The latest fashions in skin disease.

The complex nature of epidermal tissue homeostasis is borne out by the range of diseases affecting this tissue. Indeed, mutations in proteins involved in intracellular integrity and cell-cell or cell-matrix adhesion can cause disease in an appropriate epidermal compartment. The most important realization in epidermal disease in the last two years has been that point mutations in key structural genes can result in filaments collapsing, cell cytolysis, or cell adhesion defects; and that these defects can result in severe human skin disease. Now that these associations have been made, the important next step will be to alleviate the suffering of these patients. Animal models will be an important part of these investigations; many molecules including growth factors, oncogenes, and cell adhesion molecules have been targeted to the epidermis of transgenic mice to investigate their role in disease. Such animal models should also elucidate the causes of diseases like psoriasis, a very common skin disease, the molecular basis of which remains elusive. Gene therapy involving the replacement of defective genes or local delivery of therapeutic molecules will be one of the main goals in alleviating these known epidermal diseases. Such protocols in the epidermis are aided by the relative accessibility of the skin and the presence of the "stem cells" in relatively accessible compartments. Indeed, as the last few years have shed much light on the genetic causes of epidermal disease, it is hoped that the next several years will prove as illuminating in the alleviation of these diseases.     

Cell death in the anuran tadpole tail: thyroid hormone induces keratinization and tail-specific growth inhibition of epidermal cells

The mechanism of thyroid hormone-induced and glucocorticoid-modulated death of tail epidermal cells from tadpoles of bullfrog, Rana catesbeiana, was investigated by comparing tail epidermal cells with dorsal body epidermal cells. From morphological and biochemical criteria, there were two types of epidermal cells: basal cells and skein cells. The abundance of these cells was different between the tail and the body skin. Fifty percent of body cells and more than 95% of tail cells were skein cells. Effects of 3,3',5-triiodo-L-thyronine (T3, 10(-8) M) and cortisol (5 X 10(-7) M) were investigated with cultured epidermal cells. T3 differently regulated the keratinization of the tail and body cells. The keratinization of the tail epidermal cells was not observed without T3. T3 induced the keratinization dramatically. On the other hand, body epidermal cells were constantly undergoing keratinization without the hormone: T3 merely accelerated the rate of keratinization. Cortisol generally did not show any significant effect on keratinization. T3 showed opposite effects on DNA synthesis of the tail and body cells: suppression of tail cells and stimulation of body cells. Cortisol weakened the inhibitory effect of T3 on DNA synthesis in tail cells. Immunofluorescent micrographs with anti-BrdU showed that T3 decreased the number of cells in the S phase of the cell cycle in the case of tail cells but not of body cells. Thus, thyroid hormone plays dual roles for the tadpole epidermal cells: one is an induction and a promotion of keratinization in tail and body cells, respectively, and the other is an opposite regulation for the proliferation of both epidermal cells. These roles seem to have crucial connections to a tail-specific cell death induced by thyroid hormone.     

Derivation of keratinocyte progenitor cells and skin formation from embryonic stem cells

Despite numerous elegant transgenic mice experiments, the absence of an appropriate in vitro model system has hampered the study of the early events responsible for epidermal and dermal commitments. Embryonic stem (ES) cells are derived from the pluripotent cells of the early mouse embryo. They can be expanded infinitely in vitro while maintaining their potential to spontaneously differentiate into any cell type of the three germ layers, including epidermal cells. We recently reported that ES cells have the potential to recapitulate the reciprocal instructive ectodermal-mesodermal commitments, which are characteristic of embryonic skin formation. Derivation of epidermal cells from murine ES cells has been successfully established by exposing the cells to precisely controlled instructive influences normally found in the body, including extracellular matrix and the morphogen BMP-4. These differentiated ES cells are able to form, in culture, a multilayered epidermis coupled with an underlying dermal compartment similar to native skin. This bioengineered skin provides a powerful tool for studying the molecular mechanisms controlling skin development and epidermal stem cell properties.     

Clinical interest of cutaneous models reproducedin vitro for severe burn treatment: histopathological and ultrastructural study

The healing of minimal skin lesions is usually obtained by epidermal migration and proliferation from peripheral wound margins. However, cutaneous grafts or reconstituted skin are necessary for severe injuries. Various models have recently been reproduced for this purpose. The aim of this work is to report the histopathologic evolution of burn lesions treated two years ago by autologous epidermis (Genzyme Tissue Repair, Boston, USA). Fifteen patients with severe burns (more than 80% of surface) have been treated. These observations have been based exclusively on biopsies of grafted wounds. Cultured epidermis is rapidly fully differentiated after grafting with temporary hyperplasia and normal strata. At 18 months, rete ridges formation is present only in young patients. Melanocytes and Langerhans' cells repopulated grafts rapidly. The use of cultured epidermis nowadays represents an important improvement in burn treatment.Abbreviations CEA cultured epidermal autologous sheets - TBSA total burn surface area     

Establishment of epidermal cell columns in mammalian skin: computer simulation

The present investigation shows an example of tissue formation in relation to individual cell properties. Epidermis of mammalian skin has recently been shown to be organized during ontogeny into neat vertical cell columns, in which a cell approximates the flattened form of Kelvin's tetrakaidecahedron, a 14-sided body with eight hexagonal faces and six square faces. Such an epidermal architecture is compatible with the organization and turnover of stacked cells, a constant loss of surface cells and a supply that cells from a basal layer differentiate during migration to the surface. The developmental process from irregular cell aggregate into neat columns in skin is simulated on a digital electronic computer with the assumption as follows: a new cell migrating upward from a basal layer jostles and settles at the less crowded area among upper cells. After migration of many cells to the surface, the simulation demonstrates that cells have been stacked in neat columns, and the stability of the architecture is also shown.     

INFLUENCE OF HAIR PLUCKING ON THE TURNOVER TIME OF THE EPIDERMAL BASAL LAYER

The epidermal cell kinetics of male DBA-2 mice have been studied using tritiated thymidine. Liquid scintillation data from skin punches, taken after stimulation of hair growth by plucking, agree well with similar data from DBA-1 mice. A technique has been devised for obtaining sheets of epidermal cells from haired mice. Labelling index values from these sheets show that epidermal proliferation increases after plucking and they agree well with values obtained from sections. Counts of cells per unit area of epidermis show that cells are removed by plucking.
Using an estimated value for the length of S, the turnover time of the basal layer was calculated. The growth fraction and proliferative cell cycle time have also been estimated.     

Burn injury suppresses human dermal dendritic cell and Langerhans cell function

Human skin contains epidermal Langerhans cells (LCs) and dermal dendritic cells (DCs) that are key players in induction of adaptive immunity upon infection. After major burn injury, suppressed adaptive immunity has been observed in patients. Here we demonstrate that burn injury affects adaptive immunity by altering both epidermal LC and dermal DC functions. We developed a human ex vivo burn injury model to study the function of DCs in thermally injured skin. No differences were observed in the capacity of both LCs and dermal DCs to migrate out of burned skin compared to unburned skin. Similarly, expression levels of co-stimulatory molecules were unaltered. Notably, we observed a strong reduction of T cell activation induced by antigen presenting cell (APC) subsets that migrated from burned skin through soluble burn factors. Further analyses demonstrated that both epidermal LCs and dermal DCs have a decreased T cell stimulatory capacity after burn injury. Restoring the T cell stimulatory capacity of DC subsets might improve tissue regeneration in patients with burn wounds.     

A simple method for the evaluation of epidermal thickness variations in psoriasis vulgaris

OBJECTIVE: To evaluate the "variation" of the epidermal thickness and variables such as the mean, maximum, and minimum thickness for the diagnosis and follow-up of some skin diseases such as psoriasis. STUDY DESIGN: A simple quantitative method is described. A series of psoriasis vulgaris cases is provided as an example. Biopsy samples taken before and after topical treatment were used. RESULTS: The mean thickness was higher in lesions compared to the perilesional control skin (p < 0.001) and post-treatment lesional area (p < 0.001). CONCLUSION: The thickness of the epidermis varies due to a multitude of factors. Measuring the mean epidermal thickness may be important. However, this measurement may not be the best way for studies involving a series of patients and a variety of lesion sites. The natural variations in the thickness of the epidermis of different body sites and of different persons may prevent the usage of thickness measurements alone on a wider scale. It is hoped that this method can be used for evaluation of certain skin diseases affecting epidermal thickness where variation of the epidermal thickness is a potentially useful diagnostic and therapeutic variable.     

Dermal matrix proteins initiate re-epithelialization but are not sufficient for coordinated epidermal outgrowth in a new fish skin culture model

We have established a new culture system to study re-epithelialization during fish epidermal wound healing. In this culture system, fetal bovine serum (FBS) stimulates the epidermal outgrowth of multi-cellular layers from scale skin mounted on a coverslip, even when cell proliferation is blocked. The rate of outgrowth is about 0.4 mm/h, and at 3 h after incubation, the area occupied by the epidermal sheet is nine times larger than the area of the original scale skin. Cells at the bottom of the outgrowth show a migratory phenotype with lamellipodia, and “purse string”-like actin bundles have been found over the leading-edge cells with polarized lamellipodia. In the superficial cells, re-development of adherens junctions and microridges has been detected, together with the appearance and translocation of phosphorylated p38 MAPK into nuclear areas. Thus, this culture system provides an excellent model to study the mechanisms of epidermal outgrowth accompanied by migration and re-differentiation. We have also examined the role of extracellular matrix proteins in the outgrowth. Type I collagen or fibronectin stimulates moderate outgrowth in the absence of FBS, but development of microridges and the distribution of phosphorylated p38 MAPK are attenuated in the superficial cells. In addition, the leading-edge cells do not have apparent “purse string”-like actin bundles. The outgrowth stimulated by FBS is inhibited by laminin. These results suggest that dermal substrates such as type I collagen and fibronectin are able to initiate epidermal outgrowth but require other factors to enhance such outgrowth, together with coordinated alterations in cellular phenotype. This work was supported by grants from the Faculty of Science at Toho University and from Fujirebio to M.S.     

Allodynic skin in post-herpetic neuralgia: histological correlates

Most post-herpetic neuralgia (PHN) patients suffer from tactile allodynia (pain evoked by lightly touching the skin) and it is frequently the dominant clinical manifestation. The pathophysiology of tactile allodynia in PHN patients is poorly understood and this is one of the major limits to the development of appropriate therapies. Epidermal nerve fibres (ENFs) are free nerve endings of small-diameter A-delta and C primary afferents, which can easily be assessed by neurodiagnostic skin biopsy (NSB). The aim of this study was to establish the correlation between the residual epidermal innervation of the allodynic skin and the intensity of tactile allodynia in that area. Twenty-five patients (13 males and 12 females) with PHN were enrolled. Eighteen patients had PHN in the thoracic dermatome, four in the cervical, two in the trigeminal and one in the lumbar. The severity of allodynia evoked by a paintbrush was graded according to an eleven-point numerical scale. A skin biopsy was obtained from the maximal allodynia area and from the contralateral skin. Nerve fibres were labelled with indirect immunofluorescence. Results showed that epidermal innervation was lower in the allodynic skin than in the contralateral skin, although there was great variability among patients. There was no correlation between severity of allodynia and epidermal innervation of the PHN skin. In conclusion, the present study further indicates peripheral nervous system involvement in PHN but does not support a direct correlation between epidermal innervation changes and tactile allodynia.     

A suspensory jacket permitting long-term sitting in paraplegic patients

A thoracic suspensory jacket has been used in four patients with recurring ischial pressure sores to transfer the site of skin pressure from the insensate ischial area to the sensory area over the rib cage. This modality is not an alternative to surgery, but rather an adjunct to prevent an early recurrence after an ischial pressure sore has been repaired with a proper surgical procedure.     

Impaired wound healing in transgenic mice overexpressing the activin antagonist follistatin in the epidermis

Recently, we demonstrated a strong upregulation of activin expression after skin injury. Furthermore, overexpression of this transforming growth factor beta family member in the skin of transgenic mice caused dermal fibrosis, epidermal hyperthickening and enhanced wound repair. However, the role of endogenous activin in wound healing has not been determined. To address this question we overexpressed the soluble activin antagonist follistatin in the epidermis of transgenic mice. These animals were born with open eyes, and the adult mice had larger ears, longer tails and reduced body weight compared with non-transgenic littermates. Their skin was characterized by a mild dermal and epidermal atrophy. After injury, a severe delay in wound healing was observed. In particular, granulation tissue formation was significantly reduced, leading to a major reduction in wound breaking strength. The wounds, however, finally healed, and the resulting scar area was smaller than in control animals. These results implicate an important function of endogenous activin in the control of wound repair and scar formation.     

Pathogenesis of acute ulceration response (AUR) in hybrid striped bass

In a previous study, we discovered that acute confinement stress causes rapid ulceration of the fins of hybrid striped bass Morone saxatilis female x M. chrysops male (Noga et al. 1998. Vet Pathol 35:102-107). In this paper, we report the development of a reproducible model for studying this phenomenon in juvenile hybrid striped bass. We also determined how quickly ulceration could develop in acutely stressed fish and documented the sequential light microscopic and ultrastructural changes associated with this response. When hybrid striped bass were subjected to a standardized confinement protocol, the pathological response was extremely rapid (fin ulceration began to develop within 15 min of confinement). Grossly, the distal edges of the fins became blanched, and melanophores aggregated near the basement membrane and dermis after 15 min of confinement. Microscopically, the earliest detectable change in the fins, which occurred within 15 min of confinement, was swelling and loss of microridges of the outermost epidermal cells; this was followed by epidermal erosion. After 30 min of stress, epidermal ulceration developed at the distal edges of the fins. At this time, both necrotic and apoptotic epidermal cells were present. The middle and basal epidermal layers were severely spongiotic and the dermis and hypodermis were edematous. Over longer periods (up to 2 h), lesions were similar but increasingly more severe, progressing from the distal edge of the fin towards the base. The response to acute stress showed a significant correlation between confinement period and severity of the pathological changes (epidermal degeneration, epidermal ulceration and leukocyte infiltration). Also, we demonstrated that epidermal damage was not restricted to the fins but also affected the body skin and eyes. The ventral area of the body and the corneal epithelium of stressed fish were ulcerated; however, skin on the head and operculum was not affected, suggesting a site-specific mode of damage. In stressed fish, epidermal ulceration was found in 67 to 97% of all fins, 88% of skin on the ventrum, and 67% of corneas, while control fish had only very mild epidermal ulceration in the few fish in which it was present (on 5 to 10% of the fins, but not on the ventral skin or corneas). Due to the widespread damage to epidermal tissues of the body surface, we have named this the acute ulceration response (AUR). Our study indicates that acute confinement can rapidly cause significant damage to epidermal and ocular epithelium. AUR might be a primary cause of morbidity in acutely stressed fish.     

Molecular,cellular and histological changes in skin from a larval to an adult phenotype during bony fish metamorphosis

Developmental models for skin exist in terrestrial and amphibious vertebrates but there is a lack of information in aquatic vertebrates. We have analysed skin epidermal development of a bony fish (teleost), the most successful group of extant vertebrates. A specific epidermal type I keratin cDNA (hhKer1), which may be a bony-fish-specific adaptation associated with the divergence of skin development (scale formation) compared with other vertebrates, has been cloned and characterized. The expression of hhKer1 and collagen 1α1 in skin taken together with the presence or absence of keratin bundle-like structures have made it possible to distinguish between larval and adult epidermal cells during skin development. The use of a flatfish with a well-defined larval to juvenile transition as a model of skin development has revealed that epidermal larval basal cells differentiate directly to epidermal adult basal cells at the climax of metamorphosis. Moreover, hhKer1 expression is downregulated at the climax of metamorphosis and is inversely correlated with increasing thyroxin levels. We suggest that, whereas early mechanisms of skin development between aquatic and terrestrial vertebrates are conserved, later mechanisms diverge. This work was carried out within the project “Arrested development: The Molecular and Endocrine Basis of Flatfish Metamorphosis” (Q5RS-2002-01192) with financial support from the Commission of the European Communities. It does not necessarily reflect the Commission’s views and in no way anticipates its future policy in this area. This project was further supported by Pluriannual funding to CCMAR by the Portuguese Science and Technology Council. M.A. Campinho was sponsored by the Portuguese Ministry of Science (grant no. SFRH/BD/6133/2001).     

Cultured epithelial autografts for giant congenital nevi

Eight pediatric patients with giant congenital nevi confluent over 21 to 51 percent body surface area were treated by excision and grafting. The nevus was excised to the muscle fascia, and the open wound was grafted with cultured epithelial autografts and split-thickness skin grafts. The patients have been followed from 17 to 56 months. Seventeen operations were performed in the eight patients, excising a mean of 6.9 percent body surface area at each procedure. The mean duration of anesthesia was 3.7 hours, and the mean operative blood loss was 12.3 percent estimated blood volume. The mean "take" for the cultured epithelial autografts was 68 percent, and for the split-thickness skin grafts, 84 percent. Epithelialization of open wound areas adjacent to the grafts was somewhat slower for the cultured epithelial autografts than for the split-thickness skin grafts, but it led to a healed wound in all patients except one. Ten of the 17 areas grafted with cultured epithelial autografts resulted in small open wounds that required regrafting. Wound contraction under the cultured epithelial autografts and under split-thickness skin grafts was similar and depended more on the anatomic site grafted than on the type of graft employed. in 16 of 17 operations, the cultured epithelium remained as a permanent, durable skin coverage. The use of cultured epithelial autografts allowed a larger area of excision than would have been possible with split-thickness skin grafts alone and, therefore, a more rapid removal of nevus. Cultured epithelial autograft are an important new technique in the care of patients with giant congenital nevi.     

Characterization of human cytokeratin 2, an epidermal cytoskeletal protein synthesized late during differentiation.

Among the more than 30 different human proteins of the cytokeratin (CK) group of intermediate filament (IF) proteins, the significance of the epidermal polypeptide CK 2 (Moll et al., 1982, Cell 31, 11-24) has been repeatedly questioned in the literature. Here, we show, by in vitro translation and protein gel electrophoresis, that human epidermis from various body sites does indeed contain relatively large amounts of mRNA encoding a distinct polypeptide comigrating with native epidermal CK 2. We also report the isolation of a cDNA clone encoding the complete sequence of CK 2, which is a type II CK different from--but related to--epidermal CKs 1 and 5 on the one hand and corneal CK 3 on the other. The mRNA of approximately 2.6 kb encodes a polypeptide of 645 amino acids and M(r) 65,852, in good agreement with the value of 65.5 kDa previously estimated from gel electrophoresis. This human CK, the largest so far known, displays several features typical of CKs of stratified epithelia, including numerous repeats of glycine-rich tetrapeptides in the head and tail domains. Northern blot and in situ hybridizations have shown that CK 2 is expressed strictly suprabasally, usually starting in the third or fourth cell layer of epidermis, and this was confirmed at the protein level by immunohistochemistry using CK 2-specific antibodies. The protein has been detected as a regular epidermal component in skin samples from different body sites, albeit as a minor CK in "soft skin" (e.g., breast nipple, penile shaft, axilla), but not in foreskin epithelium and in other epithelia, in squamous metaplasias and carcinomas, or in cultured cell lines derived therefrom. We propose that CK 2 is a late cytoskeletal IF addition synthesized during maturation of epidermal keratinocytes which probably contributes to terminal cornification.     

Treating the abdominotorso region of the massive weight loss patient: an algorithmic approach

There has been tremendous growth in the number of patients seeking body contouring procedures after massive weight loss. Most patients desire improvement of the abdominotorso region first. After massive weight loss, there is enormous variability of body proportions, and therefore there have been many surgical options proposed based on the quality of the skin, subcutaneous fat component, and location of the lax tissue. Each area needs to be assessed to see whether there is a significant lower abdominal component, an upper midline abdominal component, or contributions from the buttocks and flanks. An algorithm for treatment is presented to simplify the decision-making process. Patient examples are also shown to demonstrate the usefulness of the algorithm.