- Introduction
- Materials and Methods
- Cells and cell culture.
- Parasite preparation and labeling with carboxyfluorescein diacetate succinimidyl ester.
- Isolation and culture of peripheral blood mononuclear cells.
- Isolation of CD8+ T cells.
- Chromium release assay to measure cytolytic activity.
- RNA isolation and real-time polymerase chain reaction to measure elimination of parasites from cell cultures.
- Inhibition studies.
- Fluorescent microscopy.
- Statistical analysis.
- Results
- Donor HLA types.
- Antigen expanded sensitized CD8+ T cells reduce the C. parvum load in infected cell cultures.
- Lysis of C. parvum-infected cells by gp15 expanded CD8+ T cells from a sensitized donor.
- Antiparasitic effect depends on antigen presentation through HLA-A and HLA-B.
- Antiparasitic effect involves release of cytolytic granules from CD8+ T cells.
- Antiparasitic effect depends on cytotoxic granules.
- Discussion
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Human CD8+ T Cells Clear Cryptosporidium parvum from Infected Intestinal Epithelial Cells
Intracellular protozoans of the genus Cryptosporidium are a major cause of diarrheal illness worldwide, especially in immunocompromised individuals. CD4+ T cells and interferon-gamma are key factors in the control of cryptosporidiosis in human and murine models. Previous studies led us to hypothesize that CD8+ T cells contribute to clearance of intestinal epithelial Cryptosporidium infection in humans. We report here that antigen expanded sensitized CD8+ T cells reduce the parasite load in infected intestinal epithelial cell cultures and lyse infected intestinal epithelial cells. These effects are most likely mediated by the release of cytotoxic granules. Elimination of parasites seems to require antigen presentation through both human leukocyte antigen (HLA)-A and HLA-B. These data suggest that cytotoxic CD8+ T cells play a role in clearing Cryptosporidium from the intestine, a previously unrecognized feature of the human immune response against this parasite.
Author Notes
Authors' addresses: Birte Pantenburg, Alejandro Castellanos-Gonzalez, and A. Clinton White Jr., Division of Infectious Diseases, Department of Internal Medicine, University of Texas Medical Branch, Galveston, TX, E-mails: birte.pantenburg@upch.pe, alcastel@utmb.edu, and acwhite@utmb.edu. Sara M. Dann and Dorothy E. Lewis, Division of Infectious Diseases, Department of Internal Medicine, and Department of Microbiology and Immunology, University of Texas Medical Branch, Galveston, TX, E-mails: smdann@utmb.edu and dolewis@utmb.edu. Rhykka L. Connelly, Center for Electromechanics, University of Texas, Austin, TX, E-mail: r.connelly@cem.utexas.edu. Honorine D. Ward, Division of Geographic Medicine and Infectious Diseases, Tufts Medical Center, Boston, MA, E-mail: hward@tuftsmedicalcenter.org.