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

Objective:To investigate the leishmanicidal effects of two antioxidants,caffeic acid and quercetin on Leishmania major (L.major) promasdgotes 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 (iNOS),tumor necrosis factor alpha (TNF-α) 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 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.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.