Specific antibodies induce apoptosis in Trypanosoma cruzi epimastigotes.
The susceptibility of Trypanosoma cruzi epimastigotes to lysis by normal or immune sera in a complement-dependent reaction has been reported. Mouse immune sera depleted complement-induced damage in epimastigotes characterized by morphological changes and death. The purpose of this work was to study the mechanism of death in epimastigotes exposed to decomplemented mouse immune serum. Epimastigotes were maintained in RPMI medium. Immune sera were prepared in mice by immunization with whole crude epimastigote extracts. Viable epimastigotes were incubated with decomplemented normal or immune sera at 37 degrees C. By electron microscopy, agglutinated parasites showed characteristic patterns of membrane fusion between two or more parasites; this fusion also produced interdigitation of the subpellicular microtubules. Apoptosis was determined by flow cytometry using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and annexin V assays. Nuclear features were examined by 4'-,6-diamidino-2'-phenylindole diHCI cytochemistry that demonstrated apoptotic nuclear condensation. Caspase activity was also measured. TUNEL results showed that parasites incubated with decomplemented immune sera took up 26% of specific fluorescence as compared to 1.3% in parasites incubated with decomplemented normal sera. The Annexin-V-Fluos staining kit revealed that epimastigotes incubated with decomplemented immune sera exposed phosphatidylserine on the external leaflet of the plasma membrane. The incubation of parasites with immune sera showed caspase 3 activity. We conclude that specific antibodies are able to induce agglutination and apoptosis in epimastigotes, although the pathway is not elucidated.
Departamento de Microbiología y Parasitología, Facultad de Medicina, Universidad Nacional Autónoma de México, Av Universidad 3000, Col Copilco Universidad, 04510 Mexico Distrito Federal, Mexico. presas@servidor.unam.mx
Ana María
AM
Fernández-Presas
Patricia
P
Tato
Ingeborg
I
Becker
Sandra
S
Solano
Natalia
N
Copitin
Natalia
N
Kopitin
Miriam
M
Berzunza
Kaethe
K
Willms
Joselin
J
Hernández
José Luis
JL
Molinari
D000818:Animals; D017304:Annexin A5 / Q000032:analysis; D000913:Antibodies, Protozoan / Q000276:immunology*; D017209:Apoptosis; D053148:Caspase 3 / Q000032:analysis; D003165:Complement System Proteins / Q000276:immunology*; D005260:Female; D020287:In Situ Nick-End Labeling; D051379:Mice; D050296:Microbial Viability; D008854:Microscopy, Electron; D014349:Trypanosoma cruzi / Q000737:chemistry / Q000276:immunology* / Q000648:ultrastructure
D017304:Annexin A5; D000913:Antibodies, Protozoan; D003165:Complement System Proteins; D053148:Caspase 3