title: In vitro activity of phospholipase A2 and of peptides from Crotalus durissus terrificus venom against amastigote and promastigote forms of Leishmania (L.) infantum chagasi.
abstract: American visceral leishmaniasis is caused by the intracellular parasite Leishmania (L.) infantum chagasi, and transmitted by the sand fly Lutzomyia longipalpis. Since treatment is based on classical chemotherapeutics with significant side effects, the search for new drugs remains the greatest global challenge. Thus, this in vitro study aimed to evaluate the leishmanicidal effect of Crotalus durissus terrificus venom fractions on promastigote and amastigote forms of Leishmania (L.) infantum chagasi.
  
  
  
  Phospholipase A2 (PLA2) and a pool of peptide fraction (<3 kDa) were purified from Crotalus venom. Furthermore, promastigotes and peritoneal macrophages of mice infected by amastigotes were exposed to serial dilutions of the PLA2 and peptides at intervals varying between 1.5625 μg/mL and 200 μg/mL. Both showed activity against promastigotes that varied according to the tested concentration and the time of incubation (24, 48 and 72 h).
  
  
  
  MTT assay for promastigotes showed IC50 of 52.07 μg/mL for PLA2 and 16.98 μg/mL for the peptide fraction of the venom. The cytotoxicity assessment in peritoneal macrophages showed IC50 of 98 μg/mL and 16.98 μg/mL for PLA2 and peptide by MTT assay, respectively. In peritoneal macrophages infected by Leishmania (L.) infantum chagasi amastigotes, the PLA2 stimulated growth of parasites, and at higher doses reduced growth by 23 %. The peptide fraction prevented 43 % of the intracellular parasite growth at a dose of 16.98 μg/mL, demonstrating the toxicity of this dose to macrophages. Both fractions stimulated H2O2 production by macrophages but only PLA2 was able to stimulate NO production.
  
  
  
  We have demonstrated the in vitro leishmanicidal activity of the PLA2 and peptide fraction of Crotalus venom. The results encourage further studies to describe the metabolic pathways involved in cell death, as well as the prospecting of molecules with antiparasitic activity present in the peptide fraction of Crotalus durissus terrificus venom.
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Gustavo A C
authors.initials: GA
authors.lastname: Barros
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Andreia V
authors.initials: AV
authors.lastname: Pereira
authors.affiliation: Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Luciana C
authors.initials: LC
authors.lastname: Barros
authors.affiliation: Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Airton
authors.initials: A
authors.lastname: Lourenço
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Sueli A
authors.initials: SA
authors.lastname: Calvi
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil ; Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Lucilene D
authors.initials: LD
authors.lastname: Santos
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil ; Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil.
authors.forename: Benedito
authors.initials: B
authors.lastname: Barraviera
authors.affiliation: Department of Tropical Diseases, Botucatu Medical School, São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil ; Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP Brazil ; CEVAP/UNESP, Caixa Postal 577, Fazenda Experimental Lageado, Rua José Barbosa de Barros, 1780, 18610-307 Botucatu, SP Brasil.
authors.forename: Rui Seabra
authors.initials: RS
authors.lastname: Ferreira
keywords: Crotalus dutissus terrificus; Leishmanicidal activity; PLA2; Peptides; Venom