Scanning electron-microscopic findings in orthoergically damaged skin surfaces of hairless mice (author's transl)

Mechanic-chemically defined irritation of the skin surface of hairless mice regularly leads to macroscopically sub-visible changes of the horny layer. Scanning electron-microscopic analysis reveals extended regular horny layer dehiscences, as well as deeper dehiscence or trench formations in the area of irritation, as compared to normal skin. The damaged horny layer sample, presented respectively, depends-to a degree-upon type, intensity and duration of irritation. Hairless mice proved to be best suited at test animals for informative studies concerning cutaneous resistance to stress.     

Perturbation in connexin 43 and connexin 26 gap-junction expression in mouse skin hyperplasia and neoplasia

To examine the possible role of gap junctions in mouse skin tumor progression, we generated a panel of mouse skin tissue samples exhibiting normal, hyperplastic, or neoplastic changes and characterized the expression of the gap-junction genes connexin 43 (Cx43) and connexin 26 (Cx26) by in situ hybridization and immunohistochemical analyses. In normal skin, these two gap junction genes were differentially expressed; Cx43 was found predominantly in the less differentiated lower spinous layers, whereas Cx26 was found in terminally differentiating upper spinous and granular layers. In hyperplastic epidermis exhibiting an expansion of the differentiated upper layer, i.e., epidermis with a thickened granular layer or in which the granular layer was replaced with keratinocytes exhibiting tricholemmal differentiation, expression of Cx43 and Cx26 remained segregated in the lower and upper spinous layers, respectively. However, in papillomas, Cx26 was localized in the lower but not upper spinous layer, an expression pattern identical to that of Cx43. In addition, the overall expression levels of both Cx43 and Cx26 appeared to be greatly elevated in the papillomas. It is interesting that such marked alteration in the pattern of Cx26 expression occurred within the context of hyperplastic changes histologically identical to those seen in the nonpapillomous hyperplasias. Interestingly, in neoplastic skin lesions containing a squamous cell carcinoma, Cx43 and Cx26 expression was extinguished. Moreover, expression of Cx43 was also significantly reduced in adjacent apparently nonneoplastic tissues. Overall, these observations show that perturbations in gap-junction gene expression are associated with skin hyperplasia and neoplasia. Such findings suggest a possible role for gap junctions in the malignant conversion of mouse epidermal cells.     

Measuring memory for source: some theoretical assumptions and technical limitations

In a search for pathogenetic mechanisms underlying retention hyperkeratosis, we examined the pH gradient over the stratum corneum in 13 male patients suffering from either x-linked recessive (XRI) or autosomal dominant ichthyosis vulgaris. For recording pH values, a flat glass electrode was repeatedly applied to the skin during tape stripping of mildly involved forearm skin. Before stripping, surface pH was higher in ichthyosis vulgaris (5.3 +/- 0.7; n = 7) than in XRI (4.6 +/- 0.4; n = 6; p < 0.05) and healthy control men (4.5 +/- 0.2; n = 7; p < 0.01). Removal of stratum corneum, which required 100-240 strippings in ichthyotic skin and 80-120 strippings in healthy control skin, disclosed markedly different pH variations in the two types of ichthyosis. The major abnormality in ichthyosis vulgaris skin was that a neutral pH was attained already halfway through the horny layer, possibly reflecting a congenital lack of acidic breakdown products from keratohyaline. By contrast, stripping of XRI skin revealed a shallow pH gradient that plateaued at 6.2-6.6, instead of about 7 as in normal and ichthyosis vulgaris skin. A likely explanation is the XRI-associated accumulation of cholesterol sulfate in lower stratum corneum. Our results suggest that the "acid mantle" of normal skin, which penetrates deep into the stratum corneum, is the combined result of cornification-associated organic acids and back-diffusion of acid material from the surface. Because corneocyte desquamation involves many pH-dependent enzymes, abnormalities in the transcorneal pH gradient might play a role in the pathogenesis of ichthyosis.     

Effects of calcium and phosphate on catecholamines, ATP and dopamine beta-hydroxylase of chromaffin medullary granules

The penetration into the single layers of human skin in vitro of 17 alpha-estradiol, 17 beta-estradiol and estriol was investigated. The radiolabeled substances were incorporated into 4 standard ointments and into an alcoholic solution mixture. After application to the skin and after different penetration periods, the horny layer was taken off by adhesive tape stripping. The epidermis and the dermis were separated by slicing them down parallel to the skin surface in a freeze microtome. In each single layer, the amount of substance was determined and calculated relative to the applied quantity and in absolute concentrations (mug per tissue weight and molarity). Besides the expected dependence of the penetration on the type of ointment, there is a distinct dependence on the chemical structure: estriol penetrates considerably slower and in less concentrations into the living layers of the human skin than the estradiols. Furthermore, estriol reaches the dermis only in low concentrations so that this substance may be termed epidermotropic. 17 alpha-estriol which has only weak sexhormone properties in humans penetrates as well as the sexhormone 17 beta-estradiol.     

Light and electron microscopy of excoriated psoriatic lesions in patients during topical treatment

This is a light- and electron-microscopic study of excoriated, recalcitrant psoriatic lesions in ambulatory patients undergoing topical treatment. During regeneration of a stratum corneum without scales, in areas showing excoriations bounded by a non-cornified epidermis, horny cell layers, designated I and II, are formed. In areas in which excoriations are delimited by an epidermis with a retained stratum corneum, the latter is composed of a horny cell layer I and/or II or III. The horny cell layers differ, as shown by light- and electron-microscopy. In the less fresh excoriations, a homogeneous substance, which includes fibrin deposits, is observed. This substance is also seen frequently in the stratum corneum. In the less fresh excoriations and in the stratum corneum there are numerous polymorphonuclear leukocytes. In these locations, however, they are only found simultaneously with the homogeneous substance containing fibrin deposits. Polymorphonuclear leukocytes are also seen in the dermal papillae and in the non-cornified epidermis, particularly in the macroscopically "old" excoriations. The changes observed in these recalcitrant lesions have been interpreted as secondary and are believed to be expressions of a non-specific reaction to the mechanical damage to which the ambulatory patients have constantly exposed the lesions. They are not interpreted as changes in the psoriatic condition per se.     

The epidermal architecture of an in vitro reconstructed human skin equivalent (Advanced Tissue Sciences Skin2 Models ZK 1300/2000)

We investigated the epidermal ultrastructure of an organotypic skin culture model over a culture period of one week. At delivery, the stratified keratinizing epidermis consists of a basal layer with hemidesmosomes, a spinous layer with desmosomes, a granular layer with keratohyalin and membrane coating granules, a horny layer enclosing many lipid droplets. At the epidermal-dermal junction laminae lucida and densa are evident. After 7 days of culture the multilayered cornifying aspect persists; the number of viable cell layers remains constant. Spinous cells display abundant glycogen deposits. In the granular layer keratohyalin granules are more abundant. Both the size and number of hemidesmosomes increase. Anchoring fibrils are observed. From a structural point of view the model remains stable and well differentiated in culture and, therefore, may be well suited for pharmacological purposes.     

Penetration, permeation, and resorption of 8-methoxypsoralen. Comperative in vitro and in vivo studies after topical application of four standard preparations

The penetration, permeation, and resorption of radioactively labelled 8-Methoxypsoralen was investigated in human skin. Siultaneously, the effects to time and ointment carrier on the penetration kinetics were ascertained. The carriers tested were: vaseline, aqueous wool-wax alcohol ointment, aqueous hydrophilic ointment and polyethylene glycol ointment. The absolute concentrations of 8-Methoxypsoralen were estimated in the horny layer, epidermis and dermis. With the most advantageous carrier, aqueous wool-wax alcohol ointment, 4-6X10(-5) M and 10(-5) M were attained in the epidermis and dermis, respectively. Moreover, it was shown that the substance penetrates rapidly (10 min) into the epidermis and dermis and the high concentrations reached constant over a period of 16 h. Only with a formulation of aqueous wool-wax alcohols is any accumulation at all achieved in the deeper areas of the horny layer. A uniform decrease in drug concentration with increasing depth of the horny layer is found with the other 3 vehicles, whereby slight variations in concentrations pertain from carrier to carrier. 4 h after local application, 8-Methoxypsoralen can be detected in the urine. Regardless of the ointment base employed, 8-Methoxypsoralen is no longer detectable in the urine 40 h after application. In comparison to the oral therapy, the same magnitude of percutaneous resorption into the central compartment is to be derived from the data, if half the body surface is treated locally.     

Gingival involvement in mucous membrane pemphigoid

A 60-year-old woman with clinical features of desquamative gingivitis had a history of painful, blistering gingival lesions for more than 2 years. There were no other accompanying mucosal or skin lesions. Clinical examination revealed erythematous and edematous gingiva with ulcerated areas and evidence of intact and ruptured bullae. White plaquelike lesions were also noted. Gingival manipulation caused epithelial desquamation. Light microscopic examination of biopsy specimens from the perilesional gingival tissue showed separation of the oral gingival epithelium and connective tissue at the margin of the collapsed bulla. A moderately intense inflammatory infiltrate was present in the connective tissue. Direct immunofluorescent microscopy revealed a continuous linear deposition of immunoglobulin G and C3 at the basement membrane zone. On the basis of clinical, histopathologic, and immunofluorescent findings, the diagnosis of mucous membrane pemphigoid was made.     

Expression of low affinity NGF (p75) receptors in rat superior colliculus: studies in vivo, in vitro, and in fetal tectal grafts

The distribution and level of expression of the low affinity nerve growth factor receptor (LNGFR) in the rat visual system has been investigated under a number of experimental conditions. The aim was to determine the cellular location of the receptor and to study the factors which influence its expression. The monoclonal antibody 192-Ig and immunohistochemical techniques were used to examine LNGFR expression in (i) developing and adult rat superior colliculus (SC), (ii) fetal collicular tissue transplanted to the midbrain of newborn host rats, (iii) the SC of rats which had been unilaterally enucleated at birth, and (iv) mixed glial cell cultures from the neonatal SC. The effect of eye removal on LNGFR immunoreactivity in other normally retino-recipient areas was also assessed. Postnatal maturation of the rat SC was associated with an increase in LNGFR immunoreactivity. At birth, weak staining was seen ventral to the superficial gray layer. Staining gradually became located more dorsally until in the adult there was a dense band of immunoreactivity that extended 50-100 microns from the surface. Immunoreactive processes and cellular-like profiles were seen. Compared to adult host SC there was considerably more LNGFR immunoreactivity in transplanted tectal tissue, irrespective of whether the grafts were connected to the host. LNGFR expression in transplants was patchy and usually contiguous with the graft surface. Staining was not obviously related to the distribution of astrocytes or microglia and very few cells were LNGFR positive in tectal glial cell cultures. In SC and in tectal grafts it is probable that LNGFR immunoreactivity was primarily associated with intrinsic neurons. In support of this, LNGFR expression in the superficial SC was unaffected by neonatal eye removal; however, LNGFR staining in the pretectum and diencephalon was reduced or absent on the side contralateral to visual deafferentation. These conflicting sets of data suggest that (i) LNGFRs in central visual pathways are sometimes, but by no means always, associated with retinal innervation and (ii) LNGFR expression in visual target areas originates from diverse sources and is influenced and regulated by a variety of factors.     

Biochemical and molecular basis of Bernard-Soulier syndrome: a review

Bernard-Soulier syndrome (BSS) is a rare hereditary recessive autosomal bleeding disorder characterized by a prolonged bleeding time, giant platelets, thrombocytopenia, normal platelet aggregation in response to ADP and no agglutination in response to ristocetin. This disease is due to absence or abnormality of the platelet membrane glycoprotein GPIb-IX-V, the receptor for von Willebrand factor. All four genes encoding the complex have been cloned and 17 forms of BSS have to date been characterized at the functional, immunological and molecular levels. The mutations can be divided into two main groups. Firstly, mutations located in leucine rich repeats (LRR), responsible for conformational modifications of the molecule, in some cases higher sensitivity to proteases and loss of adhesive function of the receptor, which is expressed at lower than normal levels at the platelet membrane. When mutations affect the LRR of GPIbalpha, the presence of the other chains varies from normal to residual amounts. When mutations affect the LRR of GPIX, expression of the other chains is strongly diminished, suggesting that GPIX plays a major role in the stability of the complex. A second type of mutations leads to synthesis of a truncated molecule lacking the transmembrane domain and absence of its expression at the platelet surface, while the other chains are present in residual amounts. Expression of recombinant proteins in eukaryotic cells has recently confirmed the results derived from studies of natural mutations. Separate expression of each chain can be obtained, although the presence of all subunits is required for full expression.     

Gas transfer and in vitro and in vivo blood compatibility of a fluorinated polyimide membrane with an ultrathin skin layer

The authors have synthesized fluorinated polyimides to develop a novel membrane oxygenator combining excellent gas transfer and blood compatibility. Gas exchange membranes of fluorinated polyimides prepared by a dry/wet process showed an asymmetric structure and consisted of an ultrathin and defect-free skin layer supported by a porous substructure. The asymmetric polyimide membranes never incurred plasma leakage because of the defect-free skin layer of the membrane surface. The calculated, apparent defect-free skin layer thickness of the asymmetric membrane was approximately 20 nm. Carbon dioxide and oxygen transfer rates through the membranes were dramatically enhanced because of the ultrathin skin layer and were 96 and 64 times larger than those determined in currently available oxygenator polymer membranes, such as polydimethylsiloxane (PDMS). For the evaluation of in vitro blood compatibility, platelet adhesion and plasma protein adsorption on the polyimide membranes were measured by using scanning electron microscopic examination and an amino acid analyzer. Deformation and aggregation of platelets adherent to the membranes were not observed, and the number of platelets was 1.6 micrograms/cm2, which was one-sixth less than the value measured in PDMS. For in vivo evaluation, the polymer tubes were implanted in the femoral vein of a mongrel dog for 7 days. Thrombus formation and fibrin were found on the surface of PDMS. However, thrombus formation was not observed on the polyimide. These results indicate that the fluorinated polyimides show excellent blood compatibility and are a promising membrane material for an oxygenator.     

Dermatophytic disease: exuberant hyperkeratosis with cutaneous horns

BACKGROUND: Dermatophytic disease is a chronic dermatophytosis involving the skin and viscera and caused by benign dermatophytes which cross the skin barrier. We present a case presenting previously unreported giant cutaneous horns. CASE REPORT: A male Algerian patient aged 29 years consulted for chronic skin disease which had progressed for 2 years. Manifestations included warts, papulonodules, eczematiform lesions, lichenifications, alopecia, onyxis, multiple node enlargement and giant cutaneous horns on the plantar aspect of the feet making walking difficult. The patient's kinhood included 10 members and the patient's parents were first cousins; no other family member had a similar disease state. Laboratory findings included eosinophilia (11%, 550 cells/mm3), an inflammatory syndrome, and normal phosphorus and calcium levels despite demineralization of the hands and feet. The tuberculin interdermal reaction was positive but the trichophytine interdermal reaction was negative. Trichophyton violaceum was found in all mycological samples from skin and nails. The histology examination of skin biopsies revealed acanthosis, considerable hyperkeratosis, and myceleal filaments invading the horny layer, the sweat glands and hair follicles. Abscesses and granulomas were also seen in the superficial dermis. Oral griseofulvin 1 g per day and topical applications with ketoconazole gave clear improvement. DISCUSSION: Dermatophytic disease is a chronic dermatophytosis observed mainly in North Africa. The locally high rate of consanguinous marriages would suggest autosomal recessive inheritance of a genetic anomaly possibly associated with deficient cellular immunity. The lesions are polymorphous and develop progressively, probably due to diminished cellular immunity. This case with giant cutaneous horns would be a new variant. Antifungals can provide definitive cure. The pathogenesis of this severe dermatophytosis remains to be established.     

Failure conditions of glass filament yarns: a contribution to the valuation of carcinogenic potentials of fiber fragments

BACKGROUND: The circumanal glands of the dog are thought to be a glandular tissue, but there is some controversy as to whether they should be classified as exocrine or endocrine. In this study, we examined the nature of the circumanal glands to determine whether they should be described as exocrine, endocrine, or something else altogether. In addition, we investigated the cell degeneration in lobules of the circumanal glands in relation to the apocrine glands. METHODS: Light microscopic observations were made of paraffin sections stained with hematoxylin and eosin, and after immunohistochemical staining with antibodies against alpha-smooth muscle actin, keratin, filaggrin, and 3beta-hydroxysteroid dehydrogenase/isomerase (3beta-HSD). Samples were also examined by electron microscopy after fixation by aldehyde perfusion. RESULTS: The lobules of circumanal glands could be divided into two types on the basis of the presence or absence of cysts. Four layers (I-IV) were detected in the lobules with cysts. The outermost layer (layer I or the basal layer) consisted of flattened cells that contained bundles of tonofilaments and were stained immunohistochemically with the antibody against keratin. Layer II (the polyhedral or "spinous" layer) consisted of polyhedral cells that contained bundles of tonofilaments. These cells were connected to adjacent cells by desmosomes, interdigitations, and gap junctions, and they were immunopositive for keratin. A small number of polyhedral cells were immunopositive for 3beta-HSD. Layer III (the granular layer) was composed of flattened cells that contained hematoxylin-stainable granules and were moderately immunopositive for filaggrin. The innermost layer (layer IV or the horny layer) consisted of keratin. Lobules without cysts consisted only of layer I (the basal layer) and layer II (the polyhedral layer). Lobules of the circumanal glands were not directly connected to apocrine glands. Polyhedral cells degenerated and were phagocytosed by basal cells at a periphery of lobules. Then, basal cells phagocytosing degenerated polyhedral cells escaped from lobules, moved into the walls of apocrine glands, and, finally, dropped into the lumen of apocrine glands. CONCLUSIONS: Lobules of the circumanal glands have many characteristics of epidermis (a basal layer, a polyhedral or "spinous layer," a granular layer, and a horny layer) and they should not be classified as glandular tissue. The cysts in lobules can be interpreted as "closed hair canals." We suggest that steroid metabolism might occur in the polyhedral cells of the lobules.     

Electromechanical properties of the single cell-layered heart of tunicate Boltenia ovifera (sea potato)

The tubular heart of the sea potato is composed of a single layer of myoepithelial cells interconnected near the extraluminal surface by specialized junctions. If these junctions are used as the border which separates the luminal from extraluminal membrane, the surface area ratio, luminal:extraluminal, is approximately 12:1. A single myofibril is located near the luminal surface in each cell. Current passed across the heart wall in the direction that depolarizes the luminal membrane and hyperpolarizes the extraluminal membrane immediately produces "all-or-none" action potentials and contractions. Current passed in the opposite direction fails to produce action potentials until after the break of the stimulus, suggesting anodal break excitation of the hyperpolarized luminal membrane. High potassium solutions depolarized the myoepithelium and produced contractions only when applied to the luminal surface of the heart. [Ca]0 increases and [Mg]0 decreases twitch tension only on the luminal surface of the heart. The transwall resistivity is low (50-100 omega/cm2) due to an extracellular shunt. Because of this shunt and the larger surface area of the luminal membrane, the extraluminal membrane is effectively clamped to the potential of the luminal membrane and is not capable of directly influencing excitation-contraction coupling. These findings suggest that only the luminal membrane of the sea potato myoepithelium is capable of generating an action potential and triggering contraction.     

The S1(1A1)-S0(1A1) Electronic Transition of Jet-Cooled o-Difluorobenzene

Retinal pigment epithelium transplantation has been proposed as adjunctive treatment for age-related macular degeneration following surgical excision of choroidal neovascular membranes. The goal of this study was to develop a model to evaluate retinal pigment epithelium transplantation onto human Bruch's membrane in vitro. We investigated the ability of cultured fetal human retinal pigment epithelium to colonize human cadaver Bruch's membrane, determined the incubation time needed to form a monolayer and to exhibit apical microvilli and tight junctions, and assessed the production of basement membrane. Freshly enucleated (less than 48 hours old) human eyes were cut through the pars plana, and the anterior segment, vitreous, and retina were removed. The native retinal pigment epithelium was debrided with a surgical sponge. Bruch's membrane and choroid at the macula were trephined with a 7.0 mm diameter trephine and then incubated with 1/2 ml of Dulbecco's modified Eagle's medium +15% fetal calf serum+basic fibroblast growth factor (1 ng ml-1), and fetal human retinal pigment epithelium at a concentration of 242,000 cells ml-1. Specimens were incubated for 1, 4, 6, 8, 12, or 24 hours. The specimens were fixed in half strength Karnovsky's fixative, processed, and analysed with scanning and transmission electron microscopy. The retinal pigment epithelium covered the debrided macular specimens to different degrees at different incubation times. After 1 hour, the cells started to attach and flatten (median percent coverage: 78%). The extent of Bruch's membrane coverage by fetal retinal pigment epithelium varied greatly between specimens. After 4-6 hours, the cells covered the entire debrided surface in a monolayer (median percent coverage: 97.2% at 4 hours, 99.8% at 6 hours). Tight junctions were observed, and the cells had few apical microvilli. The lateral cell borders were obliquely oriented with respect to Bruch's membrane, and the nuclei were elongated, exhibited prominent nucleoli, and were oriented parallel to Bruch's membrane. After 6-8 hours, cells started to become hexagonal (median percent coverage at 8 hours: 99.97%). Cells attached to the inner collagenous layer tended to be flatter than cells attached to residual native basement membrane. At 12 and 24 hours, expression of hexagonal shape, tight junctions, and apical microvilli were observed more frequently (median percent coverage: 99.87% at 12 and 100% at 24 hours). No newly formed basement membrane was observed at these time points. In separate experiments comparing attachment in the presence and absence of native RPE basement membrane, the presence of native retinal pigment epithelial basement membrane promoted the early attachment of the cells and more rapid expression of normal morphology. This in vitro system provides a reproducible way to study the adherence of retinal pigment epithelium to normal and diseased human Bruch's membrane.     

Corneal involvement in epidermolysis bullosa simplex

A 17-year-old boy and his mother represent the first reported cases of ocular (corneal) involvement of the simplex form of epidermolysis bullosa. Both had a ring-like configuration of fine bullous lesions in the midperiphery bilaterally at the level of deep corneal epithelium superficial to Bowman's membrane (basal cell layer), with the son manifesting symptoms when some of his bullae ruptured through to the corneal epithelial surface. Nonscarring blistering was present in three generations of this family, suggestive of dominantly inherited epidermolysis bullosa simplex, and was confirmed by electron microscopy of a skin specimen from the son. Thus, ocular involvement has now been observed in all of the major types of epidermolysis bullosa.     

Costa's acrokerato-elastoidosis. Ultrastructural study (author's transl)

Acrokerato-elastoidosis described by Costa is characterized by numerous little keratotic points and translucent horny wheals on the dorsal aspect of hands and (or) feet. The ultrastructural study shows lesions of the elastic tissue which is very rarefied with an abnormal structure, but also demonstrates alterations of the fibroblasts that contain dense granules at the periphery of their cytoplasm without elastic extracellular fibers, suggesting a trouble in the secretion or the excretion of the elastic material.