Papel de PI3K na invasão celular e maturação de fagolisossomos de L. (L.) amazonensis

Abstract

Cutaneous leishmaniasis caused by Leishmania (L.) amazonensis is largely distributed throughout the country and can manifest as severe and disseminated lesions in man and other animals. During microorganism invasion phagocytes are activated and one of the signs of this activation is the phosphorylation of specific substrates by the enzyme phosphatidylinositol-3- kinase (PI3K), which is in its turn activated by a variety of extracellular signals, generating lipid products that will serve as mediator in signal transduction downstream. During parasite invasion of the host cells there is recruitment of cytoskeleton proteins to form a phagosome, followed by fusion with lysosomes to generate the phagolysosome, where the parasites will thrive. Modulation of PI3K signaling pathways by parasites are important strategies for the establishment of the infection and intracellular survival of intracellular parasites. In this study we aimed to investigate the maturation of the phagolysosome of L. (L.) amazonensis through the labeling of F-actin and LAMP-1, as well as to determine the role of PI3K on the invasion process by L. (L.) amazonensis. Murine peritoneal macrophages treated or not with wortmannin (an inhibitor of the PI3K pathway) were infected with L. (L.) amazonensis promastigotes. To validate the role of this pathway by this species of Leishmania, macrophages were transfected with GFP-PLC-δ-PH plasmid, which has homology with PI(4,5)P2, a component of the PI3K pathway, after which they were infected. Our results demonstrated that, during the maturation of L. (L.) amazonensis phagosomes, there was initially F-actin recruitment to the nascent phagosome/endosome, which disappeared after fusion with lysosomes, as shown by the recruitment of LAMP-1, a lysosomal marker. The processes of invasion and lysosomal fusion of L. (L.) amazonensis demonstrated a clear dependency of PI3K activity, as it was demonstrated by the inhibition of these processes by the PI3K inhibitor wortmannin.