Increased metal allergy in patients with failed metal-on-metal hip arthroplasty and peri-implant T-lymphocytic inflammation.

BACKGROUND: In 16 patients with revised metal-on-metal arthroplasty and peri-implant lymphocytic inflammation, we verified the role of metal hypersensitivity by patch testing (PT) and lymphocyte transformation test (LTT). METHODS: In the 16 patients with lymphocyte dominated periprosthetic inflammation, allergy history was obtained by a questionnaire, specific serum IgE to aeroallergens was measured to assess atopy, PT to standard and metal series was performed and metal sensitivity was further assessed by LTT using blood mononuclear cells. RESULTS: Revision surgery was performed because of pain (8/16), osteolysis (4/16), dislocation (3/16) and loosening of the stem (1/16). Histological examination showed perivascular infiltrates of T lymphocytes, high endothelial venules, fibrin exudation and accumulation of macrophages with drop-like inclusions. Five patients had a history of cutaneous metal allergy and atopy was found in 25% of the patients. In 13/16 patients (81%), systemic metal sensitivity was found based on PT and/or LTT. Patch test reactions were seen in 11/16 patients (69%; partly multiple reactions/patient): 7/16 to Cobalt (Co), 7/16 to Chromium (Cr), 4/16 to Nickel (Ni), and one each to Molybdenum (Mo) and Manganese (Mn). Ten of 16 patients (62%) showed enhanced LTT reactivity to metals: 7/16 to Ni, 7/16 to Co, 5/16 to Cr, 5/16 to Mo and 4/16 to Mn. CONCLUSIONS: The lymphocyte dominated peri-implant inflammation may well reflect an allergic hyper-reactivity in these patients, given the high rate of concomitantly found metal allergy. Despite the overall incidence of metal implant allergy being low, allergic reactions should be included as differential diagnosis in failed metal-on-metal arthroplasty.

Adjuvant interferon alfa-2a treatment in resected primary stage II cutaneous melanoma. Austrian Malignant Melanoma Cooperative Group.

PURPOSE: Patients with primary cutaneous melanoma with a Breslow thickness > or = 1.5 mm have only a 30% to 70% probability of survival after surgery, and no adjuvant therapy has so far improved this outcome. Since interferon alfa-2a (IFNalpha2a) exhibits antitumor activity in metastatic melanoma, we investigated whether adjuvant IFNalpha2a diminishes the occurrence of metastases and thus prolongs disease-free survival in melanoma patients after excision of the primary tumor. PATIENTS AND METHODS: In a prospective randomized study, 311 melanoma patients with a Breslow thickness > or = 1.5 mm were assigned to either adjuvant IFNalpha2a treatment (n = 154) or observation (n = 157) after excision of the primary tumor. IFNalpha2a was given daily at a dose of 3 mIU subcutaneously (s.c.) for 3 weeks (induction phase), after which a dose of 3 mIU s.c. three times per week was given over 1 year (maintenance phase). RESULTS: Prolonged disease-free survival was observed in patients treated with IFNalpha2a versus those who underwent surgery alone. This difference was significant (P = .02) for all patients enrolled onto the study (intention-to-treat analysis) at a mean observation time of 41 months. Subgroup analysis showed that Breslow tumor thickness had no influence on treatment results in the groups of patients investigated. CONCLUSION: Adjuvant IFNalpha2a treatment diminishes the occurrence of metastases and thus prolongs disease-free survival in resected primary stage II cutaneous melanoma patients.

Lytic susceptibility of target cells to cytotoxic T cells is determined by their constitutive major histocompatibility complex class I antigen expression and cytokine-induced activation status.

Cytotoxic T-cell lines (TCL) were raised in vitro using stimulator cells with a defined major histocompatibility complex (MHC) mismatch and tested in a cytotoxic chromium-release assay against haemopoietic and non-haemopoietic target cells from the original stimulator. Monoclonal antibody (mAb)-blocking experiments and simultaneous determination of MHC class I, class II, lymphocyte function-associated antigen-1 (LFA-1) and intracellular adhesion molecule-1 (ICAM-1) density by quantitative radioimmunometric methods and flow cytometry on target cells demonstrated that lysis was restricted by MHC class I and dependent upon the constitutive MHC class I antigen expression. Measurements showed a high constitutive expression of class I MHC antigens on peripheral blood mononuclear cells (PBMC), but a low one on keratinocytes (K). Also, PBMC were more susceptible to lysis by TCL than K. Interferon-gamma (IFN-gamma) treatment of K resulted in increased MHC class I antigen expression and enhanced lytic susceptibility to TCL. IFN-alpha and tumour necrosis factor-alpha (TNF-alpha) treatment, which did not modulate MHC class I antigen expression on K, did not influence the amount of K lysis either. None of the cytokines tested in this analysis, however, increased the expression of MHC class I, class II, ICAM-1 and LFA-1 on PBMC. Only IFN-gamma pretreatment showed a minimal, statistically significant increase in MHC class I antigen expression. In spite of the minimal effect of IFN-gamma and no effect of IFN-alpha on class I MHC expression, pretreatment of target cells with both cytokines considerably increased their lytic susceptibility. The mechanism of cytokine-induced enhanced lytic susceptibility to TCL was not explained by increased MHC class I, LFA-1 or ICAM-1 expression, since no correlation was found between surface expression of these molecules and lytic susceptibility to TCL. These data demonstrate that: (1) the constitutive density of MHC class I antigens determines the extent of TCL lysis; (2) IFN-gamma, and not IFN-alpha or TNF-alpha controls the amount of K target cell lysis by increasing their MHC class I antigen expression; and (3) IFN-gamma and IFN-alpha control the amount of PBMC target cell lysis by a mechanism independent of MHC class I, ICAM-1 or LFA-1 expression.

[Local therapy of ulcus cruris]. / Lokaltherapie des Ulcus cruris.

The primary step of local ulcer therapy consists in debridement. Cleaning the ulcers from necrotic tissue can be achieved by surgical, mechanical, osmotic, autolytic and enzymatic means. The introduction of (semi-)occlusive wound dressings for moist wound healing has significantly improved ulcer treatment, since critical steps of wound healing, such as fibroblast proliferation, angiogenesis and epithelialisation are markedly accelerated. In contrast, topical antimicrobial therapy and traditional local treatment with "wound ointments" are of diminishing value because of frequent side effects such as inhibition of granulation tissue or allergic sensitization.

Correlation of minor histocompatibility antigen-specific cytotoxic T lymphocytes with graft-versus-host disease status and analyses of tissue distribution of their target antigens.

Peripheral blood mononuclear cells (PBMC) from 17 patients receiving HLA-identical sibling bone marrow grafts were stimulated with host pretransplant PBMC. Cytotoxic T-cell lines (TCL) with specificity for host pretransplant PBMC were obtained from 9 of these patients, all presenting with severe graft-versus-host disease (GVHD), but from none of the remaining cases lacking evidence of disease. Cytotoxic TCL were specific for host targets and failed to lyse donor cells. Monoclonal antibodies (MoAbs) blocking experiments and donor population screening analyses demonstrated that minor histocompatibility antigen (MiHA)-specific lysis of host targets was restricted by class I major histocompatibility complex (MHC) determinants. Whereas hematopoietic cells such as phytohemagglutinin (PHA) blasts or lymphoblastoid cell lines were susceptible to lysis by MiHA-specific TCL, keratinocytes (K) representing the natural targets of GVHD were quite resistant. Quantitative radioimmunometric measurements indicated very low constitutive expression of class I MHC antigens on K targets, which was readily increased by treatment with interferon-gamma (IFN-gamma). IFN-gamma treatment at the same time rendered these cells susceptible to lysis by MiHA-specific TCL. Host leukemic cells of 3 patients were recognized by MiHA-specific TCL in a chromium release assay and in one experiment host leukemic cells were effectively killed and their growth specifically inhibited in a leukemia colony assay by a clone. These data demonstrate that (1) host-specific cytotoxic TCL are detected exclusively in the PB of patients with acute GVHD grades II through IV after allogeneic matched bone marrow transplantation, and (2) their target antigens are simultaneously expressed on several host cell lines, including lymphoblastoid cell lines, PHA blasts, leukemic cells, and K. We also extend previous findings by showing that, besides the expression of the nominal MiHA, the density of the restricting class I MHC elements also crucially determines the extent of TCL lysis. Because of its capacity to enhance class I MHC antigen expression, IFN-gamma represents a key cytokine for determining the susceptibility of MiHA targets for lysis by TCL and clones, and in one patient an MiHA-specific clone recognized host leukemic cells and also inhibited host leukemic cell growth in a colony inhibition assay.

Acquired perforating dermatosis. Transepidermal elimination of DNA material and possible role of leukocytes in pathogenesis.

A patient had acquired perforating dermatosis and suffered from renal disease, diabetes mellitus, and lupus vulgaris. Histologic and immunohistochemical studies revealed that the bulk of the coarse granular basophilic material being extruded by transepidermal elimination was of nuclear origin obviously derived from polymorphonuclear leukocytes that were particularly abundant in an early, nonperforated lesion. At the lower boundary of the material being eliminated transepidermally, leukocytes were seen to accumulate, to undergo pyknosis and karyorrhexis, and to transform into nuclear debris. As a minor component, the material contained collagen fibers with altered staining qualities and, in an early lesion, elastic fibers. We speculate that accumulation, disintegration, and enzyme release from polymorphonuclear leukocytes may represent an important, hitherto disregarded driving force in transepidermal elimination. Lysosomal enzymes may later be responsible for the alteration of staining properties in collagen fibers, the degradation of elastic fibers, and for opening up the transepidermal route by impairing intercellular keratinocyte cohesion.

Amyloid P component binds to keratin bodies in human skin and to isolated keratin filament aggregates in vitro.

Dermal keratin bodies, consisting mainly of keratin intermediate filament aggregates (KIFA) coated with IgM anti-KIF autoantibodies, are present in normal human skin and occur in increased quantities in certain skin diseases. Keratin bodies are normally rapidly removed, but in primary localized cutaneous amyloidosis (PLCA) they are converted by an unknown mechanism to amyloid. Amyloid P component (AP), a glycoprotein identical to, and derived from, the normal plasma protein serum amyloid P component (SAP), is present in all forms of amyloid including PLCA. We investigated the interaction between SAP, keratin bodies, and KIFA. Immunofluorescence staining of normal skin using fluoresceinated anti-SAP and rhodamine-conjugated anti-IgM, or AE-1/AE-3 anti-keratin antibodies followed by Texas Red-conjugated anti-mouse immunoglobulin, showed that 52% +/- 4 (mean +/- sem, n = 6) of keratin bodies bound anti-SAP. Similar findings were present in a biopsy from a patient with lichen planus. Isolated KIFA, prepared by 8M urea extraction of normal human epidermis or cultured keratinocytes, were preincubated with normal human serum as a source of SAP and then stained with fluoresceinated anti-SAP. Bright fluorescence seen when the incubation medium contained Ca++ was absent in the presence of ethylenediamine tetraacetic acid. Specific Ca++-dependent binding of SAP to KIFA was confirmed using immunoblotting. Binding of SAP to KIFA did not prevent their degradation following exposure to trypsin or alpha-chymotrypsin. Similarly, partial enzymatic digestion of KIFA did not abrogate their ability to bind SAP. Our findings, that SAP is associated with keratin bodies in skin and exhibits Ca++-dependent binding to KIFA in vitro, identify keratin filaments as a newly recognized ligand for SAP.

[Skin replacement with cultured keratinocytes]. / Hautersatz durch kultivierte Keratinozyten.

Epidermal skin of only a few cm2 in size can be expanded several thousand times in cell cultures within 3 or 4 weeks and then be used to cover wounds. There have been reports on some patients with severe burns, whose skin was successfully replaced by autologous cultured epidermis in up to 50% of their body surface. The major drawbacks of this technique of skin replacement are the long culture period as well as the inferior mechanical stability and increased rate of wound contractures due to the lack of the dermal component. Since allogenic epidermal cell cultures are immunologically rejected, they are not suitable for the permanent coverage of wounds.

Rejection, after a slightly prolonged survival time, of Langerhans cell-free allogeneic cultured epidermis used for wound coverage in humans.

Autologous cultured epidermis (CE) grown from small skin biopsies in vitro has been successfully applied for wound grafting in humans. Since it has been reported recently that allogeneic CE might be tolerated as permanent wound cover, we investigated the properties of CE and its use as autologous and allogeneic grafts. Except for some differences, such as the absence of Langerhans cells and the lack of a stratum corneum, CE resembled its natural analogue. Autologous CE applied for grafting of leg ulcers and various surgical skin defects adhered firmly and permanently to the wound bed within 2 weeks, became regularly stratified, and formed a stratum corneum. Langerhans cells gradually entered the grafts; the dermis contained no inflammatory infiltrate. Allogeneic CE unmatched for MHC and blood group antigens used to partially cover tangentially excised third-degree burns, donor sites of split-thickness skin, and a defect after tumor excision initially survived well like the autografts. However, they were completely rejected after 10-22 (mean, 14.5) days, which is 4-5 days later than reported for split-thickness skin allografts. Clinically, rejection presented as "melting" of the graft. (Immuno)histologically, we found a dense mononuclear dermal infiltrate consisting predominantly of activated T cells, vacuolization, and single-cell necrosis of keratinocytes, as well as HLA-DR expression on keratinocytes, and finally separation and lysis of the epidermis. Limiting dilution analysis in 2 out of 4 allograft recipients revealed a considerable increase of circulating donor-specific cytotoxic T cell precursors during graft rejection. We conclude that grafting of allogeneic CE does not lead to permanent but to slightly prolonged graft survival.

IFN-mediated induction of MHC antigen expression on human keratinocytes and its influence on in vitro alloimmune responses.

The MHC Ag expression on the surface of keratinocytes is altered after treatment with IFN. IFN-gamma induces, as expected, a strong increase in class I MHC Ag expression as well as de novo expression of class II MHC Ag, whereas IFN-alpha 2 only slightly increases class I MHC Ag and does not induce keratinocytes to express class II MHC Ag. We used untreated and IFN-pretreated keratinocytes as stimulators and also as targets to study whether IFN-induced MHC Ag changes would alter the immunogenicity of keratinocytes in alloimmune responses. It was found that class II MHC Ag-carrying keratinocytes were unable to induce the proliferation of resting lymphocytes, but did stimulate T blasts. Untreated keratinocytes were virtually resistant to the lysis by classical CTL but became susceptible after exposure to IFN-gamma, but not IFN-alpha 2 at physiologic doses. These data demonstrate mechanisms by which the release of IFN-gamma might contribute to the development of disease such as the graft vs host disease.

Interferons (IFNs) and tumor necrosis factors (TNFs) in T cell-mediated immune responses against alloantigens. I. Influence on the activation of resting and antigen-primed T cells.

The aim of this study was to investigate the influence that endogenous IFNs released in response to antigenic or viral stimuli has on recognition of alloantigens in MLC. Results indicated that both the magnitude and the kinetics of response can be modified by IFNs. Neutralizing antibodies with specificity for IFN-gamma inhibit early and enhance late proliferative responses in MLC. Addition of physiological concentrations of IFN-gamma enhanced both early and peak proliferation, whereas IFN-alpha markedly inhibited alloantigen-induced lymphocyte proliferation. Further experiments revealed that IFN effects in MLC are not caused by direct interaction with responder cells: pretreatment with IFNs neither failed to alter their subsequent proliferative reactivity, nor did it influence production of IL 2 in MLC. IFN-gamma mainly affected MLC responses by direct interaction with stimulator cells. These influences on hemopoietic and non-hemopoietic stimulator cells were complex and could not simply be explained on the basis of an altered expression of class II MHC antigens. When induced by IFN-gamma to maximally express class II antigens, pbmnc, LCL or homogeneous populations of macrophages showed a marked deficiency to induce primary or secondary proliferative T cell responses. Resting unsensitized or sensitized T cells were not stimulated by class II MHC antigens constitutively expressed or induced by IFN-gamma on cell types other than dendritic cells or LCL. Class II antigens on the former cells were, however, readily recognized by T helper blasts, and this process involved the T4 epitope of the T cell receptor. IFN-gamma treatment also influenced the intrinsic suppressive capacity of macrophages or keratinocytes without involving prostaglandin synthesis or inducing expression of IL 2 receptors on non T cells.

[Cultivated epidermis as a skin replacement--improved technics using mesh silastic sheets]. / Kultivierte Epidermis als Hautersatz--Verbesserung der Technik mittels gemeshter Silastic-Folie.

The cover of cutaneous wounds with cultured epidermis can be significantly improved by prior backing of the epithelium with meshed silastic sheets, whereas other materials, such as vaseline gauze, foam rubber etc., are not as satisfactory. Meshed silastic is affixed to the epithelium surface just before the enzymatic detachment from the culture flask is completed. Thus it protects the delicate epithelial membrane from injury during transplantation and helps to control its polar orientation. In addition, the silastic mesh prevents the graft, which initially does not possess any horny layer, from drying up and allows adequate drainage of wound secretions. Cultured epidermal grafts prepared by this method take very well (more than 90%) attaching to the wound bed quite firmly within a week.

Arachidonic acid metabolism in guinea pig Langerhans cells: studies on cyclooxygenase and lipoxygenase pathways.

Epidermal Langerhans cells are macrophage-like la+ leukocytes that are critically involved in cutaneous immune reactions. Because macrophages exert their immunoregulatory activity in part by generation of oxygenated arachidonic acid metabolites, we systematically studied arachidonic acid transformations by purified guinea pig Langerhans cells and compared them with mixed epidermal cells and Langerhans cell-depleted keratinocytes. Products formed from arachidonic acid by cell homogenates were measured after thin-layer or reverse-phase high-pressure liquid chromatographic separation. In addition, leukotriene B4 and C4 formation was assessed in supernatants of Ca ionophore A23187-challenged intact cells by radioimmunoassay. Mixed epidermal cells converted arachidonic acid predominantly via cyclooxygenase and 12-lipoxygenase pathways. The main products were prostaglandin D2 (PGD2) and 12-hydroxyeicosatetraenoic acid (12-Hete), although significant amounts of PGE2, PGF2 alpha, and 6-keto-PGF1 alpha were formed as well. PGD2 synthesis was dependent on the presence of reduced glutathione. The product spectrum formed by Langerhans cell-depleted keratinocytes was virtually indistinguishable from mixed epidermal cells. In contrast, Langerhans cells showed a markedly different metabolism of arachidonic acid. They exhibited an exceedingly high PGD2-generating capacity, whereas only minor amounts of 12-HETE and very low amounts of other prostaglandins were synthesized. The PGD2/12-HETE ratio was 1.22 for mixed epidermal cells and 4.37 for Langerhans cells. Leukotriene production from exogenous or endogenous arachidonic acid could not be demonstrated by either radioenzymatic or radioimmunologic detection methods. We conclude that guinea pig Langerhans cells transform arachidonic acid predominantly to PGD2, which might mediate significant immunoregulatory, inflammatory, and antitumoral activity in the skin.

HLA-DR expression on keratinocytes is a common feature of diseased skin.

Biopsy specimens from 185 patients with 52 different skin disorders were investigated by indirect immunofluorescence staining for the presence of HLA-DR bearing keratinocytes and their association with an underlying inflammatory infiltrate and in particular with activated (HLA-DR-positive, Leu-4-positive) T lymphocytes. HLA-DR expression on keratinocytes was demonstrated in 38 dermatoses, including lymphocytic vasculitis, lupus erythematosus, morphea, vitiligo, lichen planus, cutaneous T-cell lymphoma, various infectious dermatoses, allergic contact dermatitis, granulomatous dermatoses, Sweet's syndrome, lichen sclerosus and erythema nodosum. In 27 of these this had not previously been reported. Occurrence of HLA-DR on keratinocytes was invariably linked to the presence of a lymphocytic infiltrate containing numerous activated T-cells (Leu-4 +, HLA-DR +) whereas such infiltrates were not accompanied by HLA-DR expression on keratinocytes in all the dermatoses investigated, as in pseudolymphoma and erythema anulare centrifugum. However, HLA-DR positive keratinocytes were consistently absent in skin disorders lacking any significant lymphocytic infiltration (e.g. leukocytoclastic vasculitis, bullous autoimmune dermatoses, genodermatoses and mastocytosis). Although it has been suggested that HLA-DR-positive keratinocytes are involved in various immune responses of the skin, their exact functional significance is, as yet, unknown.

[Mycetoma caused by Petriellidium boydii: treatment with ketoconazole]. / Myzetom durch Petriellidium boydii: Behandlung mit Ketoconazol.

A case of mycetoma of the lower leg (Madura foot) with bone involvement caused by Petriellidium boydii is presented. After an initially favourable therapeutic response to ketoconazole (2 X 200 mg/die) the process recurred despite proven sensitivity of the fungus in vitro. It could not be controlled with conservative treatment and ultimately necessitated amputation of the lower leg. A striking, hitherto unreported feature of mycetoma was the episodic occurrence of circulating immune complexes with febrile monoarthritis of the adjacent ankle.