The Effect of Artemether-induced Apoptosis in Promastigotes of Leishmania major (MRHO/IR/75/ER) under In-vitro Conditions

Authors
P. Ebrahimisadr 1
F. Ghaffarifar 2
Z. M. Hassan 3
N. Beheshti 1
1 M.Sc, Department of Parasitilogy, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
2 Associated Professor, Department of Parasitilogy, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
3 Professor, Department of Immunology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran
Abstract
Objective: Leishmaniasis is one of the significant causes of morbidity and mortality in several countries. It is an important problem in endemic areas such as Iran. The goal in treatment of leishmaniasis is to reduce the disease period and leave no evidence of any remaining scars or lesions. A derivative of artemisinin is artemether. Scientists believe that the strong action of artemether against parasites is due to the presence of an endoperoxide bridge. Due to problems in the treatment of Leishmania major, in this research we have studied the effect of artemether on Leishmania major under in vitro conditions. Methods: Parasites were cultured in NNN and RPMI, after which artemether at concentrations of 5, 10, 25, 50 and 100 μg/ml were used for the promastigote assay. Apoptosis was detected by flow cytometry and DNA ladder assay. Results: The inhibitory concentration (IC50) of artemether was determined to be 25 μg/ml. The percentage of apoptotic promastigotes at 25 μg/ml of artemether was 42.28. The results of DNA fragmentation show that exposure of Leishmania major promastigote cells to 25 μg/ml of artemether lead to DNA fragmentation. Conclusion: We have proven the effect of artemether on apoptosis of Leishmania major by flow cytometry and the DNA ladder assay.

Keywords

 [1]     Akhondzadeh S. Encyclopedia of Iranian Medicinal Plants.Tehran: Arjomand Press 2000; p: 199-209. (Persian)
[2]     Desjeux P. Leishmaniasis: current situation and new perspectives. Comp Immunol Microbiol Infect Dis 2004; 27(5): 305-18.
[3]     Sundar S, Agrawal NK, Sinha PR, Horwith GS, Murray HW. Short-course, low-dose amphotericin B lipid complex therapy for visceral leishmaniasis unresponsive to antimony. Ann Intern Med 1997; 127(2): 133-7.
[4]     Urbina JA. Lipid biosynthesis pathways as chemotherapeutic targets in kinetoplastid parasites. Parasitology 1997; 114 Suppl: S91-9.
[5]     Soussi N, Milon G, Colle JH, Mougneau E, Glaichenhaus N, Goossens PL. Listeria monocytogenes as a short-lived delivery system for the induction of type 1 cell-mediated immunity against the p36/LACK antigen of Leishmania major. Infect Immun 2000; 68(3): 1498-506.
[6]     Mauël J. Vaccination against Leishmania infections. Curr Drug Targets Immune Endocr Metabol Disord 2002; 2(3): 201-26.
[7]     Dietze R, Fagundes SM, Brito EF, Milan EP, Feitosa TF, Suassuna FA, Fonschiffrey G, Ksionski G, Dember J. Treatment of kala-azar in Brazil with Amphocil (amphotericin B cholesterol dispersion) for 5 days. Trans R Soc Trop Med Hyg 1995; 89(3): 309-11.
[8]     Das N, Mahato SB, Naskar K, Ghosh DK, Basu MK. Targeting of urea stibamine encapsulated in liposomes to reticulo-endothelial system for the treatment of experimental leishmaniasis. Biochem Med Metab Biol 1990; 43(2): 133-9.
[9]     Abahusein A, Larbi EB, al-Khawajah A, al-Gindan Y, Jain S. Evaluation of topical ketoconazole in cutaneous leishmaniasis. East Afr Med J 1992; 69(1): 14-7.
[10]  Méndez S, Gurunathan S, Kamhawi S, Belkaid Y, Moga MA, Skeiky YA, Campos-Neto A, Reed S, Seder RA, Sacks D. The potency and durability of DNA- and protein-based vaccines against Leishmania major evaluated using low-dose, intradermal challenge. J Immunol 2001; 166(8): 5122-8.
[11]  Cowan MM. Plant products as antimicrobial agents. Clin Microbiol Rev 1999; 12(4): 564-82.
[12]  Kayser O, Kiderlen AF, Laatsch H, Croft SL: In vitro leishmanicidal activity of monomeric and dimeric naphthoquinones. Acta Trop 2000; 77(3):307-314.
[13]  Berman JD, Ksionski G, Chapman WL, Waits VB, Hanson WL. Activity of amphotericin B cholesterol dispersion (Amphocil) in experimental visceral leishmaniasis. Antimicrob Agents Chemother 1992; 36(9): 1978-80.
[14]  Muñoz V, Moretti C, Sauvain M, Caron C, Porzel A, Massiot G, Richard B, Le Men-Olivier L. Isolation of bis-indole alkaloids with antileishmanial and antibacterial activities from Peschiera van heurkii (syn. Tabernaemontana van heurkii). Planta Med 1994; 60(5): 455-9.
[15]  van Agtmael MA, Eggelte TA, van Boxtel CJ. Artemisinin drugs in the treatment of malaria: from medicinal herb to registered medication. Trends Pharmacol Sci 1999; 20(5): 199-205.
[16]  Falade C, Makanga M, Premji Z, Ortmann CE, Stockmeyer M, de Palacios PI. Efficacy and safety of artemether-lumefantrine (Coartem) tablets (six-dose regimen) in African infants and children with acute, uncomplicated falciparum malaria. Trans R Soc Trop Med Hyg 2005; 99(6): 459-67.
[17]  Chen T, Li M, Zhang R, Wang H. Dihydro-artemisinin induces apoptosis and sensitizes human ovarian cancer cells to carboplatin therapy. J Cell Mol Med 2009; 13(7): 1358-70.
[18]  Chen H, Sun B, Pan S, Jiang H, Sun X. Dihydroartemisinin inhibits growth of pancreatic cancer cells in vitro and in vivo. Anticancer Drugs 2009; 20(2): 131-40.
[19]  Golenser J, Waknine JH, Krugliak M, Hunt NH, Grau GE. Current perspectives on the mechanism of action of artemisinins. Int J Parasitol 2006; 36(14): 1427-41.
[20]  Hamann A, Brust D, Osiewacz HD. Apoptosis pathways in fungal growth, development and ageing. Trends Microbiol 2008; 16(6): 276-83.
[21]  Keiser J, Rinaldi L, Veneziano V, Mezzino L, Tanner M, Utzinger J, Cringoli G. Efficacy and safety of artemether against a natural Fasciola hepatica infection in sheep. Parasitol Res 2008; 103(3): 517-22.
[22]  Keiser J, Vargas M. Effect of artemether, artesunate, OZ78, praziquantel, and tri-bendimidine alone or in combination chemotherapy on the tegument of Clonorchis sinensis. Parasitol Int 2010; 59(3): 472-6.
[23]  Xiao S, Utzinger J, Chollet J, Endriss Y, N'Goran EK, Tanner M. Effect of artemether against Schistosoma haematobium in experimentally infected hamsters. Int J Parasitol 2000; 30(9): 1001-6.
[24]  Xiao SH, Booth M, Tanner M. The prophylactic effects of artemether against Schistosoma japonicum infections. Parasitol Today 2000; 16(3): 122-6.
[25]  Shuhua X, Chollet J, Weiss NA, Bergquist RN, Tanner M. Preventive effect of artemether in experimental animals infected with Schistosoma mansoni. Parasitol Int 2000; 49(1): 19-24.
[26]  Shalaby HA, El Namaky AH, Kamel RO. In vitro effect of artemether and triclabendazole on adult Fasciola gigantic. Vet Parasitol 2009; 160(1-2): 76-82.
[27]  Olliaro PL, Haynes RK, Meunier B, Yuthavong Y. Possible modes of action of the artemisinin-type compounds. Trends Parasitol 2001; 17(3): 122-6.
[28]  Shaha C. Apoptosis in Leishmania species & its relevance to disease pathogenesis. Indian J Med Res 2006; 123(3): 233-44.
Pathobiology Reserach
Volume 15, Issue 2
June 2012
Pages 1-10