Prognosis of primary melanoma.

MATERIALS AND METHODS: To investigate the prognosis of primary melanoma, we studied a Finnish population of 298 primary melanoma patients, the majority with stage I or II tumours. The median clinical follow-up (4.8 years) was acquired from the patients' records, and the overall survival thereafter was collected from patient registries. The median follow-up for overall survival was 9.5 years. RESULTS: The overall survival rate was 66.8%. 24.5% developed metastasis, 17.8% died of melanoma, and 15.4% died of some other cause. Surgical margins had no effect on survival. In univariate analysis the most significant prognostic factors for disease-free and overall survival were stage of tumour (p < 0.0001), thickness of tumour (p < 0.0001), depth of tumour invasion (p < 0.0001) and tumour ulceration (p = 0.0005, p < 0.0002). Ulceration was an unfavorable prognostic marker. Younger patients had better survival outcomes than older ones (p = 0.04). Accordingly, in the multivariate Cox model the independent prognostic factors for both disease-free and overall survival were stage of tumour and thickness of tumour. Tumour location on trunk was an independent adverse prognosticator for overall survival. CONCLUSION: We conclude that the prognosis of primary melanoma has improved in Finland in the last decades being in line with a global tendency.

Metalloelastase (MMP-12) and 92-kDa gelatinase (MMP-9) as well as their inhibitors, TIMP-1 and -3, are expressed in psoriatic lesions.

In skin biology, matrix metalloproteinases (MMPs) have been implicated in inflammatory matrix remodeling, neovascularization, wound healing and malignant transformation. Psoriasis is histologically characterized by keratinocyte hyperproliferation, infiltration of inflammatory cells, neoangiogenesis and production of cytokines, such as TNF-alpha, IL-1beta, TGF-alpha, and IFN-gamma, also capable of regulating MMP transcription. To investigate the role of stromelysins-1 and -2, matrilysin, metalloelastase, collagenases-1 and -3 and 92-kDa gelatinase as well as their inhibitors, TIMPs-1 and -3, in psoriasis, we performed in situ hybridization using 35S-labeled cRNA probes on 29 psoriatic lesions and 9 samples of normal looking skin from psoriatic patients. Metalloelastase mRNA was detected in 21/27 samples in macrophages that had migrated into the epidermis or in the inflammatory infiltrates of the superficial dermis. A quantity of 92-kDa gelatinase was found in macrophages and neutrophils (25/27). Stromelysin-1 mRNA was detected in basal keratinocytes in 4/21 lesions. Intracellular laminin-5 immunosignal in basal keratinocytes of the same samples, suggested that stromelysin-1 might participate in remodeling of the basement membrane zone. No signal for stromelysin-2 or collagenase-3 was found and only sweat glands were positive for matrilysin. TIMP-1 was more abundantly expressed than TIMP-3 in the inflammatory infiltrates and endothelial cells of dermal papillae (22/29). TIMP-3 was expressed perivascularly in 9/16 samples. Our results suggest that overexpression of the investigated MMPs by keratinocytes is not associated with psoriasis. However, macrophages express MMPs in psoriatic skin. Also TIMPs, particularly TIMP-1, were abundantly expressed, suggesting that mere MMP overexpression is unlikely to contribute to psoriatic tissue changes.

Vascular changes in erythropoietic protoporphyria: histopathologic and immunohistochemical study.

BACKGROUND: Erythropoietic protoporphyria (EPP) is an inherited disease caused by deficient activity of ferrochelatase in the heme biosynthetic pathway. Accumulation of protoporphyrins and light exposure results in acute phototoxic skin reactions. The histopathologic findings of the light-exposed skin are thickening of the superficial dermal vessel walls and amorphous deposits around the vessels, but the origin and detailed composition of the perivascular material have been unclear. OBJECTIVE: The vascular morphology and composition of the perivascular material were studied in the skin samples of patients with EPP. METHODS: Skin biopsy specimens of 8 patients with EPP representing 7 Finnish EPP families with different genotypes were studied by means of light and electron microscopy and immunohistochemical methods. RESULTS: The characteristic finding was thickened, periodic acid-Schiff-positive vessel walls caused by concentric reduplication of basal lamina and excess of fine granular material at the basal membrane zone in the superficial dermis. The perivascular deposits in the vicinity of vessel walls had a homogeneous or fine granular appearance without filaments. Direct immunofluorescence showed constant IgG deposits together with IgA, IgM, and C3 in the vessel walls. In immunohistochemistry, collagen IV and laminin could be demonstrated at the vascular basal membrane together with serum amyloid P protein, kappa and lambda light chains, and a 90-kd glycoprotein. CONCLUSION: The vascular involvement indicates that the blood vessel walls in the papillary dermis are the primary tissues affected during an acute photoreaction. The repeated acute damage and repair processes in the basement membrane zone result in thickening of the vessel walls. Perivascular deposits are a secondary and irreversible phenomenon resulting from the leakage and accumulation of different serum components. These changes were not found in the nonexposed skin, indicating that an increased level of erythrocyte protoporphyrin per se is not responsible for the cutaneous manifestations, but the interaction of solar radiation is mandatory. Amorphous deposits distinguish EPP from variegate porphyria and porphyria cutanea tarda; a histopathologic examination may be a helpful tool in differentiating porphyric and nonporphyric photosensitivity.

Cryptococcosis during systemic glucocorticosteroid treatment.

Cryptococcosis is an opportunistic infection caused by a fungus, Cryptococcus neoformans. It is usually seen in immunocompromised patients with AIDS, leukaemia, lymphoma, sarcoidosis or immunosuppressive treatments. We describe a patient who was treated with systemic glucocorticosteroids for 4 years because of lung sarcoidosis. During the last year of treatment, a papular eruption developed which later became ulcerative. In a histopathological examination of a skin biopsy, there was granulomatous inflammation, and the disease was treated as sarcoidosis without success. After 1 year's unsuccessful treatment, another skin biopsy and skin fungal culture revealed C. neoformans. Cryptococcal antigen was found in blood and cerebrospinal fluid, too. The patient was successfully treated first with an amphotericin-B-flucytosine combination and later with fluconazole.

Prognostic value of tumour vascularity in primary melanoma.

To investigate the prognostic value of tumour vascularity we studied 84 patients with primary melanomas ranging in tumour thickness (Breslow) from 0.37 to 7 mm and in depth of tumour infiltration (Clark) from II to V. Vascularization was assessed by immunohistochemistry with a CD-31 antibody recognizing endothelial cells. The CD-31-positive vessels were counted and the degree of vascularization was correlated with the survival of the patients. In addition, the relationship between blood vessel density and some histopathological data is discussed. In our study, the multivariate Cox model showed that the only independent variable in disease-free survival was tumour thickness (Breslow classification) and the only one in overall survival was depth of tumour infiltration (Clark classification). In disease-free survival, tumour thickness (Breslow classification) was a clear prognostic factor (P = 0.004) after 4 years' follow-up, as were depth of tumour infiltration (Clark classification) (P = 0.04) and ulceration (P = 0.04). In overall survival, tumour vascularity was the strongest prognostic factor at 4 years, high vascularity being associated with a good prognosis (P = 0.06). Clark classification was also a prognostic factor (P = 0.02) in overall survival. We conclude that high vascularization is associated with a better prognosis but is not an independent prognostic indicator.

Hemidesmosomal molecular changes in dermatitis herpetiformis; decreased expression of BP230 and plectin/HD1 in uninvolved skin.

Recent BP230-knockout experiments with subsequent blistering and recently identified plectin/HD1 mutations in epidermolysis bullosa simplex patients suggest that defective expression of BP230 and plectin/HD1 may predispose to blister formation in human skin. We have studied the expression of the epithelial adhesion complex as well as the basement membrane and anchoring fibril antigens in uninvolved dermatitis herpetiformis skin to find out if alterations can be detected in these structures predisposing to the blister formation typical of the disease. Ten uninvolved dermatitis herpetiformis skin specimens, which all showed clear granular deposits of IgA under the basement membrane in direct immunofluorescence and five normal skin specimens, were studied by indirect immunofluorescence technique. Six uninvolved dermatitis herpetiformis skin specimens showed distinctly decreased immunoreaction for BP230 and four uninvolved dermatitis herpetiformis skin specimens showed distinctly decreased immunoreaction for plectin/HD1. All five skin controls showed strong immunoreactions for BP230 and plectin/HD1. Other hemidesmosomal proteins including BP180 and integrin alpha6beta4, as well as basement membrane proteins laminin-5, laminin-1, nidogen and type IV collagen, and the anchoring fibril protein type VII collagen showed a normal strong expression. Our results suggest that alterations in BP230 and plectin/HD1 may contribute or predispose to blister formation in dermatitis herpetiformis skin.

Expression of collagenases-1 and -3 and their inhibitors TIMP-1 and -3 correlates with the level of invasion in malignant melanomas.

Since proteolysis of the dermal collagenous matrix and basement membranes is required for local invasive growth and early metastasis formation of cutaneous melanomas, we have analysed the activities/expression levels of certain metalloproteinases in melanomas and cultured melanoma cells by in situ hybridization and Northern analysis. In addition to collagenases-1 and -3 that have been implicated in invasive growth behaviour of various malignant tumours, we analysed the levels of 72-kDa gelatinase and its activators MT1-MMP and TIMP-2 in cultured melanoma cells. The lesions examined included three cases of lentigo maligna and 28 cases of Clark grade I-V melanomas. The premalignant as well as the grade I tumours were consistently negative for collagenase-1 and -3 and TIMP-1 and -3. The collagenases were predominantly expressed in the cancer cells of Clark grade III and IV tumours. TIMP-1 and -3 were abundantly expressed in the cancer and/or stromal cells of grade III and IV melanomas, while TIMP-2 protein was detected also in melanomas representing lower invasive potential. Northern analysis of seven melanoma cell lines showed that the expression of collagenase-1 and TIMPs-1 and -3 was associated with 72-kDa gelatinase positivity. All melanoma cell lines were positive for MT1-MMP and TIMP-2 mRNAs. Our results suggest that overexpression of collagenases-1 and -3 and TIMPs-1 and -3 is induced during melanoma progression. Expression of TIMPs may reflect host response to tumour invasion in an effort to control MMP activity and preserve extracellular matrix integrity.

Enhanced expression of human metalloelastase (MMP-12) in cutaneous granulomas and macrophage migration.

Accumulation of inflammatory cells such as macrophages may lead to degeneration of connective tissue matrix in various skin diseases. Macrophage metalloelastase, is a matrix metalloproteinase (MMP-12) capable of degrading elastin as well as various basement membrane components. To investigate the role of human macrophage metalloelastase in skin, we assessed by in situ hybridization and immunohistochemistry 66 specimens representing skin diseases characterized either by changes in elastic fibers or by pronounced infiltrations of extravasating and migrating macrophages. CD68 immunostaining was performed to identify the human macrophage metalloelastase-positive cells and Weigert's Resorcin-Fuchsin staining to reveal the status of elastic fibers. We found abundant expression of human macrophage metalloelastase mRNA in macrophages in areas devoid of normal elastic fibers in granulomatous skin diseases sarcoidosis, necrobiosis lipoidica diabeticorum, and granuloma annulare. Positive cells for human macrophage metalloelastase protein could be detected in the same regions as well as positive immunostaining for urokinase plasminogen activator. Of the other matrix metalloproteinases capable of degrading elastin, 92 kDa gelatinase colocalized with human macrophage metalloelastase, while 72 kDa gelatinase was produced by surrounding fibroblast-like cells. Furthermore, human macrophage metalloelastase was expressed by macrophages in areas with disrupted basement membrane, as assessed by type IV collagen staining, in pityriasis lichenoides and dermatitis herpetiformis. Specimens of anetoderma, acrodermatitis chronica atrophicans and pseudoxanthoma elasticum showed no signal for human macrophage metalloelastase. Matrilysin was not detected in any of the samples investigated. Our study suggests that human macrophage metalloelastase may contribute to elastin degradation occurring in granulomatous skin diseases and may aid macrophage migration through the epidermal and vascular basement membranes in inflammatory disorders.

Angiosarcoma. A rare secondary malignancy after breast cancer treatment.

Life-saving mastectomy and radiation therapy are established in the treatment of early stage breast cancer. Angiosarcoma, i.e. malignant angioendothelioma, is a rare tumor which can develop after several years of such treatment. The number of post-operative and post-irradiation angiosarcomas has increased in recent years. We report four cases of angiosarcoma which occurred after treatment of breast cancer and review the literature. In two of these cases the angiosarcoma developed on the irradiated breast skin after partial mastectomy and radiation therapy, in the other two cases the angiosarcoma appeared on a chronically edematous arm after radical mastectomy and radiation therapy.

Demodex mites in acne rosacea.

The hair follicle mites Demodex folliculorum and Demodex brevis and their role in the pathogenesis of rosacea have been the subject of much debate in the past. We studied the prevalence of Demodex mites in facial skin biopsies obtained from 80 patients with rosacea, 40 with facial eczematous eruption and 40 with lupus erythematosus discoides. The mite prevalence in the rosacea group (51%) was significantly higher than in the rest of the study population (eczema 28% and lupus discoides 31%). Demodex mites were found on all facial sites. The most infested areas in the whole study group were the forehead (49%) and the cheeks (44%). Males were more frequently infested (59%) than females (30%). We did not find any significant difference in mite counts of infested follicles between rosacea and the control group. A lympho-histiocytic cell infiltration was seen around the infested hair follicles. Our results suggest that Demodex mites may play a role in the inflammatory reaction in acne rosacea.

Down-regulation of transforming growth factor-beta receptors I and II is seen in lesional but not non-lesional psoriatic epidermis.

Transforming growth factor-beta s (TGF-beta s) are a family of growth factors with inhibitory effects on epithelial cell proliferation. Their effects are mediated by two interacting receptors, of which type I (T beta R-I) mediates signal transduction after interaction with type II (T beta R-II) carrying the TGF-beta ligand. We have studied the expression of T beta R-I and T beta R-II in psoriatic and normal human skin by using polyclonal rabbit antisera and immunohistochemistry. Immunohistochemical analysis revealed an intense immunoreactivity for both receptors in the basal and often also suprabasal layer of normal and non-lesional psoriatic epidermis. In contrast, all psoriatic lesions studied lacked detectable immunoreactivity of either receptor in the epidermis. The results suggest that lack of TGF-beta-mediated growth inhibition by down-regulation of TGF-beta receptor expression may play an important part in the pathogenesis of psoriasis.

Human collagenase-3 is expressed in malignant squamous epithelium of the skin.

Co-expression of several members of the matrix metalloproteinase (MMP) family is a characteristic of human carcinomas. To investigate the role of the recently cloned collagenase-3 (MMP-13) in epidermal tumors, we studied samples representing malignant (basal and squamous cell carcinoma, Paget's disease), pre-malignant (Bowen's disease, solar keratosis), and benign (keratoacanthoma, seborrheic keratosis, linear epidermal nevus) tumors. Basal cell carcinomas expressed collagenase-3 mRNA in focal areas of keratinized cells, the squamous differentiation of which was confirmed by positive immunostaining for involucrin. Apoptosis was observed in central parts of these foci. In squamous cell carcinomas, collagenase-3 expression was detected at the epithelial tumor front and less frequently in the surrounding stromal cells. Collagenase-3 mRNA co-localized with immunostaining for laminin-5, an adhesion molecule suggested to participate in the migration of tumor cells. The pre-malignant and benign tumors were mostly negative for collagenase-3. Stromelysin-1, a potential activator of latent collagenases, was frequently expressed by stromal cells surrounding the malignant tumors, and the two MMPs occasionally co-localized in keratotic foci. Our results demonstrate that in basal cell carcinomas, expression of collagenase-3 is associated with terminal differentiation of epithelial cells. Furthermore, the gene is activated during skin carcinogenesis, and we suggest a role for collagenase-3 in degradation of the extracellular matrix associated with malignant epithelial growth.

Expression of collagenase-3 (matrix metalloproteinase-13) in squamous cell carcinomas of the head and neck.

Squamous cell carcinomas (SCCs) of the head and neck are malignant tumors with high capacity to invade and metastasize. We have examined expression of the new collagenase, collagenase-3 (MMP-13), in SCCs of the head and neck. MMP-13 mRNAs were detected in 22 of 29 SCC cell lines: in 14 of 15 primary SCC cell lines and in 8 of 14 SCC cell lines from recurrent tumors or metastases. MMP-13 mRNAs were expressed by all 6 cell lines from highly invasive primary tumors and in all 4 cell lines from small aggressive tumors. Using in situ hybridization, MMP-13 mRNAs were detected in 15 of 17 SCC tumor samples. In most tumors, MMP-13 was expressed by tumor cells at the invading front of the tumors, but in a subset of SCCs, MMP-13 mRNA was also expressed by stromal fibroblasts. No MMP-13 expression was detected in intact skin or oral mucosa. MMP-13 mRNA levels in SCC cells were enhanced by transforming growth factor-beta, tumor necrosis factor-alpha, transforming growth factor-alpha, and keratinocyte growth factor. Specific expression of MMP-13 by SCC cells in vitro and in vivo strongly suggests a role for MMP-13 in the high invasion capacity of SCC cells.