Treatment of severe autoimmune blistering skin diseases with combination of protein A immunoadsorption and rituximab: a protocol without initial high dose or pulse steroid medication.

BACKGROUND: Skin blistering diseases due to autoantibodies are typically treated with high dose systemic corticosteroids and other conventional immunosuppressants. However, in severe cases, this treatment may not be sufficient to achieve disease control or contraindicated because of comorbidity. METHODS: We describe 15 patients (pts.) with such diseases: 6 pts. with pemphigus vulgaris, 3 pts. with bullous pemphigoid, 3 pts. with mucous membrane pemphigoid (MMP), one being anti-laminin-332-MMP (AL332-MMP), 2 pts. with pemphigus foliaceus and 1 pt. with epidermolysis bullosa acquisita (EBA). Patients were treated with a combination of protein A immunoadsorption (PAIA, 3-21 treatments) and rituximab (3-6 treatments) in addition to low dose conventional immunosuppression. RESULTS: All patients showed rapid clinical improvement starting within the first 4 weeks and decline of circulating autoantibody levels. Complete/partial remission was 88%/12% in pemphigus and 71%/29% in subepidermal blistering diseases. Overall relapse rate was 13% with an average follow-up of 22 months. In the AL332-MMP pt. the PAIA/rituximab treatment was stopped because of an oesophagus cancer considered as the paraneoplastic cause of the skin disease. CONCLUSION: Combined treatment with PAIA and rituximab showed rapid and long-lasting response, thereby allowing substantial reduction of dosage of concomitant immunosuppressive medication. We hereby confirm data from other investigators that PAIA/rituximab treatment is a promising therapeutical modality for pemphigus, pemphigoids and EBA, characterized by a favourable ratio of beneficial efficacy and minimized long-term adverse effects.

Progressive lipo-lymphedema associated with increased activity of dermal fibroblasts in monoclonal gammopathy of undetermined significance: is there a causal relationship?

The pathophysiology of skin diseases associated with monoclonal gammopathies is generally unknown. Our aim was to investigate whether a monoclonal gammopathy could be a causal factor in progressive lymphedema. We describe a 75 year old patient with a rapidly progressive lipo-lymphedema and a monoclonal gammopathy of unknown significance (MGUS) suspected as a key etiological factor. Dermal fibroblasts were cultured from lesional lower leg skin and non-lesional abdominal skin and compared to healthy control fibroblasts. We found 10-fold elevated basic fibroblast growth factor 2 (FGF-2) in the patient's serum and significantly increased basal FGF-2 production of lesional and non-lesional fibroblasts compared to healthy controls. Upon restimulation with patient or healthy control serum, lesional fibroblasts showed significantly increased proliferation rates and FGF-2 production in vitro. Non-lesional abdominal fibroblasts showed an intermediate phenotype between lesional and control fibroblasts. Our findings provide the first evidence that lesional dermal fibroblasts from lipo-lymphedema with plasma cell infiltration show increased proliferation and FGF-2 production and that both local tissue factors and altered FGF-2 serum levels associated with monoclonal gammopathies might contribute to this phenotype. Thus we propose a possible pathophysiologic link between the gammopathy-associated factors and the generation of lymphedema with initial fibrogenesis aggravating pre-existing lipedema.

Selective regional perfusion of the bilateral external carotid arteries with pegylated liposomal doxorubicin and melphalan to treat metastatic malignant melanoma of the scalp.

We present the case of a 79-year-old patient with extensive metastatic malignant melanoma (MM) of the scalp. Cutaneous MM of the head and neck often presents a therapeutic challenge. Radical surgical procedures and conventional chemotherapy are often unfeasible and contraindicated because of the difficult anatomy, the extent of the tumour process, and systemic toxicity. In our patient, selective intra-arterial perfusion with pegylated liposomal doxorubicin (PLD) and melphalan was performed after catheterization of both bilateral external carotid arteries with an arterial port system. PLD 4.5 mg/m(2) and melphalan (1.35 mg/m(2), followed by 2.7 mg/m(2) after reaching tolerance) were given as short-term infusions at two-weekly intervals into the right and left external carotid arteries, respectively. After eight applications with tolerable side-effects, no MM cells were detected; however, infiltrates of lymphocytes and melanophages were seen. This case suggests that intra-arterial chemotherapy may be a useful treatment for metastatic melanoma of the scalp.

Topo-proteomic in situ analysis of psoriatic plaque under efalizumab treatment.

In a pilot study 6 psoriasis patients were treated over 12 weeks with efalizumab targeting the CD11a subunit of LFA-1. The treatment was well tolerated. Five of these patients proved to be responders with an average decrease in psoriasis area and severity index (PASI) from 21.3 +/- 5.4 (day 0) to 3.9 +/- 0.6 (week 12). The nonresponder was subsequently successfully treated with cyclosporin. Skin biopsies were taken before and after efalizumab treatment and subjected to Multi-Epitope Ligand Cartography (MELC) robot microscopy. A MELC library of 46 antibodies including FITC-labeled efalizumab was chosen focusing upon inflammatory epitopes. Quantification of marker expression was performed using a special adaptation to the needs of skin tissue in terms of pixel events normalized to a standardized horizontal skin width of 100 mum. The before-versus-after comparison for the responders revealed at the 'single epitope level' of MELC analysis a significant decrease (p < 0.05) in epidermal thickness (represented by pan-cytokeratin, CD71, CD138), of the expression of common leukocyte antigen (CD45), T-cell markers (CD2, CD4, CD8, CD45R0), CD11a, efalizumab binding site (EfaBS), and CD58. At the 'EfaBS-centered, double colocation level' a corresponding decrease was observed for CD2, CD3, CD4, CD8, CD11a, CD13, CD26, CD44, CD45, CD45R0, CD54, CD62L, HLA-DR, and TIA-1. MELC analysis at the 'multicombinatorial level' revealed predominant combinatorial molecular phenotype (CMP) motifs, which showed an efalizumab treatment-dependent significant decrease. These CMP motifs were defined as toponomic combinations of lead markers for (i) leukocytes in general (CD45), (ii) T cells (CD2, CD3, CD4, CD45R0, CD45RA), (iii) macrophages (CD68), (iv) cell activation (CD13, CD26, HLA-DR), and (v) cell adhesion (CD11a, EfaBS). Thirty-five of the most relevant 50 CMP motifs were directly related to the T-cell type. A descriptive statistical analysis of the nonresponder before treatment showed a below-responder range degree of expression for CD4, CD8, CD44 (H-CAM), CD56, CD62L, HLA-DQ, and also for these epitopes in colocation with EfaBS. In the nonresponder and before treatment we observed an above-responder range degree of expression for CD54 (ICAM-1) as LFA-1 ligand. In conclusion, the topo-proteomic data provide new diversified insights into the pleiotropic cellular dynamics in psoriatic skin lesions under effective efalizumab treatment. Moreover, the data may be relevant to the future development of possible strategies for individual prediction of efalizumab treatment response or nonresponse.

The CD11a binding site of efalizumab in psoriatic skin tissue as analyzed by Multi-Epitope Ligand Cartography robot technology. Introduction of a novel biological drug-binding biochip assay.

Efalizumab (Raptiva) is an immunomodulating recombinant humanized IgG1 monoclonal antibody that binds to CD11a, the alpha-subunit of leukocyte function antigen-1 (LFA-1). By blocking the binding of LFA-1 to ICAM-1, efalizumab inhibits the adhesion of leukocytes to other cell types and interferes with the migration of T lymphocytes to sites of inflammation (including psoriatic skin plaques). Analysis of the response in patients treated with efalizumab to date shows that distinct groups of responders and nonresponders to the drug exist. It would therefore be of great practical value to be able to predict which patients are most likely to respond to treatment, by identifying key parameters in the mechanism of action of efalizumab. Detailed investigation and detection of multiple epitopes in microcompartments of skin tissue has until recently been restricted by the available technology. However, the newly developed technique of Multi-Epitope Ligand Cartography (MELC) robot technology combines proteomics and biomathematical tools to visualize protein networks at the cellular and subcellular levels in situ, and to decipher cell functions. The MELC technique, which is outlined in this paper, was used to help characterize the binding of efalizumab to affected and unaffected psoriatic skin as compared to normal control skin under ex vivomodel conditions. Efalizumab was labeled with fluorescein isothiocyanate and integrated into a MELC library of more than 40 antibodies. These antibodies were selected for their potential to detect epitopes which may be indicative of (a) various cell types, (b) structural components of the extracellular matrix, or (c) the processes of cell proliferation, activation and adhesion. Efalizumab bound to CD11a in affected psoriatic skin by a factor 15x and 32x higher than in unaffected psoriatic skin and normal control skin, respectively. CD11a and the efalizumab binding site were primarily expressed in the extravascular dermis, whereas CD54 (ICAM-1) as its ligand was most prevalent in the dermal vessels. T lymphocytes (for which the markers were CD3, CD8, CD4, and CD45R0) were the major cellular targets of efalizumab. In contrast, NK cells were only a minor target of efalizumab. Our study demonstrated that efalizumab represents a treatment for psoriasis that primarily targets memory CD4+ and CD8+ T cells and has a high specificity for psoriatic disease activity. Moreover, we hereby introduce the novel principle of a biological drug-binding biochip assay being especially useful for the future monitoring of psoriatic skin lesions under efalizumab treatment conditions.

[Interdisciplinary training program for adults with psoriaisis: six months follow-up]. / Interdisziplinäre psoriasisschulung in blockform für erwachsene patienten: ein halbjahres-follow-up.

BACKGROUND: The aim of our pilot study was to evaluate effects of an interdisciplinary training program for adult psoriasis patients after six months follow-up. SUBJECTS AND METHODS: Eleven patients with psoriasis participated in an interdisciplinary training program over a weekend taught by dermatologists, psychologists/psychiatrists, and dieticians. Six months follow-up was performed with a questionnaire. RESULTS: The knowledge acquired improved the cooperation with the treating dermatologist (7 of 11), the patients' ability to cope with their disease (11 of 11), and their ability to improving their health status (8 of 11). Their general well-being was increased (9 of 11) and they could better care for their skin disease because they better understood the need for care (9 of 11), and could better judge the best approach for various levels of disease activity (8 of 11). CONCLUSION: These data show early benefits and suggest such a longer lasting effect of this type of psoriasis training and prevention program. Further studies with larger samples and control parameters will have to examine if these results can be confirmed.

Interferon-gamma-dependent in vitro model for the putative keratin 17 autoimmune loop in psoriasis: exploration of pharmaco- and gene-therapeutic effects.

In 1999, A.S. Gudmundsdottir et al. have envisaged an epitope on keratin 17 (K17) as a putative psoriasis major autoantigen recognized by T cells. In a HaCaT keratinocyte model, we now demonstrate that IFN-gamma and to a less extent also TNF-alpha and TGF-alpha are able to induce K17 protein expression, in contrast to IL-1alpha, IL-1beta, IL-6, IL-8 and IL-18. This supports our hypothesis of an existing proinflammatory cytokine/K17 autoimmune loop as a presumptive positive feedback mechanism driving psoriasis etiopathogenesis. K17 overexpression was now found to also coincide with suppression of keratinocyte proliferation, e.g. induced by NF-kappa B inhibitors (Bay 11-7082 and Bay 11-7085), and thereby correlated hyperapoptosis to be encountered in psoriatic epidermis. Acitretin as an established antipsoriatic drug and the tyrosine kinase inhibitor imatinib decreased, whereas hydrocortisone as well as dexamethasone increased the IFN-gamma-induced K17 overexpression. The latter might be another mechanism explaining the well-known rebound phenomena after abrupt withdrawal of corticosteroids in psoriasis treatment. Finally, we defined a K17-directed and effective antisense oligodesoxynucleotide which may hold promise for future gene-therapeutic approaches in psoriasis.

Prognostic significance of detecting micrometastases by tyrosinase RT/PCR in sentinel lymph node biopsies: lessons from 322 consecutive melanoma patients.

This prospective study was performed to determine the prognostic value of tyrosinase mRNA detection in sentinel lymph nodes (SLN) of melanoma patients. About 847 SLNs from 322 consecutive patients were assessed by histopathology and immunohistochemistry as well as tyrosinase-reverse transcriptase-polymerase chain reaction (RT/PCR) for the presence of micrometastases. The results were correlated with the prognostic parameters employing a multivariate analysis after a median follow-up of 37 months. Histopathological analysis revealed metastases in 34/322 patients (10.6%). Among the 288 patients with histopathologically negative SLN, tyrosinase-mRNA was detected in 39 patients. A relapse of the tumour occurred in 44.1% of the patients with histopathologically positive SLN, in 25.6% with histopathologically negative, but tyrosinase-RT/PCR-positive SLN, and 8.0% with "double-negative" SLN. A multivariate analysis identified tumour thickness, the histopathological SLN status, and the ulceration of the primary tumour as independent prognostic factors. Thus, by assessing tyrosinase mRNA in the SLN of melanoma patients, we identified a subgroup with histopathologically negative, but Tyr-RT-PCR-positive SLN who have a high risk of disease relapse.

Complete remission of a primary cutaneous B-cell lymphoma of the lower leg by first-line monotherapy with the CD20-antibody rituximab.

BACKGROUND: Rituximab is a genetically engineered antibody recognizing the CD20 antigen known to be expressed by more than 95% of B-cell lymphomas. Recently the antibody has been approved for routine administration in primary extracutaneous, treatment-refractory or relapsed low-grade, follicular non-Hodgkin B-cell lymphomas. With regard to the pathogenetically related primary cutaneous lymphomas, the so-called large B-cell lymphoma of the leg represents a distinct, but rare subentity. In an 89-year-old, multimorbid patient who was affected by such a non-resectable CD20+ large B-cell lymphoma limited to the skin of both lower legs, rituximab was used as a first-line monotherapy in order to avoid local or systemic toxicities inevitably linked to conventional treatment modalities, i.e., radio- or chemotherapy. METHODS: Rituximab was administered at a dosage of 375 mg/m(2) i.v. eight times in weekly intervals. As a premedication we used prednisolone 150 mg i.v. as well as loratadine 10 mg p.o. 1 h before each rituximab infusion. RESULTS: The treatment was well tolerated without any adverse reactions, but was accompanied by a mild transient blood eosinophilia. The histologically proven, exophytic, multi-nodular lymphoma showed a substantial regression already at 2 weeks after the onset of the rituximab treatment. At 8 weeks we observed a complete clinical remission which is now stabile for a follow-up period of 6 months without any maintenance therapy. CONCLUSIONS: Our case observation demonstrates that an intensified, i.e. eightfold, rituximab application in weekly intervals may be a highly effective, tumor target cell-specific first-line monotherapy in the management of primary cutaneous large B-cell lymphoma of the leg. Given the rareness of the disease, the result as well as the possible contribution of the prednisolone premedication will have to be evaluated in a future, controlled, multi-centre study.

Arthus reaction to lepirudin, a new recombinant hirudin, and delayed-type hypersensitivity to several heparins and heparinoids, with tolerance to its intravenous administration.

The pathogenesis of allergic reactions to heparin is poorly understood. Clinically, this phenomenon is relevant because of its increasing incidence and the resulting therapeutic challenges due to various cross-reactions between unfractionated and low-molecular weight heparins as well as between heparins and heparinoids. A 44-year-old female patient had developed a delayed-type hypersensitivity to certoparin-sodium. Diagnostic allergy testing revealed various cross-reactions between different heparins as well as an intolerance to heparinoids. After subcutaneous challenge with the recombinant hirudin lepirudin (Refludan) the patient developed a local Arthus reaction at the injection site. In general, recombinant hirudins do not cross-react with high- or low-molecular weight heparins and heparinoids because of a different molecular structure and are therefore an alternative in case of adverse reactions to heparins and heparinoids. Whereas a local Arthus reaction has already been described twice for low-molecular weight heparins, this is to the best of our knowledge the first observation of a superficial leukocytoclastic vasculitis due to s.c. applied lepirudin. Intravenous administration of heparins and heparinoids in case of hypersensitivity to these drugs following topical application risks a generalized eczematous reaction in patients with delayed-type allergy to both groups of substances. In our patient with delayed-type hypersensitivity to heparins and heparinoids and superficial vasculitis due to lepirudin, the intravenous challenge with heparin and a heparinoid was justified as an ultima ratio measure and proved to be the useful therapeutical alternative.

[Disseminated actinic keratoses. Comparison of topical photodynamic therapy with 5-aminolevulinic acid and topical 5% imiquimod cream]. / Ausgedehnte aktinische Keratosen. Vergleich einer topischen photodynamischen Therapie mit 5-Aminolävulinsäure und topischer 5%iger Imiquimod-Creme.

We report on a 75-year-old woman with skin type I suffering for 20 years from multiple actinic keratoses of areas exposed to the sun. The skin of her forearms was treated on one side with imiquimod 5% cream over 4 weeks and on the contralateral side with photodynamic therapy (PDT) using topical 5-aminolevulinic acid (ALA). Both sides showed good clinical response after 3-months follow-up, suggesting that these modalities can be effectively used for treating widespread multiple actinic keratoses.

Perforin granule release from cytotoxic lymphocytes ex vivo is inhibited by ciclosporin but not by methotrexate.

The 70-kD plasma membrane pore-forming protein perforin is a key component of lymphocyte cytotoxicity mediated by lytic granules. It represents a major player in the regulation of various immune reactions like immunoglobulin synthesis, T-cell activation and homeostasis, and in the elimination of virus-infected and tumor cells. Dysregulation of the perforin-granule system, i.e. an increase of perforin-containing lymphocytes, was recently demonstrated in exacerbated psoriasis and generalized drug reactions. In contrast, in patients with exacerbated atopic dermatitis or unsymptomatic rhinitis allergica, a severe perforin depletion in cytotoxic T cells was demonstrated. In addition, these cells displayed a remarkable transport defect of lytic granules, i.e. a perforin hyperreleasability. Thus, the process of perforin-granule release may represent an attractive target for therapeutic immune modulation in various dermatological diseases. Ficoll isolated peripheral blood mononuclear cells (PBMCs) of healthy volunteers were preincubated with different concentrations of ciclosporin or methotrexate (MTX) for 1 h. A newly developed flow cytometry based perforin release assay was used to quantify the velocity of ionomycin/phorbol 12-myristate 13-acetate stimulated perforin-granule release in the presence or absence of pharmacological agents. The immunosuppressant MTX did not influence perforin-granule release. Ciclosporin, in contrast, was found to inhibit perforin-granule release significantly and dose dependently: whereas release from CD8(+) lymphocytes was almost maximal for the untreated control after 60 min (41% of CD8(+) perforin(+) cells at time zero), ciclosporin at 20, 4 and 2 microg/ml elevated the aforementioned parameter up to 73, 65 and 53%, respectively. Our data demonstrate that (i) perforin-granule release can be targeted efficiently by pharmacological agents which can be monitored directly in a newly developed perforin-granule release assay, and (ii) suppression of perforin-granule based cytotoxicity by ciclosporin might contribute to the beneficial therapeutic effects of this drug as an immunomodulating and immunosuppressant target.

UV erythema reducing capacity of mizolastine compared to acetylsalicylic acid or both combined in comparison to indomethacin.

UV light exerts hazardous effects such as induction of skin cancer and premature skin aging. In this study we evaluated an assumptive anti-inflammatory effect of the nonsedative histamine H1-receptor antagonist, mizolastine, on UV-induced acute sunburn reaction. Therefore, a clinical, randomized, double-blind, four-arm, crossover study was conducted in healthy young female volunteers (skin type II) comparing the UV sensitivity under mizolastine, acetyl-salicylic acid (ASA), indomethacin or a mizolastine/ASA combination. Moreover, HaCaT keratinocytes were incubated with mizolastine under various UV treatment modalities in vitro to study its effect on the release of inflammatory cytokines, i.e. interleukin (IL)-1 alpha, IL-6 and tumor necrosis factor alpha (TNF-alpha). All three drugs were effective in suppressing the UVB-, UVA- and combined UVA/UVB-erythema. However, the strongest effects were observed using the combined treatment with both 250 mg ASA and 10 mg mizolastine. An inhibitory effect in vitro of 10 nM mizolastine upon UV-induced cytokine release from HaCaT keratinocytes was observed for IL-1 alpha at 24 h after 10 J/cm2 UVA1, for IL-6 at 48 h after 10 J/cm2 UVA1 and 30 mJ/cm2 UVB, and also for TNF-alpha at 4 h after 10 J/cm2 UVA, 10 J/cm2 UVA1 and 30 mJ/cm2 UVB, respectively. The combination of mizolastine and ASA can be strongly recommended as a protective measure against UV erythema development with a lower unwanted side effect profile than that of the hitherto treatment modality, i.e. indomethacin.

Dithranol and dimethylfumarate suppress the interferon-gamma-induced up-regulation of cytokeratin 17 as a putative psoriasis autoantigen in vitro.

In psoriasis an etiopathogenetic vicious circle has been hypothesized in which disease manifestation is triggered by skin-specific autoantigen structures. Autoreactive T cells are supposed to mediate inflammation and hyperproliferation in the epidermopapillary compartment, positively feeding back the expression and accessibility of decisive antigen structures. Recently an epitope within cytokeratin 17 (K17) has been described as such a putative psoriasis autoantigen, which is moreover known to be up-regulated under the influence of proinflammatory interferon-gamma (IFN-gamma), which is abundantly detected in psoriatic plaques. The present study proposes an in vitro model for this presumptive IFN-gamma/K17 autoimmune loop, i.e. the incubation of hyperproliferative human HaCaT keratinocytes with 25 U IFN-gamma/ml for 72 h. This treatment led to a significant up-regulation of K17 protein expression (> or =300%) measured by flow immunocytometry as compared to the untreated control (100%, p < or = 0.05). Preincubation with a subcytotoxic and antiproliferative dithranol concentration as low as 0.3 microM for 2 h prior to the IFN-gamma exposure resulted in a K17 expression that was significantly lower than the IFN-gamma-induced K17 expression reference level. The IFN-gamma-induced K17 expression was also significantly lowered by coincubation with a subcytotoxic and nonantiproliferative concentration of 3 microM dimethylfumarate. The data indicate for dithranol and dimethylfumarate that a part of their antipsoriatic mode of action may be related to a direct down-regulation of putative psoriasis autoantigen structures.

[Successful symptomatic tazarotene treatment of juvenile acanthosis nigricans of the familial obesity-associated type in insulin resistance]. / Erfolgreiche symptomatische Tazarotentherapie der juvenilen Acanthosis nigricans vom familiär adipositasassoziierten Typ bei Insulinresistenz.

An 11-year-old boy suffering from morbid obesity since infancy developed at age 9 a progressive brown-black hyperpigmentated and hyperkeratotic eruption in the neck and axillary region with minor involvement of both groins. Based on this clinical picture, and confirmed by histopathology, we diagnosed acanthosis nigricans. Following a thorough endocrinological examination and because the patient's obese mother showed similar skin lesions, the disease was subclassified as a familial obesity-associated type of acanthosis nigricans associated with insulin resistance. In a right-left-comparison the affected skin of one body side was treated with tazarotene 0.05% versus urea 10%, once daily each. A great benefit for the tazarotene-treated over the opposite side could already be seen after three weeks which was also verified by dermatohistopathology. Three months after topical tazarotene treatment had been extended to both sides, the residual lesions were significantly improved. The highly satisfying, good result has been maintained up to now by a continuous topical retinoid treatment over 18 months, usually with an interval application regimen, i.e., 3 x per week.

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. CONCLUSION: 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.