Effect of Intra-Peritoneal Injection of Chitosan Nanoparticle Loaded with Leishmania major Excretory-Secretory Antigens on Macrophage Function in Exposure to Parasites in BALB/c Mice.

Leishmania major (L.major) is a causative agent of leishmaniasis. The disease has now become a public health problem. Modulation of the immune responses to inhibit the infection or the proliferation of the parasite is an effective way to overcome the leishmaniosis. Leishmania parasite secretory-excretory antigens are responsible for immune system modulation; therefore, they are used in vaccine design and immunotherapy. Nanoparticles have been considered in recent years due to the features that stimulate the immune system. The purpose of the present study was to produce a combination of nanoparticles and secretory-excretory antigens to induce immune responses. L.major secretory-excretory antigens were isolated from supernatant of parasite culture using ultracentrifugation and protein molecular weight and concentration were determined using SDS-PAGE and Bradford methods. The chitosan nanoparticle were loaded with antigens which confirmed by FTIR. MTT test was performed to determine the cytotoxic effect of nanoparticles on macrophages. Five groups of female BALB/c mice were sensitized with nanoparticles and nanoparticles loaded with the protein on days 0.10 and 21. The macrophages were isolated from the mice and infected with L.major for one hour. After 72 hours of incubation, NO production and inhibition of parasite growth were measured in response to L.major. In the present study, the excretory-secretory antigens of the L.major were in the range of 75-110 kDa, and loaded with chitosan nanoparticle by %76 efficacy. The results showed that the excretory-secretory antigens of the L.major parasites significantly increased the production of nitric oxide in the macrophages and the presence of chitosan induced this increase (P<0.01). In addition, the group that received chitosan loaded with the excretory secretory antigens of the L.major showed the least proliferation of the parasites (P<0.05). If chitosan nanoparticles loaded with L.major excretory-secretory antigens can increase the activity of macrophage killing function by increasing nitric oxide production; therefore, they can be nominated as candidates for vaccine development and disease improvement.