RESUMEN
Leishmaniasis is a widespread tropical infection; cutaneous leishmaniasis (CL) is the most common form of this disease known to cause significant morbidity. Trace metals, including selenium, zinc, iron, and copper, are required for the activity of several enzymes involved in immune system responses. The aim of this research was to measure the serum levels of selenium (Se), zinc (Zn), copper (Cu), iron (Fe), and Zn/Cu ratio in patients with CL. In this case-control study, 80 patients with CL and 80 healthy volunteers (not exposed to CL) from a CL endemic region in southwest Iran agreed to participate. Both clinical and parasitological verifications were made to include each subject as a CL-positive case. A questionnaire was completed for each participant which included the following criteria: age (year), height (cm), weight (kg), body mass index (kg/m2), and duration of disease (day). The biochemical assays were performed according to the standard protocols, and the values of Zn, Cu, Se, and Fe were expressed in micrograms per deciliter (µg/dl). All results were expressed as mean ± standard deviation (SD), and the statistical significance level was defined to be less than 0.05 (P < 0.05). There were no statistically significant differences in terms of mean values of age, body weight, height, and body mass index between CL patients and the control group (P > 0.05). The mean ± SD concentrations of Zn, Fe, and Se in the control group were found to be 118.87 ± 6.35 µg/dl, 123.00 ± 8.40 µg/dl, and 11.26 ± 1.88 µg/dl, respectively. These trace elements (TEs) were statistically lower (P < 0.001) in patients with CL (case group) with values of 83.05 ± 7.32 µg/dl for Zn, 86.51 ± 10.09 µg/dl for Fe, and 3.83 ± 1.20 µg/dl for Se. We have also observed that serum levels of Cu in CL-positive group were significantly higher than in the controls (P < 0.001). Furthermore, CL patients had significantly lower Zn/Cu ratio than controls (0.63 ± 0.05 µg/dl vs. 1.11 ± 0.10; P < 0.001). The alternation in serum levels of TEs may be a part of the defense strategy of the organism. Based on these results, it can be suggested that serum levels of these TEs can be a useful marker to estimate the prognosis of CL infection.
Asunto(s)
Cobre , Hierro , Leishmaniasis Cutánea , Selenio , Oligoelementos , Zinc , Estudios de Casos y Controles , Cobre/análisis , Cobre/sangre , Humanos , Irán/epidemiología , Hierro/análisis , Hierro/sangre , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/epidemiología , Selenio/sangre , Zinc/análisis , Zinc/sangreRESUMEN
Background and Objectives: The live non-pathogenic Leishmania tarantolae has recently provided a promising approach as an effective vaccine candidate against experimental leishmaniasis (ILL). Here, we evaluated the immunoprotective potential of the live Iranian Lizard Leishmania mixed with CpG adjuvant against L. major infection in BALB/c mice. Methods: Four groups of female BALB/c mice were included in the study. The first and second groups received PBS and CpG, respectively. The immunized groups received 2 × 105 ILL promastigotes and the CpG-mixed ILL (ILL+CpG). Injections were performed subcutaneously in the right footpad. Three weeks later, all mice were challenged with 2 × 105 metacyclic promastigotes of Leishmania majorEGFP ; inoculation was done in the left footpad. The measurement of footpad swelling and in vivo fluorescent imaging were used to evaluate disease progress during infection course. Eight weeks after challenge, all mice were sacrificed and the cytokines levels (IFN-γ, IL-4, and IL-10) and sera antibodies concentrations (IgG2a and IgG1) using ELISA assay, nitric oxide production using Griess assay, and arginase activity in cultured splenocytes, were measured. In addition, direct fluorescent microscopy analysis and qPCR assay were used to quantify the splenic parasite burden. Result: The results showed that mice immunized with ILL+CpG were protected against the development of the dermal lesion. Moreover, they showed a significant reduction in the parasite load, in comparison to the control groups. The observed protection was associated with higher production of IFN-γ, as well as a reduction in IL-4 level. Additionally, the results demonstrated that arginase activity was decreased in ILL+CpG group compared to other groups. Conclusion: Immunization using ILL+CpG induces a protective immunity; indicating that ILL with an appropriate adjuvant would be a suitable choice for vaccination against leishmaniasis.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Leishmania major/inmunología , Vacunas contra la Leishmaniasis/farmacología , Leishmaniasis Cutánea/prevención & control , Lagartos/parasitología , Oligodesoxirribonucleótidos/farmacología , Piel/efectos de los fármacos , Vacunas Vivas no Atenuadas/farmacología , Animales , Anticuerpos Antiprotozoarios/sangre , Arginasa/metabolismo , Células Cultivadas , Citocinas/sangre , Modelos Animales de Enfermedad , Femenino , Inmunización , Inmunogenicidad Vacunal , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Macrófagos/parasitología , Ratones Endogámicos BALB C , Carga de Parásitos , Piel/inmunología , Piel/parasitología , Bazo/efectos de los fármacos , Bazo/inmunología , Bazo/metabolismo , Bazo/parasitología , Vacunas Vivas no Atenuadas/inmunologíaRESUMEN
BACKGROUND: Cutaneous leishmaniasis lacks effective and well-tolerated treatments. The current therapies mainly rely on antimonial drugs that are inadequate because of their poor efficacy. Traditional medicine offers a complementary alternative for the treatment of various diseases. Additionally, several plants have shown success as anti-leishmanial agents. Therefore, we sought to evaluate the in vitro and in vivo activity of MEBA against Leishmania mexicana. MATERIALS AND METHODS: Methanolic extract of B. aptera was obtained by macetration, after we determined in vitro anti-leishmanial activity of MEBA by MTT assay and the induced apoptosis in promastigotes by flow cytometry. To analyze the in vivo anti-leishmanial activity, we used infected mice that were treated and not treated with MEBA and we determined the levels of cytokines using ELISA. The phytochemical properties were determined by CG-MS and DPPH assay. RESULTS: We determined of LC50 of 0.408 mg/mL of MEBA for in vitro anti-leishmanial activity. MEBA induced apoptosis in promastigotes (15.3% ± 0.86). Treated mice exhibited smaller lesions and contained significantly fewer parasites than did untreated mice; in addition, we found that IFN-γ and TNF-α increased in the sera of MEBA-treated mice. GC-MS analysis showed that podophyllotoxin was the most abundant compound. Evaluation of the activity by DPPH assay demonstrated an SC50 of 11.72 µg/mL. CONCLUSION: Based on the above data, it was concluded that MEBA is a good candidate in the search for new anti-leishmanial agents.
Asunto(s)
Bursera/química , Leishmania mexicana , Leishmaniasis Cutánea/tratamiento farmacológico , Fitoterapia , Extractos Vegetales/uso terapéutico , Animales , Femenino , Interferón gamma/sangre , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/parasitología , Medicina Tradicional , Ratones Endogámicos BALB C , Corteza de la Planta , Extractos Vegetales/farmacología , Podofilotoxina/análisis , Podofilotoxina/farmacología , Podofilotoxina/uso terapéutico , Factor de Necrosis Tumoral alfa/sangreRESUMEN
Cutaneous leishmaniasis triggers a varied immune response depending on parasite and host factors, which in turn can be influenced by nutrients. The resistance to the infection is associated with the Th1 type of cytokine production. The Th1 type can be reduced as a consequence of zinc deficiency, which may increase the risk for chronicity of the infection. Using in vitro and ex vivo models, we studied the influence of zinc supplementation on the immune response in patients with cutaneous leishmaniasis treated with antimony and the data were also compared to those of matched controls. Twenty-nine patients with cutaneous leishmaniasis (n=14 in zinc-supplemented group [45mg/day] and n=15 in placebo group) were treated by intramuscular injections of antimony for 20 days and took supplements for 60 days. Immunoglobulins in plasma and cell proliferation, IFN-γ production and CD markers of isolated peripheral blood mononuclear cells (PBMC) were measured. It was found that the cellular immune response of the patients maintained its activity as assessed by the ability of the PBMC to proliferate and produce IFN-γ in response to concanavalin A. Moreover, there was no difference in these variables between the zinc-supplemented and placebo groups after 60 days. The addition of zinc sulphate in vitro to PBMC reduced the IFN-γ production in the placebo group only. It is concluded that the cellular immune response of the cutaneous leishmaniasis patients remained active during treatment by antimony when compared to that of controls. It was not possible to document an additional effect of zinc supplementation for 60 days on the immune response.
Asunto(s)
Suplementos Dietéticos , Inmunidad , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/inmunología , Zinc/uso terapéutico , Estudios de Casos y Controles , Proliferación Celular/efectos de los fármacos , Concanavalina A/farmacología , Humanos , Inmunidad/efectos de los fármacos , Inmunoglobulinas/sangre , Interferón gamma/metabolismo , Leishmaniasis Cutánea/tratamiento farmacológico , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/metabolismo , Recuento de Linfocitos , Zinc/farmacologíaRESUMEN
Evolutionary and closer structural relationships are demonstrated by phylogenetic analysis, peptide prediction and molecular modelling between Solanum tuberosum apyrase, Schistosoma mansoni SmATPase 2 and Leishmania braziliensis NDPase. Specific protein domains are suggested to be potentially involved in the immune response, and also seem to be conserved during host and parasite co-evolution. Significant IgG antibody reactivity was observed in sera from patients with American cutaneous leishmaniasis (ACL) and schistosomiasis using potato apyrase as antigen in ELISA. S. mansoni adult worm or egg, L. braziliensis promastigote (Lb) and Trypanosoma cruzi epimastigote (EPI) have ATP diphosphohydrolases, and antigenic preparations of them were evaluated. In ACL patients, IgG seropositivity was about 43% and 90% for Lb and potato apyrase, respectively, while IgM was lower (40%) or IgG (100%) seropositivity for both soluble egg (SEA) and adult worm (SWAP) antigens was higher than that found for potato apyrase (IgM=10%; IgG=39%). In Chagas disease, IgG seropositivity for EPI and potato apyrase was 97% and 17%, respectively, while the IgM was low (3%) for both antigens. The study of the conserved domains from both parasite proteins and potato apyrase could lead to the development of new drug targets or molecular markers.
Asunto(s)
Apirasa/inmunología , Secuencia Conservada/inmunología , Mapeo Epitopo , Parásitos/enzimología , Parásitos/inmunología , Solanum tuberosum/enzimología , Secuencia de Aminoácidos , Animales , Anticuerpos Antiprotozoarios/inmunología , Apirasa/química , Enfermedad de Chagas/sangre , Enfermedad de Chagas/inmunología , Humanos , Leishmania braziliensis/enzimología , Leishmania braziliensis/genética , Leishmania braziliensis/inmunología , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/inmunología , Datos de Secuencia Molecular , Parásitos/genética , Filogenia , Estructura Terciaria de Proteína , Schistosoma mansoni/enzimología , Schistosoma mansoni/genética , Schistosoma mansoni/inmunología , Esquistosomiasis/sangre , Esquistosomiasis/inmunología , Alineación de SecuenciaRESUMEN
We evaluated the relationship between the trace elements copper, zinc and selenium and the response to the Montenegro skin test (MST) in 172 volunteers vaccinated against American cutaneous leishmaniasis. The MST diameter was categorized as negative and in quartiles of positive response, constituting five groups. Trace element serum levels were analyzed by coupled plasma atomic emission spectrometry and hydride generation atomic absorption spectrometry, with study subjects classified into two groups depending on low or high levels of trace elements observed. MST-positive subjects had an MST diameter (mean+/-SD) of 10.35+/-4.64mm, with copper, selenium and zinc serum levels of 1433.7+/-665.7microg/l, 88.6+/-39microg/l and 999.2+/-366microg/l, respectively. The MST diameter was significantly different in the selenium groups only. The selenium levels also differed with the quartiles of the MST diameters of individuals testing positive (P<0.05). Our findings suggest that trace elements, particularly selenium, should be measured in future vaccine trials so that vaccine immunogenicity and response can be assessed and compared between different studies.
Asunto(s)
Cobre/sangre , Vacunas contra la Leishmaniasis/inmunología , Leishmaniasis Cutánea/prevención & control , Selenio/sangre , Zinc/sangre , Adolescente , Adulto , Brasil , Niño , Preescolar , Femenino , Humanos , Leishmaniasis Cutánea/sangre , Masculino , Persona de Mediana Edad , Pruebas Cutáneas/métodos , Estadística como AsuntoRESUMEN
Plasma essential trace elements, selenium, copper, zinc, and iron concentrations and the levels of immunoregulatory cytokines, interleukin-1beta (IL-1beta), interleukin-2 receptor (IL-2r), IL-6, IL-8, and tumor necrosis factor-alpha (TNF-alpha) were evaluated in patients with cutaneous leishmaniasis (CL) to investigate a possible role of these cytokines on selenium, zinc, copper, and iron homeostasis in CL patients. Plasma albumin levels were measured as an index of nutritional status. Plasma selenium, zinc, and iron concentrations, and IL-2r levels were significantly lower, and copper concentrations and IL-1beta, IL-8, IL-6 and TNF-alpha levels were significantly higher in patients with CL than those of healthy controls. There was no significant difference in plasma albumin levels between two groups. There were positive important correlations between plasma selenium and IL-2r, copper and IL-6, and copper and IL-1beta, and negative correlations between selenium and IL-8, iron and TNF-alpha, and zinc and IL-1beta contents in patients with CL. Our results showed that plasma trace element contents change in patients with CL. These changes may not be a result of a specific deficiency from dietary inadequacies or imbalances, but, probably, a result of a part of the defense strategies of an organism that is regulated by immunoregulatory cytokines.
Asunto(s)
Cobre/sangre , Citocinas/sangre , Citocinas/inmunología , Hierro/sangre , Leishmaniasis Cutánea/sangre , Selenio/sangre , Zinc/sangre , Adolescente , Adulto , Animales , Índice de Masa Corporal , Niño , Femenino , Humanos , Interleucinas/sangre , Masculino , Factor de Necrosis Tumoral alfa/análisis , PavosRESUMEN
BACKGROUND: Safe, effective and inexpensive vaccines may be the most practical tool for control of any form of leishmaniasis. Leishmaniasis produces a state of pre-immunition which is the underlying mechanism for prolonged immunity to re-infection. Low doses of parasites has been shown to be able to induce protection in mice. It is not known, however, how immune sera from a susceptible host immunised with Leishmania-derived antigens when taken in by the sandfly affects the development and the subsequent transmission of the parasite to naive hosts. OBJECTIVE: To monitor the course of disease in BALB/c mice following challenge using L. major infected P. duboscqi which had previously fed on immunised mice. METHODS: BALB/c mice were immunised adequately with Leishmania major-derived antigens namely, crude whole parasite (WPA), recombinant 63 kilodalton glycoprotein (rgp63), lipophosphoglycan (LPG) and a cocktail composed of rgp63 plus LPG antigens. Laboratory reared Phlebotomus duboscqi sandflies, the natural vector for L. major were later allowed to feed on immunised animals, interrupted and allowed to continue feeding on infected animals for an equal amount of time until they became fully engorged. The sandflies were maintained on apples as a carbohydrate source in an insectary maintained at a temperature of 25 degrees C and 80% relative humidity. On the seventh day these sandflies were used to infect naive BALB/c mice and the course of infection followed for a period of at least three months. RESULTS: Mice infected using sandflies which had previously fed on WPA or rgp63-immunized mice showed disease exacerbation as the infection progressed, whereas those infected using sandflies which had previously fed on LPG-immunised mice had the least lesion sizes compared to control mice infected using sandflies which had fed on saline immunised mice (p < 0.05). CONCLUSIONS: Results from this study indicate that the course of L. major infection in BALB/c mice was dependent on the infective dose of parasites transmitted by the sandflies. Results from this study suggests that sub-infective doses of the parasite from sandflies previously fed on animals immunised with Leishmania-derived antigens needs to be evaluated for their potential in vaccine development against Leishmania infections.
Asunto(s)
Antígenos de Protozoos/inmunología , Modelos Animales de Enfermedad , Glicoesfingolípidos/inmunología , Leishmania major/inmunología , Leishmaniasis Cutánea/prevención & control , Leishmaniasis Cutánea/transmisión , Metaloendopeptidasas/inmunología , Vacunas Antiprotozoos/inmunología , Animales , Progresión de la Enfermedad , Evaluación Preclínica de Medicamentos , Insectos Vectores/parasitología , Leishmaniasis Cutánea/sangre , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/parasitología , Ratones , Ratones Endogámicos BALB C , Phlebotomus/parasitología , Factores de TiempoRESUMEN
BACKGROUND: New strategies for control of leishmaniasis is needed as chemotherapy using antimonial drugs is prolonged, expensive, associated with side effects and relapses. Vector control has limitations and a vaccine which may be the best approach is not available. OBJECTIVES: To assess the level of inhibition of promastigote development and gut morphology in infected Phlebotomus duboscqi sandflies fed on different groups of BALB/c mice immunised with rgp63, lipophosglycan (LPG) or their cocktail and whole parasite antigens prepared from L. major culture-derived promastigotes. METHODS: BALB/c mice were immunised adequately with Leishmania major-derived antigens namely, crude whole parasite (WPA), recombinant 63 kilodalton glycoprotein (rgp63), LPG and a cocktail composed of rgp63 plus LPG antigens. Laboratory reared Phlebotomus duboscqi sandflies, the natural vector for L. major were later allowed to feed on immunised animals, interrupted and allowed to continue feeding on infected animals for an equal amount of time until they became fully engorged. The sandflies were maintained on apples as a carbohydrate source in an insectary maintained at a temperature of 25 degrees C and 80% relative humidity. Some of the sandflies were dissected on days 2, 4 and 6 after feeding and observed using the light and the transmission electron microscopy for any changes in their gut morphology. The remaining sandflies were all dissected on the sixth day post-feeding and examined for procyclics, nectomonads, haptomonads and metacyclic promastigote forms of Leishmania. RESULTS: Sandflies which had previously fed on WPA, LPG plus rgp63 cocktail and LPG-immunised mice showed the lowest infection rates compared to control sandflies fed on saline immunised mice (p < 0.05). A significant number of procyclic promastigotes, the first developmental form of the parasite in culture as well as in the sandfly was observed in sandflies which fed on LPG-immunised mice (p < 0.05). The dominant parasite form in sandflies which fed on rgp63 or LPG-immunised mice was the nectomonad form but very few of the infective metacyclic forms (p < 0.05). Control sandflies fed on saline immunised or infected mice alone displayed a normal pattern of parasite development up to the metacyclic stage. Studies showed that two possible mechanisms through which immune sera from immunised mice may cause inhibition of parasite development is by exflagellation of nectomonad forms and degeneration of the sandfly midgut epithelium as revealed by light and electron microscopy studies respectively. CONCLUSIONS: This study has shown that immune-mediated transmission blocking may be applied to Leishmania infections. Based on observation of the procyclic promastigotes, the dominance of the nectomonad forms, low infectivity rates in sandflies fed on LPG-immunised mice, we concluded that LPG stands out to be a promising transmission blocking vaccine candidate in leishmaniasis.