Epidermal expression of interleukin-6 and tumour necrosis factor-alpha in normal and immunoinflammatory skin states in humans.

The in vivo presence of cytokines in human skin was examined by ABC- and IF-techniques, and by using antisera to human rIL-6, rIL-1-alpha, rIL-1-beta, rTNF-alpha and an antiserum to crude supernatants of Staph. albus activated human blood monocytes (anti-MK-antiserum) before and after absorption with rIL-6. The following normal and immunoinflammatory human skin states were examined: Normal skin. Examination of biopsies from 9 healthy individuals and single cell preparations from 3 healthy individuals revealed a granular intercellular/membrane associated staining for IL-6 and TNF-alpha in the upper epidermal layers. A cytoplasmic staining was also detected, most pronounced using the anti-rIL-6 antiserum. Membrane associated IL-6 and TNF-alpha were detected in epidermal single cell preparations, but CD1 positive LC failed to express any of the cytokines examined. Psoriasis skin. Biopsies from lesional and unaffected skin of 11 patients with chronic, nummular/plaque type psoriasis were investigated for IL-6, and in addition 5 of these, moreover, for TNF-alpha. Biopsies from lesional psoriatic skin continually revealed increased staining for IL-6 compared with non-lesional skin, where the staining was similar to that observed in healthy individuals. Epidermal TNF-alpha expression did not differ from that observed in normal skin. AIDS related Kaposi's sarcom. Epidermal staining patterns were similarly increased for IL-6 and TNF-alpha in biopsies obtained from nodular KS tumours compared with unaffected skin of 6 homosexual male AIDS patients, whereas the endotheloid cell of the tumour did not stain for any of the cytokines. Staining of unaffected skin was similar to that observed in healthy individuals. The allergic and irritant patch test reaction. Investigation of biopsies from 5 patients with positive APR and 5 healthy individuals with IPR disclosed increased epidermal staining for IL-6, but not for TNF-alpha, in both reactions, while staining from petrolatum tested and non-tested skin was similar to that observed in normal individuals. CD1 positive LC remained negative in all biopsies when stained for IL-6 and TNF-alpha. In vivo UV-irradiated skin. Using anti-MK and anti TNF-alpha antisera skin biopsies from 5 healthy subjects 24 h after UVB irradiation revealed markedly increased epidermal staining for both cytokines compared with non-irradiated skin. Furthermore, a gradual decrease in epidermal staining for IL-6 was observed in specimens from lesional skin of 6 patients with psoriasis taken before and during PUVA therapy, while staining of non-lesional skin remained unchanged.

Keratinocyte-expression of interleukin-6 but not of tumour necrosis factor-alpha is increased in the allergic and the irritant patch test reaction.

The cytokine expression on epidermal cells in the allergic patch test reaction (APR) and irritant patch test reaction (IPR) was studied using antibodies against rIL-6, rTNF alpha, rIL-1 alpha and rIL-1 beta in a histochemical biotin-avidin technique. Nickel sulphate was used for APR in 5 nickel allergic patients and sodium lauryl sulphate for IPR in 5 healthy individuals. The individuals served as their own control. Enhanced keratinocyte expression of IL-6 was observed in APR and IPR, whereas staining for TNF alpha remained unaltered compared with non-tested and petrolatum-tested skin. Staining for IL-1 alpha and IL-1 beta proved negative in all specimens. Double-staining experiments demonstrated that epidermal and dermal OKT-6 (CD1) positive Langerhans cells (LC) remained negative for all cytokines. These results demonstrate that enhancement of keratinocyte-bound IL-6 does not induce TNF alpha, IL-1 alpha/beta or IL-6 expression by LC during APR or IPR, and that enhanced keratinocyte expression of IL-6 fails to distinguish between these two reactions.

Interleukin-6 and tumour necrosis factor alpha are expressed by keratinocytes but not by Langerhans cells.

The presence of human cytokines was examined in parallel skin biopsies and epidermal single cell preparations obtained from normal individuals. Using biotin-avidin-peroxidase and immunofluorescence techniques and antibodies against recombinant cytokines, a granular intercellular/membrane-associated staining for interleukin-6 (IL-6) and tumour necrosis factor alpha (TNF alpha), but not IL-1 alpha or beta, was observed. An epidermal cytoplasmic staining pattern was also detected, which was most pronounced using the anti-rIL-6 antiserum. In the epidermal single cell preparations, membrane-associated staining was detected for both IL-6 and TNF alpha. Double staining revealed that CD1-positive Langerhans cells (LC) failed to express any of the examined cytokines. In vitro binding of rIL-6 or rTNF alpha to skin sections and epidermal single cell preparations indicated that the cell surface-associated IL-6 and TNF alpha originally demonstrated on keratinocytes were truly membrane-bound. Finally, co-cultivation of epidermal cells with an IL-6 responsive cell line, B9, and testing of epidermal cell supernatants in this assay, indicated that the in vivo membrane-bound IL-6 had biological activity.

Abnormal essential fatty acid metabolism in Darier's disease.

Fatty acid levels in plasma and erythrocyte cell membranes were determined in 13 Danish patients with Darier's disease and 21 Danish controls. Concentrations of the main dietary essential fatty acids, linoleic acid (18:2n-6) and alpha-linolenic acid (18:3n-3), were consistently modestly above normal; concentrations of the delta 6-desaturase metabolites of both linoleic and alpha-linolenic acids, however, were consistently and often significantly below normal. These results suggest that the capacity of the enzyme delta 6-desaturase activity is inadequate in patients with Darier's disease.

Epidermal cell surface-associated IgG in patients with primary Sjögren's syndrome: in vitro evidence for immune complex binding.

In vivo deposits of IgG have previously been demonstrated in the epidermal intercellular area of clinically unaffected skin from 68% of patients with primary Sjögren's syndrome (primary SS). This study compared circulating IgG from patients with primary SS with that from secondary SS in their ability to bind normal human epidermal cells in vitro. We observed a granular pattern of IgG binding to the normal epidermal cell surfaces with 9 of 18 sera from patients with primary SS (50%), 3 of 19 sera from patients with SS secondary to rheumatoid arthritis (16%) (p = 0.025), and none of 24 normal control sera (p less than 0.001). In a subsequent analysis of polyethylene glycol separated sera from two normal controls and two primary SS patients, the epidermal IgG binding capacity was found only in the precipitates of the patients. These findings support our previous hypothesis that the in vivo intraepithelial IgG deposits in primary SS patients are due, at least in part, to cell surface-bound immune complexes.

Epidermal tumour necrosis factor alpha and interleukin 6-like activities in AIDS-related Kaposi's sarcoma. An immunohistological study.

Biopsies from 6 patients with AIDS and Kaposi's sarcoma (KS) in the tumour stage, and 6 healthy controls, were immunohistologically examined for the presence of tissue-bound tumour necrosis factor alpha (TNF alpha) and interleukin 6 (IL-6) in the skin. TNF alpha was demonstrated using specific polyclonal antiserum to human recombinant TNF alpha. IL-6 was visualized indirectly using a polyclonal antiserum to partially purified human crude supernatants of activated human blood monocytes, followed by absorption with recombinant human IL-6. The cytokines were found identically located in epidermal cell membranes in stratum granulosum and spinosum of the epidermis from unaffected skin in both AIDS patients and in controls. Biopsies from KS elements showed markedly increased epidermal staining for both TNF delta and IL-6. It was not possible to detect TNF alpha or IL6 in the endothelial cells of the tumour. The observation of increased amounts of epidermal-bound TNF alpha and IL-6 in AIDS-related KS elements supplements previous studies indicating that the skin plays an active immunoinflammatory role in patients with AIDS.

Interleukin-6 in the epidermis of patients with psoriasis before and during PUVA treatment.

Biopsies from lesional and unaffected skin of 6 patients with psoriasis, taken before and during treatment with psoralen plus UVA (PUVA) were examined immunohistologically, using partially purified polyclonal antibodies to crude supernatants of activated human blood monocytes. By absorption with recombinant derived human monokines, we were able to demonstrate that interleukin-6 (IL-6) (but not IL-1 alpha or IL-1 beta) was located in a laminar and granular pattern in stratum corneum, and on epidermal cell membranes in the viable cellular epidermis. Before PUVA treatment, the intensity and the extension of staining for IL-6 were both markedly increased in lesional skin compared with uninvolved skin. A weaker staining for IL-6 was observed in lesional skin, simultaneous with the clinical improvement of psoriasis. The staining patterns for IL-6 in biopsies from cleared lesional skin and uninvolved psoriatic skin were identical at the conclusion of therapy.

Expression of interleukin-6-like molecules and tumour necrosis factor after topical treatment of psoriasis with a new vitamin D analogue (MC 903).

Skin biopsies from 5 patients with chronic plaque psoriasis were examined to test the effect of topical treatment with a new synthetic vitamin D3 analogue, MC 903, on epidermal interleukin 6 (IL-6) and tumour necrosis factor alpha (TNF alpha). Skin biopsies, taken before therapy and after one and 2 weeks of therapy were examined immunohistologically. IL-6 was visualized using a partially purified polyclonal antiserum to crude supernatants of activated human blood monocytes before and after absorption with recombinant human IL-6. TNF alpha was demonstrated using a specific polyclonal antiserum to human recombinant TNF alpha. Both the intensity and extension of staining for IL-6, but not for TNF alpha, were increased in lesions compared with unaffected skin. During therapy with MC 903 a decline in the staining intensity for IL-6, but not for TNF alpha, was observed in both lesional and unaffected skin. Placebo-treated lesions, however, remained unchanged.

Squamous cell carcinomas in relation to cyclosporin therapy of non malignant skin disorders.

Two patients with psoriasis and one with pityriasis rubra pilaris developed squamous cell carcinomas in relation to cyclosporin A therapy. Dermatological patients previously treated with ultraviolet radiation and other kinds of immunosuppressive therapy may represent a special risk group with respect to cyclosporin. A related malignancies.

Immunohistological detection of interleukin I-like molecules and tumour necrosis factor in human epidermis before and after UVB-irradiation in vivo.

Skin biopsies from five healthy subjects, taken before and after UVB irradiation, were examined using immunohistological techniques for the cytokines interleukin-I (IL-I) and tumour necrosis factor (TNF). Using polyclonal specific antibodies against IL-I and TNF, the two cytokines appeared identically located on the epidermal cell membranes of the stratum granulosum and stratum spinosum in unexposed skin. After UVB-exposure, the staining intensity for both IL-I/epidermal cell derived thymocyte-activating factor (ETAF) and TNF was markedly increased, and the epidermal staining included the basal cell layer. Immunohistological investigation of tissue-bound epidermal cytokines may be valuable in the study of skin diseases.

Is the effect of phototherapy in psoriasis partly due to an impact on vitamin D metabolism?

To elucidate the effect of phototherapy on vitamin D metabolism in psoriatics, the serum concentrations of the major vitamin D metabolites (25-hydroxy-vitamin D (25(OH)D), 1,25-dihydroxy-vitamin D (1,25(OH)2D), and 24,25-dihydroxy-vitamin D (24,25(OH)2D)) were studied in 10 patients with disseminated psoriasis, both before and after phototherapy. Some 3-4 weeks of Goeckerman therapy induced significantly increased serum levels of 25(OH)D (mean: 24.6 ng/ml versus 54.4 ng/ml; (p less than 0.001] and 24,25(OH)2D (mean: 2.01 ng/ml versus 3.49 ng/ml; (p less than 0.001)). After phototherapy the mean serum level of 1,25(OH)2D increased nearly to the level found in healthy controls (mean: 23.8 vs. 32.2 pg/ml). However, this increase was not significant. It is shown that conventional phototherapy does have an impact on vitamin D metabolism in psoriatics. Since previous investigations have indicated an abnormal vitamin D metabolism in patients with psoriasis, it is possible that the beneficial effect of phototherapy in this disease might be due partly to an impact on vitamin D metabolism.

Circulating IgG from patients with primary Sjögren's syndrome deposited in the epidermis of normal human skin transplanted to athymic nude mice.

Sera from 7 patients with primary Sjögren's syndrome and from two control persons were administered intraperitoneally to athymic nude mice transplanted with normal human skin. Seven days after transfer of serum from 5 of the patients, intra-epidermal IgG1 and IgG3 deposits were demonstrated in the skin grafts by immunofluorescence. The deposits closely resembled in vivo deposits found in the skin of these patients. No correlation was found between the presence of epidermal deposits and levels of IgG1 and IgG3 in serum. No IgG deposits were found in skin grafts on animals given control serum, and neither could human IgG be detected in mouse skin adjacent to the grafts. Epidermal deposits of human-IgM, -IgA, -fibrinogen, -C3c and mouse-Ig were not demonstrated in biopsies from grafts or mouse skin. The results support the hypothesis that epidermal in vivo deposits of IgG in patients with primary Sjögren's syndrome are the result of Fc-receptor-mediated binding to epidermal cells.

Immunohistochemical characterization of intraepidermal in vivo IgG deposits in patients with primary Sjögren's syndrome.

68% of patients with primary Sjögren's syndrome have previously been found to have intra-epidermal in vivo IgG deposits in clinically unaffected skin. In this investigation, we examined immunohistologically skin biopsies from 5 patients with primary Sjögren's syndrome and from 5 normal controls in order to characterize further the intra-epidermal IgG deposits. Employing direct immunofluorescence and peroxidase-antiperoxidase (PAP) techniques, IgG was localized to epidermal cell surfaces. Double-labelling immunofluorescence experiments showed IgG to be bound to OKT6-positive Langerhans cells, and to some degree also to keratinocytes. Only IgG1, IgG3 and in one patient IgA were deposited, whereas IgG2, IgG4, IgM, IgD, IgE, C1q, C3c, C3d, C4, beta-2 microglobulin, albumin, fibrinogen and C-reactive protein were not found deposited in the intra-epidermal area. The epidermal IgG deposits were reactive with anti-Fc-fragment antibodies and with staphylococcal protein-A. These results are compatible with, although not definitive proofs of, the hypothesis that the in vivo deposited IgG is found in the form of IgG-containing immune complexes.

In vitro binding of IgG from patients with primary and secondary Sjögren's syndrome to different components of keratinized stratified squamous epithelia.

The ability of serum IgG from patients with Sjögren's syndrome (SS) to bind to keratinized stratified squamous epithelium of human skin or rat oesophagus was examined, using an indirect immunofluorescence technique. No in vitro binding of serum IgG to the cell membranes of normal human epidermis was demonstrated in 7 patients with primary SS and two normal controls. Thus, the intraepidermal in vivo IgG deposits previously found in 5 of the 7 patients could not be imitated in vitro. Examination for in vitro binding to rat oesophageal epithelium of serum IgG from 21 consecutive patients with primary SS, 19 consecutive patients with rheumatoid arthritis and secondary SS and 22 normal controls showed that antikeratin antibodies occur more frequently (p less than 0.001) in the patients with secondary SS compared to patients with primary SS and to normal controls.

Epidermal IgG deposits in patients with chronic inflammatory connective tissue diseases: diagnostic value and correlation to clinical and immunological parameters in patients with primary Sjögren's syndrome.

Thirty-seven patients with primary Sjögren's syndrome (Primary SS), 12 patients with incipient primary SS, 100 patients with other chronic inflammatory connective tissue diseases (CTD) and 20 healthy controls had a punch biopsy taken from clinically unaffected skin from the buttock. Direct immunofluorescence examinations revealed intraepidermal IgG deposits in 68% of patients with primary SS, in 42% of patients with incipient primary SS, in 13% of patients with rheumatoid arthritis (RA, n = 31), in 15% of patients with systemic lupus erythematosus (SLE, n = 13), in 24% of patients with other well-defined CTD (n = 41), in 40% of patients with ill-defined CTD (n = 15) and in 10% of healthy controls. Immunoglobulin deposits along the dermoepidermal junction zone (DEJ) were not found in any of the patients with primary SS, but were present in 16% of the patients with RA and in 23% of the patients with SLE. In the patients with primary SS, no correlation was found between intraepidermal IgG deposits and age, disease duration, extraglandular manifestations, P-IgG concentration, ANA, rheumatoid factors or circulating immune complexes. Examination for intraepidermal IgG deposits seems valuable in the differential diagnosis between primary SS and RA or SLE, and it could supplement the lupus band test.