Tehran University of Medical Sciences Journal of Arthropod-Borne Diseases 2322-1984 0 0 2026 09 08 1991 1991 Alireza Khorashad Infectious Diseases and Tropical Medicine Research Center, Research Institute of Cellular and Molecular Science in Infectious Diseases, Zahedan University of Medical Sciences, Zahedan, Iran Ehsan Salarkia Leishmaniasis Research Center, Kerman University of Medical Sciences, Kerman, Iran Seyed Mohammad Mousavi Research Center for Hydatid Disease in Iran, Kerman University of Medical Sciences, Kerman, Iran Abbas Adimi Infectious Diseases and Tropical Medicine Research Center, Research Institute of Cellular and Molecular Science in Infectious Diseases, Zahedan University of Medical Sciences, Zahedan, Iran Hadi Mirahmadi Clinical Immunology Research Center, Zahedan University of Medical Sciences, Zahedan, Iran Ahmad Mehravaran Infectious Diseases and Tropical Medicine Research Center, Research Institute of Cellular and Molecular Science in Infectious Diseases, Zahedan University of Medical Sciences, Zahedan, Iran 2026 08 05 2026 08 31 Background: Leishmaniasis remains a major public health concern due to treatment toxicity, prolonged regimens and emerging drug resistance. Green-synthesized metal oxide nanoparticles are promising candidates for alternative an­tileishmanial therapies. This study synthesized copper oxide nanoparticles (CuO NPs) using hydroalcoholic Calotropis procera leaf extract and evaluated their in vitro activity against Leishmania tropica. Methods: CuO NPs were synthesized using C. procera extract as a biological reducing and stabilizing agent and char­acterized by dynamic light scattering, field-emission scanning electron microscopy, energy-dispersive X-ray spectros­copy and Fourier-transform infrared spectroscopy. The antileishmanial activity of C. procera extract (CPE), CuO NPs, meglumine antimoniate (MA) and their combinations was assessed against promastigotes and intracellular amastigotes. Cytotoxicity was evaluated in THP-1-derived macrophages and selectivity indices (SI) were calculated. Apoptosis-related markers and oxidative stress parameters were also assessed. Results: CuO NPs had a mean hydrodynamic diameter of 185.26±19 nm and a zeta potential of −15.93±2.6 mV. The CuO NPs+MA combination showed the strongest activity against amastigotes (IC₅₀= 52.19 µg/mL) and promastigotes (IC₅₀= 281.63 µg/mL), with a CC₅₀ of 603.1 µg/mL and the highest SI (11.56). Treatment increased Bax, caspase-3, caspase-8 and ROS levels while reducing Bcl-2, SOD and CAT. Conclusion: Green-synthesized CuO NPs, particularly in combination with MA, demonstrated promising antileishma­nial activity and selectivity against L. tropica. These findings support further evaluation in vivo. https://jad.tums.ac.ir/index.php/jad/article/view/1991 https://jad.tums.ac.ir/index.php/jad/article/download/1991/740