RESUMO
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.
Assuntos
Leishmania braziliensis , Leishmania braziliensis/enzimologia , Leishmania braziliensis/genética , Leishmania braziliensis/efeitos dos fármacos , Animais , Macrófagos/parasitologia , Western Blotting , Citometria de Fluxo , Fatores de Virulência , Peptídeo Hidrolases/metabolismo , Fenótipo , NADH NADPH OxirredutasesRESUMO
BACKGROUND: Lutzomyia longipalpis-derived cell line (Lulo) has been suggested as a model for studies of interaction between sandflies and Leishmania. OBJECTIVES: Here, we present data of proteomic and gene expression analyses of Lulo cell related to interactions with Leishmania (Viannia) braziliensis. METHODS: Lulo cell protein extracts were analysed through a combination of two-dimensional gel electrophoresis and mass spectrometry and resulting spots were further investigated in silico to identify proteins using Mascot search and, afterwards, resulting sequences were applied for analysis with VectorBase. RESULTS: Sixty-four spots were identified showing similarities to other proteins registered in the databases and could be classified according to their biological function, such as ion-binding proteins (23%), proteases (14%), cytoskeletal proteins (11%) and interactive membrane proteins (9.5%). Effects of interaction with L. (V.) braziliensis with the expression of three genes (enolase, tubulin and vacuolar transport protein) were observed after an eight-hour timeframe and compared to culture without parasites, and demonstrated the impact of parasite interaction with the expression of the following genes: LLOJ000219 (1.69-fold), LLOJ000326 (1.43-fold) and LLOJ006663 (2.41-fold). CONCLUSIONS: This set of results adds relevant information regarding the usefulness of the Lulo cell line for studies with Leishmania parasites that indicate variations of protein expression.
Assuntos
Leishmania braziliensis , Leishmania , Proteômica , Psychodidae , Animais , Linhagem Celular , Leishmania/genética , Leishmania braziliensis/genética , Psychodidae/parasitologia , TranscriptomaRESUMO
Leishmania (Leishmania) amazonensis has adaptive mechanisms to the host environment that are guided by its proteinases, including cysteine proteinase B (CPB), and primarily its COOH-terminal region (Cyspep). This work aimed to track the fate of Cyspep by surface plasmon resonance (SPR) of promastigotes and amastigotes to gain a greater understanding of the adaptation of this parasite in both hosts. This strategy consisted of antibody immobilization on a COOH1 surface, followed by interaction with parasite proteins and epoxysuccinyl-L-leucylamido(4-guanidino)butane (E-64). Pro-CPB and Cyspep were detected using specific polyclonal antibodies against a recombinant Cyspep in both parasite forms. The parasitic supernatants from amastigotes and promastigotes exhibited higher anti-Cyspep recognition compared with that in the subcellular fractions. As the supernatant of the promastigote cultures exhibited resonance unit values indicative of an effective with to E-64, this result was assumed to be Pro-CPB detection. Finally, after using three sequential SPR assay steps, we propose that amastigotes and promastigotes release Cyspep into the extracellular environment, but only promastigotes release this polypeptide as Pro-CPB.
Assuntos
Adaptação Fisiológica/fisiologia , Cisteína Proteases/metabolismo , Leishmania mexicana/metabolismo , Leishmaniose Cutânea/patologia , Animais , Anticorpos Antiprotozoários/imunologia , Cisteína Proteases/imunologia , Inibidores de Cisteína Proteinase/farmacologia , Imunoglobulina G/imunologia , Leishmania mexicana/crescimento & desenvolvimento , Leishmaniose Cutânea/parasitologia , Leucina/análogos & derivados , Leucina/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Ressonância de Plasmônio de SuperfícieRESUMO
Leishmania (Leishmania) amazonensis is a protozoan that causes infections with a broad spectrum of clinical manifestations. The currently available chemotherapeutic treatments present many problems, such as several adverse side effects and the development of resistant strains. Natural compounds have been investigated as potential antileishmanial agents, and the effects of epoxy-α-lapachone on L. (L.) amazonensis were analyzed in the present study. This compound was able to cause measurable effects on promastigote and amastigote forms of the parasite, affecting plasma membrane organization and leading to death after 3 h of exposure. This compound also had an effect in experimentally infected BALB/c mice, causing reductions in paw lesions 6 weeks after treatment with 0.44 mM epoxy-α-lapachone (mean lesion area, 24.9 ± 2.0 mm(2)), compared to untreated animals (mean lesion area, 30.8 ± 2.6 mm(2)) or animals treated with Glucantime (mean lesion area, 28.3 ± 1.5 mm(2)). In addition, the effects of this compound on the serine proteinase activities of the parasite were evaluated. Serine proteinase-enriched fractions were extracted from both promastigotes and amastigotes and were shown to act on specific serine proteinase substrates and to be sensitive to classic serine proteinase inhibitors (phenylmethylsulfonyl fluoride, aprotinin, and antipain). These fractions were also affected by epoxy-α-lapachone. Furthermore, in silico simulations indicated that epoxy-α-lapachone can bind to oligopeptidase B (OPB) of L. (L.) amazonensis, a serine proteinase, in a manner similar to that of antipain, interacting with an S1 binding site. This evidence suggests that OPB may be a potential target for epoxy-α-lapachone and, as such, may be related to the compound's effects on the parasite.
Assuntos
Antiprotozoários/farmacologia , Leishmania mexicana/efeitos dos fármacos , Leishmania mexicana/enzimologia , Naftoquinonas/farmacologia , Inibidores de Serina Proteinase/farmacologia , Animais , Antipaína/farmacologia , Simulação por Computador , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Cutânea/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Ligação Proteica , Serina Endopeptidases/metabolismoRESUMO
In this work, we analyze the leishmanicidal effects of epoxy-α-lapachone on Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis. Promasigotes and amastigotes (inhabiting human macrophages) from both species were assayed to verify the compound's activity over the distinct morphological stages. The incubation with epoxy-α-lapachone led to a significant decrease in the numbers of promastigotes from both species in the cultures, in a dose-and time-dependent fashion. The survival of amastigotes inhabiting human macrophages was also drastically affected by the compound, as shown by the variations in the endocytic index. Our results indicate that the epoxy-α-lapachone has an antiparasitic effect over Leishmania in both morphological stages and may potentially affect a range of species in two distinct subgenera of this parasite.
Assuntos
Antiprotozoários/farmacologia , Leishmania braziliensis/efeitos dos fármacos , Leishmania mexicana/efeitos dos fármacos , Naftoquinonas/farmacologia , Antiprotozoários/química , Relação Dose-Resposta a Droga , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia , Naftoquinonas/química , Fatores de TempoRESUMO
Leishmaniasis is a disease caused by Leishmania spp., affecting millions of people around the world. For decades, its treatment has been based on pentavalent antimonials, which notoriously cause toxic side effects in patients. In this study, epoxy-α-lapachone incorporated into an oil-in-water-type microemulsion (ELAP-ME) and meglumine antimoniate (MA) were assayed in monotherapy and in combination (ELAP-ME/MA) in BALB/c mice infected with Leishmania (Leishmania) amazonensis. In general, there was a reduction in paw lesion size (up to 37% reduction) and decreases of parasite loads in the footpad (â¼40%) and lymph nodes (â¼31%) of animals treated with ELAP-ME/MA, when compared to the non-treated control groups. Analyses of serum biochemical parameters revealed that the ELAP-ME/MA showed lower renal and hepatic toxicity when compared to MA 2-doses/week monotherapy. These findings indicate that the ELAP-ME/MA combination may be a promising approach for the treatment of cutaneous leishmaniasis.
Assuntos
Antiprotozoários , Leishmania , Leishmaniose Cutânea , Naftoquinonas , Compostos Organometálicos , Humanos , Animais , Camundongos , Antimoniato de Meglumina/uso terapêutico , Leishmaniose Cutânea/tratamento farmacológico , Leishmaniose Cutânea/parasitologia , Meglumina/uso terapêutico , Compostos Organometálicos/uso terapêutico , Camundongos Endogâmicos BALB CRESUMO
Degrons are short peptide sequences that signalize target sites for protein degradation by proteases. Herein, we bring forth the discussion on degrons present in proteins related to the immune system of Mus musculus that are potential targets for cysteine and serine proteases of Leishmania spp. and their possible roles on host immune regulation by parasites. The Merops database was used to identify protease substrates and proteases sequence motifs, while MAST/MEME Suite was applied to find degron motifs in murine cytokines (IFN-y, IL-4, IL-5, IL-13, IL-17) and transcription factors (NF-kappaB, STAT-1, AP-1, CREB, and BACH2). STRING tool was used to construct an interaction network for the immune factors and SWISS-MODEL server to generate three-dimensional models of proteins. In silico assays confirm the occurrence of degrons in the selected immune response factors. Further analyses were conducted only in those with resolved three-dimensional structures. The predicted interaction network of degron-containing M. musculus proteins shows the possibility that the specific activity of parasite proteases could interfere with the trend of Th1/Th2 immune responses. Data suggest that degrons may play a role in the immune responses in leishmaniases as targets for parasite proteases activity, directing the degradation of specific immune-related factors.
RESUMO
Leishmania parasites are able to interfere with host immune responses on many levels, as T cell responses balance, as observed in the murine model of infection. In the present study, we analyzed genes expression in both host and parasite during the progression of infection. Host genes associated to T-lymphocytes responses, MHC classes I and II, as well as parasite enzymes genes, cysteine-proteinases (CP) B and C, were examined in mice along evolution of infection by Leishmania (Leishmania) amazonensis. Murine strains with distinct levels of susceptibility to infection presented different patterns of MHC genes expression: MHC class I genes tend to have higher expression levels in CBA mice, whereas MHC class II genes expression predominates in BALB/c mice. CPB genes expression in the parasites was shown to predominate over CPC in both mice strains tested. Understanding genes expression patterns during infection may lead to new and more efficient treatments for leishmaniasis.
Assuntos
Expressão Gênica , Interações Hospedeiro-Patógeno , Leishmania mexicana/genética , Leishmania mexicana/patogenicidade , Animais , Cisteína Proteases/biossíntese , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Antígenos de Histocompatibilidade/biossíntese , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/parasitologia , Leishmaniose Cutânea/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos CBARESUMO
Leishmaniasis is a vector-borne disease and an important public health issue. Glycosaminoglycan ligands in Leishmania parasites are potential targets for new strategies to control this disease. We report the subcellular distribution of heparin-binding proteins (HBPs) in Leishmania (Viannia) braziliensis and specific biochemical characteristics of L. (V.) braziliensis HBPs. Promastigotes were fractionated, and flagella and membrane samples were applied to HiTrap Heparin affinity chromatography columns. Heparin-bound fractions from flagella and membrane samples were designated HBP Ff and HBP Mf, respectively. Fraction HBP Ff presented a higher concentration of HBPs relative to HBP Mf, and SDS-PAGE analyses showed 2 major protein bands in both fractions (65 and 55 kDa). The 65 kDa band showed gelatinolytic activity and was sensitive to inhibition by 1,10-phenanthroline. The localization of HBPs on the promastigote surfaces was confirmed using surface plasmon resonance (SPR) biosensor analysis by binding the parasites to a heparin-coated sensor chip; that was inhibited in a dose-dependent manner by pre-incubating the parasites with variable concentrations of heparin, thus indicating distinct heparin-binding capacities for the two fractions. In conclusion, protein fractions isolated from either the flagella or membranes of L. (V.) braziliensis promastigotes have characteristics of metallo-proteinases and are able to bind to glycosaminoglycans.
Assuntos
Moléculas de Adesão Celular/metabolismo , Regulação da Expressão Gênica/fisiologia , Leishmania braziliensis/fisiologia , Moléculas de Adesão Celular/genética , Fracionamento Celular , Leishmania braziliensis/ultraestrutura , Peptídeo Hidrolases/metabolismo , Transporte ProteicoRESUMO
Cysteine proteinases are an important virulence factor in Leishmania parasites. In this study we analyzed the cysteine proteinase expression of infective Leishmania (Viannia) braziliensis promastigotes, examining the expression induced by successive in vitro passages in culture. We observed that this parasite presents a decrease in its virulence over BALB/c macrophages, after successive passages in culture, but still they present proteinase activity, being capable of hydrolyzing the substrate pGlu-Phe-Leu-p Nitroanilide at pH 7.0. This proteinase activity also decreases in the course of the successive passages. Additionally, the decrease in the amount of CPB proteins following successive passages of promastigotes was verified by immunoblotting assays, using an anti-CPB antiserum. Real-time PCR assays were performed to assess the relative cpb expression when compared to a housekeeping gene in promastigote cDNA preparations from the first, fourth and seventh passages. Interestingly, the data indicate a relative increase in cpb gene transcripts as the promastigotes were maintained under in vitro culture: 2.2 times higher for fourth and 2.7 times higher for seventh passages when compared to the first passage. Thus, the information gathered here shows that the expression of cysteine proteinases is modified during in vitro cultivation of L. (V.) braziliensis promastigotes.
Assuntos
Cisteína Proteases/biossíntese , Regulação Enzimológica da Expressão Gênica/fisiologia , Leishmania braziliensis/enzimologia , Fatores de Virulência/biossíntese , Animais , Cisteína Proteases/genética , Immunoblotting , Leishmania braziliensis/crescimento & desenvolvimento , Leishmania braziliensis/patogenicidade , Macrófagos Peritoneais/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Inoculações Seriadas , Virulência , Fatores de Virulência/genéticaRESUMO
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.
RESUMO
The murine models of Leishmania infection are well-studied and suitable models for studying this disease, which, despite its incidence of nearly 2 million new cases worldwide per year and its prevalence of 12 million cases, has been a somewhat neglected disease. Data obtained using such models are important for a better understanding of the disease in humans due to similarities in physiology and the advantage provided by the uniform infection profile within each mouse strain. In this review, we focus on studies of experimental murine infection with Leishmania (Leishmania) amazonensis, a species that has been associated with infections exhibiting various clinical features in humans. Mainly, we point out and discuss reports on: the effects of variations of the inoculum (such as strain, site, and size) in the establishment and development of the infection; characteristics of the infection in distinct mouse strains; and, the effects and subversions of the infection on components of the host innate and adaptive immune responses. The results obtained in these studies show that L. (L.) amazonensis infection in mice presents some unique features and immunoregulatory mechanisms, making it an interesting model for obtaining further knowledge of potential drugs targets and immunotherapy in Leishmania infection.
Assuntos
Leishmania/imunologia , Leishmaniose/imunologia , Leishmaniose/parasitologia , Animais , Antiprotozoários/uso terapêutico , Leishmania/classificação , Leishmaniose/tratamento farmacológico , CamundongosRESUMO
The present study demonstrates that the Leishmania (Viannia) braziliensis strain MCAN/BR/1998/R619 is composed of multiple subpopulations with measurable distinctions. Single parasites were separated from a culture of promastigotes in stationary phase by cell sorting and then cultivated as subpopulations. Subsequently, these subpopulations were evaluated for features of in vitro growth, infectivity to murine macrophages and proteinase gene expression. The first evidence of distinct characteristics was observed during the in vitro cultivation of isolated subpopulations, as distinct clusters of patterns were formed among the cultures, indicating the existence of quantifiable fluctuations in metrics. Further, when infecting murine macrophages, the subpopulations induced distinct patterns of production of immune response mediators. While some subpopulations mainly induced the production of IL-1ß, IL-6 and TNF-α, others induced the production of IL-12p70 and nitric oxide. Finally, amastigotes of these subpopulations had higher expression of proteinase genes than promastigotes. Additionally, cysteine proteinase, serine proteinase, metalloproteinase and aspartic proteinases were differentially expressed in promastigote and amastigote forms. These data suggest the existence of distinct profiles for the L. (V.) braziliensis MCAN/BR/1998/R619 strain and subpopulations that could drive the success of parasite adaptation to the environments that they inhabit.
Assuntos
Leishmania braziliensis/crescimento & desenvolvimento , Animais , Humanos , Interleucina-1beta/imunologia , Interleucina-6/imunologia , Leishmania braziliensis/classificação , Leishmania braziliensis/genética , Leishmania braziliensis/isolamento & purificação , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/parasitologia , Macrófagos/imunologia , Macrófagos/parasitologia , Camundongos , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/metabolismo , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Fator de Necrose Tumoral alfa/imunologiaRESUMO
A surface plasmon resonance- (SPR-) based recognition method applying H-2 Ld:Ig/peptides complexes for ex vivo monitoring cellular immune responses during murine infection with Leishmania (Leishmania) amazonensis is described. Lymphocytes from lesion-draining popliteal lymph nodes were captured on a carboxylated sensor chip surface previously functionalized with H-2 Ld:Ig (DimerX) protein bound to synthetic peptides derived from the COOH-terminal region of cysteine proteinase B of L. (L.) amazonensis. In computational analysis, these peptides presented values of kinetic constants favorable to form complexes with H-2 Ld at neutral pH, with a Gibbs free energy ΔG° < 0. The assayed DimerX:peptide complexes presented the property of attaching to distinct T lymphocytes subsets, obtained from experimentally infected BALB/c mice, in each week of infection, thus indicating a temporal variation in specific T lymphocytes populations, each directed to a different COOH-terminal region-derived peptide. The experimental design proposed herein is an innovative approach for cellular immunology studies of a neglected disease, providing a useful tool for the analysis of specific T lymphocytes subsets.
Assuntos
Imunidade Celular , Leishmania/imunologia , Leishmaniose Cutânea/imunologia , Linfócitos T/patologia , Sequência de Aminoácidos , Animais , Cisteína Proteases/química , Cisteína Proteases/imunologia , Modelos Animais de Doenças , Humanos , Leishmania/patogenicidade , Leishmaniose Cutânea/parasitologia , Leishmaniose Cutânea/patologia , Camundongos , Complexos Multiproteicos/imunologia , Complexos Multiproteicos/isolamento & purificação , Peptídeos/imunologia , Peptídeos/isolamento & purificação , Ressonância de Plasmônio de Superfície , Linfócitos T/imunologiaRESUMO
Leishmania (Viannia) braziliensis presents adaptive protease-dependent mechanisms, as cysteine proteinases B (CPB). This study investigates the expression of three cpb gene isoforms and CPB enzymatic activity during the parasite differentiation. Relative expression levels of LbrM.08.0810 gene were assessed, exhibiting a higher quantity of transcripts in the logarithmic promastigotes phase than in the stationary promastigotes phase (>1.5 times). The cbp gene tends to decrease during acid pH shock and increases when the temperature rises (>1.3 times). LbrM.08.0820 and LbrM.08.0830 genes exhibited similar expression profiles to LbrM.08.0810 gene, with lower levels being observed overall. The proteolytic activity exhibits a gradual increase during the parasite's differentiation with low levels in samples of logarithmic promastigotes phase (3.2 ± 0.08 mmol min-1 mg protein-1) to a peak of activity after 72 h of incubation at 32 °C (4.2 ± 0.026 mmol min-1 mg protein-1) followed by a subsequent decrease of 68 % of peak activity levels after 96 h of incubation at 32 °C (2.8 ± 0.37 mmol min-1 mg protein-1). These activities were also measured in the presence of selective inhibitors for cysteine proteinases, such as Z-Phe-Phe-fluoromethyl ketone and trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane, demonstrating their source as cathepsin-like proteinases. To the best of our knowledge, this report presents the first description of a modulation of cathepsin L-like expression during the L. (V.) braziliensis in vitro differentiation induced by acid pH and high temperature.
Assuntos
Catepsinas/biossíntese , Diferenciação Celular/efeitos dos fármacos , Cisteína Proteases/biossíntese , Leishmania braziliensis/enzimologia , Animais , Catepsinas/genética , Catepsinas/metabolismo , Diferenciação Celular/genética , Cisteína Proteases/genética , Cisteína Proteases/metabolismo , Inibidores de Cisteína Proteinase/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Leishmania braziliensis/crescimento & desenvolvimento , Proteólise/efeitos dos fármacos , TemperaturaRESUMO
The use of proteinases as targets to develop novel chemotherapies against Leishmania spp. infections is a very promising strategy. Based on a previous study by Goyal et al. [J Mol Model (2014) 20:2099], we discuss herein the idea that only a combined treatment with distinct proteinase inhibitors would be an effective antileishmanial therapy.
Assuntos
Antiprotozoários/farmacologia , Desenho de Fármacos , Leishmania/efeitos dos fármacos , Terapia de Alvo Molecular , Peptídeo Hidrolases , Inibidores de Proteases/farmacologia , Proteínas de Protozoários/antagonistas & inibidores , Antiprotozoários/química , Antiprotozoários/metabolismo , Leishmania/classificação , Leishmania/enzimologia , Estrutura Molecular , Peptídeo Hidrolases/química , Peptídeo Hidrolases/metabolismo , Inibidores de Proteases/química , Inibidores de Proteases/metabolismo , Conformação Proteica , Proteínas de Protozoários/química , Proteínas de Protozoários/metabolismo , Relação Estrutura-AtividadeRESUMO
BACKGROUND: The genus Leishmania includes protozoan parasites that are able to infect an array of phlebotomine and vertebrate species. Proteases are related to the capacity of these parasites to infect and survive in their hosts and are therefore classified as virulence factors. FINDINGS: By analyzing protease genes annotated in the genomes of four Leishmania spp [Leishmania (Leishmania) infantum, L. (L.) major, L. (L.) mexicana and L. (Viannia) braziliensis], these genes were found on every chromosome of these protozoa. Four protease classes were studied: metallo-, serine, cysteine and aspartic proteases. Metalloprotease genes predominate in the L. (V.) braziliensis genome, while in the other three species studied, cysteine protease genes prevail. Notably, cysteine and serine protease genes were found to be very abundant, as they were found on all chromosomes of the four studied species. In contrast, only three aspartic protease genes could be detected in these four species. Regarding gene conservation, a higher number of conserved alleles was observed for cysteine proteases (42 alleles), followed by metalloproteases (35 alleles) and serine proteases (15 alleles). CONCLUSIONS: The present study highlights substantial differences in the organization of protease genes among L. (L.) infantum, L. (L.) major, L. (L.) mexicana and L. (V.) braziliensis. We observed significant distinctions in many protease features, such as occurrence, quantity and conservation. These data indicate a great diversity of protease genes among Leishmania species, an aspect that may be related to their adaptations to the peculiarities of each microenvironment they inhabit, such as the gut of phlebotomines and the immune cells of vertebrate hosts.
Assuntos
Regulação Enzimológica da Expressão Gênica/fisiologia , Genoma/fisiologia , Leishmania/enzimologia , Leishmania/genética , Peptídeo Hidrolases/metabolismo , Peptídeo Hidrolases/genética , Especificidade da EspécieRESUMO
This review presents and discusses the current status and perspectives of leishmaniasis treatment, with a special focus on the use of proteinase inhibitors. The history of treatment development, the first- and second-choice modern drugs and the advantages and disadvantages of using proteinases inhibitors as leishmanicidal treatments are presented and discussed. The reports gathered herein confirm the potential usefulness of proteinases inhibitors as an alternative or complement to the current leishmaniasis treatments. They also support the hypothesis that a combined treatment with multiple proteinase inhibitors may be efficient against Leishmania infections in vertebrate hosts.
Assuntos
Antiprotozoários/administração & dosagem , Inibidores de Cisteína Proteinase/administração & dosagem , Leishmaniose/tratamento farmacológico , Animais , Antiprotozoários/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Quimioterapia Combinada , HumanosRESUMO
Leishmania parasites cause human tegumentary and visceral infections that are commonly referred to as leishmaniasis. Despite the high incidence and prevalence of cases, leishmaniasis has been a neglected disease because it mainly affects developing countries. The data obtained from the analysis of patients' biological samples and from assays with animal models confirm the involvement of an array of the parasite's components in its survival inside the mammalian host. These components are classified as virulence factors. In this review, we focus on studies that have explored the role of proteinases as virulence factors that promote parasite survival and immune modulation in the mammalian host. Additionally, the direct involvement of proteinases from the host in lesion evolution is analyzed. The gathered data shows that both parasite and host proteinases are involved in the clinical manifestation of leishmaniasis. It is interesting to note that although the majority of the classes of proteinases are present in Leishmania spp., only cysteine-proteinases, metalloproteinases and, to a lesser scale, serine-proteinases have been adequately studied. Members from these classes have been implicated in tissue invasion, survival in macrophages and immune modulation by parasites. This review reinforces the importance of the parasite proteinases, which are interesting candidates for new chemo or immunotherapies, in the clinical manifestations of leishmaniasis.