[Frontiers in Bioscience E5, 662-675, January 1, 2013]

Cell apoptosis induced by hookworm antigens: a strategy of immunomodulation

Pedro Henrique Gazzinelli-Guimarães1,2, Elaine Maria de Souza-Fagundes3, Guilherme Grossi Lopes Cancado1,2,4, Virgillio Gandra Martins1,2, Lucas de Carvalho Dhom-Lemos1,2, Natasha Delaqua Ricci1,2, Jacqueline Araujo Fiuza5, Lilian Lacerda Bueno1, Rodrigo Rodrigues Cambraia de Miranda1, Silvia Guatimosim3, Andrea Gazzinelli2,6, Rodrigo Correa-Oliveira2,5, Daniella Castanheira Bartholomeu1, Ricardo Toshio Fujiwara1,2,5

1Department of Parasitology, Institute of Biological Sciences, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil, 2Instituto Nacional de Ciencia e Tecnologia em Doenças Tropicais (INCT-DT), Salvador, Bahia, Brazil, 3Department of Physiology and Biophysics, Institute of Biological Sciences, Universidade Federal de Minas Gerais, Minas Gerais, Brazil, 4Clinical Hospital, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil, 5Laboratory of Cellular and Molecular Immunology, Rene Rachou Institute, Oswaldo Cruz Foundation, Belo Horizonte, Minas Gerais, Brazil, 6Nurse School, Institute of Biological Sciences,, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brazil


1. Abstract
2. Introduction
3. Materials and Methods
3.1. Jurkat T cell culture
3.2. Parasites and experimental infection
3.3. Isolation of mesenteric lymph nodes cells of hamsters
3.4. Isolation and preparation of hookworm excretory-secretory (ES) products and adult crude extract (HEX)
3.5. Study population
3.6. Cell viability and cell proliferation assay
3.7. DNA labeling and flow cytometry analysis
3.8. Annexin V and Propidium iodide cell staining
3.9. Confocal microscopy
3.10. Apoptotic pathways triggered by hookworm antigens
3.11. Statistical analysis
4. Results
4.1. Hookworm excretory-secretory (ES) products induce apoptosis in Jurkat T cells
4.2. Cell apoptosis induced by hookworm antigens is also observed in a controlled experimental primary infection in hamsters
4.3. Induction of apoptosis by hookworm antigens is confirmed in peripheral lymphocyte population of chronic N. americanus-infected patients
4.4. Hookworm antigens might induce apoptosis by an intrinsic mitochondrial pathway
5. Discussion
6. Acknowledgments
7. References


While several mechanisms of immunoregulation have been demonstrated for hookworm and other neglected tropical infections, the influence of apoptosis in the immunomodulation of hookworm infection is still poorly understood. In this study, we demonstrate the cytotoxic and pro-apoptotic activity of hookworm antigens in Jurkat T cells, mesenteric lymph nodes lymphocytes of healthy and hookworm-infected hamsters and during human natural infection. Our results showed that in vitro stimulation of Jurkat T cells with antigens induces a significant decrease of cell viability leading to a relevant increase of apoptotic cells. Similar results were also observed in experimental conditions, for both healthy and hookworm-infected hamsters` lymphocytes. Flow cytometric analysis demonstrated that hookworm-infected patients presented a significant increase of CD4+, CD8+, and CD19+ lymphocytes in early and/or late apoptosis when compared with non-infected individuals. The downmodulation of TNF receptors, as well as the up-regulation of the pro-apoptotic genes belonging to the BCL-2 and P53 families, suggest that hookworm antigens induced apoptosis by an intrinsic mitochondrial pathway, acting as a sophisticated strategy to safeguard parasite long-term survival in their hosts.