Studying the role of chitosan nanoparticle loaded with Leishmania major Secretory and excretory antigens on the number of apoptotic macrophages in parasite sensitive mouse.

Leishmaniasis is a public health problem caused by different types of Leishmania. The induction of apoptosis in infected macrophages is one of the main mechanisms for the escape of the parasite from the immune system. Preventing apoptosis and enhancing the ability to kill macrophages can be effective in treating or controlling leishmaniasis.  In this study, the effect of chitosan nanoparticle loaded with Leishmania major secretory and excretory antigens on the number of apoptotic macrophages on exposure to Leishmania was investigated.   Secretory and excretory proteins were isolated from the supernatant of the Leishmania major. The protein molecular weight range and its concentration were determined by SDS-PAGE and Bradford method. Proteins were coupled with chitosan nanoparticles. The coupled proteins and nanoparticles were confirmed by FTIR. The nanoparticle toxicity on macrophages was determined by MTT assay. Rats were treated intraperitoneally with protein, nanoparticle, protein coupled with nanoparticles on days 0, 10, and 21. After 28 days, the macrophages were isolated and their apoptosis percentage in presence and absence of parasites was investigated by flow cytometry.   The maximum production of Leishmania major secretory and excretory proteins was 72 hours after parasite culture. Proteins were in the range of 35-110 kDa. At a concentration of 250 μg/ml, the highest percentage of nanoparticles was 76%. The results of the MTT confirmed non-toxicity of all chitosan concentrations coupled with Leishmania major secretory and excretory proteins. The results of apoptotic measurements by Annexin showed that apoptosis in macrophages treated with chitosan and chitosan coupled with Leishmania major secretory and excretory proteins was significantly (p<0.05) less than untreated macrophages.   Chitosan coupled with Leishmania major secretory and excretory proteins can increase the ability of infected macrophages to remove parasites by reducing apoptosis.