Pars Journal of Medical Sciences

Investigating the Effect of L-arginine on Blastocystis SubType 3 In Vitro

Document Type : Original Article

Authors

Department of Parasitology and Entomology, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran

Abstract

Introduction: Blastocystis is an extracellular and noninvasive unicellular enteric parasite with zoonotic potential.
Intestinal epithelial cells produce nitric oxide (NO), primarily on the apical side, in order to target luminal pathogens. L-arginine acts as a substrate for inducible nitric oxide synthesis (iNOS), which leads to the synthesis of nitric oxide. The current study was designed to assess the effect of L-arginine on Blastocystis ST3 in vitro conditions.
Materials and Methods: The parasite was cultivated in DMEM F-12`s medium and was then identified by polymerase chain reaction (PCR) and the subtype of the parasite was determined which was subtype 3. Then, the methyl thiazolyl tetrazolium (MTT) and flow cytometry methods were used to evaluate the cytotoxicity and probable apoptosis of the prepared drugs /substances on CaCO2 cells. This study investigated the concentrations of L-arginine ( 0.1, 0.2, 0.4, 0.8 and 1.6 mM ) and Metronidazole (1, 2, 4, 8 and 16 μg/ml), and their effect on 24 and 48 hour time points after exposure to the parasite. Then, the final number of parasites was counted after staining with trypan blue by a Neubauer slide and the values of IC50 were calculated for each substance.
Results: It was found that after 48 hours the number of live Blastocystis trophozoites decreases with the increase in L-arginine concentration and At the concentration of 1.6 mM the number of live trophozoites was 33.83 (P< 0.05).
Conclusion: L-arginine, in high concentrations‚ is capable of inhibiting Blastocystis trophozoites In vitro.

Keywords

1.Mirza H, Wu Z, Kidwai F, Tan KS. A metronidazole-resistant isolate of Blastocystis spp. is susceptible to nitric oxide and downregulates intestinal epithelial inducible nitric oxide synthase by a novel parasite survival mechanism. Infect Immun 2011; 79 (12): 5019–26
2.Deng L, Wojciech L, Gascoigne NRJ, Peng G, and Tan KSW. New insights into the interactions between Blastocystis, the gut microbiota, and host immunity. PLoS Pathog 2021; 17 (2): e1009253
3.Stensvold CR, Tan KSW, Clark CG. Blastocystis.Trends Parasitol 2020; 36 (3): 315-316
4.Coyle CM, Varughese J, Weiss LM, Tanowitz HB. Blastocystis: to treat or not to treat. Clin Infect Dis 2012; 54 (1); 105-110
5.Tan KS. New insights on classification, identification, and clinical relevance of Blastocystis spp. Clin Microbiol Rev 2008; 21 (4): 639-665
6.Hernández Ceruelos A, Romero-Quezada LC, Ruvalcaba Ledezma JC, López Contreras L.Therapeutic uses of metronidazole and its side effects: an update. Eur Rev Med Pharmacol Sci 2019; 23 (1):397-401
7.Stadelmann B, Merino MC, Persson L, Svard SG. Arginine Consumption by the Intestinal Parasite Giardia intestinalis Reduces Proliferation of Intestinal Epithelial Cells. PLoS One2012; 7 (9): e45325
8.Badirzadeh A. Arginase activity and Its effects on Pathogenesis of Leishmania Parasites. J Ardabil Univ Med Sci 2019; 19 (2): 126-136 . (Persian)
9.Schairer DO, Chouake JS, Nosanchuk JD, Friedman AJ. The potential of nitric oxide releasing therapies as antimicrobial agents. Virulence 2012; 3 (3): 271–279
10. Stadelmann B, Hanevik k, Andersson M, Bruserud O, and Svard SG. The role of arginine and arginine –metabolizing enzymes during Giardia –host cell interactions in vitro. BMC Microbiol 2013; 13 (1): 1-11
11. Pazoki R, Ghasemi Nejad Raeeni M, Ghaffarifar F, Karimipour Saryazdi A, Bineshian F.The effect of aqueous extract of Artemisia aucheri seed on Acanthomoeba in vitro. J Pharm Rest Int 2019; 31 (6):1-10
12. Rajamanikam A, Hooi HS, Kudva M, Samudi C, and Kumar S. Resistance towards metronidazole in Blastocystis sp.: A pathogenic consequence. PLoS One 2019; 14 (2): e 0212542
13. Helmy MM, Hussein EM, El- moamly AA, Eida AM, Eida OM, Salem AM. Effect of exogenous adminstration of anti-oxidants on Blastocystis hominis In vivo. J Egypt Soc Parasitol 2008; 38 (1): 103-114
14. Costa ISF, Souza GFP, Oliveira MG, Abrahamsohn IA. S-nitrosoglutathione (GSNO) is cytotoxic to intracellular amastigotes and promotes healing of topically treated Leishmania major or Leishmania braziliensis skin lesions.J Antimicrob Chemother 2013;68 (11): 2561-2568
15. Peng BW, Lin J, Lin JY, Jiang MS, Zhang T. Exogenous nitric oxide induces apoptosis in Toxoplasma gondii tachyzoites via a calcium signal transduction pathway. Parasitol Open 2003; 126 (6): 541-550
16. Vespa GN, Cunha FQ, Silva JS. Nitric oxide is involved in control of Trypanosoma cruzi induced parasitemia and directly kills the parasite in vitro. Infect Immun1994; 62 (11): 5177-5182
17. Eida OM, Hussein EM, Eida AM, El-moamly AA, and Salem AM. Evaluation of the nitric oxide activity against Blastocystis hominis in vitro and in vivo. J Egypt Soc Parasitol 2008; 38 (2): 521-536
18. Eckmann L, Laurent F, Langford TD, Hetsko ML, Smith JR, Kagnoff MF, Gillin FD. Nitric oxide production by human intestinal epithelial cells and competition for arginine as potential determinants of host defense against the lumen-dwelling pathogen Giardia lamblia. J Immunol 2000;164 (3): 1478-1487
19. Hadas E, Derda M, and Wandurska-Nowak E. Effect of exogenous nitric oxide in experimental trichinellosis. Parasitol Res 2002; 88: 86-88
20. Abdeltawab MS, Abdel-Shafi IR, Aboulhoda BS, Wanas H, El-Din Sh, Amer SA, Hamed AM. Investigating the effect of the nitric oxide donor L-arginine on albendazole efficacy in Trichinella spiralis induced myositis and myocarditis in mice.
Parasitol United J 2022; 15 (1): 60-70
21. Michiels C, Raes M, Toussaint O, Ramacle J. Importance of Se-glutathione peroxidase, catalase and Cu,Zn-SOD for cell survival against oxidative stress. Redox Biol 1994; 17 (3): 235-248
22. EL-Khazragy N, and Saad GhA. Blastocystis sp. Enhances oxidative stress-induced carcinogenesis in colorectal cancer: In vivo experimental study. Int J Adv Res 2019; 7 (9): 1231-1238