Regina Zimmermann

Mutational analysis of the N-ras,p53,p16INK4a,CDK4, and MC1R genes in human congenital melanocytic naevi

Eighteen human congenital melanocytic naevi (CMN) from 17 patients were screened for activating point mutations in the oncogenesN-ras andCDK4 and for sequence variants in the MC1R gene by combined RFLP-PCR/SSCP analysis. In addition, all lesions were screened for deletions and point mutations in the tumour suppressor genesp53 andp16INK4a (CDKN2A) by combined multiplex PCR/SSCP analysis. Positive screening data were specified by sequencing of the corresponding PCR product. Activating point mutations in theN-ras gene (nine CAA (Gln) to AAA (Lys) transversions and one CAA (Gln) to CGA (Arg) transition at codon 61) were detected at high frequency (56%). Furthermore, three missense mutations (V92M) and two silent mutations (CGA (Arg) to CGG (Arg), codon 213, exon 6) were found in theMC1R andp53 genes, respectively. No mutations were found in p16 orCDK4. The activatedN-ras oncogene, which is also found in human cutaneous melanomas, may constitute a potential risk factor for melanoma formation within CMN.

Prostaglandin-H-synthase isozyme expression in normal and neoplastic human skin

Expression of prostaglandin‐H‐synthase (PGHS) isozymes was analyzed in 50 biopsies of normal human skin and of pre‐malignant and malignant skin lesions, by means of quantitative RT‐PCR, immunoprecipitation and Western blotting, as well as immunohistochemistry. Normal skin constitutively expressed PGHS‐1 in all cell layers of the epidermis, in endothelial cells of small blood vessels and in sweat‐gland epithelium. PGHS‐2 expression was very low and restricted to a few keratinocytes of the interfollicular and follicular epidermis. Steady‐state concentrations of PGHS‐1 and PGHS‐2 mRNA were similar in normal skin and in basal‐cell carcinomas, but PGHS‐1 mRNA was reduced and PGHS‐2 mRNA was elevated in actinic keratoses, squamous‐cell carcinomas and keratoacanthomas. PGHS‐1 protein was detected in all tumor biopsies, being occasionally increased in basal‐cell carcinomas. High amounts of PGHS‐2 protein were found in actinic keratoses, squamous‐cell carcinomas and keratoacanthomas, but not in basal‐cell carcinomas. Four malignant melanomas included in this study contained PGHS‐1 but no PGHS‐2 protein. Immunohistochemical analysis of the biopsies identified keratinocytes, in addition to cells of inflammatory infiltrates and of dendritic morphology, as the major PGHS‐expressing cell types. PGHS‐2‐specific signals were spread throughout the epidermal part of actinic keratoses and squamous‐cell carcinomas. These data suggest that constitutive up‐regulation of PGHS‐2 expression is a consistent pre‐malignant event in squamous‐cell cancer development in man, as it is in animal models of skin carcinogenesis. Thus, pre‐cancerous lesions such as actinic keratoses present a likely target for chemoprevention of skin cancer by selective PGHS‐2 inhibitors. Int. J. Cancer 82:648–656, 1999.

Mutational analysis of the BRAF gene in human congenital and dysplastic melanocytic naevi.

Eighteen congenital melanocytic naevi (CMN) from 17 patients and 18 dysplastic melanocytic naevi (DMN) from 18 patients were screened for mutations in the BRAF oncogene (present study) and the N-ras oncogene (in the course of two foregoing studies) by single-strand conformational polymorphism (SSCP)/sequencing analysis. BRAF mutations were demonstrated in both types of lesion. As a whole, 17 of 18 CMN (94.4%) and five of 18 DMN (27.7%) harboured either BRAF or N-ras mutations. As the BRAF oncogene is frequently found to be mutated in human cutaneous melanomas, it may constitute a risk factor for melanoma formation within CMN and DMN.

Mutational analysis of N-ras , p53 , CDKN2A ( p16 INK4a ), p14 ARF , CDK4 , and MC1R genes in human dysplastic melanocytic naevi

In order to detect possible dysplastic melanocytic naevi (DMN) associated melanoma risk factors and lesion specific differences in the mutation spectrum of dysplastic and congenital melanocytic naevi (CMN), we screened 19 specimens of human sporadic DMN derived from 19 patients for the presence of mutations in five genes, which we had investigated in a former study in 19 CMN1 and which have been reported to be associated with human cutaneous melanoma ( N-ras ,2 p53 ,3 CDKN2A ,4 CDK4 ,5 and MC1R 6,7). DNA was extracted from selected paraffin embedded DMN resection specimens using the QIAamp DNA Mini Kit (Qiagen) according to the recommendations of the supplier. The relative number of atypical melanocytes in the DMN and the histological subtype of the DMN were determined in parallel slides by an experienced dermatologist (Dr Regina Zimmermann) (table 1). View this table: Table 1 N-ras mutations and MC1R variants in human spontaneous dysplastic naevi The screening strategy for the detection of activating point mutations in the oncogenes N-ras and CDK4 as well as for germline sequence variants in the MC1R gene by combined RFLP-PCR/SSCP analysis, and the screening strategy for the detection of homozygous deletions and point mutations in the tumour suppressor genes p53 and CDKN2A by combined multiplex-PCR/SSCP analysis, have been described previously.1 In order to …

Detection of Silver Sulfide Deposits in the Skin of Patients with Argyria After Long-term Use of Silver-containing Drugs

Five patients with generalized slate-gray discoloration of the skin have been diagnosed histologically as argyria in the last 35 years in the Department of Dermatology and Venereology of Rostock and Halle. Light microscopically, there was visible black pigmentation in histiocytes, fibroblasts, and multinucleated giant cells of the dermis. In the transmission electron microscope (TEM), the authors observed electron-dense deposits inside lysosomes and residual bodies of phagocytes as well as outside the cells in the connective matrix. These deposits were identified by elemental analysis in TEM and electron energy loss spectroscopy (EELS) as well as scanning electron microscope (SEM) and energy dispersive x-ray analysis (EDX) containing silver and sulfur. Therefore, they seem to consist of silver sulfide. Argyria is of low medical relevance and is very rarely induced because of silver-containing drugs. Nevertheless, there are still a lot of silver products on the market, easily available over-the-counter. Therefore, argyria should not be forgotten or missed in the diagnostics of human dermis.

Successful Treatment of Nodular Actinic Reticuloid with Tacrolimus Ointment

Actinic reticuloid (AR) is the severest clinical variant of chronic actinic dermatitis (CAD) and describes a persistent photoinduced skin disorder often associated with delayed-type hypersensitivity reactions. Histopathologically, AR resembles pseudolymphoma and may also show features of cutaneous lymphoma. In contrast to other UV-induced skin diseases, AR patients show persistent photosensitivity to UV radiation and visible light parallel to permanent changes of skin texture with infiltrated papules and thickened plaques. Therapeutically, CAD is a problematic condition. Daily application of sun protection has to be combined with local or systemic immunosuppression. However, various therapeutic approaches including systemic corticosteroids and other systemic immunosuppressive agents are limited by severe side effects. Tacrolimus, a Streptomyces-derived immunosuppressive macrolide antibiotic and inhibitor of calcineurin, has been proven successful in vari ous inflammatory skin diseases including atopic eczema, allergic contact dermatitis and photodermatoses without undesired side effects. We present a case of severe recalcitrant and nodular chronic actinic dermatitis responding to topically applied tacrolimus (Protopic®). To avoid further relapses we continued this therapy twice a day over a period of 2 years.

Detection of gold particles in the neck skin after lightning stroke with evaporation of an ornamental chain

A German couple was struck by lightning. Both patients survived this event. Whereas the husband was unconscious for only a few minutes, his wife fell into coma for 24 h. The lightning stroke entered the body of the woman behind the left ear and left it at the left shoe. The stroke caused a partial evaporation of a gold ornamental chain on the neck, resulting in a tattoo of the neck skin. A biopsy of the skin 6 months after the event showed the accumulation of gold particles of different size in the dermis down to the subcutaneous fatty tissue. In semithin sections, histiocytes, multinucleated foreign giant cells, and fibroblasts were visible with uptaken metallic particles. In transmission electron microscopy, gold globules of up to 30 µm in diameter were visible outside the cells in the collageneous matrix of the connective tissue besides smaller metallic particles up to 5 nm inside lysosomes and residual bodies of phagocytic cells. Four different kinds of gold particles could be differentiated: globules, granular irregular particles, tubules, and tanglelike tracks. In scanning electron microscopy, gold particles were demonstrated by backscatter detection in the connective tissue of subcutis, where the EDX elemental analysis showed strong signals of aurum (Au), copper (Cu), and argentum (Ag). The detected metals were quantified by AAS as 70% gold, 21% silver, and 9% copper, which demonstrates the composition of gold alloy of the neck chain of the patient. Tanglelike tracks and elongated gold deposits represent crystals of gold salts, as detected by electron diffraction and polarization microscopy. Attempts to remove the gold particles from the skin to remove the tattoo should not be undertaken because the gold is deep and widespread.

Treatment of severe erythrodermic acute graft‐versus‐host disease with photochemotherapy

Sir, graft-versus-host disease (GVHD) is a frequent complication occurring after allogenic bone marrow transplantation. Skin changes may be divided into acute and chronic GVHD, both displaying a particular pattern. Acute GVHD may develop into generalized erythroderma and even further into skin necrosis and epidermal blistering. Photochemotherapy (PUVA) is highly effective for the treatment of patients with acute GVHD. However, since the overall prognosis of patients with severe manifestations of GVHD is poor, treatment of these patients with PUVA is still challenging. It is further complicated by severe immunosuppression. We report here on the successful treatment of a patient suffering from overall grade IV GVHD with PUVA therapy. A 34-year-old male patient had received allogenic bone marrow transplantation due to myeloid leukaemia. Despite GVHD prophylaxis with cyclosporin and methotrexate, the patient developed severe mucositis after several days. After a further 3±4 weeks, GVHD of the liver developed which required additional systemic glucocorticoids (prednisolone 5 mg kg body weight daily). Two weeks later the patient developed a rapidly progressing erythematous rash, which finally led to a generalized erythroderma (Fig. 1). Histopathological examination revealed an acanthotic epidermis with many dyskeratotic cells and scattered inflammatory lymphocytes (Fig. 2). The basal epidermal layer showed extensive vacuolization, and the dermal compartment a marked oedema and a moderate inflammatory infiltrate of lymphocytes accompanied by erythrocyte extravasation. These findings were characteristic of acute cutaneous GVHD. The prednisolone dosage was increased and the immunosuppressive agent mycophenolate mofetil (2 g daily) was added. Because skin lesions did not respond to this treatment regimen, oral PUVA therapy was started. 8-methoxypsoralen (0.6 mg kg body weight) was administered 2 h before UVA exposure. After 3 weeks of treatment the skin lesions had dramatically improved. PUVA therapy was stopped for 2 weeks due to bone marrow re-transplantation. After that, PUVA therapy was continued with local application of 8-methoxypsoralen cream combined with UVA irradiation. Skin lesions cleared completely after a further 3 weeks of treatment and an overall dosage of 12 J cm (Fig. 3). Histopathological evaluation of skin biopsy showed a flattened epidermal layer and slight fibrosis in the dermal compartment, features which might indicate early chronic GVHD. Although a series of reports on PUVA treatment of acute GVHD have been published, treatment of more severe cases is less common. In the present report the severe skin involvement of an overall grade IV GVHD did not respond to a combination of three different high-dose immunosuppressive agents (prednisolone, cyclosporin and mycophenolate