Toll like receptor 2 and CC chemokine receptor 5 cluster in the lipid raft enhances the susceptibility of Leishmania donovani infection in macrophages.

In experimental visceral leishmaniasis the causative obligate protozoan parasite, L. donovani invades and multiplies inside of macrophages, one of the sentries of the mammalian immune system. The initial host-parasite interaction between the Leishmania promastigote and the macrophage takes place at the plasma membrane interface. To trace any possible interaction between Toll-like receptor 2 (TLR2) and CC chemokine receptor 5 (CCR5) during early Leishmania-macrophage interactions, it was observed that the expression of both TLR2 and CCR5 were significantly increased, along with their recruitment to the lipid raft. TLR2 silencing attenuates CCR5 expression and restricts L. donovani infection, indicating a regulatory role of TLR2 and CCR5 during infection. Silencing of CCR5 and TLR2 markedly reduced the number of intracellular parasites in macrophages by host protective cytokine responses, while raft disruption using β-MCD affected TLR2/CCR5 cross-talk and resulted in a significant reduction in parasite invasion. In vivo RNA interference of TLR2 and CCR5 using shRNA plasmids rendered protection in Leishmania donovani-infected mice. Thus, this study for the first time demonstrates the importance of TLR2/CCR5 crosstalk as a significant determinant of Leishmania donovani entry in host macrophages.

Corpúsculos e mediadores lipídicos na resposta inflamatória a saliva de Lutzomiya longipalpis durante a infecção com Leishmania chagasi / Corpuscles and lipid mediators in the inflammatory response to the saliva of Lutzomyia longipalpis during infection with Leishmania chagasi

Lutzomyia longipalpis é o principal transmissor da Leishmania chagasi, agente etiológico da leishmaniose visceral (LV), a forma mais letal da doença. A saliva de L. longipalpis possui moléculas capazes de modular as repostas hemostática, inflamatória e imunológica do hospedeiro, favorecendo o estabelecimento da infecção. Entretanto, poucos trabalhos têm abordado o papel da saliva de L. longipalpis na indução de mediadores lipídicos e sua implicação nos momentos iniciais da infecção por Leishmania. Neste trabalho, investigamos o papel do sonicado de glândula salivar (SGS) de Lutzomyia longipalpis na indução da formação de corpúsculos lipídicos (CL) e produção de mediadores lipídicos durante os estágios iniciais da infecção por Leishmania chagasi. O SGS foi capaz de induzir in vitro a formação de CLs e a produção de PGE2 em macrófagos murinos. Durante a infecção de camundongos C57BL/6 por L. chagasi, a presença de saliva de L. longipalpis aumentou a formação de CLs em macrófagos após 3 horas de estímulo, o que esteve correlacionado com o aumento do influxo de células para o sítio inflamatório, principalmente neutrófilos. A infecção de L. chagasi em presença do SGS de L. longipalpis induziu a produção de LTB4 e PGE2 em leucócitos peritoneais ex vivo. Após 3 horas, macrófagos e neutrófilos infectados, bem como a interação entre essas células foram observadas. O percentual de macrófagos infectados e o número de parasitas por macrófago esteve reduzido durante a infecção em presença de SGS. O conjunto dos nossos resultados indica que a saliva do vetor desempenha um papel importante na modulação da inflamação durante os estágios iniciais da infecção por L. chagas i e abre novas perspectivas para o entendimento da imunopatogênese da leishmaniose visceral.

Monoclonal antibody affinity purification of a 78 kDa membrane protein of Leishmania donovani of Indian origin and its role in host-parasite interaction.

Monoclonal antibodies were raised against pathogenic promastigotes of Leishmania donovani of Indian origin. Among these, one was used for immuno-affinity purification of a 78 kDa membrane protein present in both the amastigote and promastigote forms of the parasite. Results of immunoblot experiments with the anti-78 kDa antibody revealed that the protein was present only in parasites belonging to the L. donovani complex. The expression of the protein was observed to be the same during different phases of growth of the promastigotes. Therefore, the 78 kDa protein is neither stage-specific nor differentially regulated. Surface iodination and subcellular fractionation of the promastigotes indicated that the protein was localized on the cell surface. The 78 kDa protein was found to inhibit the binding of promastigotes to macrophages significantly, suggesting that it may play a role in the process of infection. Thus, here we report the purification of a surface protein of L. donovani of Indian origin, which may play an important role in the process of infection.

Status of glutathione in lymphoid tissues of golden hamster during Leishmania donovani infection.

During Leishmania donovani infection of golden hamsters, changes in the glutathione levels of lymphoid tissues were observed. While in liver and thymus there was significant decrease in the levels of reduced and oxidised glutathione, that in spleen, bone marrow cells, peritoneal exudate cells and lymph nodes increased, indicating a role for host glutathione in establishing the infection.

Lipid peroxidation in hepatic microsomal membranes isolated from mice in health and in experimental leishmaniasis.

Normal hepatic microsomal membranes when exposed in vitro to different free radicals, cause membrane damage by lipid peroxidation which could be monitored by the analysis of malonaldehyde formation and measurement of membrane microviscosity. Lipid peroxidation in vivo, when examined in hepatic microsomal membranes in experimental Leishmaniasis, reveals a direct relationship between membrane microviscosity and the extent of lipid peroxidation. Scavengers of free radicals and peroxides such as superoxide dismutase (SOD) for O2.-, mannitol for (OH.) and catalase for H2O2 in modest amounts were used for preventing the membrane damage caused by lipid peroxidation.

Gamma interferon production by lymphocytes from children infected with L. chagasi

The present study was performed to evaluate the ability of lymphocytes from 18 children living in an endemic area of visceral leishmaniasis (VL) to produce gamma-interferon. These children had no previous history of VL and were considered to be infected with Leishmania chagasi based on leishamnial seroconversion. The gamma IFN levels were determined by radioimmunoassay on supernatants of lymphocyte cultures (3 x 10**6/ml). stimulate with PHA (final dilution 1:10) and Leishamnia chagasi antigen (10µg/ml). The gamma-IFN production by lymphocytes from seroconverting children stimulated with PHA (178 ñ 151 U/ml) and Leishmania chagasi (47 ñ 77 U/ml) was significantly higher than that observed in visceral leishmaniasis. For clinical follow-up, these 18 seroconverting children were divided into three groups: asymptomatic infection (N=4); self-healing subclinical illness (N=9), and sublinical infection progressing to VL (N=5). Gamma IFN levels inchildren with either asymptomatic or subclinical infection (65 ñ 85 U/ml were significantly higher (P < 0.003) than those observed in children progressing to VL (9 ñ 6 U/ml). The data demonstrate that there is an association between gamma IFN levels and the clinical course of Leishmania infection