RESUMO
Background: Periodontitis is a chronic infectious disease, characterized by an exacerbated inflammatory response and a progressive loss of the supporting tissues of the teeth. Porphyromonas gingivalis is a key etiologic agent in periodontitis. Cystatin C is an antimicrobial salivary peptide that inhibits the growth of P. gingivalis. This study aimed to evaluate the antimicrobial activity of this peptide and its effect on cytokine production, nitric oxide (NO) release, reactive oxygen species (ROS) production, and programmed cell death in human macrophages infected with P. gingivalis. Methods: Monocyte-derived macrophages generated from peripheral blood were infected with P. gingivalis (MOI 1:10) and stimulated with cystatin C (2.75 µg/ml) for 24 h. The intracellular localization of P. gingivalis and cystatin C was determined by immunofluorescence and transmission electron microscopy (TEM). The intracellular antimicrobial activity of cystatin C in macrophages was assessed by counting Colony Forming Units (CFU). ELISA assay was performed to assess inflammatory (TNFα, IL-1ß) and anti-inflammatory (IL-10) cytokines. The production of nitrites and ROS was analyzed by Griess reaction and incubation with 2',7'-dichlorodihydrofluorescein diacetate (H2DCFDA), respectively. Programmed cell death was assessed with the TUNEL assay, Annexin-V, and caspase activity was also determined. Results: Our results showed that cystatin C inhibits the extracellular growth of P. gingivalis. In addition, this peptide is internalized in the infected macrophage, decreases the intracellular bacterial load, and reduces the production of inflammatory cytokines and NO. Interestingly, peptide treatment increased ROS production and substantially decreased bacterial-induced macrophage apoptosis. Conclusions: Cystatin C has antimicrobial and immuno-regulatory activity in macrophages infected with P. gingivalis. These findings highlight the importance of understanding the properties of cystatin C for its possible therapeutic use against oral infections such as periodontitis.
Assuntos
Cistatina C , Macrófagos , Óxido Nítrico , Porphyromonas gingivalis , Espécies Reativas de Oxigênio , Porphyromonas gingivalis/imunologia , Humanos , Macrófagos/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Macrófagos/microbiologia , Cistatina C/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Óxido Nítrico/metabolismo , Citocinas/metabolismo , Periodontite/microbiologia , Periodontite/imunologia , Periodontite/tratamento farmacológico , Periodontite/patologia , Apoptose/efeitos dos fármacosRESUMO
Leishmaniases are a group of vector-borne diseases with different clinical manifestations caused by parasites transmitted by sand fly vectors. In Mexico, the sand fly Lutzomyia olmeca olmeca is the only vector proven to transmit the parasite Leishmania mexicana to humans, which causes leishmaniasis. Other vector species with potential medical importance have been obtained, but their geographic distributions and relation to transmission areas have never been assessed. We modeled the ecological niches of nine sand fly species and projected niches to estimate potential distributions by using known occurrences, environmental coverages, and the algorithms GARP and Maxent. All vector species were distributed in areas with known recurrent transmission, except for Lu. diabolica, which appeared to be related only to areas of occasional transmission in northern Mexico. The distribution of Lu. o. olmeca does not overlap with all reported cutaneous leishmaniasis cases, suggesting that Lu. cruciata and Lu. shannoni are likely also involved as primary vectors in those areas. Our study provides useful information of potential risk areas of leishmaniasis transmission in Mexico.
Assuntos
Insetos Vetores/fisiologia , Leishmania/isolamento & purificação , Leishmaniose/epidemiologia , Leishmaniose/transmissão , Psychodidae/fisiologia , Animais , Demografia , Humanos , Insetos Vetores/parasitologia , México/epidemiologia , Psychodidae/parasitologia , Especificidade da EspécieRESUMO
Leishmania mexicana is an intracellular protozoan parasite that infects macrophages and dendritic cells and causes a chronic cutaneous disease. Although many enzymatic activities have been reported in this parasite, the presence of kinases and phosphatases has been poorly studied. These enzymes control the phosphorylation and dephosphorylation of proteins. Specifically, protein tyrosine kinases phosphorylate tyrosine residues and protein tyrosine phosphatases (PTPases) dephosphorylate tyrosine residues. PTPase activities have been reported as pathogenic factors in various infectious microorganisms such as viruses, bacteria, and parasites. Also, it has been shown that the induction of one or more PTPase activities in macrophages represents an important pathogenicity factor in Leishmania. Recently, we reported a membrane-bound PTPase activity in promastigotes of Leishmania major. In the present work, we give evidence that promastigotes of L. mexicana are able to secrete a PTPase into the culture medium. Two antibodies: one monoclonal against the catalytic domains of the human placental PTPase 1B and a polyclonal rabbit anti-recombinant protein Petase7 from Trypanosoma brucei cross-reacted with a 50-kDa molecule. The anti-human PTPase 1B antibody depleted the enzymatic activity present in the conditioned medium. The pattern of sensitivity and resistance to specific PTPase and serine/threonine inhibitors showed that this enzyme is a protein tyrosine phosphatase.
Assuntos
Leishmania mexicana/enzimologia , Proteínas Tirosina Fosfatases/metabolismo , Proteínas de Protozoários/metabolismo , Anticorpos Monoclonais/imunologia , Anticorpos Antiprotozoários/imunologia , Meios de Cultura/química , Proteínas Tirosina Fosfatases/imunologia , Proteínas de Protozoários/imunologia , Fatores de Virulência/imunologia , Fatores de Virulência/metabolismoRESUMO
The surveillance of prevalent Leishmania sand fly vectors is an important issue for epidemiological studies in populated areas where leishmaniasis is endemic. In this study, we collected sand flies from a peri-urban area in the southeast of Mexico. Natural infection with Leishmania (L.) mexicana was studied by PCR using a Leishmania internal transcribed spacer of the ribosomal RNA gene for amplification. Infected Lutzomyia olmeca olmeca, Lu. shannoni and Lu. cruciata sand flies were collected mainly during the high transmission season (November to March), coinciding with the highest sand fly densities. Additionally, positive specimens of Lu. olmeca olmeca were also captured during July and August. The infected sand flies were from primary forest (subperennial forest) and secondary forest (18-25 years old and 10-15 years old respectively). Sand flies collected with Disney and Shannon traps were the ones found to be infected with L. (L.) mexicana. We conclude that the high-risk period in which L. (L.) mexicana is transmitted in the peri-urban area of Chetumal City is from July to March and that transmission is associated with both the subperennial forest and the secondary forest.