Perforin hyperreleasability and depletion in cytotoxic T cells from patients with exacerbated atopic dermatitis and asymptomatic rhinoconjunctivitis allergica.
BACKGROUND
As a plasma membrane pore-forming protein, perforin is essential for T-cell cytotoxicity mediated by lytic granules. Recent studies on the immune system of perforin knockout mice demonstrated striking similarities to the immunopathology of atopic diseases.


OBJECTIVE
We sought to investigate the perforin system of atopic patients.


METHODS
Monoclonal antibodies were used to characterize perforin-positive PBMCs of patients with exacerbated atopic dermatitis (AD) and asymptomatic rhinoconjunctivitis allergica (RCA) by means of immunoflow cytometry. In addition, a perforin release assay was developed to quantify the velocity of ionomycin and phorbol 12-myristate 13-acetate-induced secretion of lytic granules.


RESULTS
In atopic patients significantly fewer lymphocytes contained perforin-positive lytic granules compared with those of healthy control subjects (patients with AD: 14% +/- 5%, n = 13, P <.0001; patients with RCA: 24% +/- 5%, n = 9, P <.01; healthy control subjects: 33% +/- 11%, n = 13). Of all CD8(hi+) cytotoxic T lymphocytes (CTLs), only 18% +/- 9% and 17% +/- 12% were perforin-positive in patients with AD and RCA, respectively, compared with 44% +/- 13% in control subjects (P <.0001). In addition, perforin-positive CD8(hi+) CTLs of atopic patients released their perforin twice as fast and more completely than control CTLs. This means that 50% of initially perforin-positive CD8(hi+) CTLs from patients with AD and RCA released their perforin completely within 32 +/- 16 and 36 +/- 19 minutes, respectively, and an over 85% release was reached within 113 +/- 41 and 118 +/- 60 minutes, respectively. In CTLs of healthy control subjects, however, it took 64 +/- 40 minutes to achieve a 50% release of lytic granules, and an 85% depletion was not reached in 60% of healthy control subjects, even after 180 minutes.


CONCLUSIONS
The perforin hyperreleasability explains, at least in part, the decreased percentage of perforin-positive CD8(hi+) CTLs in atopic patients. These distortions in the system of lytic granules of atopic patients may contribute to the functional defects observed in T-cell cytotoxicity in vivo and in vitro in patients with AD and RCA.
Ambach
A
A
Department of Dermatology and Venereology, Otto-von-Guericke-University, Magdeburg, Germany.
Bonnekoh
B
B
Gollnick
H
H
D000911:Antibodies, Monoclonal / Q000276:immunology; D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm; D016516:CD4-CD8 Ratio; D003233:Conjunctivitis, Allergic / Q000276:immunology* / Q000473:pathology; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000378:metabolism; D003602:Cytotoxicity, Immunologic; D003876:Dermatitis, Atopic / Q000276:immunology* / Q000473:pathology; D005434:Flow Cytometry; D006801:Humans; D006969:Hypersensitivity, Immediate / Q000276:immunology / Q000473:pathology; D007073:Immunoglobulin E / Q000097:blood; D015759:Ionomycin / Q000494:pharmacology; D007694:Killer Cells, Natural / Q000187:drug effects / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000097:blood / Q000378:metabolism*; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D012221:Rhinitis, Allergic, Perennial / Q000276:immunology* / Q000473:pathology; D006255:Rhinitis, Allergic, Seasonal / Q000276:immunology* / Q000473:pathology; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D018417:Th1 Cells / Q000276:immunology; D018418:Th2 Cells / Q000276:immunology; D013997:Time Factors
D000911:Antibodies, Monoclonal; D000945:Antigens, Differentiation, T-Lymphocyte; D000951:Antigens, Neoplasm; C063419:CTAGE1 protein, human; D008562:Membrane Glycoproteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin; D007073:Immunoglobulin E; D015759:Ionomycin; D013755:Tetradecanoylphorbol Acetate