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

Objective: To investigate the leishmanicidal effects of two antioxidants, caffeic acid and quercetin on Leishmania major(L.major) promastigotes in vitro, and their immunomodulatory effects on infected phagocytes derived from susceptible BALB/c mice.Methods: Caffeic 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(i NOS), tumor necrosis factor alpha(TNF-a) and nuclear factor kappa B in the air pouch membrane were therefore evaluated using appropriate methods.Results: Caffeic 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 p65^(RelA) subunit, and persisted by inhibiting TNF-a and i NOS expression and reactive oxygen species generation.Caffeic acid or quercetin restored reactive oxygen species production and TNF-a-induced i NOS activity, and abrogate apoptosis delay of infected phagocytes.Conclusions: The 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.