Leishmania braziliensis: differential lipid profiles of promastigotes and amastigotes.

GIMÉNEZ, GUADALUPE; LAMMEL, ESTELA MARÍA; BOTT, EMANUEL; BELAUNZARÁN, MARIA LAURA; DURANTE DE ISOLA, ELVIRA LUISA

Río de Janeiro

Congreso; XVIII International Congress for Tropical Medicine and Malaria and XLVIII Congress of the Brazilian Society for Tropical Medicine; 2012

Sociedad Brasilera de Medicina Tropical

Leishmania spp. is an intracellular parasite that belongs to the Trypanosomatidae family. This protozoa is transmitted by phlebotomine sandflies as metacyclic promastigotes that infect macrophages in the vertebrate host where they multiply as amastigotes. In Argentina, the coexistence of L. braziliensis, L. guyanensis y L. amazonensis has been detected, being the major causative agents of human leishmaniasis. The limitation of initial infection depends on the activation of the innate immunity. Since there is a growing body of evidence pointing to an important modulatory role of microorganisms’ lipids in this process, herein we focused on the study of the lipid profiles of L. braziliensis promastigotes (PRO) and intact or autolysing amastigotes (AMA). Material and Methods Parasites. L. braziliensis, clon MHOM/BR/75M2904, PRO and AMA were axenically cultured in LIT media at 24ºC or 37ºC, respectively. Autolysing AMA were prepared by freezing-thawing and subsequent overnight incubation at 37ºC. Lipid profile analysis. Lipids were extracted from PRO and intact or autolysing AMA according to Bligh & Dyer, analyzed by thin layer chromatography and identified by comparison with authentic standards after charring. Densitometric analysis was performed with Gel-Pro® Analyzer 4.0 software. Nitric oxide (NO) detection. NO production was determined in J774 cell line stimulated with L. braziliensis lipids by the Griess assay. Results Quantitative differences in the phospholipid (PL) fraction of L. braziliensis PRO and AMA were determined. The latter presented higher amounts of lysophosphatidylcholine and phosphatidylethanolamine with respect to PRO (~ 4 and 1.2 fold higher, respectively). Besides, PRO presented higher levels of sphingomyelin and phosphatidic acid with respect to AMA (~ 2-fold). Considering that autolytic processes can generate inflammatory factors, we also performed the lipid analysis of autolysing AMA. Results showed a significant parasite PL degradation with an important increase in free fatty acids (FFA), thus suggesting the action of Phospholipase A, enzymatic activity previously described in this protozoa by our group. On the other hand, we determined that total lipids from each parasite stage were able to induce nitric oxide, a soluble factor implicated in parasite growth inhibition. Conclusion The quantitative differences observed between the lipid profiles of L. braziliensis PRO and AMA, could be related to the biological function of each stage. In autolysing AMA, PL degradation generated FFA, second messenger that might participate in several signaling pathways related to inflammatory processes. Supported by FONCYT/CONICET.