Growth and characterization of a cell line from a human primary neuroendocrine carcinoma of the skin (Merkel cell carcinoma) in culture and as xenograft.

The primary neuroendocrine carcinoma of the skin or Merkel cell carcinoma (MCC) is a skin tumor with aggressive biological behaviour. Experimental models for investigating the biological properties of the tumor are prerequisite for developing new therapeutic approaches. In this study, we report the establishment and characterisation of a cell line derived from the lymph-node metastasis of a patient with highly aggressive MCC. Merkel carcinoma cells (MCC-1) grew as floating aggregates in suspension cultures for more than two years and over 70 subcultures. The proliferation rate in suspension cultures was rather moderate with a population doubling time of 69 h. The immunocytochemical pattern of the cultured MCC-1 was similar to that of the original tumor with expression of cytokeratin 18, neuron-specific enolase, neurofilaments, and synaptophysin. In addition, reverse transcriptase polymerase chain reaction (RT-PCR) revealed presence of chromogranin A mRNA in the MCC-1 cell line. Furthermore, electron microscopy yielded the rare finding of neuroendocrine granules in the cytoplasm of the cultured cells. The cell line MCC-1 was able to form colonies in soft agar. Nude mice developed solid tumors with similar histology to the original tumor after subcutaneous and intravenous injections of cultured MCC-1, and malignant ascites was seen after intraperitoneal injection. Also, two MCC-1 sublines were established by reculturing cells from the xenografts grown in vivo and immunocytochemistry confirmed their neuroendocrine origin. The MCC-1 line may thus serve as a model for studying the biology and the metastatic potential of Merkel cell carcinoma.

[Granulomatosis disciformis Miescher]. / Granulomatosis disciformis Miescher.

We describe granulomatosis disciformis Miescher in a 57-year old female patient with typical clinical manifestation: disc-shaped plaques with slight central sclerosis and lesions resembling necrobiosis lipoidica and granuloma anulare. Histology revealed an infiltrate mainly composed of macrophages and lymphocytes and small foci of necrobiotic collagen and of new collagen formation. We review the literature on granulomatosis disciformis and nosology, concluding--as do most authors--that granulomatosis disciformis is a characteristic variant within the spectrum of cutaneous noninfectious granulomatous disease lying in between generalized granuloma anulare and necrobiosis lipoidica.

Monitoring of isotretinoin therapy by measuring the plasma levels of isotretinoin and 4-oxo-isotretinoin. A useful tool for management of severe acne.

BACKGROUND: Isotretinoin for oral therapy in severe acne conglobata and acne nodulocystica represents a significant achievement; however, the drug exerts several mucocutaneous and systemic adverse effects, besides its teratogenic potency. OBJECTIVE: The aim of this study was to investigate the plasma levels of isotretinoin and of 4-oxo-isotretinoin over long-term treatment of severe acne and to assess any correlation with the given dose, the clinical improvement and the occurrence of side effects. METHODS: Forty-one patients with severe acne and acne-related disorders were studied under long-term oral intake of isotretinoin. Therapeutic effects and side effects were evaluated prior, during and at the end of therapy. The plasma levels of isotretinoin and of its major metabolite 4-oxo-isotretinoin were measured by reversed-phase HPLC and were correlated with the administered oral dose and the number and frequency of side effects. RESULTS: Dose-dependent plasma levels of isotretinoin and its metabolite were observed. At a mean dosage of 0.75-1.0 mg/kg/day, 404 +/- 142 ng/ml were measured, whereas the plasma levels of 4-oxo-isotretinoin were 1-2x higher. The plasma levels correlated well with the orally administered dose of isotretinoin and the observed mucocutaneous side effects. CONCLUSION: The study demonstrates that measuring of the plasma levels may be a helpful tool to monitor the individual therapeutic dose regimen in patients with severe acne in order to minimize undesired side effects and to control oral intake.

[Psoriatic onycho-pachydermo-periostitis (POPP)]. / Psoriatische Onycho-Pachydermo-Periostitis (POPP).

We describe a patient with painful enlargement and severe nail deformity of both great toes. The patient furthermore presented with some minor diagnostic criteria for psoriasis such as nail pitting and had a positive family history of the disease. In the recent literature, such cases have been recognized as a new entity termed psoriatic onycho-pachydermo-periostitis (POPP). In this case report, POPP is discussed in relationship to other forms of psoriatic arthropathy and with special emphasis regarding its diagnostic criteria and therapeutic implications.

Merkel cell carcinoma: report of ten cases with emphasis on clinical course, treatment, and in vitro drug sensitivity.

BACKGROUND: Merkel cell carcinoma (MCC) is an uncommon primary neuroendocrine skin tumor most often seen in the elderly. The clinical course varies. Treatment is controversial and few data on drug sensitivity are available. OBJECTIVE: We evaluated the clinical course and treatment of 10 MCC patients and determined MCC chemosensitivity. METHODS: Clinical records as well as laboratory and histopathologic data from 10 patients with MCC treated in our department were examined. Chemosensitivity to various chemotherapeutic agents and interferons of MCC cells from four patients was determined in a soft agar clonogenic assay. RESULTS: MCC behaved as an aggressive tumor with early and frequent local relapses (4 of 10 patients at a 2.2-month average), regional (4 of 10 patients at 2.5 months), and distant metastases (5 of 10 patients 9.6 months after excision of the primary tumor). In all but one patient, regional metastases preceded distant ones. Metastatic spread was associated with an average survival of 21 months from the initial diagnosis. Long-term survival (53+ and 65+ months) was observed in two women. Wide excision of the primary tumor, alone or combined with adjuvant chemotherapy and radiotherapy, was the most effective treatment. In advanced disease, chemotherapy and radiotherapy were not able to induce long-term remission. In vitro assays for MCC drug sensitivity revealed cisplatin, doxorubicin, and vindesine to be the most active. CONCLUSION: MCC has a poor prognosis in advanced stages; therefore the primary tumor should be aggressively treated. The in vitro clonogenic assay may help to identify the chemosensitivity profile of MCC and to optimize chemotherapy protocols.

Current use and future potential role of retinoids in dermatology.

Since their introduction 15 years ago, retinoids have been increasingly used for topical and systemic treatment of psoriasis and other hyperkeratotic and parakeratotic skin disorders, keratotic genodermatoses, severe acne and acne-related dermatoses, and also for therapy and/or chemoprevention of skin cancer and other neoplasia. Oxidative metabolites of vitamin A (retinol) are natural retinoids present at low levels in the peripheral blood. Synthetic retinoids are classified into 3 generations including nonaromatic, monoaromatic and polyaromatic compounds. They are detectable in plasma 30-60 minutes after systemic administration, and reach maximum concentrations 2 to 4 hours later. Elimination half-life is 10 to 20 hours for isotretinoin, 80 to 175 days for etretinate and 2 to 4 days for, trans-acitretin; the latter, however, partially converts into etretinate. Retinoid concentrations in skin are rather low in contrast to subcutaneous fat tissue. Intracellularly, retinoids interact with cytosolic proteins and specific nuclear receptors. Two classes of nuclear receptors have been suggested to mediate retinoid activity at the molecular level, RARs and RXRs. The expression of retinoid receptors is tissue specific; skin mainly espresses RAR gamma and RXR alpha. Retinoids affect epidermal cell growth and differentiation as well as sebaceous gland activity and exhibit immunomodulatory and anti-inflammatory properties. Current retinoid research targets the development of receptor-selective retinoids for tailoring and/or improving their therapeutic profile. Currently, tretinoin is used systemically for acute promyelocytic leukaemia, etretinate and acitretin for psoriasis and related disorders, as well as other disorders of keratinisation and isotretinoin for seborrhoea, severe acne, rosacea and acneiform dermatoses. Systemic retinoids are also applied for chemoprevention of epithelial skin cancer and cutaneous T cell lymphoma. The major adverse effect of retinoids is teratogenicity; all other adverse effects are dose-dependent and controllable. Contraception is, therefore, essential during retinoid treatment in women of child-bearing age. Clinical monitoring requires physical examination for adverse effects every 3 to 4 weeks and proper laboratory investigations, also including analysis of retinoid bioavailability in selected cases. Topical retinoids are rapidly developing at present and seem promising for the future; their clinical application includes acne, aging, photodamage, precanceroses, skin cancer and disorders of skin pigmentation. The development of receptor-specific retinoids for topical treatment of psoriasis and/or acne may lead to interesting new compounds based on our current concepts of retinoid function.

The phospholipid analogue hexadecylphosphocholine (HePC) inhibits proliferation of keloid fibroblasts in vitro and modulates their fibronectin and integrin synthesis.

Hexadecylphosphocholine (HePC), a topically effective compound, exerts a strong antiproliferative effect on neoplastic cells. In the present study we investigated (1) the antiproliferative effect of HePC on benign mesenchymal cells in vitro, using as examples normal and keloid fibroblasts, and (2) the influence of HePC on various functional properties of these cells, including phosphatidylcholine biosynthesis, their capacity to reorganize a three-dimensional collagen-I matrix, and their expression and synthesis of fibronectin and subunits of the beta 1 integrin family. Fibroblasts derived from keloids (kelfib) and from normal skin (nfib) were cultured in serum-containing medium and treated in the third passage with 50 mumol/l HePC. Proliferative activity was significantly (P < 0.05) more strongly inhibited in kelfib than in nfib under HePC treatment, whereas their phosphatidylcholine synthesis was inhibited to a similar extent. However, the ability of fibroblasts to contract a three-dimensional collagen-I lattice was significantly (P < 0.05) enhanced only in kelfib treated with HePC. These functional differences following treatment with HePC were paralleled by an upregulation of the alpha 2- and beta 1-integrin chains, and a downregulation of fibronectin synthesis and the alpha 5-subunit. Our results indicate a differential modulation of kelfib metabolism by HePC, suggesting a possible new therapeutic approach for keloid and hypertrophic scars in vivo.

Inhibition of plasma membrane NADH oxidase activity and growth of HeLa cells by natural and synthetic retinoids.

Several retinoids, both natural and synthetic, were evaluated for their ability to modulate NADH oxidase activity of plasma membranes of cultured HeLa cells and the growth of HeLa cells in culture. Both NADH oxidase activity and the growth of cells were inhibited by the naturally-occurring retinoids all trans-retinoic acid (tretinoin) and retinol as well as by the synthetic retinoids, trans-acitretin, 13-cis-acitretin, etretinate and arotonoid ethylester (Ro 13-6298). For all retinoids tested, inhibition of NADH oxidase activity and inhibition of growth were correlated closely. With tretinoin, etretinate and arotonoid ethylester, NADH oxidase activity and cell growth were inhibited in parallel in proportion to the logarithm of retinoid concentration over the range of concentrations 10(-8) to 10(-5) M. Approximately 70% inhibition of both NADH oxidase activity and growth was reached at 10 microM. With retinol, trans-acitretin and 13-cis-acitretin, inhibition of NADH oxidase activity and growth also were correlated but maximum inhibition of both was about 40% at 10 microM. The possibility is suggested that inhibition of the plasma membrane NADH oxidase activity by retinoids may be related to their mechanism of inhibition of growth of HeLa cells in culture.

Cultured dermal papilla cells of the rat vibrissa follicle. Proliferative activity, adhesion properties and reorganization of the extracellular matrix in vitro.

The dermal papilla of the mammalian hair follicle plays an important role in regulating and controlling the hair cycle. Distinct functional stages of dermal papilla cells (DPC) are involved in this process, thus suggesting that the dermal papilla is a highly specialized suborgan of the pilosebaceous unit. The aim of the present study was to investigate the functional properties of cultured DPC in various assays and to compare their functional properties with those of dermal fibroblasts (DFB). In monolayer cell cultures DPC showed an aggregative growth pattern, different to that of DFB, and lower proliferation rates, as compared to the controls. Adhesion assays performed using a 51[Cr]labeling method showed strong adhesion of both cell populations to collagen types I and IV, fibronectin and laminin, but DPC in vitro showed significantly higher adhesiveness to collagen type IV, a major component of the basement membrane of dermal papillae in vivo. The capacity of DPC to reorganize extracellular matrix components, as measured by gel contraction with three-dimensional collagen type I lattices, proved to be significantly lower than that of DFB and, moreover, DPC lysed the collagen lattices completely after 48 h in culture. The functional differences between DPC and DFB were paralleled by higher surface expression and synthesis levels of the beta 1, alpha 1, and alpha 5 chains of integrin adhesion receptors in DPC, as detected by fluorescence-activated cell-sorter analysis and radioimmunoprecipitation. These findings provide evidence that DPC are a highly specialized cell population, which clearly differs from another mesenchymal cell type, DFB. After their isolation and cultivation in vitro, DPC still preserve functional properties related to important steps of cell-matrix interaction involved in the hair cycle.

[Trans-acitretin is metabolized back to etretinate. Importance for oral retinoid therapy]. / Trans-Acitretin wird in Etretinat rückmetabolisiert. Bedeutung für die orale Retinoidtherapie.

The oral retinoid etretinate (Tigason) has recently been replaced by trans-acitretin (Neotigason) in the treatment of severe psoriasis, primarily because of its short half-life and the assumption that a shorter period of subsequent contraception would be required. After introduction of the new drug, however, circulating quantities of etretinate were detected in patients treated with trans-acitretin. Thus far, re-esterification has been detected in at least 83 cases. The 2-month period of strict contraception initially recommended after oral intake of trans-acitretin has been extended to 2 years, as with etretinate. We review some aspects of trans-acitretin metabolism, with special emphasis on etretinate formation. Re-esterification of trans-acitretin into etretinate takes place under varying conditions in volunteers and patients, as well as in animal models. Ethanol is a co-factor for the enzymatic re-esterification of trans-acitretin. It is unclear whether the introduction of trans-acitretin has been of any significant benefit to patients. Monitoring of plasma retinoid levels during and after retinoid therapy remains of decisive importance in managing difficult cases and or in approving decisions for pregnancy.

[Contact allergies to corticosteroids. Prevalence, cross-reactions and methods of detection]. / Kontaktallergien auf Kortikosteroide. Prävalenz, Kreuzsensibilisierung und Nachweismöglichkeiten.

Topical corticosteroids (CS) are frequently used to treat of allergic contact dermatitis, so that their own potential for sensitization is rarely detected. However, increasing numbers of hypersensitivity reactions to corticosteroids have been reported during the last 10 years, and allergic skin reactions to over 50 particular compounds have been observed. CS must now be regarded as a rather frequent contact allergen, the prevalence being twice as high in women as in men. Contact allergy to CS is clinically characterized by a chronic and persistent, corticosteroid-resistant dermatitis. In general, hypersensitivity to CS is a type I and/or type IV mediated allergic reaction. Cross-sensitivity between individual corticosteroid preparations has been described owing to their closely related chemical structures (e.g. hydrocortisone and tixocortol pivalate). Contact allergy to CS can be confirmed by patch-tests, intradermal injection tests or application tests; because of the delayed appearance of positive skin reactions a prolonged evaluation period of up to 120 h is recommended. For routine screening the use of the patient's own material and of tixocortol pivalate and budesonide as test marker substances seems appropriate. The development of further marker substances is a target for ongoing research.

[Transformation of multiple basaliomas to squamous cell carcinomas in an HIV patient with Gorlin-Goltz syndrome]. / Ubergang multipler Basaliome in Plattenepithelkarzinome bei einem HIV-Patienten mit Gorlin-Goltz-Syndrom.

We report on a 32-year-old male patient with Gorlin-Goltz syndrome, who presented with excessive numbers of superinfected basal cell carcinoma. This led us to suspect an underlying HIV infection, which was confirmed by ELISA and Western blotting. Laboratory investigation of the immunological state revealed severe immunosuppression with 267 CD4+ lymphocytes and a CD4/CD8 ratio of 0.3. The histological picture showed multiple basal cell carcinomas, some of them transforming into squamous cell carcinomas. We suspect that the excessive number and the unusual clinical and histological picture of the basal cell carcinomas in this patient were probably influenced by the underlying HIV infection.

Tumors of the pilosebaceous unit.

Appendage tumors of the pilosebaceous apparatus are relatively rare, and their clinical aspect is usually nonspecific. In most cases pilosebaceous tumors are benign; however, malignant neoplasias (e.g. trichilemmal carcinoma, malignant pilomatricoma) have been described. In the present review we propose the classification of pilosebaceous tumors by degree and by direction of differentiation and by histological configuration. Three different histological patterns have to be distinguished: tumors with central dilated hair follicle, tumors consisting of epithelial islands and keratotic cysts in the corium and neoplasias with predominantly mesenchymal components surrounding epithelial formations. Careful histopathological examination of pilar appendage tumors will help to classify the wide range of clinically and histologically different tumors and to establish the exact diagnosis.