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Notes: We have studied the expression of involucrin in a variety of keratinization disorders, mostly of genetic origin using an avidin-biotin-peroxidase technique. In normal human epidermis 25% of the living epidermis was labelled. The diseases studied fell into two groups. Diseases with greatly increased involucrin staining including collodion baby (38%), Darier's disease (49%), Flegel's disease (56%), erythrokeratoderma variabilis (6o%), epidermal naevus with epidermolytic hyperkeratosis (45%) and congential bullous (58%) and non-bullous (44%) ichythyosiform erythroderma; and diseases with normal or slightly increased staining, including ichthyosis vulgaris (27%), X-linked ichthyosis (25%), confluent and reticulate papillomatosis (27%) and simple epidermal naevus (28%). These results demonstrate that involucrin exression is altered in some keratinization disorders and suggest that in such conditions cellular functions other than keratin metabolism are also affected.

Notes: Background  Junctional epidermolysis bullosa with pyloric atresia (PA-JEB) is a rare autosomal recessive genodermatosis that manifests with neonatal mucocutaneous blistering and gastric outlet obstruction. The disease, which is caused by mutations in the α6β4 integrin genes (ITGA6, ITGB4), is usually lethal. However, nonlethal cases have also been reported. Mutation database analysis has suggested that premature termination codons predominantly result in lethal forms while missense mutations frequently associate with nonlethal variants. Nevertheless, it is becoming more and more evident that the disease phenotype is also influenced by the position of the mutation in the protein functional domains.Objective  To investigate the molecular basis of a novel PA-JEB lethal case.Methods  Reverse transcriptase–polymerase chain reaction and direct sequencing-based mutation screening were performed. Mutation consequences in the patient's keratinocytes were then analysed by Northern blot and immunoprecipitation. Immunofluorescence analysis of cultured keratinocytes treated with protein intracellular degradation pathway inhibitors was also carried out.Results  The phenotype was caused by the presence, in the homozygous state, of a novel 33 bp in-frame deletion (nucleotides 175–207) in the ITGB4 coding sequence. Despite the normal steady-state level of integrin β4 mRNA, the mutation, designated ΔR59-A69, results in the almost complete absence of α6β4 integrin in the patient's skin and cultured keratinocytes. Exposure of the patient's keratinocytes to the proteasomal inhibitor clasto-lactacystin β-lactone increased the expression of the mutated β4 integrin chains indicating that the proteasome complex is involved in the degradation of the internally deleted β4 polypeptides.Conclusions  We report for the first time a homozygous in-frame deletion in the ITGB4 gene. Our results suggest that the deletion of amino acids R59-A69 interferes with the biosynthetic folding of the protein, leading to a rapid degradation of the mutated β4 chains. These findings provide new insight into the pathogenic effects of mutations affecting different functional domains of the β4 integrin molecule and their prognostic implications in PA-JEB patients.

Notes: Background  Disseminated superficial actinic porokeratosis (DSAP) is the most common porokeratosis and is characterized by multiple keratotic lesions which tend to occur at sun-exposed sites. A mild hypersensitivity to X-rays has been reported for DSAP-derived fibroblasts and frequent over-expression of p53 has been found in lesional epidermis.Objectives  In order to clarify whether genome maintenance mechanisms might be compromised in this disease the following approaches were undertaken: (i) primary cultured keratinocytes and fibroblasts from DSAP patients were characterized for ultraviolet (UV) B and X-ray response; (ii) 15 lesions were studied for p53 mutations, and (iii) the differentiation status of DSAP-derived keratinocytes was evaluated.Methods  Primary cultures of keratinocytes and fibroblasts were established from lesional and nonlesional skin biopsies of two subjects with DSAP. p53 mutations were analysed by DNA sequencing of the conserved region of the TP53 gene. Differentiation was evaluated both in stratified epithelial sheets from confluent keratinocyte cultures and in organotypic skin cultures.Results  The cytotoxic and apoptotic response to UVB or X-irradiation was similar in DSAP-derived keratinocytes and fibroblasts when compared with normal cells. Two of 15 lesions examined presented p53 mutations located at nondipyrimidine sites. A strikingly decreased expression of filaggrin was observed both in reconstructed epidermis and in reconstructed skin.Conclusions  The UVB and X-ray response of DSAP-derived keratinocytes and fibroblasts indicates that the actinic character of this skin pathology is not due to radiation hypersensitivity. In agreement with this finding, mutations in the p53 gene, which are often associated with UV-related skin carcinogenesis, were rarely detected in DSAP lesions and were not UV-specific. Reconstructed epidermis and reconstructed skin models successfully reproduced the main features of this genodermatosis, showing that DSAP-derived keratinocytes bear an inherent defect in the terminal differentiation programme.

Notes: Melkersson–Rosenthal syndrome (MRS) is a complex neuromucocutaneous disorder characterized by localized orofacial oedema and cranial nerve dysfunction, frequently associated with minor signs, including furrowed tongue. Complete forms are rare whereas mono- and oligosymptomatic variants are more common. A 71-year-old man presented with a 2-year history of relapsing and progressively persistent oedema of the right eyelids and periorbital region. A fissured tongue and telangiectatic rosacea had been present since the age of 50 and 60 years, respectively. The patient was also affected by essential hypertension and diabetes mellitus. A skin biopsy showed a marked upper dermal oedema, and small epithelioid cell granulomas arranged in perivascular and perilymphatic location. Collections of small epithelioid cells were occasionally observed within lymphatic spaces. No acid-fast bacteria, fungi or foreign bodies were detected. Intralesional corticosteroids induced transient improvement, whereas minocycline, clofazimine and dapsone have been ineffective. MRS may present with unilateral eyelid and periorbital swelling. Differential diagnoses of such cases may include a variety of cutaneous, ophthalmic and systemic diseases.

Notes: In this work the role of trypsin in revealing epidermal cell surface antigens, with the use of immunological markers, was investigated. Two monoclonal antibodies (MCA) were used, the first: D47, belongs to the first cluster of differentiation and recognizes a. membrane antigen of human thymocytes; the second HLA-ABC-m3, is an anti-HLA-B27 MCA. Preliminary treatment with various concentrations of trypsin was performed on frozen skin sections and followed by indirect immunofiuorescence. D47 reacted with epidermal dentritic cells only after trypsin pre-treatment of skin sections. In addition a mild preliminary trypsinization was shown to increase in situ immunoreactivity of MCA HLA-ABC-m3 with epidermal cells. Best results were obtained when trypsin concentrations ranging from 2.5 to 5 μg/ml were applied for 10 min at 37°C. Preliminary trypsinizalion may be of interest for a better exposure of some surface antigens to immunohistochemical markers.

Notes: p29 is a cytoplasmic serine phosphoprotein of 29 kD MW, closely linked to estrogen receptors. In this work we studied the expression of p29 protein in normal human skin and a group of cutaneous benign and malignant tumors by using a monoclonal antibody (ERD5) that specifically recognizes p29. In normal skin, p29 reactivity was observed in epidermal and some adnexal keratinocytes, as well as in smooth muscle cells of dermal arterioles and arrector pili muscles. p29 was also detected in most, but not all, epithelial tumors studied. The expression of p29 was generally stronger in the more differentiated (keratinized) normal and neoplastic keratinocytes; however, no correlation could be noted between immunochemical staining for p29 and either benignity of the lesion or sex of the patient considered. Whereas, in breast cancer, the expression of p29 is reported to correlate with endocrine response, the precise relationship between epithelial tumors of the skin and the action of estrogens remains to be elucidated.

Notes: Granular-cell tumour (GCT) of the skin is an uncommon tumour of disputed histogenesis, that has been subjected to several immunohistochemical studies. The controversy existing in the literature concerning the expression of carcinoembryonic antigen (CEA) by GCT prompted us to study a series of 17 cases of cutaneous GCT by using an avidin-biotin-immunoperoxidase technique on routinely-processed tissue sections. No CEA activity was detected in any of the tumours screened. The reasons for this controversy are discussed.

Notes: ICAM-3 is a newly recognized adhesion molecule, which is a member of the immunoglobulin supergene family of ICAMs. and has been shown to be identical with the CD50 antigen. Recent functional studies have shown that ICAM-3 is a ligand for LFA-1, and plays an important part in immune reactions. To date, very few data exist in the literature concerning its expression in the skin. In the present study, we investigated the expression of ICAM-3 in normal skin and in 98 biopsy specimens of various inflammatory and neoplastic dermatoses. ICAM-3 was found to be expressed by epidermal CD la+ Langerhans cells, by cells of Langerhans cell hisliocytosis, by T and B lymphocytes infiltrating the dermis in cutaneous lymphomas and in a wide spectrum of inflammatory dermaloses. Epidermal keralinocytes were consistently negative: endothelial expression of ICAM-3 was observed in six of the 48 cases. These results show that ICAM-3 is constitutively and widely expressed by cells participating in inflammatory dermaloses (including Langerhans cells and T and B lymphocytes), and that it can be albeit rarely, induced on endothelial cells and dermal dendrocytes. These results highlight the important part that ICAM-3 may play in cutaneous inflammatory and immune reactions.