infected in the foot pad with 1 × 106 amastigotes of L. braziliensis and treated with 5-aminolevulinic acid (5-ALA), visible light, or 5-ALA-PDT. The ex vivo mRNA expression levels of interleukin-10, tumor necrosis factor-α (TNF-α), arginase-1, heme oxygenase ( Hmox), and induced nitric oxide synthase (iNOS) were quantities as markers of macrophage activation with distinct ability to kill intracellular parasite. Results: The parasite load decreased significantly in the group treated with PDT compared with the other groups. The iNOS relative mRNA was higher in the group treated with PDT and light only compared with the group without treatment, whereas iNOS/arginase ratio was significantly higher only in the PDT group. The expression of TNF-α was significantly higher in 5-ALA and light compared with PDT and control group. No significant difference was observed in the expression of the other markers evaluated. Conclusions: Both, light and 5-ALA-PDT were able to upregulate iNOS expression only; 5-ALA-PDT was able to reduce parasite burden. The increase in the iNOS levels suggests it might participate in the antimicrobial mechanisms triggered by 5-ALA-PDT; although parasite death mechanism was not completely clarified, the results presented in this study suggest that macrophage activation may contribute to parasite control.


Assuntos
Ácido Aminolevulínico/uso terapêutico , Leishmaniose Cutânea/terapia , Ativação de Macrófagos/efeitos da radiação , Fotoquimioterapia , Fármacos Fotossensibilizantes/uso terapêutico , Animais , Modelos Animais de Doenças , Interleucina-10/metabolismo , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Carga Parasitária , Fator de Necrose Tumoral alfa/metabolismo
9.
Immun Inflamm Dis ; 7(3): 95-104, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-30997749

RESUMO

INTRODUCTION: Zoonotic cutaneous leishmaniasis (ZCL), due to infection by Leishmania (L). major, is characterized by polymorphic clinical manifestations which could be attributed to the host's immune response. In this study we investigated the involvement of cytotoxic cells on the outcome of the disease. METHODS: Expression of granzyme B (GrB), granulysine (Grly), and interferon (IFN)-γ was evaluated within ZCL lesion specimens using the technique of real-time quantitative polymerase chain reaction (RT-qPCR). Immunohistochemical staining was performed using anti-CD3, CD4, CD8, CD56, GrB, and IFN-γ antibodies to identify the phenotype of GrB and IFN-γ-producing cells. RESULTS: GrB and Grly mRNA was detected within 75% and 80% of ZCL lesions, respectively. Statistical analysis demonstrated a significant correlation between levels of GrB and Grly. Interestingly, expression of these molecules correlates negatively with the lesion's age. The highest levels were measured in early lesions (E-ZCL) (lesion age ≤1 month) comparing to late lesions (L-ZCL) (lesion age >1 month). Otherwise, IFN-γ mRNA was detected only within 56% and a positive correlation was found between levels of this cytokine and those of GrB. Immunohistochemical analysis showed that GrB is produced essentially by CD8+ T cells whereas IFN-γ is produced by both CD4+ and CD8+ T cells. CONCLUSION: Together our results demonstrate the presence of cytotoxic cells producing GrB and Grly within leishmaniasis cutaneous lesions.


Assuntos
Leishmania major/imunologia , Leishmaniose Cutânea/imunologia , Linfócitos T Citotóxicos/imunologia , Zoonoses/imunologia , Adulto , Animais , Antígenos de Diferenciação de Linfócitos T/genética , Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos de Diferenciação de Linfócitos T/metabolismo , Criança , Pré-Escolar , Citocinas/imunologia , Citocinas/metabolismo , Feminino , Expressão Gênica/imunologia , Granzimas/genética , Granzimas/imunologia , Granzimas/metabolismo , Humanos , Lactente , Recém-Nascido , Interferon gama/genética , Interferon gama/metabolismo , Leishmania major/parasitologia , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/parasitologia , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Pele/imunologia , Pele/parasitologia , Linfócitos T Citotóxicos/metabolismo , Zoonoses/parasitologia
10.
Methods Mol Biol ; 1971: 315-349, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30980313

RESUMO

Experimental cutaneous leishmaniasis of mice is a valuable model to study the immune response to the protozoan pathogen Leishmania and to define mechanisms of parasite control and resolution of inflammation as well as of parasite evasion and chronicity of disease. In addition, over many years Leishmania-infected mice have been successfully used to analyze the function of newly discovered immune cell types, transcription factors, cytokines, and effector mechanisms in vivo. In this chapter we present detailed protocols for the culture, propagation, and inoculation of Leishmania promastigotes, the monitoring of the course of cutaneous infection, the determination of the tissue parasite burden and for the phenotyping of the ensuing immune response. The focus lies on the L. major mouse model, but an overview on other established models of murine cutaneous leishmaniasis is also provided.


Assuntos
Modelos Animais de Doenças , Leishmania/crescimento & desenvolvimento , Leishmaniose Cutânea/parasitologia , Estágios do Ciclo de Vida , Carga Parasitária , Animais , Doença Crônica , Inflamação/metabolismo , Inflamação/parasitologia , Inflamação/patologia , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/patologia , Camundongos , Camundongos Endogâmicos BALB C
11.
Molecules ; 24(7)2019 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-30934998

RESUMO

Leishmaniasis is a neglected tropical disease caused by members of the Leishmania genus of parasitic protozoa that cause different clinical manifestations of the disease. Current treatment options for the cutaneous disease are limited due to severe side effects, poor efficacy, limited availability or accessibility, and developing resistance. Essential oils may provide low cost and readily available treatment options for leishmaniasis. In-vitro screening of a collection of 52 commercially available essential oils has been carried out against promastigotes of Leishmania amazonensis. In addition, cytotoxicity has been determined for the essential oils against mouse peritoneal macrophages in order to determine selectivity. Promising essential oils were further screened against intracellular L. amazonensis amastigotes. Three essential oils showed notable antileishmanial activities: frankincense (Boswellia spp.), coriander (Coriandrum sativum L.), and wintergreen (Gualtheria fragrantissima Wall.) with IC50 values against the amastigotes of 22.1 ± 4.2, 19.1 ± 0.7, and 22.2 ± 3.5 µg/mL and a selectivity of 2, 7, and 6, respectively. These essential oils could be explored as topical treatment options for cutaneous leishmaniasis.


Assuntos
Antiprotozoários/química , Leishmania/química , Óleos Voláteis/química , Animais , Antiprotozoários/farmacologia , Boswellia/química , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Coriandrum/química , Avaliação Pré-Clínica de Medicamentos/métodos , Concentração Inibidora 50 , Leishmaniose Cutânea/metabolismo , Macrófagos Peritoneais/química , Camundongos Endogâmicos BALB C , Óleos Voláteis/farmacologia
12.
J Immunol ; 202(5): 1453-1464, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30665936

RESUMO

Arginase (Arg) 1 is expressed by hematopoietic (e.g., macrophages) and nonhematopoietic cells (e.g., endothelial cells) and converts l-arginine into ornithine and urea. The enzyme is implicated in tissue repair but also antagonizes the production of NO by type 2 NO synthase in myeloid cells and thereby impedes the control of intracellular parasites such as Leishmania major In this study, we tested whether Arg1 is required for spontaneous healing of acute cutaneous leishmaniasis in C57BL/6 mice and for lifelong parasite persistence in draining lymph nodes (dLNs) of healed mice. In vitro, bone marrow-derived macrophages and lymphoid endothelial cells readily expressed Arg1 in response to IL-4 and/or IL-13, whereas skin or dLN fibroblasts failed to do so, even during hypoxia. In vivo, Arg1 was found in skin lesions and, to a much lower extent, also in dLNs of acutely infected C57BL/6 mice but became undetectable at both sites after healing. Deletion of Arg1 in hematopoietic and endothelial cells using Tie2Cre+/-Arg1fl/fl C57BL/6 mice abolished the expression of Arg1 in skin lesions and dLNs but did not affect development and resolution of skin lesions, parasite burden, NO production, or host cell tropism of L. major during the acute or persistent phase of infection. Similar to wild-type controls, parasites persisting in Arg1-deficient mice favored NO synthase 2-negative areas and mainly resided in myeloid cells and fibroblasts. We conclude that Arg1 expression by hematopoietic and endothelial cells is completely dispensable for clinical resolution of cutaneous leishmaniasis and for long-term persistence of L. major.


Assuntos
Arginase/metabolismo , Leishmania major/metabolismo , Leishmaniose Cutânea/metabolismo , Animais , Arginase/genética , Células Cultivadas , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
13.
Biomed Microdevices ; 21(1): 8, 2019 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-30617619

RESUMO

Current therapeutic options against cutaneous leishmaniasis are plagued by several weaknesses. The effective topical delivery of an antileishmanial drug would be useful in treating some forms of cutaneous leishmaniasis. Toward this end, a microneedle based delivery approach for the antileishmanial drug amphotericin B was investigated in murine models of both New World (Leishmania mexicana) and Old World (Leishmania major) infection. In the L. mexicana model, ten days of treatment began on day 35 post infection, when the area of nodules averaged 9-15 mm2. By the end of the experiment, a significant difference in nodule area was observed for all groups receiving topical amphotericin B at 25 mg/kg/day after application of microneedle arrays of 500, 750, and 1000 µM in nominal length compared to the group that received this dose of topical amphotericin B alone. In the L. major model, ten days of treatment began on day 21 post infection when nodule area averaged 51-65 mm2 in the groups. By the end of the experiment, there was no difference in nodule area between the group receiving 25 mg/kg of topical amphotericin B after microneedle application and any of the non-AmBisome groups. These results show the promise of topical delivery of amphotericin B via microneedles in treating relatively small nodules caused by L. mexicana. These data also show the limitations of the approach against a disseminated L. major infection. Further optimization of microneedle delivery is needed to fully exploit this strategy for cutaneous leishmaniasis treatment.


Assuntos
Anfotericina B/farmacologia , Sistemas de Liberação de Medicamentos , Leishmania mexicana/metabolismo , Leishmaniose Cutânea/tratamento farmacológico , Agulhas , Animais , Sistemas de Liberação de Medicamentos/instrumentação , Sistemas de Liberação de Medicamentos/métodos , Feminino , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/patologia , Camundongos , Camundongos Endogâmicos BALB C
14.
Cell Immunol ; 335: 76-84, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30424873

RESUMO

Neutrophils are short-lived phagocytic cells equipped with several receptors for pathogen recognition and phagocytosis and have intracellular and extracellular effector mechanisms that can inactivate pathogens. Leishmaniases are diseases caused by different species of Leishmania that mainly afflicts poorer populations of tropical and subtropical regions and immunocompromised individuals. Thus, the present study aims to investigate the effector response of murine neutrophils to species of Leishmania causing American cutaneous leishmaniasis and zoonotic visceral leishmaniasis by evaluating pattern recognition receptors (PRR) and intracellular and extracellular effector microbicide activity. When exposed to Leishmania parasites, mouse neutrophils produced superoxide, released enzymes in the extracellular space and generated neutrophil extracellular traps, although PRR gene expression is negatively regulated. L. infantum, L. guyanensis, and L. shawi inhibited enzymatic activity, whereas L. amazonensis reduced the emission of extracellular structures. These findings indicate that although neutrophils trigger several microbicide mechanisms, Leishmania parasites can manipulate extracellular effector mechanisms. The present study also provides evidence that neutrophils can internalize parasites by coiling phagocytosis.


Assuntos
Leishmaniose/imunologia , Neutrófilos/imunologia , Receptores de Reconhecimento de Padrão/imunologia , Animais , Linhagem Celular , Citoplasma , Imunidade Inata/imunologia , Leishmania/imunologia , Leishmania/patogenicidade , Leishmaniose/metabolismo , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Leishmaniose Visceral/imunologia , Leishmaniose Visceral/metabolismo , Camundongos , Neutrófilos/metabolismo , Parasitos , Fagocitose
15.
J Cell Physiol ; 234(8): 13145-13156, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-30584667

RESUMO

OBJECTIVES: In our research, we aimed to investigate the roles of CC-chemokine receptor 7 (CCR7) and relevant signaling pathways in Leishmania major-infected human dendritic cells (DCs). METHODS: Differentially expressed genes (DEGs) in L. major-infected human DCs were selected out and visualized using R program. Kyoto Encyclopedia of Genes and Genomes pathway analysis was conducted for investigation of significantly enriched signaling pathways and Gene Ontology enrichment analysis was carried out for the unveiling of enriched Molecular Functions and Biological Processes in L. major-infected human DCs. Besides, Hub gene was screened out using weighted gene coexpression network analysis and Cytoscape. In addition, enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction were used for detection of relative expression of CCR7, interleukin-12 (IL-12), and interferon-γ (IFN-γ) in L. major-infected human DCs and western blot analysis was used for detection of relative expression of CCR7 and other proteins in JAK-STAT signaling pathway in L. major-infected human DCs. RESULTS: CCR7 was upregulated and both chemokine and JAK-STAT signaling pathway were activated in L. major-infected human DCs. During the L. major infection, total number of L. major-infected human DCs were increased, as well as the relative expression levels of CCR7, IL-12, and IFN-γ and proteins in the JAK-STAT signaling pathway. Overexpression of CCR7 not only increased expression levels of IL-12 and IFN-γ but also activated the JAK-STAT signaling pathway to affect the leishmaniasis progression. CONCLUSION: L. major infection-induced activation of CCR7, as well as JAK2 and STAT1, might well upregulate the expression of BAX yet suppress the expression of both Bcl2 and c-Jun to affect leishmaniasis progression.


Assuntos
Células Dendríticas/metabolismo , Leishmaniose Cutânea/metabolismo , Receptores CCR7/metabolismo , Transdução de Sinais/fisiologia , Células Dendríticas/imunologia , Humanos , Janus Quinases/imunologia , Janus Quinases/metabolismo , Leishmania major , Leishmaniose Cutânea/imunologia , Receptores CCR7/imunologia , Fatores de Transcrição STAT/imunologia , Fatores de Transcrição STAT/metabolismo
16.
Immunity ; 49(4): 654-665.e5, 2018 10 16.
Artigo em Inglês | MEDLINE | ID: mdl-30266340

RESUMO

Recruitment of immune cells with antimicrobial activities is essential to fight local infections but has the potential to trigger immunopathology. Whether the immune system has the ability to sense inflammation intensity and self-adjust accordingly to limit tissue damage remains to be fully established. During local infection with an intracellular pathogen, we have shown that nitric oxide (NO) produced by recruited monocyte-derived cells was essential to limit inflammation and cell recruitment. Mechanistically, we have provided evidence that NO dampened monocyte-derived cell cytokine and chemokine production by inhibiting cellular respiration and reducing cellular ATP:ADP ratio. Such metabolic control operated at the tissue level but only when a sufficient number of NO-producing cells reached the site of infection. Thus, NO production and activity act as a quorum sensing mechanism to help terminate the inflammatory response.


Assuntos
Citocinas/imunologia , Inflamação/imunologia , Monócitos/imunologia , Óxido Nítrico/imunologia , Animais , Células Cultivadas , Citocinas/metabolismo , Células HEK293 , Interações Hospedeiro-Parasita/imunologia , Humanos , Inflamação/metabolismo , Inflamação/parasitologia , Leishmania major/imunologia , Leishmania major/fisiologia , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/parasitologia , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/parasitologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Monócitos/metabolismo , Monócitos/parasitologia , Óxido Nítrico/metabolismo , Percepção de Quorum/imunologia
17.
Mediators Inflamm ; 2018: 9787128, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30150896

RESUMO

Certain cytokines modulate the expression of insulin-like growth factor- (IGF-) I. Since IL-4 and IGF-I promote growth of the protozoan Leishmania major, we here addressed their interaction in downregulating the expression of Igf-I mRNA using small interfering RNA (siRNA) in Leishmania major-infected macrophages. Parasitism was decreased in the siRNA-treated cells compared with the nontreated cells, reversed by the addition of recombinant IGF-I (rIGF-I). In IL-4-stimulated macrophages, parasitism and the Igf-I mRNA amount were increased, and the effects were nullified upon siRNA transfection. IGF-I downregulation inhibited both parasite and macrophage arginase activation even in IL-4-stimulated cells. Searching for intracellular signaling components shared by IL-4 and IGF-I, upon siRNA transfection, phosphorylated p44, p38, and Akt proteins were decreased, affecting the phosphatidylinositol-3-kinase (PI3K)/Akt pathway. In L. major-infected C57BL6-resistant mice, the preincubation of the parasite with rIGF-I changed the infection profile to be similar to that of susceptible mice. We conclude that IGF-I constitutes an effector element of IL-4 involving the PI3K/Akt pathway during L. major infection.


Assuntos
Fator de Crescimento Insulin-Like I/metabolismo , Interleucina-4/farmacologia , Leishmania major/metabolismo , Leishmania major/patogenicidade , Leishmaniose Cutânea/metabolismo , Animais , Western Blotting , Eletroforese em Gel de Poliacrilamida , Fator de Crescimento Insulin-Like I/genética , Leishmaniose Cutânea/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células RAW 264.7
18.
J Dermatol Sci ; 92(1): 78-88, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30037731

RESUMO

BACKGROUND: Cutaneous leishmaniasis (CL) skin lesions are the result of a deregulated immune response, which is unable to eliminate Leishmania parasites. The control of both, parasites and host immune response, is critical to prevent tissue destruction. The skin ulceration has been correlated with high TNF-α level. OBJECTIVE: Because human anti-TNF-α antibodies (Ab) have been successfully assayed in several mice inflammatory diseases, we hypothesized that their anti-inflammatory effect could optimize the healing of CL lesions achieved after topical application of paromomycin (PM), the current chemotherapy against CL. METHODS AND RESULTS: We first compared the in vitro efficacy of PM and Ab alone and the drug given in combination with Ab to assess if the Ab could interfere with PM leishmanicidal activity in L. major-infected bone marrow-derived macrophages. The combination therapy had similar antileishmanial activity to the drug alone and showed no influence on NO production, which allows macrophage-mediated parasite killing. Next, we demonstrated in an in vivo model of Imiquimod®-induced inflammation that topical Ab and PM inhibit the infiltration of inflammatory cells in the skin. In the efficacy studies in L. major-infected BALB/c mice, PM combined with Ab led to a sharp infection reduction and showed a stronger anti-inflammatory activity than PM alone. This was confirmed by the down-regulation of TNF-α, IL-1ß, iNOS, IL-17, and CCL3 as well as by a decrease of the neutrophilic infiltrate during infection upon treatment with the Ab. CONCLUSIONS: In terms of parasite elimination and inflammation reduction, topical application of Ab in combination with PM was more effective than the drug alone.


Assuntos
Anticorpos/farmacologia , Antiprotozoários/farmacologia , Dermatite/tratamento farmacológico , Mediadores da Inflamação/antagonistas & inibidores , Leishmania major/efeitos dos fármacos , Leishmaniose Cutânea/tratamento farmacológico , Paromomicina/farmacologia , Pele/efeitos dos fármacos , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Animais , Células Cultivadas , Dermatite/etiologia , Dermatite/imunologia , Dermatite/metabolismo , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Interações Hospedeiro-Patógeno , Imiquimode , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Leishmania major/imunologia , Leishmania major/patogenicidade , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Leishmaniose Cutânea/parasitologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/parasitologia , Camundongos Endogâmicos BALB C , Infiltração de Neutrófilos/efeitos dos fármacos , Pele/imunologia , Pele/metabolismo , Pele/parasitologia , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo
19.
Iran J Immunol ; 15(2): 74-83, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29947337

RESUMO

BACKGROUND: Toll like receptors play a major role in immune responses against Leishmania parasites. OBJECTIVE: To evaluate the efficacy of vaccination with live attenuated L. major and TLR4 agonist in protection against L. major infection. METHODS: Attenuated L. major was prepared by continuous sub-culturing of the parasite. A total of 90 mice were assigned to 9 groups including 6 groups of BALB/c (G1-6) and 3 groups (G7-9) of C57BL/6 mice. Group 1 was the control groups, group 2 received the wild-type L. major promastigotes, group 3 the attenuated line, group 4 the TLR4 agonist, group 5 the wild-type L. major and TLR4 agonist, and group 6 the attenuated line along with TLR4 agonist. Group 7 was control, group 8 received wild-type L. major and group 9 the wild-type along with TLR4 agonist. Vaccinated mice were then challenged with wild-type of L. major. Lesion size, parasite burden, and the expression levels of IL-4, IFN-γ, IL-2, 1L-17A, IL-10, TGF-ß and TLR4 were evaluated before the challenge while parasite burden and lesion size were evaluated. RESULTS: Vaccinated mice with a TLR4 agonist or attenuated L. major plus TLR4 agonist produced the highest levels of IFN-γ, IL-2, and IL-17A. Post-challenge analysis revealed that mice vaccinated with the attenuated line along with TLR4 agonist displayed the lowest lesion size and parasite load. These mice developed a predominant Th1 immune response. CONCLUSION: Vaccination with the attenuated L. major along with TLR4 agonist promotes a Th1-mediated immune response which leads to the protection of BALB/c mice against L. major infection.


Assuntos
Leishmania major/imunologia , Vacinas contra Leishmaniose/imunologia , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Células Th1/imunologia , Receptor 4 Toll-Like/agonistas , Vacinas Atenuadas/imunologia , Animais , Anticorpos Antiprotozoários/sangue , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Citocinas/genética , Citocinas/metabolismo , Feminino , Expressão Gênica , Imunização , Vacinas contra Leishmaniose/administração & dosagem , Leishmaniose Cutânea/genética , Leishmaniose Cutânea/prevenção & controle , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Células Th1/metabolismo , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/metabolismo , Transcrição Genética , Vacinas Atenuadas/administração & dosagem
20.
Mediators Inflamm ; 2018: 3487591, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29743809

RESUMO

In Honduras, Leishmania (L.) infantum chagasi causes both visceral leishmaniasis (LV) and nonulcerated or atypical cutaneous leishmaniasis (NUCL). NUCL is characterized by mononuclear inflammatory infiltration of the dermis, composed mainly of lymphocytes followed by macrophages with discrete parasitism. Considering that little is known about the pathogenesis of NUCL, the aim of this study was to evaluate the regulatory response in situ in skin lesions of patients affected by NUCL. Biopsies (n = 20) from human cutaneous nonulcerative lesions were collected and processed by usual histological techniques. The in situ regulatory immune response was evaluated by immunohistochemistry using antihuman CD4, FoxP3, IL-10, and TGF-ß antibodies. CD4+, FoxP3+, TGF-ß+, and IL-10+ cells were observed in the dermis with inflammatory infiltration in all studied cases and at higher densities compared to the normal skin controls. A positive and strong correlation was observed between CD4+ and FoxP3+ cells, and a positive and moderate correlation was observed between FoxP3+ and TGF-ß+ but not with IL-10+ cells. The data suggest that T regulatory FoxP3+ cells and the regulatory cytokines, especially TGF-ß, play an important role in the immunopathogenesis of NUCL, modulating a cellular immune response in the skin, avoiding tissue damage, and leading to low tissue parasitic persistence.


Assuntos
Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/metabolismo , Pele/metabolismo , Pele/patologia , Antígenos CD4/metabolismo , América Central , Fatores de Transcrição Forkhead/metabolismo , Honduras , Humanos , Imuno-Histoquímica , Interleucina-10/metabolismo , Pele/imunologia , Dermatopatias/imunologia , Dermatopatias/metabolismo , Dermatopatias/patologia , Fator de Crescimento Transformador beta/metabolismo
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