Caffeic acid and quercetin exert caspases-independent apoptotic effects on Leishmania major promastigotes, and reactivate the death of infected phagocytes derived from BALB/c mice

Abstract

ObjectiveTo investigate the leishmanicidal effects of two antioxidants, caffeic acid and quercetin on Leishmania major (L. major) promastigotes in vitro, and their immuno-modulatory effects on infected phagocytes derived from susceptible BALB/c mice. MethodsCaffeic acid and quercetin-induced cell death was examined by Pi-Hoechst double staining of L. major promastigotes and MTT assay, in the presence or absence of protease inhibitors in vitro. Caffeic acid or quercetin were administered subcutaneously to BALB/c mice infected with L. major promastigotes through a dorsal air pouch. Nitric oxide and superoxide anion production by phagocytes infiltrating the air pouch and the expression of inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α) and nuclear factor kappa B in the air pouch membrane were therefore evaluated using appropriate methods. ResultsCaffeic acid and quercetin displayed a dose-dependent cytotoxic effect against L. major promastigotes, and induced cell death via caspases-independent pathways. In vivo, L. major promastigotes inoculation into air pouch cavity of BALB/c mice leads to a sequential influx of neutrophils (hours), followed by macrophages (days). Results showed that L. major delayed apoptosis of infected neutrophils and macrophages by the cleavage of the nuclear factor kappa B p65RelA subunit, and persisted by inhibiting TNF-α and iNOS expression and reactive oxygen species generation. Caffeic acid or quercetin restored reactive oxygen species production and TNF-α-induced iNOS activity, and abrogate apoptosis delay of infected phagocytes. ConclusionsThe leishmanicidal effect of caffeic acid and quercetin on promastigotes and amastigotes, as well as reactivation of infected phagocytes apoptosis, suggested a potential therapeutic role against cutaneous leishmaniasis.