Background: Leishmania major, an intracellular protozoan, is the primary cause of cutaneous leishmaniasis (CL). Although polyvalent antimony compounds are commonly used to treat CL, treatment failure is frequent. Immunometabolic reprogramming of macrophages—the main host cells for L. major—toward the M1 phenotype may inhibit parasite growth. Pyrimethamine, an antimalarial antibiotic, inhibits dihydrofolate reductase (DHFR), affecting both host and parasite metabolism. Since pyrimethamine interferes with the folate pathway and phagocytosis in L. major, this study investigated its effect on key immunometabolites in infected macrophages.
Materials and Methods: RAW264.7 macrophages were divided into six groups: Sham, L. major-infected, LPS-stimulated, pyrimethamine-treated (L. major+pyrimethamine, LPS+pyrimethamine), and pyrimethamine alone (5 μg/mL for 24 hours). Supernatants and cells were collected to measure lactate, nitric oxide (NO), reactive oxygen species (ROS), infection rate and index, as well as cytokine concentrations.
Results: Pyrimethamine significantly increased intracellular ROS and NO levels in L. major-infected macrophages (P≤0.05), correlating with a significant reduction in infection rate and index (P≤0.05). TNF-α production was also significantly elevated in pyrimethamine-treated groups. However, no significant change in lactate concentration was observed.
Conclusion: Pyrimethamine enhances the immunometabolic response of macrophages against L. major by increasing ROS, NO, and TNF-α levels. These changes contribute to a reduced infection rate and lower macrophage infection index, suggesting pyrimethamine’s potential as an immunomodulatory agent in leishmaniasis treatment.