Evidence that DNA damage is a mediate in ultraviolet B radiation-induced inhibition of human gene expression: ultraviolet B radiation effects on intercellular adhesion molecule-1 (ICAM-1) expression.

Expression of intercellular adhesion molecule-1 (ICAM-1) is a prerequisite for the capacity of cells to physically interact with leukocytes. Ultraviolet B radiation previously was found to inhibit interferon gamma-induced ICAM-1 expression in human keratinocytes by suppressing interferon gamma-mediated upregulation of ICAM-1 mRNA levels. Because ultraviolet B radiation induces photoproducts in cellular DNA, the potential role of ultraviolet B radiation-induced DNA damage in this system was assessed. For this purpose, cells from a normal donor were compared with cells from patients with xeroderma pigmentosum from complementation groups C and D. Xeroderma pigmentosum cells are defective in the removal of ultraviolet B radiation-induced DNA lesions, and thus lower ultraviolet B radiation doses are required to retain equivalent numbers of DNA photoproducts at a given time point after irradiation. In the present study, ultraviolet B radiation inhibited interferon gamma-induced ICAM-1 mRNA expression in primary human skin fibroblasts in a manner identical to that previously observed for keratinocytes. Comparative studies employing normal versus xeroderma pigmentosum fibroblasts revealed that in xeroderma pigmentosum fibroblasts, two- to threefold lower ultraviolet B radiation doses were required to achieve inhibition equivalent to that observed in normal fibroblasts. In irradiated normal cells, inhibition of interferon gamma-induced ICAM-1 mRNA expression was transient and restored 12 h after ultraviolet B radiation exposure. In contrast, in xeroderma pigmentosum complementation group D cells, no restoration could be observed for up to 48 h, but responsiveness was restored in xeroderma pigmentosum complementation group C cells after 24 h. These studies indicate that ultraviolet B radiation-induced inhibition of interferon gamma-mediated ICAM-1 expression involves the generation of DNA photo-products.

Establishment and characterization of two Merkel cell tumor cultures.

Two Merkel cell tumor cultures (MC-MA1, MC-MA2) have been established from metastases of typical Merkel cell tumors. The mestastases in vivo were characterized by co-expression of cytokeratins 8, 18, 19, 20 and neurofilaments, presence of intermediate filament whirls, expression of synaptophysin, neuron-specific enolase, and chromogranin A, rare and weak immunostaining for plakoglobin but absence of cadherins and desmoplakins. Both cultures grow, using supplemented RPMI medium on human irradiated fibroblast feeder layers, as loosely arranged floating small aggregates. Their karyotypes are mostly hyperdiploid. The mean doubling times were about 84 h in the first 8 months and later increased. Ultrastructural and immunoelectron microscopic studies of the Merkel cell tumor cells in vitro (MC-MA1, MC-MA2) revealed sparse membrane-bound neuroendocrine granules and typical IFs that were partly arranged in paranuclear whirls and were labeled by antibodies against cytokeratins and neurofilaments. In immunocytochemical studies using antibodies to cytokeratins 8, 18, 19, and 20 and neurofilament protein NF-L, Merkel cell tumor cells in vitro showed a uniform staining appearing as paranuclear whirls and cytoplasmic fibrils as well. Double-labeling experiments showed a co-localization of both intermediate filament types in most cells. Biochemically we found cytokeratins 8, 18, 19, and 20, and NF-L in tumor cells in vitro. Immunocytochemical staining was negative for desmoplakins, various cadherins, and cell adhesion molecules, whereas plakoglobin was only rarely detectable in some Merkel cell tumor cells in vitro. By immunoluminometric assay chromogranin A was detected in cell homogenates and culture supernatants as well. Immunocytochemically, synaptophysin and neuron-specific enolase were detectable additionally in some of the cells. These established cell cultures will allow further studies devoted to the biology, differentiation, and hormone secretion of Merkel cell tumors that may also increase our knowledge about normal Merkel cells.

[DNA-repair of lymphocytes during PUVA-treatment (author's transl)]. / DNA-Reparatur von Lymphocyten unter PUVA-Behandlung.

In 20 psoriatic patients under PUVA maintenance treatment blood samples had been collected imediately before and after the UVA irradiation. The DNA-repair activity has been determined in the isolated lymphocytes by the incorporation of 3H-thymidine during the first 3 h (inhibition of the semiconservative replication by hydroxyurea). There is evidence for an increased repair activity in the lymphocytes after PUVA treatment whereas the lymphocytes exposed only to 8-MOP (before UVA irradiation) show lower values. The difference is statistically significant 45 and 90 min after irradiation, but not any more after 3 h.

Risk factors of the cutaneous melanoma phenotype.

Of 20 melanoma patients 85% show a prolonged erythema persistence after a marked test erythema of 8 MED with 300 nm +/- 10 nm (control group only 34%). This phenomenon does not correlate with the skin type and is useful in identifying high-risk patients prone to melanoma and light-induced skin cancer (92%). The spontaneous and the UV-C-induced number of sister chromatid exchanges (SCE) per metaphase was significantly higher in peripheral leukocytes of melanoma patients than in normal controls. The attempt was made to establish a "risk-spectrum" of cutaneous melanoma phenotype.

Light-induced skin cancer and prolonged uv-erythema.

The individual minimal erythema dose (MED) and the persistence of a marked erythema (8 MED) was monitored over 3 weeks (300 nm +/- 10 nm) in 4 groups: White students with fair complexion compared with students of homogenous pigmentation as well as skin carcinoma patients compared with a control group of the same age, i.e., older than 50 years. The MED of the 4 groups gives no significant differences, while the skin carcinoma group shows in 80% a prolonged erythema persistence (control group only 28%). This phenomenon does not seem to correlate with the skin type and may be useful in identifying high-risk patients prone to light-induced skin cancer.

A ninth complementation group in xeroderma pigmentosum, XP I.

A new complementation group of excision-deficient xeroderma pigmentosum (XP) is described in 2 patients living in the F.R.G. Dermatological, ophthalmological and neurological symptoms of XP are presented together with DNA repair characteristics such as unscheduled DNA synthesis, colony-forming ability and alkaline elution studied in cultured fibroblasts. The results are compared to normal controls.

[Excision repair in lymphocytes of psoriasis (author's transl)]. / Excisions-Reparatur in Lymphocyten von Psoriatikern

Psoriasis is a autosomal hereditary disease of the skin, which has been treated with a great number of possibly carcinogenic substances. No increase of maligonomies has been observed except for the tumors induced by arsenic. The question may be asked whether the psoriatic cell has additional or more effective mechanisms to eliminate onocogenic somatic mutations. In this study the capacity of the excision repair of circulating blood lymphocytes from 12 psoriatic patients has been investigated and compared to a control group of equal size. After irradiation with ultraviolet light, the lymphocytes are incubated with 3 H-thymidine and at various intervals the radioactivity in the cell material precipitated with perchloric acid is measured. It appears, that the excision reapir in lymphocytes from patients with psoriasis and normal persons is equal. In addition, the influence of hydroxyurea on the thymidine incorporation in lymphocytes has been studied.

Heterogeneity of xeroderma pigmentosum (XP); variability and stability within and between the complementation groups C, D, E, I and variants.

Thirty-two cases of xeroderma pigmentosum (XP) of the complementation groups C (7), D (12), E (3), I (2) and 8 variants are analyzed biochemically and clinically. There is some congruence of the cellular defects (UDS, CFA, SCE) and the clinical severity of the skin symptoms. Despite the large clinical variability within and between the complementation groups, several clinical features are to be attributed to one group or another. The most striking observation is the predominance of LMM in the D group and BCC in the mild E group as well as in the variants. This observation might stimulate research to find a cellular characteristic of the melanoma risk.

Dysplastic nevus syndrome: ultraviolet hypermutability confirmed in vitro by elevated sister chromatid exchanges.

The dysplastic nevus syndrome (DNS) is a clinical and genetic entity in which affected individuals have increased numbers of dysplastic nevi and a markedly increased risk of developing one or more cutaneous melanomas. Sister chromatid exchanges (SCE) are one of the cytogenetic end points that are positively correlated with the mutation rate and may therefore be used to estimate the spontaneous and the UV-induced mutagenesis in cultured and stored fibroblasts. SCE were presented and stained according to the Hoechst-Giemsa method. The normal control fibroblasts showed 9.48 +/- 1.74 SCE per metaphase (n = 23) with an UV-C-induced increase (delta SCE) of 1.61 +/- 0.53 SCE per mJ/cm2 in the range of 0-5 mJ/cm2. Fibroblasts from DNS patients with melanoma (n = 12) showed normal values of spontaneous SCE but a significant increase (p = 0.01) of 2.43 +/- 0.68 SCE per mJ/cm2. As in xeroderma pigmentosum, UV-C-induced delta SCE appears as a valuable tool for measuring the individual hypermutability in DNS. delta SCE indicates the increased susceptibility to UV-induced somatic mutations and may be etiologically related to the increased melanoma incidence in DNS.

Effects of ultraviolet B radiation on cytoskeletal and adhesion molecules in human epidermis.

The expression of cytokeratins (CK), various adhesion molecules and growth factor receptors were investigated in ultraviolet (UV) erythemas 24 h and 48 h after exposure to 4 times the minimal erythema dose. Skin biopsies were analysed immunohistochemically using a battery of antibodies and biochemically by gel-electrophoresis. The CK pattern was shown to change within 24 h. CK typical of basal keratinocytes were detectable heterogeneously within the suprabasal compartment, and the staining for suprabasal CK was heterogeneous in prickle cells. Interestingly, CK 17, not detectable in normal epidermis, was induced within a few hours in suprabasal cells but not in basal ones. In addition, CK 6 and 16, typical for hyperproliferation, were intensively synthesized in all epidermal layers already 24 h after UV exposure. Moreover, integrin expression was studied and surprisingly integrins were heterogeneously detectable, the staining being patchy in suprabasal keratinocytes and reduced within the basal layer. Receptors of epidermal growth factor and nerve growth factor were distributed in UV erythema very irregularly and weakly. Our findings argue for profound changes in composition of cytoskeleton and of cell adhesion molecules in the epidermis shortly after UV exposure.

[Effects of UV-C and of 8-MOP + UV-A on the T- and B-population of human lymphocytes in vitro (author's transl)]. / Wirkung von UV-C und von 8-MOP + UV-A auf die T- und B-Population menschlicher Lymphocyten in vitro.

The transformation of UV-C irradiated leucocytes and lymphocytes by the mitogens Con A, PHA and PWM is measured by the 3H-Tdr. incorporation after 72 h incubation. Furthermore T- and B-cells are determined by the method of rosette formation. A clear inhibition of the cell activity is seen after irradiation of leucocytes with 450 mJ/cm(2) and of a lymphocyte suspension with 5-10 mJ/cm(2). There are no significant differences between the effects of the various mitogens. The number of T-cells decreases proportionality to the various intensity, the number of B-cells remain constant. Irradiation with UV-A + 8-MOP cause, equally for all mitogens, a dosis dependent inhibition of thymidine incorporation.

Effects of 8-methoxypsoralen (8-MOP) and UVA on human lymphocytes.

Peripheral lymphocytes of 37 psoriatic patients are tested before and under PUVA treatment using as parameter the non specific stimulation effect of HgCl2 (10 microgram/ml) in culture, measuring the 3H-thymidine incorporation after the last 16 h of a 5-days culture. Oral 8-MOP in therapeutic doses is decreasing the lymphocyte stimulation as well as 8-MOP together with UVA irradiation during the first week of treatment. After 1 week, the stimulation is, on the contrary, significantly enhanced after irradiation. Lymphocytes isolated by centrifugation over Lymphoprep are submitted to PUVA conditions in petri dishes (Hank's solution 8-MOP 1 microgram/ml, irradiation with 350 nm, 93-372 mJ/cm2). The total cell number, the E-rosette formation (as marker for T-lymphocytes) and the EAC-rosette formation (as marker for B-lymphocytes) are determined. PUVA conditions have an energy dependent decreasing effect on the cell number, while the T- and B-cell proportions remain constant. UVA irradiation alone has such an effect only with high energies. 8-MOP without UVA has no significant influence on the cell number.

De Sanctis-Caccione syndrome: xeroderma pigmentosum with oligophrenia, short stature and neurologic disorders. / De-Sanctis-Cacchione-Syndrom: Xeroderma pigmentosum mit Oligophrenie, Minderwuchs und neurologischen Störungen.

A 6 year-old boy with de Sanctis-Cacchione syndrome is reported. This syndrome is characterized by the triad xeroderma pigmentosum (XP), mental deficiency and neurological disturbances. The patient's cells were assigned to genetic complementation group A by use of the cell fusion technique. DNA repair capacity as measured by unscheduled DNA synthesis (UDS) was drastically reduced to 7.5%, compared with 100% of the controls. The rate of sister chromatid exchange (SCE), an indicator of the hypermutability in XP, was clearly elevated after ultraviolet radiation of skin fibroblasts of the patient.

Effect of pentoxifylline on sunburn cell formation in a novel supravital human skin model.

BACKGROUND/AIM: The aim of this study was to determine whether pentoxifylline (Pf) has an effect on sunburn cell (SBC) formation in humans. METHODS: A novel supravital human skin model was used. Normal skin samples were placed on gauze in completed RPMI 1640 Medium and remained vital for 48 h. Three concentrations of Pf (7.5, 15.0 and 30.0 microg/ml) were tested. After 2 h, each skin sample was irradiated with 120 mJ/cm2 of UVB. After 24-h incubation, the samples were formalin fixed, paraffin embedded and sections were stained with haematoxylin-eosin. RESULTS: The mean count of SBC (10.43 +/- 1.35 (SEM)) was significantly higher in the control group (without Pf) compared with its mean count in 7.5 microg/ml Pf (5.18 +/- 0.62, P < 0.001), or 15.0 microg/ml Pf (5.79 +/- 1.70, P < 0.001), or 30.0 microg/ml Pf (4.37 +/- 1.47, P < 0.001). CONCLUSION: Pentoxifylline reduced SBC formation in our supravital human skin model. It presumably acts as an antioxidant agent.

Dysplastic nevus syndrome: intrafamilial identification of carriers by cytogenetics.

Seven members of a family with dysplastic nevus syndrome (DNS) were examined clinically; skin biopsies of unaffected skin from 6 were taken. Biopsy-derived cultivated fibroblasts were examined by cytogenetic methods, i.e. by measuring the spontaneous and the UVB- and UVC-driven increase in sister chromatid exchange (SCE). A male patient with malignant melanoma, his son and his nephew, both with multiple dysplastic nevi, showed a distinctive elevation of UV-induced SCE, whereas the other, unaffected members of the family showed normal values. These results give evidence that in siblings with DNS the affected members can be identified not only on clinicopathological grounds but also by UV-induced elevated SCE at the cytogenetic level.

Nephrotic syndrome, hypertension, and adrenal failure in atypical Cockayne syndrome.

This report describes a boy with an atypical severe from of Cockayne syndrome type II manifesting in infancy. He developed nephrotic syndrome at the age of 4.7 years and a hypertensive crisis with hemiparesis at 5.4 years. Renal biopsy revealed focal segmental glomerulosclerosis, which was confirmed at autopsy. Adrenocortical failure was also present. The course was characterized by frequent infections and an episode of myocarditis. The boy died at the age of 6.0 years after rapid neurological deterioration accompanied by renal insufficiency. Autopsy disclosed cerebral leukodystrophy compatible with Cockayne syndrome.