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Gardner syndrome (GS) is caused by mutations in the APC and besides adenomatous colorectal polyps includes such manifestations as osteomas, epidermoid cysts (ECs) and occasionally multiple pilomatricomas. More than 50 % of ECs in patients with GS exhibit pilomatricoma-like ghost cell keratinization. The latter may be explained by the fact that the development of both GS and pilomatricoma is driven by activation of the Wnt/β-catenin signaling pathway. A 62-year-old, Caucasian male with history of GS presented with a unilocular, mixed radiopaque/radiolucent mandibular lesion causing divergence and external root resorption of involved teeth. Histopathologically, the lesion was composed of two cystic components, an orthokeratinized odontogenic cyst (OOC) and a smaller one with characteristics of keratocystic odontogenic tumor (KCOT) featuring, focally, ghost cells and an epithelial morule-like structure. Dystrophic calcifications essentially similar to those seen in pilomatricomas were observed in the fibrous connective tissue wall. The KCOT and OOC epithelia revealed strong and diffuse cytokeratin (AE1/AE3) and β-catenin immunoreactivity. CD10 positive immunostaining was seen in the keratin and superficial spinous cell layers in both OOC and KCOT. The intraepithelial and mural ghost cells showed a cytokeratin (+), β-catenin and CD10 (−) immunophenotype. The diagnosis of OOC with ghost cell calcifications in association with KCOT was rendered. The patient was lost to follow-up. Although a coincidental co-existence cannot be excluded, ghost cell calcifications mimicking pilomatricoma-like changes in an unusual odontogenic cyst combining OOC and KCOT features as seen in this patient with GS may be explained by the common molecular mechanisms underlying the pathogenesis of cutaneous pilomatricomas and GS.

The routine diagnostic procedure of actinic keratosis (AK) and invasive squamous cell carcinoma (SCC) is a histological examination after taking a biopsy. In the past decades, non-invasive optical methods for skin examination have been developed. Patients with clinical diagnosis of AK or SCC were examined. The morphological criteria were determined for healthy, AK and SCC skin and compared for statistically significant differences. In this study, the applicability of multiphoton tomography (MPT) as an in vivo diagnostic tool for AK and SCC was evaluated. Changes in the morphology of the keratinocytes such as broadened epidermis, large intercellular spaces, enlarged nucleus and a large variance in cell shape could easily be recognized. The cell nuclei of AK and SCC were significantly larger compared to healthy skin cells in all cell layers. The nucleus-cytoplasm ratio was also significantly higher for AK and SCC than for the healthy skin cells. It was even higher in SCC compared to spinous and basal cell layer of AK. The cell density in AK and SCC was significantly lower than in the basal and spinous cell layers of healthy skin. In SCC, the cell density was significantly lower than in AK. Concerning the intercellular spaces, significant differences were found for AK and healthy skin in spinous and basal cell layer and for SCC compared to AK and healthy skin. In this study, MPT proved to be a valuable non-invasive imaging method for in vivo detection and discrimination of AK and SCC from healthy skin.

Objective The presence in the brain of α-synuclein containing Lewy neurites, or bodies, is the histological hallmark of Parkinson's disease (PD). The discovery of α-synuclein aggregates in nerve endings of the heart, digestive tract, and skin has lent support to the concept of PD as a systemic disease. Our goals were, first, to demonstrate the presence of α-synuclein inclusions in the skin and, second, to detect quantitative differences between patients with PD and atypical parkinsonism (AP). Methods Skin biopsies were taken from 67 patients and 20 controls. The biopsies underwent immunohistochemistry (IHC) and immunofluorescence (IF) testing for α-synuclein, whereupon its presence was quantified as the percentage of positive cells. Patients were divided into those with PD and those with AP. AP patients included AP with neurodegenerative disease (proteinopathies) and secondary AP. Results Sixty-seven patients (34 with PD) and 20 controls were recruited. In the PD group, α-synuclein was detected in 58% of the cells in the spinous cell layer (SCL), 62% in the pilosebaceous unit (PSU), and 58% in the eccrine glands (EG). The AP-proteinopathies group showed 7%, 7%, and 0% expression of α-synuclein, respectively. No expression was found in the skin of the control group. Conclusions The expression of α-synuclein in the skin was relatively high in the PD group, scarce in AP, and null for the individuals in the control group. While these findings require further confirmation, this minimally invasive technique may aid in the improvement of the accuracy of PD diagnoses.

Localization of each keratin isoform differs among epidermal layers. Proliferating basal cells synthesize keratin 14 (K14) and suprabasal cells express keratin 10 (K10) in normal skin. Notch signaling is essential for keratinocyte differentiation. Notch1 is expressed in all epidermal layers, Notch2 in the basal cell layer and Notch3 in basal cell and spinous cell layers in normal epidermis. It has been poorly elucidated how localization and expression levels of Notch molecules are related to epidermal molecular markers K10 and K14 in psoriatic skin with abnormal differentiation of epidermal tissue. This study aimed to investigate the relationship between abnormal differentiation of epidermal cells in psoriatic skin and expression of Notch molecules. We investigated keratins (K14 and K10) and Notches (1, 2, 3 and 4) using immunohistochemistry in psoriatic skin (n=30) and normal skin (n=10). In normal skin, K14 and K10 were discretely observed in the basal cell layer and suprabasal layer, respectively. In psoriatic skin, K14 was expressed in the pan epidermal layer while it and K10 were co-expressed in some middle suprabasal layer cells. Notch1, 2, 3, and 4 localized in all epidermal layers in normal skin. In psoriatic skin, Notch1, 2, and 4 mainly localized in suprabasilar layers and Notch3 is lacalized in pan epidermal, suprabasilar, and basilar layers. Protein and mRNA of Notch1, 2, and 3 isoforms decreased in psoriatic epidermis compared with normal epidermis. These data suggest that decrements in these Notch molecules might cause aberrant expression of K10 and K14 leading to anomalous differentiation of the epidermis in psoriatic lesions.

Twenty-five dogs with signs of atopic dermatitis were included in this study. Additionally, 10 healthy dogs were chosen as healthy skin controls. Skin biopsy specimens were taken from these dogs and evaluated for the following cells: basal cell layer including the number of mitotic figures in this layer, spinous cell layer, macrophages, melanocytes, mast cells and dendritic cells. Identification of mast cells and dendritic cells was performed by means of immunohistochemistry. Histological and statistical investigations showed that the number of mitotic figures in the basal cell layer as well as the number of mast cells, melanocytes, dendritic cells and macrophages was significantly higher in the skin of dogs with atopic dermatitis compared to the healthy dogs (p < 0.01). This finding indicates multilateral quantitative activation within the cellular elements of the skin immune system. Furthermore, marked morphological heterogeneity and distinct degranulation patterns observed among mast cells in atopic skin points to their significant functional activation and confirms that canine atopic dermatitis is still notably a mast cell-dependent disease. Canine, SIS, atopic dermatitis, mast cells, epidermal dendritic cells

Background Recent developments in molecular genetics may lead to re-examination of the histopathology of inherited palmoplantar keratodermas (PPKs) based on more precise groupings of the various entities and syndromes. Objective We sought to characterize the histopathological findings in PPKs associated with mutations in DSG1 , which encodes desmoglein 1. Methods We studied the histopathology of 3 cases of keratosis palmoplantaris striata type I and one case of diffuse PPK, all associated with autosomal-dominant mutations in DSG1 . Our cases for comparison included 4 cases with Mal de Meleda PPK associated with autosomal-recessive SLURP1 mutations, one case with pachyonychia congenita type II PPK associated with an autosomal-dominant KRT17 mutation, and one case with focal PPK associated with an autosomal-dominant KRT16 mutation. Results The distinguishing histopathological features of the 3 keratosis palmoplantaris striata type I cases and the diffuse PPK case associated with DSG1 mutation were: varying degrees of widening of the intercellular spaces and partial disadhesion of keratinocytes in the mid and upper epidermal spinous cell layers, often extending to the granular cell layer. These findings, which are associated with haploinsufficiency of desmoglein 1, were not observed in any of the other 6 PPK cases. Mild perinuclear eosinophilic condensations and cytoplasmic vacuolizations were observed in the spinous cell layer keratinocytes of the pachyonychia congenita type II PPK and the nonspecified focal PPK cases. Limitations There were a limited number of patients and control patients with hereditary PPKs. Conclusion Widening of the intercellular spaces and disadhesion of epidermal keratinocytes may serve as a histologic clue to PPKs caused by DSG1 mutations.

Osteopontin (OPN) is an adhesive, matricellular glycoprotein, whose expression is elevated in many types of cancer and has been shown to facilitate tumorigenesis in vivo. To understand the role of OPN in human skin cancer, this study is designed to determine whether OPN is expressed in premalignant [solar/actinic keratosis (AK)] and in malignant skin lesions such as squamous cell carcinomas (SCC) and basal cell carcinomas (BCC), as well as in normal skin exposed or not exposed to sunlight. Immunohistochemical analyses showed that OPN is expressed in SCC (20/20 cases) and in AK (16/16 cases), which are precursors to SCC, but is absent or minimally expressed in solid BCC (17 cases). However, positive staining for OPN was observed in those BCC that manifest differentiation toward epidermal appendages such as keratotic BCC. In sunlight-exposed normal skin, OPN is minimally expressed in the basal cell layer, but in contrast to those not exposed to sunlight, OPN is more prominent in the spinous cell layer with increasing intensity toward the granular cell layer. Additionally, OPN is expressed in the hair follicles, sebaceous glands, and sweat glands of normal skin. In conclusion, these data suggest that OPN is associated with keratinocyte differentiation and that it is expressed in AK and SCC, which have metastatic potential, but minimally expressed in solid BCC. (J Histochem Cytochem 56:57–66, 2008)

Keratinocyte apoptosis mediated by Fas/Fas ligand molecular interactions and subsequent caspase activation is believed to play an important role in the pathogenesis of atopic dermatitis (AD), in particular for the formation of spongiosis. To estimate epidermal caspase activation in normal and AD skin under in vivo conditions, we analysed caspase-3 cleavage by immunohistology. In normal skin as well as non-lesional AD skin, we detected caspase-3 cleavage in single cells of the basal layer. In contrast, in acute lesional AD skin, we not only obtained evidence for increased expression of cleaved caspase-3 in keratinocytes of the basal layer but also observed caspase-3 cleavage in one or more layers of the spinous cell layer, in particular in spongiotic areas. Short-term topical treatment of the skin lesions with tacrolimus or pimecrolimus abolished the expression of cleaved caspase-3 in the spinous layer. Moreover, epidermal caspase-3 cleavage correlated with the numbers of dermal interferon-gamma (IFN-gamma)-expressing CD4+ and CD8+ lymphocytes in skin lesions of AD patients, supporting the view that IFN-gamma is important for the activation of proapoptotic pathways in keratinocytes. This is also confirmed by the observation of increased Fas expression on keratinocytes in acute AD lesions that was markedly reduced following topical calcineurin inhibitor treatment. These data suggest that caspase-3 cleavage in the spinous layer of the epidermis is a pathologic event contributing to spongiosis formation in AD, whereas cleavage of caspase-3 in basal cells might represent a physiologic mechanism within the process of epidermal renewal.

Psoriasis is a skin disorder of chronic keratinization characterized by epidermal hyperplasia, hyperkeratosis, and inflammation. However, little is known about the mechanism (s) underlying the hyperplasia with elongated rete ridges characteristic of psoriasis. The p63 transcription factor, a homologue of the p53 tumor suppressor, has been implicated in the maintenance of epidermal stem cells and the stratification of the epidermis. p63 is up-regulated in squamous cell carcinomas with anaplasia, suggesting that it is also associated with epidermal hyperplasia. In this study, we examined the expression of p63 in the remodeling of psoriatic epidermis. Lesional tissues from 17 psoriasis patients in various stages of plaque-type psoriasis and normal skin tissues from five healthy subjects were examined by immunohistochemistry using a monoclonal anti-p63 antibody. Normal epidermis stained positively for p63 in the basal cell layer and in 2 to 4 layers of the spinous cell layer. p63 was positive in the thickened rete ridges of the epidermis even in early psoriatic lesions. As the epidermis elongated, p63-positive cells moved down and were localized in the lower parts of the rete ridges where keratinocytes densely proliferated. From these results, we suggest that p63 may be involved in the early stage of the remodeling process of the psoriatic epidermis as well as in the elongation of the rete ridges.

Summary Background Molecular characterization of tight junction proteins during the past few years has provided novel methods for studying these specialized junctions. Tight junctions have recently been characterized in the granular cell layer of human epidermis, and the role of these junctions in the epidermal barrier is now being re-evaluated. Objectives To investigate the expression of tight junction components during the re-epithelialization of suction blisters and the regeneration of the corneal layer after tape stripping. Methods Suction blisters were induced in eight healthy volunteers, and skin biopsies were taken 4 or 6 days afterwards. The restoration of epidermal barrier function was evaluated by measuring water evaporation (WE) from the wound area. Tape stripping was performed on three volunteers to remove the corneal layer. The tissues were immunolabelled using indirect immunofluorescence or the avidin–biotin method. Results Prior to the biopsies, WE from the blister wounds was markedly elevated in comparison with normal skin. In the epidermis surrounding the blister, occludin and ZO-1 were expressed in the granular cell layer only. In the hyperproliferative zone adjacent to the border of the blister, the expression of ZO-1 was redistributed into several spinous cell layers, while occludin expression was restricted to the upper epidermis. In the leading edge of migrating keratinocytes, both proteins were expressed exclusively in the most superficial layer of keratinocytes. Double labelling for ZO-1 and involucrin showed expression of both proteins in the same layers of hyperproliferative keratinocytes, while the expression patterns were clearly different in the migrating keratinocytes. Conclusions Tight junctions of regenerating epidermis may provide a functional barrier prior to regeneration of the corneal layer.

Contains fulltext : 186533.pdf (Publisher’s version ) (Closed access) BACKGROUND: Cystatins are natural and specific inhibitors of endogenous mammalian lysosomal cysteine proteinases and exogenous microbial cysteine proteinases. Cystatins were shown to provide regulatory and protective functions against uncontrolled proteolysis in several disease processes. Recently we reported that cystatin M/E, which is a novel member of the cystatin gene family, has an unusually restricted expression pattern that is limited to skin. Although cystatin M/E possesses two distinct biochemical properties (it is a proteinase inhibitor and a substrate for transglutaminase) its physiological function is unknown. Disturbance of the balance between proteinases and their inhibitors can lead to irreversible damage as in chronic inflammatory reactions and tumour invasion. OBJECTIVES: To examine the expression pattern of cystatin M/E in inflammatory conditions and neoplastic skin disorders in order to obtain possible clues on its function. Furthermore, we wished to determine whether cystatin M/E expression could discriminate between various types of neoplasia. METHODS: Biopsy material of normal skin, atopic dermatitis and psoriatic lesional skin, healing excisional wounds in healthy volunteers, and several types of epidermal neoplasia (keratoacanthoma, actinic keratosis, basal cell carcinoma and squamous cell carcinoma) were used in this study. For comparison we studied the expression of cystatin M/E in squamous neoplasias from non-cutaneous origin. Affinity-purified polyclonal antibodies against cystatin M/E were used for immunohistochemical detection. RESULTS: Cystatin M/E is constitutively expressed in the stratum granulosum of normal skin, sebaceous glands, eccrine sweat glands and the infundibular epithelium of hair follicles. Expression in atopic dermatitis and psoriasis was found to extend to several layers of the stratum spinosum. In wound healing, cystatin M/E was not found in the edge of migrating keratinocytes, but it was strongly expressed in the suprabasal layers of the neo-epidermis. In epidermal neoplasias cystatin M/E expression was only found in differentiated cells and keratinized cell nests. CONCLUSIONS: Inflammation causes cystatin M/E to be expressed in the spinous cell layers where it colocalizes with transglutaminase for which it serves as a substrate. Speculatively, increased expression of cystatin M/E is compatible with a role in controlling increased levels of cysteine proteinases during inflammation and infection. Cystatin M/E expression in neoplastic epidermis is confined to well-differentiated cells and as such does not discriminate between benign and (pre)malignant epidermal neoplasias.

Epidermolysis bullosa (EB) is a group of hereditary autosomal dominant bullous diseases. EB is divided into four major phenotypes: intraepidermal EB (or EB simplex), junctional EB, dermolytic EB and mixed EB (Kindler syndrome). EB simplex is further divided into three subtypes: localized EB simplex, Dowling-Meara EB simplex and other generalized EB simplex. We report a 28-year-old man with EB simplex with a missense keratin 14 mutation p.Arg125Cys associated with clumping of keratin filaments and acantholysis in mainly the spinous cells and basal cells. Immunohistochemistry revealed that the broader expression of keratin 5 and 14 was observed in the epidermis, while the expression of keratin 1/10 was quite normal. Dysregulated expression of keratin 5/14 may hinder some functions or roles of keratin 1/10, namely filament assembly of keratin 1/10 in spinous cell integrity, although the expression of keratin 1/10 was not affected and this has not been demonstrated before.

The mechanism of carcinogenesis induced by 4-nitroquinoline 1-oxide remains poorly understood.We studied cell kinetics (BrdU, cyclin D 1) and immunohistochemically examined the relation to the cell cycle and morphogenesis (P-cadherin) to determine the mechanism of carcinogenesis. P-cadherin is a cell adhesion molecular essential for morphogenesis. It also has an important role in cell proliferation. However, the role of P-cadherin in carcinogenesis remains unclear.During the epithelial dysplasia stage of carcinogenesis, BrdU and cyclin D 1 were overexpressed significantly as compared with normal epithelial tissue (p=0.0005). The localization of cyclin D 1 and Pcadherin extended from the basal cells to the spinous cell layer. This localization of cyclin D 1 indicated that cells in the spinous cell layer were in the G 1 phase of their cell cycle. P-cadherin simultaneously appeared on their cell membrane.Our results suggest that overexpression of P-cadherin is closely related to the G 1 phase of the cell cycle and that carcinogenesis is induced by an uncontrolled cell cycle with overexpression of P-cadherin.

We investigated human papillomavirus type 1 (HPV1)-specific transcription, viral DNA replication, and viral protein expression in naturally occurring benign tumors by in situ hybridization, 5-bromodeoxyuridine (BrdU) incorporation, and immunohistochemistry and obtained results different from other HPV-infected benign tumors characterized so far. Moderate amounts of transcripts with a putative coding potential for E6/E7, E1, and E2 were demonstrated from the first subrabasal cell layer throughout the stratum spinosum and granulosum. In addition very large amounts of E4 and L1 transcripts were present in the same epithelial layers. This finding was substantiated by the demonstration of L1 and E4 protein already in the bottom-most spinous cell layer. Furthermore massive amplification of the viral DNA as measured by BrdU incorporation and different methods of in situ hybridization took place in the lowest 5 to 10 suprabasal cell layers. These findings are in contrast to the assumption that late gene expression and viral DNA synthesis are restricted to the more differentiated cell layers of the epithelium and point to differences in the regulation of the vegetative life cycle between different papillomavirus types.

Recently we demonstrated a strong induction of activin expression after skin injury, suggesting a function of this transforming growth factor-beta family member in wound repair. To test this possibility, we generated transgenic mice that overexpress the activin betaA chain in the epidermis under the control of a keratin 14 promoter. The transgenic mice were significantly smaller than control littermates, and they had smaller ears and shorter tails. In their skin, the fatty tissue was replaced by connective tissue and a severe thickening of the epidermis was found. The spinous cell layer was significantly increased, and the epidermal architecture was highly disorganized. These histological abnormalities seem to result from increased proliferation of the basal keratinocytes and abnormalities in the program of keratinocyte differentiation. After skin injury, a significant enhancement of granulation tissue formation was detected in the activin-overexpressing mice, possibly as a result of premature induction of fibronectin and tenascin-C expression. These data reveal novel activities of activin in the regulation of keratinocyte proliferation and differentiation as well as in dermal fibrosis and cutaneous wound repair.

A lectin histochemical study was carried out on the dorsal skin of Wistar-derived hypotrichotic WBN/Ila-Ht rats (HtRs) and Wistar rats (WRs) at 3, 7 and 24 weeks of age to clarify the lectinhistochemical characteristics of the skin during their development. The lectins examined were Concanavalia ensiformis (Con A), Dolichos biflorus agglutinin (DBA), Griffonia simpliciolia (GS-I), Helix pomatia agglutinin (HPA), Arachis hypogaea (PNA), Glycine maximus agglutinin (SBA), Ulex europeus agglutinin (UEA-I) and Triticum vulgaris agglutinin (WGA). None of the nucleated cell layers of the epidermis had DBA-binding sites, but they were all stained intensely with HPA and weakly with Con A irrespective of the strain and age of the rats. As to the other 5 lectins, the intensity of binding activity was generally weaker in HtRs than in WRs and at 3 weeks of age than at 7 or 24 weeks of age, respectively. Among them, UEA-I mainly bound to the spinous cell layer but not to the basal cell layer, suggesting that alpha-L-fucose would be expressed on the cell surface according to the differentiation of keratinocytes. In addition, GS-I, HPA and UEA-I bound to the hair follicle epithelium and many lectins stained sebaceous gland epithelial cells. In conclusion, except for the binding intensity of some lectins, there were no specific differences between HtRs and Wrs in the lectinhistochemical characteristics of the dorsal skin epidermis. The present data on the rat skin would be useful from the viewpoint of comparative lectinhistochemistry.

Oral keratinocytes originate from basal cells, differentiate during migration to the surface, and finally are shed. Apoptosis occurs at the end of differentiation, but the precise relationship between terminal differentiation and apoptosis is not clear. In the present study, Bcl-xL was expressed in the basal cell and spinous cell layers, and Bax was expressed in the spinous cell and granular cell layers. In cultured keratinocytes, Bcl-xL was expressed under conditions of 0.1 mM calcium (low Ca2+) but disappeared under conditions of 1.0 mM calcium (high Ca2+); the latter induces keratinocyte differentiation. Bax was not expressed in keratinocytes with low Ca2+ but was expressed in cells with high Ca2+. Finally keratinocytes with high Ca2+ underwent apoptosis, which was detected by the TUNEL method and by 180-bp DNA fragmentation. These results suggest that the process of terminal differentiation in gingival epithelium is a pathway to apoptosis.

The environmental pollutant 2,3,7,8-tetrachlorodibenzo- p -dioxin (TCDD) causes chloracne in humans by mechanisms that are as yet poorly understood. Because TCDD is known to affect keratinocyte differentiation in vitro , we have studied TCDD-dependent morphologic changes and the expression of murine keratin I (MK1; differentiation associated) and keratin 17 (MK17; presumably hyperproliferation associated) in HRS/J hr/hr hairless mouse skin. TCDD (0.2 μ g in acetone) applied topically to the dorsal skin caused epidermal acanthosis and hyperkeratosis of the dermal cysts as well as an involution of the utricles and the sebaceous glands. By means of in situ hybridization with digoxigenin-labeled riboprobes of sections from untreated and vehicle (control)-treated skin, we localized MK1 mRNA to the epidermal spinous cell compartment. MK17 transcripts were detected only in the derivatives of the hair follicleutricle epithelium and dermal cysts. No spatial overlap was observed between MK1 and MK17 expression. After TCDD application, MK17 was newly expressed in the upper spinous cell layers of the interfollicular epidermis, although it was suppressed in the involuting utricles. In contrast, MK1 expression in the interfollicular epidermis was not affected by TCDD. Furthermore, MK1 expression was induced in the epithelium of the utricle remnants and in some dermal cysts. These data suggest that increased keratinization of the part of the follicular epithelium corresponding to the dermal cyst epithelium of hairless mice most probably explains the pathogenesis of TCDD-induced chloracne. The results demonstrate, furthermore, that TCDD can differentially affect keratinocyte differentiation in vivo as well as in vitro .

We have developed a new skin transplantation method, called the skin fenestrating transplantation method, in which the nude mouse skin covering the lesion was removed one week after the transplant was embedded. One week later, transplanted psoriatic skin was biopsied and specimens were examined by electron microscopy. The basal cells and the nuclei of transplanted psoriatic epidermis were long and slender. Projections of basal cells extended to the dermis. Erythrocytes and lymphocytes were seen in the intercellular space. In the spinous cell layer, the cytoplasm and the nuclei were round. In the cytoplasm, mitochondria and endoplasmic reticuli were abundant, and tonofibrillar formation was poor. In the upper part of the epidermis, the cells were horizontally elliptic and, in the cytoplasm, short tonofibril bundles were formed. In the uppermost layers, in the cytoplasm of flat cells, degenerated flat nuclei, low dense lipid droplets and clumps of ribosomes were observed. These findings resembled the findings of intact psoriasis lesions more closely than those obtained by the skin embedding transplantation method.

New ultrastructural observations are described in skin lesions of two brothers with Richner-Hanhart's syndrome (RHS). Physical examination of the two patients showed painful skin lesions of palms and soles combined with denderitic corneal ulceration and mental retardation. The diagnosis of RHS was confirmed biochemically with high tyrosine levels in both blood and urine. Examination by transmission electron microscopy revealed several abnormal ultrastructural changes in the epidermal cells. The horny cells contained heterogeneously, electron-dense cytoplasm with many lipid droplets. The granular cell cytoplasm contained abundant tonofibrils and keratohyaline granules. The spinous cell cytoplasm was vacuolated due to the presence of minute tyrosine crystals, which are known to have a lytic effect. The surrounding keratinocytes contained multilobed nuclei. The basal epidermal cells appeared normal except for Merkel cells, which were severely damaged by vacuolatio, also due to the presence of tyrosine crystals. This study showed that high tyrosine levels can induce several ultrastructural pathological changes in the epidermal cells, including the skin chemoreceptor Merkel cells.

A series of 90 excised cutaneous warts (verrucae vulgaris) were studied for the presence of HPV (human papillomavirus) DNA using in situ hybridization (ISH) with biotinylated full genomic DNA probes of HPV types 1, 2, 3, and 4. The expression of PCNA (proliferating cell nuclear antigen) was examined using conventional immunohistochemistry. The aim was to test the hypothesis that HPV can reactivate PCNA, including in the host replication machinery. HPV DNA of the above types was detected in 60 of 90 verruca biopsies studied (66.7%): HPV 2 in 56 cases, HPV 1 in 2 cases, and HPV 3 in 2 cases. PCNA was expressed in all samples except two. The signal distribution of HPV DNA markedly differed from that of PCNA expression. ISH revealed strong HPV DNA signals in both the granular and the upper spinous cell layers, the most intense signals being detected in the upper epidermis. On the other hand, nuclear PCNA staining was present in the majority of parabasal and basal cells. Although strong PCNA signals within the wart lesions were found in the areas where HPV DNA was present, the PCNA positivity was almost invariably localized in the differentiated cells of the spinous cell layers, just below the HPV DNA-expressing cells. At the margins of the lesions, PCNA expression was still strong but disappeared abruptly towards the normal epidermis. HPV DNA-positive warts showed more intense expression of PCNA than did the HPV DNA-negative ones in this study. Our results indicate that PCNA induction is associated with the presence of HPV DNA, suggesting that HPV can reactivate PCNA, thus interfering with the host cell DNA replication machinery.

The present study deals with 114 cases of oral leukoplakia and clinicopathological findings evaluated with respect to epithelial differentiation and keratinization using keratin antibodies, expression of epidermal growth factor receptor (EGF-r), and proliferating cell nuclear antigen (PCNA). Oral leukoplakias were most often found in men (57%) and women (43%) aged 40-60 years. The commonest sites were the gingiva (32.4%), tongue (25.4%), and cheek (23.7%). Hyperorthokeratinization of the epithelium was most commonly seen in the masticatory mucosa (gingiva and hard palate) and hyperparakeratinization in non- or lesskeratinized mucosa (buccal mucosa and floor of the mouth). Epithelial dysplasia was observed in 19 cases (tongue 9, buccal mucosa 5, gingiva 3, palate 2). Cytokeratins (CK) detected by polyclonal TK were present in all epithelial layers except the keratinized layer; CK detected by monoclonal KL 1 in the spinous and granular cell layer; K8.12 in the spinous layer; and PKK 1 in the basal layer. Involucrin was present in the upper-spinous and granular cell layer and filaggrin in the granular cell and hyperkeratinized layer. EGF-r was detected in the basal and spinous cell, and dysplastic cells were unreactive. PCNA immunoreactivity was distributed in the basal and suprabasal cells. Leukoplakia on the lateral border of the tongue was characterized by a lesser degree of hyperkeratinization but with an increased frequency of epithelial dysplasia. Leukoplakia on the gingiva and hard palate, on the other hand, had more advanced hyperorthokeratinization, but epithelial dysplasia was rarely observed. The hyperkeratinization in oral leukoplakia is influenced primarily by sitewise variations in keratinization of the normal oral mucosa and less frequently by the extent of epithelial dysplasia.

Biotinylated hyaluronan (HA) binding complex (HABC) from bovine articular cartilage proteoglycan was used as a histological probe to study the localization of HA in human skin. The distribution of HA was compared with its presumptive cell surface receptor, CD44, using monoclonal antibodies. In epidermis both HA and CD44 were found in the basal and spinous cell layers, but neither was present in the stratum granulosum and stratum corneum. In the keratinizing parts of hair follicles, i.e. in the outer and inner epidermal root sheath, pilosebaceous duct and the actual hair, HA and CD44 were found between the vital but not the terminally differentiated cells. In the sebaceous glands a small amount of HA was found around all cells, whereas CD44 was restricted to the basal cell layer. The secretory acini of the sweat glands stained intensively with anti-CD44 antibodies but only weakly with HABC. In the sweat gland, CD44 was localized on the basal and lateral surfaces of the clear cells, whereas the dark cells and the myoepithelial cells were negative. Both the lower and upper layers of the sweat gland ducts showed a faint but constant staining for CD44 and only minor amounts of HA. While in the keratinizing skin epithelia both HA and its CD44 receptor showed an intense staining with a close co-distribution, in the sweat and sebaceous glands their distribution patterns were not similar. It is suggested that in epithelia with divergent differentiation programs the functions of CD44 and HA may be different.

Eighteen oral mucosal biopsies with Candida infection were studied with light and electron microscopy. Under light microscopy, candidal infected oral mucosa was classified with epithelial hyperplasia, 15 cases and epithelial dysplasia, three cases. Four of 15 epithelial hyperplasias showed marked parakeratosis, and high grade acanthosis with many eosinophilic cells in the spinous cell layers. Epithelial dysplasia was characterized by atrophy of the spinous cell layers and increased nucleocytoplasmic ratio in the basal cell layers. Ultrastructurally, candidal infected oral mucosa showed numerous small desmosomes and the interdigitation of cytoplasmic membranes between spinous cells in both epithelial hyperplasia and epithelial dysplasia. Moreover, eosinophilic spinous cells, observed predominantly in epithelial hyperplasia showed intricate arrangement of dense tonofibrils. These ultrastructural findings seemed to give rise to mechanical strength between spinous cells in oral mucous epithelium with Candida infection. Results in this study suggest that excessive hyperplasia of candidal infected oral mucosa might be a protective reaction to the invasion of candidal pseudohyphae, but not associated with precancerous conditions.

To investigate the histological distribution and the morphology of the fungi and the tissue reactions in chromoblastomycosis, especially in the process of trans-epidermal elimination, cutaneous lesions of two patients with this disease were studied morphometrically and ultrastructurally. In the dermis, most of the fungal elements appeared as sclerotic cells and their cell wall showed an irregular, worm-eaten leaf-like appearance; they seemed to be continuously attacked by polymorphonuclear neutrophils. The epidermis eliminated 10–20% of all the organisms in the skin lesions, and the hypha-forming activity tended to be higher in the epidermis than in the dermis. Ultrastructurally, basal keratinocytes facing the dermal abscess containing fungal elements frequently appeared as dark cells, suggesting an increased proliferation activity. Spinous keratinocytes facing intraepidermal microabscesses containing fungal elements showed an abnormal accumulation of tonofilaments and further early keratinization in the spinous cell layer. All of the morphological changes of the dermis and epidermis are regarded as defence reactions against the fungi existing in the skin lesions. There is a close relationship between tissue reactions and morphological changes of fungi in chromoblastomycosis.

As it is impossible to perform a longitudinal experiment of carcinogenesis of human samples, experimentally induced carcinoma has been used for this purpose. DMBA induced carcinoma on hamster buccal pouch mucosa has been frequently cited for its stability and high induction rate. We have been attending to this aspect of research. This article presents the findings from PAS stained sections on the process of DMBA induced hamster buccal pouch carcinogenesis.The PAS-positive granules in the cytoplasm of epithelial cells, regarded as glycogen, were consumed by digestion of amylase. There was no difference between the experimental groups painted DMBA for no more than 6 weeks and the control groups which contained very little glycogen. But on the groups painted for more than 8 weeks, papillomas accumulated a large amount of glycogen in the cytoplasm of the spinous cell layer, and carcinomas did a little in the keratinized part. Epithelium with atypia revealed less glycogen than that of the other part of the epithelium. Carcinoma in situ contained extremely less.Concerning on the PAS-basement membrane, that is PAS positive membraneous structure at the epithelium-connective tissue junction, there was no distinct difference between the controls and the up to 6-week-painted groups. In the groups painted more than 8 weeks, papillomas occasionally had thin membranes, and carcinomas were absent or faint.Glycocalyx in the intercellular spaces was clearly observed in the controls and the up to 6-week-painted groups. Papillomas reduced the intercellular cohesion, and carcinomas showed wide intercellular spaces without any glycocalyx. The epithelium with atypia and carcinoma in situ also reduced intercellular cohesion.Although PAS staining on the DMBA induced carcinogenesis could not predict the malignant potentiality of the epithelium, it is clarified that the induced tumors and epithelia with atypia closely resembled human oral carcinomas and precancerous lesions.

Background: The transcription factor GATA-3 was recently identified as a master regulator in the specification of the inner root sheath. Additionally, it seems to play a role in skin barrier physiology. p63 binds and transactivates the GATA-3 promoter. While the expression profile of GATA-3 is delineated for the mouse, little is known about its expression in the adult human hair follicle and no studies are published about its distribution during human cutaneous embryogenesis. Methods: We examined samples from embryonic, fetal and adult human skin for the expression of GATA-3 using immunohistochemistry. Results: GATA-3 is expressed late during human skin development. Its expression pattern is comparable to the mouse and confined to the Huxley layer and inner root sheath cuticle but sparing the Henle layer. In addition, GATA-3 localizes to the spinous cell layer of the interfollicular epidermis. Conclusions: From the described expression pattern, it is highly probable that GATA-3 plays a role in follicular and epidermal morphogenesis. What the anatomically confined expression of GATA-3 to the spinous layer means biologically for the physiology of the skin is still unclear. Likewise, it still needs to be shown if GATA-3 could be exploited in the diagnosis of adnexal neoplasms.

The term pemphigus refers to a group of cutaneous diseases that are characterized by the development of intra-epidermal blisters and, sometimes, mucosal erosions [l] (Table 1). All forms of pemphigus are characterized by epidermal cell-cell detachment (acantholysis) which leads to (intra-epidermal) vesicle formation, and by the presence of IgG autoantibodies directed against antigenic determinants present on the cell surfaces of differentiating keratinocytes [2]. The most severe form of pemphigus is pemphigus vulgaris (PV) (Figure l), which may occur at any age, although its most common onset is in the fourth, fifth and sixth decades. PV is characterized by the presence of flaccid, exceedingly fragiIe noninflammatory bullae which usually arise on normal appearing skin. These bullae have a tendency to coalesce and rupture easily resulting in large denuded areas which are, in fact, the predominant clinical feature of this type of pemphigus. PV involves both the skin and mucous membranes and, in virtually all cases, the initial lesion affects the oral mucosa. Prior to the introduction of corticosteroids in the 195Os, PV was considered almost uniformly fatal mainly due to protein, fluid and electrolyte losses and/ or uncontrollable sepsis. Acantholysis, the histological hallmark of PV, starts with the development of epidermal intercellular edema leading to dissolution of intercellular ‘bridges’ and widening of intercellular spaces (ICS), finally ending in cell to cell detachment. These microscopic changes occur in the suprabasilar area, i.e. between the basal and spinous cell layers. Basal cells remain attached to the dermis, but are laterally detached, resembling a ‘row of tombstones’ [ 11. Endemic pemphigus foliaceus (PF) or fogo selvagem (FS) (Figure 2) is clinically and immunopathologically similar to the non-endemic form seen in other parts of the world. Clinically, PF is characterized by superficial blistering and erosive lesions that affect the skin and rarely involve mucosal surfaces [3-61. Histologically, PF is characterized by acantholysis involving the subcorneal layers of the epidermis. In

Cell kinetics and distribution of Involucrin in fifteen cases of squamous cell carcinoma (SCC), eleven cases of leukoplakia, and five cases of normal mucosa in the oral region have been detected immunohistocliemically. The cell kinetics were evaluated by identifying the DNA synthetic cells (S-phase cells) by in vitro labelling method using bromodeoxyuridine (BrdU) and its monoclonal antibody. The following results were obtained:1. BrdU positive cells were localized in one or two layers of the basal cell in both normal and hyperplastic epithelium without atypism. They were also demonstrated in the periphery of cancer nest in highly and moderately differentiated SCC. However, they were scattered diffusely in poorly differentiated SCC.2. In normal epithelium Involucrin, there were positive to the superior forms of spinous cell layers. In highly and moderately differentiated SCC, Involucrin displayed in the cancer nest with an irregular and patchy pattern. But in poorly differentiated SCC, Involucrin were negative. In spite of the different stain ability of Involucrin between normal epithelium and SCC, the relation between Involucrin-positive cell and BrdU-positive cell was kept both normal epithelium and SCC.3. The labelling indexes (L. I.) for BrdU were 4.8% in normal epithelium, 7.5% in the hyperplastic one without atypism, and 11.6% in that with atypism, respectively. As the degree of the epithelial dysplasia increased, the L. I. was high.4. The L. I. was 19.2% in a highly differentiated SCC, 19.5% in the moderate one, and 19.1% in the poor one. There are, however, no correlation between the degree of cell differentiation and the L. I.

Clinically healthy human gingivae from deciduous molar regions were transplanted to subcutaneous sites of nude mice (nu/nu NC). Transplants were harvested after posttransplantation periods of 5, 6, 7, 8.5, 10.5 and 12 weeks and examined histologically after staining with hematoxylin-eosin (H.E.), bisbenzimide, and a panel of mouse monoclonal anti-keratin antibodies in an indirect fluorescence technique. Central parts of transplants contained human connective tissue covered by human stratified squamous epithelium which were unkeratinized in 5- to 7-wk-old transplants and most frequently (75%) parakeratinized in 8.5-wk to 12-wk transplants. Comparison of keratin expression before and after transplantation revealed a progressive keratin reconstitution, i.e., keratin markers of basal/suprabasal cells preceded those of suprabasal/spinous cell layers and immunohistochemical markers of keratinization preceded routine histologically observed parakeratinization. Original keratin staining and essential features of histodifferentiation were reconstituted and maintained after 8.5 wk but graft recovery rate decreased drastically 12 wk after transplantation. This study shows that the human gingiva/nude mouse model is useful in experimental studies of the gingival keratin profile in the period 8.5 to 10.5 wk after transplantation.

Electron probe microanalysis was used to analyze elemental content of human epidermis. The results revealed that the calcium content of the basal keratinocyte layer was higher than that of the lowest spinous cell layer in normal epidermis. This was surprising, as it is generally accepted that the calcium level increases with cellular differentiation from the proliferative basal layer to the stratum corneum. Hailey-Hailey disease (HHD) and Darier disease (DD) are caused by mutations in Ca(2+)-ATPases with the end result of desmosomal disruption and suprabasal acantholysis. The results demonstrated three major aberrations in HHD and DD lesions. First, in HHD and DD lesions the calcium content in the basal layer was lower than in the normal skin. Second, adenosine triphosphate (ATP) receptor P2Y2 was not localized to plasma membrane in acantholytic cells, whereas P2X7 appeared in the plasma membrane, potentially mediating apoptosis. Third, transition of keratin 14 to keratin 10 was abnormal as demonstrated by the presence of keratinocytes expressing both cytokeratins, which are usually exclusive in normal epidermis. Our results provide to our knowledge previously unreported elements for understanding how the disturbed calcium gradient is linked to the alterations in ATP receptors and keratin expression, leading to the clinical findings in HHD and DD.

Cell surface carbohydrates are excellent markers of cellular differentiation and maturation processes due to their great structural and antigenic diversity as well as their known biosynthetic precursor/product relationships. Using a panel of monoclonal antibodies with well-defined carbohydrate specificities we have studied the expression of biosynthetically related antigens in normal and psoriatic skin. Two "families" of carbohydrate structures were investigated. One series of structures based on N-acetyllactosamine chains (type 2 chain: N-acetyllactosamine and fucosylated derivates hereof of H, Lex, Ley and sialyl-Lex) and another based on the simple mucin type core structures (type 3 chain: Tn, T and sialylated derivates hereof as well as the fucosylated derivative, H). Previously we have found these carbohydrate structures define distinct cell layers in stratified squamous epithelia of mucosa of the cheek, esophagus and uterine cervix. In normal and uninvolved epidermis, N-acetyllactosamine and T carbohydrates were found in the spinous cell layer, whereas the fucosylated derivates, H structures, were found in the granular cell layers above. The fucosylated and sialylated derivate of N-acetyllactosamine, sialylated Lex, had the same distribution as N-acetyllactosamine and T structures. This sequential expression of carbohydrates is similar to our previous findings in mucosa. However, in contrast to mucosa, normal skin basal cells did not label. The glycosylation pattern in psoriatic epithelium was changed in two ways. 1) Some carbohydrates (types 2 and 3 chain H and T) were expressed at an earlier stage of cell maturation. 2) The biosynthetic precursors to T structures, Tn and sialyl-Tn, which are not expressed in normal skin, and are often considered cancer-associated antigens, appeared in psoriatic skin. The Tn-antigen was expressed on basal and lower spinous cells, whereas the sialyl-Tn was only found on basal cells above the dermal papillae. The findings in the present work support previous studies of changes in cell surface glycosylation in psoriatic epidermis and demonstrate the appearance of tumor-associated antigens in highly proliferative, but benign, stratified epithelium.

UNLABELLED The mucous epithelium of the rat palatal rugae was observed by light and electron microscopy. The shape of the epithelial-connective tissue junction (ECJ) of the rat palatal mucosa was found to be composed of ridge-type papillae (RTP) which ran in an antero-posterior direction (APD). The purpose of the study is to investigate the differentiation of the basal and spinous layer in the palatal mucosa which consists of RTP by the observation of the shape of the ECJ and the morphology and structure of the constituent cells. RESULTS (1) The RTP which ran in the APD appeared to be related to the direction of the blood vessels and collagen bundles within the submucous and papillary layers. (2) The basal cells were classified into B1, B2 and B3 cells for the sake of convenience from the morphological and structural characteristics. Polyhedral spinous cells located within the lower third of the spinous layer were also classified into S1P and S11 cells. (3) The B1 and S11 cells were mainly distributed in the inter-papillary area (1-area), the B2 and S1P cells in the papillary area (P-area) and the B3 cells between the P- and I-area (PI-area). (4) The cell division was observed chiefly in the I- or PI-area in the APD and the divided cells appeared to become B1 cells. The B1 cells transitionally changed into B2 or B3 cells and finally into S1P cells in the P-area, and S1I cells in the I-area.

Biochemical data suggest that gingival epithelium contains hyaluronate, but there is little histochemical information about its localization. Hyaluronate was here visualized in gingival and buccal mucosa using a specific probe derived from the hyaluronate binding region of cartilage proteoglycan. Hyaluronate was found both in the gingival and buccal epithelium, but its localization was correlated with the type of keratinization. In the keratinized epithelium of gingiva, whether ortho- or parakeratotic, the intercellular spaces from basal to upper spinous layers displayed strong staining, most intense in the middle spinous cell layer. The uppermost vital cell layers as well as the cornified cell layer remained unstained. In the non-keratinized epithelium of buccal mucosa and the local non-keratinized areas of gingiva, only the basal cells and the lowermost spinous cell layers stained for hyaluronate, whereas the majority of the upper epithelium was negative. Electron microscopic examination of the basal and spinous cell layers displayed hyaluronate, both associated with the cell surface and free in the intercellular space. The subepithelial connective tissue showed positive but diffuse staining in all specimens.

Surgically excised specimens of sulcular wall with minimal inflammatory response as judged by clinical then histological criteria were processed for electron microscopy. The specimens were divided into crestal, middle and cervical areas of the sulcular epithelium. The highest concentration of membrane-coating granules was found in the upper spinous cell layers of sulcular epithelium. The profiles of these granules showed examples of both classical keratinized (lamellated) and non-keratinized (non-lamellated) forms but also other appearances that were not derived from them through differences in the plane of section. The population of granules decreased between the crestal and cervical zones, and the decrease in number was marked for the lamellated granules. This decrease in numbers of membrane-coating granules, together with the wider intercellular spaces, may be the reason why the sulcular epithelium is most permeable in the cervical region.

Defensins are known to play an important role in defense against bacteria. It is also known that immunity against infection is compromised with age. The purpose of this study was to evaluate the localization of human beta-defensin (HBD)-2 in human gingival epithelia according to age. Gingival epithelia in maxillary premolar buccal normal regions was immunohistochemically stained for HBD-2. Specimens were divided into two groups: 6 cases in a young group (20 years old) and 7 cases in an elderly group (50 years old). Expression of HBD-2 in gingival epithelium in young subjects was mostly detected in the superficial layer of the parakeratinized layer, while some areas of the spinous cell layer in elderly subjects were positive for HBD-2, as was the superficial layer. Two cases in the young group and 5 cases in the elderly group immunoreacted with HBD-2 in the spinous cell layer. Furthermore, immunoreaction was stronger in the elderly group. The results revealed HBD-2 positive cells in spinous cells in the elderly group and in the parakeratinized layer in the young group.

Summary Background Cells of the granular layer are interconnected by tight junctions (TJs) in normal epidermis. The structural proteins of epidermal TJs include occludin, ZO-1, and claudin-1 and -4. Objectives Our aim was to correlate the expression of TJ components with keratinocyte differentiation using psoriasis as a model of premature keratinization. Methods The distribution of TJ proteins was evaluated in the skin of nine patients with psoriasis. Punch biopsies were taken from perilesional skin, from active psoriasis plaques, and from healed, previously lesional locations. The punch biopsies were analysed using indirect immunolabelling for ZO-1, occludin and claudin-1, -4 and -5. In addition, epidermal samples were analysed by reverse transcription–polymerase chain reaction for claudin-1, -4 and -5 mRNAs. Results Claudin-5 was localized to the granular cell layers of normal control skin as well as perilesional and lesional psoriatic epidermis. This was unexpected, as previous studies have not detected claudin-5 in the epidermis. Occludin and ZO-1 were expressed in the granular cell layer in psoriatic perilesional epidermis. In the psoriasis plaques, ZO-1 and occludin were detected in a wider zone extending from the granular layer to the middle spinous cell layers. In healed psoriasis plaques, the expression of occludin and ZO-1 resumed a normal-looking profile, being restricted to the upper epidermis only. Claudin-1 and -4 did not show marked changes in psoriasis compared with normal skin. Conclusions The results demonstrate claudin-5 in normal epidermis and psoriatic skin, and abnormal distribution of occludin and ZO-1 in psoriasis plaques. Clinical healing of aberrant keratinization is associated with restoration of the normal distribution of occludin, ZO-1 and also involucrin.

Background Pemphigus vegetans, a variant of pemphigus vulgaris, constitutes a rare form of all pemphigus cases, and oral involvement is common. Two clinical subtypes of pemphigus vegetans exist, characterized initially by flaccid bullae and erosions (Neumann) or pustules (Hallopeau). Both subtypes subsequently develop into hyperpigmented vegetative plaques with pustules and hypertrophic granulation tissue at the periphery. Methods We report three cases of pemphigus vegetans with oral manifestations exclusively. Two patients were male aged 30 and 45 years old, respectively, while one was a 51-year-old female. Conclusion Oral lesions in all cases consisted of erosions and whitish, vegetating plaques. The histopathological characteristics were in all cases identical. The spinous cell layer was characterized by intense acanthosis and by the presence of vesicles between the spinous and basal cell layers. Inside the vesicles exudative elements were observed consisting mainly of eosinophils. In the upper lamina propria severe inflammatory reaction was observed. Streptavidin-biotin immunoperoxidase technique showed in all cases intercellular epithelial deposition of IgG and C3.

Objective: The purpose of this study was to determine the clinical and histopathologic presentation of actinic cheilitis. Study design: A retrospective study on 65 patients attending an Oral Medicine clinic in Greece over a 10 year period. For each case the demographic, clinical and histopathologic information were evaluated. Results: The mean age at the time of diagnosis was 53.1 ± 11.4 years. Thirty-nine patients (60%) used tobacco in any form. An outdoor occupation was indicated for 43 (66.2%) patients. The location of the lesions of actinic cheilitis was in all cases on the lower lip. Actinic cheilitis appeared in three forms; white non-ulcerated lesions (29%), erosions or ulcers of the lip (48%), mixed white and erosive (23%). The histopathologic characteristics included increased thickness of keratin layer, alterations of the thickness of spinous cell layer, epithelial dysplasia, connective tissue changes, perivascular inflammation and basophilic changes of connective tissue. In 11 cases (16.9%) the presence of squamous cell carcinoma was observed. Conclusions: This case-series highlights varied clinical presentation of actinic cheilitis among whom a high proportion developed squamous cell carcinoma.

The expression of c-myc oncogene products in oral papillomas was studied by using an immunohistochemical method. The oncogene products were detected in 17(70.8%) of the 24 oral papillomas under study. The expression of the products was evaluated, and the histologic localization in proliferating epithelial cells of the oral papillomas was determined. In the basal cell layer, the products were detected in the nuclei of 16 oral papillomas, and in the cytoplasm of 12 oral papillomas. In the nuclei of cells in the spinous cell layer, the products were detected in 4 oral papillomas, and in the cytoplasm of 13 oral papillomas. In the keratinized cells, the products were not detected in the nuclei, but they were identified in the cytoplasm of three oral papillomas. The results suggested that the c-myc oncogene product might play an important role for proliferation and differentiation of the oral papilloma.

A rat epidermal keratinocyte (REK) line which exhibits histodifferentiation nearly identical to the native epidermis when cultured at an air-liquid interface was used to study the metabolism of hyaluronan, the major intercellular macromolecule present in basal and spinous cell layers. Two different support matrices were used: reconstituted collagen fibrils with and without a covering basal lamina previously deposited by canine kidney cells. REKs formed a stratified squamous, keratinized epithelium on both support matrices. Hyaluronan and its receptor, CD44, colocalized in the basal and spinous layers similar to their distribution in the native epidermis. Most (approximately 75%) of the hyaluronan was retained in the epithelium when a basal lamina was present while most (approximately 80%) diffused out of the epithelium in its absence. While REKs on the two matrices synthesized hyaluronan at essentially the same rate, catabolism of this macromolecule was much higher in the epithelium on the basal lamina (half-life approximately 1 day, similar to its half-life in native human epidermis). The formation of a true epidermal compartment in culture bounded by the cornified layer on the surface and the basal lamina subjacent to the basal cells provides a good model within which to study epidermal metabolism.

This study describes the clinical, radiographic and histopathological features of seven glandular odontogenic cysts. These cysts comprised 0.012% of 5800 jaw cysts diagnosed in a 19-year period. There was strong predilection for the mandible (five of the seven cases). Both clinical and radiographic features were nonspecific. The main histological findings were a nonkeratinized, stratified squamous epithelium lining to the cyst cavity which varied in thickness with superficial eosinophilic cuboidal cells and mucous pools within the spinous cell layer. Daughter cysts were found in the wall of 2 cysts. At surgery, most walls were found to be thin and lumen to contain a serous, low viscosity exudate. Because of the high rate of recurrence found in three cases out of the seven after conservative surgical treatment, careful clinical and radiographic follow-up is recommended.

The expression of SPRR (small proline-rich protein) was investigated in normal human skin and in diseased skin from patients with psoriasis, squamous cell carcinoma, basal cell epithelioma, naevus pigmentosus, ichthyosis vulgaris and several inflammatory skin diseases, by immunohistochemical staining. A polyclonal antibody was raised against a synthetic peptide for a C-terminal common region for SPRR1 and SPRR3. In immunoblot analysis, a positive band of 18 kDa was detected, which showed the presence of SPRR1 in human epidermal keratinocytes. In normal epidermis, positive staining for SPRR was observed in keratinocytes in the granular layer and the uppermost or two spinous cell layers, with no staining of the other spinous or basal layers. The staining was obvious at the cell periphery, weak at the cytoplasm, and absent in the nucleus. Staining was observed in several outer layers of the follicular infundibulum to the isthmus. No staining was detected in the inner root sheath of the hair follicles, hair matrix, sebaceous gland, eccrine gland, eccrine duct, melanocytes, Langerhans cells or fibroblasts. The arrectores pilorum, striated muscles, muscle layers of vessels, and myoepithelia of eccrine gland, were weakly stained. In psoriatic skin, stained keratinocytes were distributed in the spinous cell layers except for the basal layer. In ichthyosis vulgaris, SPRR was barely expressed in the uppermost living cell layers of the epidermis. In epidermolytic hyperkeratosis, degenerated squamous cells widely expressed SPRR. In Darier's disease, dyskeratotic cells were clearly stained. In squamous cell carcinoma, staining was observed in keratotic cells around horny pearls. In basal cell epithelioma, naevus pigmentosus, and malignant melanoma, the tumour cells or naevus cells were not stained. The distribution of SPRR was similar to that of involucrin in normal and several diseased skin, except for ichthyosis vulgaris. We conclude that SPRR is expressed in close association with epidermal differentiation in normal skin and skin diseases. The alteration of the expression of the proteins correlated to terminal differentiation, and differs from disease to disease.

This study was undertaken to analyze keratin gene expression at both the mRNA and protein level in oral hairy leukoplakia (OHL). Comparisons were made with normal lingual epithelium from a similar site, tongue biting, normal buccal mucosa and another condition which disturbs oral epithelial differentiation, white sponge nevus. Combined immunocytochemical and in situ hybridization studies for keratins 14 and 19 were carried out on 2 specimens of OHL from HIV-positive males and one sample each of the other cases. Keratin 14 protein expression was uniform throughout all the epithelia. In normal epithelia and in lesions other than OHL, keratin 14 mRNA was most strongly expressed in basal cells with weaker but still significant amounts in the spinous cell layer. In both cases of OHL there was weaker basal cell expression of keratin 14 mRNA and frequent absence in koilocytoid cells. Keratin 19 protein expression was heterogeneous in the basal layer of all specimens with suprabasal staining of occasional groups of cells. Its mRNA was uniformly distributed in all cases. The findings indicate the keratin mRNA expression does not always parallel that of protein and that, in the case of keratin 14, expression may be influenced by the presence of EBV.

The phenotypic alterations produced in mouse skin cells during the multistage development of squamous cancer have been well documented. In normal skin, all proliferating cells are confined to the basal cell compartment where less than 10% of the cells are in S phase when pulse-labeled with DNA precursors. Two keratins, K5 (M r 60 000) and K14 (M r 55 000), are transcribed largely in basal cells, although the proteins persist in the upper layers (Roop et al. 1988). The commitment to differentiate is associated with the loss of proliferative potential, the commencement of suprabasal migration, and the expression of two suprabasal keratins, K1 (M r 67 000) and K10 (M r 59 000) in the first spinous cell layer (Roop et al. 1988). Proliferating cells do not express K1 or K10 in normal epidermis. As cells migrate into the granular cell layer, K1 and K10 transcripts diminish and the genes for filaggrin, a M r 27 000 interfilamentous matrix protein, and loricrin, a major component of the cornified envelope, are activated and the proteins synthesized (Mehrel et al. 1990; Roop et al. 1989).

Twenty-four oral papillomas were fixed in a 10% formalin solution, and 5 mu paraffin sections were prepared by normal procedure. Using monoclonal antibody NCC-RAS-001 as the primary antibody, the expression of ras p21 in the oral papilloma was searched immunohistochemically by the Histostain SP kit. The ras p21 was detected in 10 (41.7%) of 24 oral papillomas. In the histologic expression, it was less frequently detected in keratinized cell layer and basal cell layer, and it was most often detected in the cytoplasm of the cells in the spinous cell layer. These findings suggested the close relation between the expression of ras p21 and the epithelial proliferation and differentiation in the oral papilloma.

Snuff-induced lesions from 21 snuff users were studied clinically, histologically and by electron microscopy. Clinically most lesions appeared coarsely wrinkled, grayish white and slightly elevated. All lesions were found in the upper vestibular area. Histologically, epithelial thickening, acanthosis, vacuolation of surface cells and a slight subepithelial inflammatory reaction were seen. The electron microscopic findings revealed an intact basement lamina, increased tonofilament concentration moving up towards the surface of the epithelium, widening of intercellular spaces in the spinous cell layer and partial keratinization of surface cells. The salivary flow of resting saliva showed a slightly significant increase compared to controls. That of stimulated saliva was not significantly increased among snuff users.