6'-substituted naphthalene-2-carboxylic acid analogs, a new class of retinoic acid receptor subtype-specific ligands.

The biological response to retinoic acid (RA) and synthetic derivatives (retinoids) is mediated by three nuclear retinoic acid receptors, RAR alpha, beta and gamma. To explore the potential of retinoids as receptor subtype selective activators, we employed a transcriptional activation assay. Hybrid receptors that recognize an estrogen response element were used to avoid measuring activities of endogenous retinoic acid receptors.

Changes in glutathione content in human hair follicle keratinocytes as a function of age of donor: relation with glutathione dependent enzymes.

Synopsis Glutathione (GSH) plays an important role in cellular protection during ageing. The hair plucking technique is a non-invasive method for the direct biochemical study of keratinocytes. Hair was taken from the suboccipital area of 63 volunteers (men and women whose ages ranged from 13 to 103 years).

[General process of aging].

Genetic instability is considered to be a fundamental factor in the ageing process. This instability results from the accumulation of damage caused to DNA, and this is demonstrated by the relationship between longevity and DNA repair. However, the accumulation of simple somatic mutations does not explain all the experimental findings.

Unscheduled DNA synthesis: a quantitative indicator of residual immunodetectable pyrimidine dimers in human fibroblasts after ultraviolet-B irradiation.

We have addressed the question whether the level of UV-B induced DNA damage can be accurately assessed by the measurement of the rate of unscheduled DNA synthesis (UDS). Cultured human fibroblasts were irradiated with UV radiation at 290, 313 or 365 nm. The LD50 was 85 J/m2 at 290 nm, 4500 J/m2 at 313 nm, and 70 kJ/m2 at 365 nm.

Evidence for an age-correlated change in glutathione metabolism enzyme activities in human hair follicle.

In this investigation, glutathione peroxidase (GSH-PX), glutathione reductase (GSSG-RD), glutathione-S-transferase (GSH-S-T), gamma-glutamyl transpeptidase (gamma-GT) and glucose-6-phosphate dehydrogenase (G6PDH) were measured in human hair follicle obtained by plucking as source of keratinized cells. This non-invasive method was used on 27 men and women volunteers ranging from 19 to 102 years. Our results show that GSSG-RD, GSH-S-T, gamma-GT and G6PDH activities decrease as a function of age, whereas GSH-PX activity does not vary.

Assessment of surfactant cytotoxicity: comparison with the Draize eye test.

Synopsis We have compared the in vitro cellular toxicity and the in vivo ocular irritation potency of 16 surfactants (7 non ionics, 3 anionics, 2 amphoterics, 4 cationics) ranking from very weakly irritant to strongly irritant. In vitro, the cellular toxicity was estimated on Chinese hamster lung fibroblasts (V79) using a cell mortality test and a cell growth inhibition test. For each surfactant, a lethal concentration 50% without foetal calf serum (LC.

An immunohistological study of the revascularization process in human skin transplanted onto the nude mouse.

The revascularization of human skin transplanted onto the nude mouse was studied by performing double-labeling immunofluorescence microscopy on human skin before transplantation and at different stages, ranging from one week to six months after grafting. With a crossreacting anti-factor-VIII antigen antibody used to identify the endothelial cells, and with human-specific monoclonal antibodies directed against vimentin or HLA-DR antigen, it appeared that the original human endothelial cells of transplanted skin progressively disappear, while murine endothelial cells invade the graft. Moreover, in double-labeling experiments with a crossreacting anti-factor-VIII antibody and a human-specific anti-type-IV-collagen antibody, anastomosis between host and graft vessels and a constant codistribution of graft endothelial cells with human type IV collagen were observed.

Wound healing of human skin transplanted onto the nude mouse. II. An immunohistological and ultrastructural study of the epidermal basement membrane zone reconstruction and connective tissue reorganization.

The reconstruction of human epidermis during healing of human skin wounded after grafting onto the nude mouse was described in a previous paper (M. Démarchez, P. Sengel, and M.

Alternative method for checking toxicity of hair dyes.

Synopsis We have compared the in vitro and in vivo toxicities of 19 hair dyes. In vitro, the toxic effect was estimated in Chinese hamster lung fibroblasts (V79), using a cell growth inhibition test. For each compound, the concentration required to reduce 50% of the growth (CI(50)) was determined.

Onset of epidermal differentiation in rapidly proliferating basal keratinocytes.

Stratified epithelia such as epidermis are classically considered to comprise 2 cell compartments, one consisting of undifferentiated proliferative cells occupying the basal layer, and the other consisting of differentiated postmitotic cells occupying the suprabasal layers. It is also generally assumed that the 58K basic-50K acidic couple of keratins is expressed in basal cells, while the 67K basic-56K acidic couple appears in suprabasal cells. In the present work we demonstrate that the population of basal keratinocytes is heterogeneous, since 8% of them are found to express the 67-56K "suprabasal" set of keratins.

Eye-derived growth factor isolated from bovine retina and used for epidermal wound healing in vivo.

Eye-derived growth factor (EDGF) has been found in several ocular tissues and shown to be able to stimulate the in vitro proliferation of cells from various tissues and organisms. It had already been shown that EDGF differs biochemically and biologically from other growth factors such as epidermal growth factor (EGF) and fibroblast growth factor in that it is the only one that can stimulate the in vitro growth of human adult keratinocytes. Moreover EDGF stimulates reepithelialization and neovascularization.

Expression of high molecular weight (67K) keratin in human keratinocytes cultured on dead de-epidermized dermis.

The influence of living dermal tissue upon epidermal differentiation during embryonic development as well as in vitro culture has been documented. Living dermal tissue contains both cellular and matricial elements. In the present study, third-passage subcultured adult human keratinocytes were either seeded on plastic dishes or recombined with dead de-epidermized dermis and further cultured for 3 weeks.

Adult human keratinocytes migrating over nonviable dermal collagen produce collagenolytic enzymes that degrade type I and type IV collagen.

Human adult keratinocytes migrating on a nonviable dermal substrate in cultures without fibroblasts induce thinning and degradation of the collagen substrate beneath the migrating epithelium. Further, unconcentrated conditioned medium from the cultures exhibit collagenolytic activity against both type I and type IV collagen which is inhibited by EDTA but not by phenylmethylsulfonyl fluoride or N-ethylmaleimide. Since the migrating epithelium and dermal substrate do not contain fibroblasts, this study shows that migratory keratinocytes in contact with interstitial collagen are capable of producing collagenases against type I and type IV collagen.

Human epidermis reconstructed by culture: is it "normal"?

Human keratinocytes were grown on a dermal equivalent (or lattice) at the liquid-air interface in an attempt to reconstitute a functional epidermis in vitro. Although the multilayered epithelium thus obtained is well differentiated, as shown by the presence of keratohyaline granules and horny layer, several differences from its in vivo counterpart were also observed: In the reconstructed epidermis, basal keratinocytes do not have the cuboidal shape found in vivo; they synthesize bullous pemphigoid antigen and laminin, but the distribution of these antigens is not linear as in vivo; they contain the plasma-membrane antigens restricted to the basal layer in vivo (VM1, BC1), but these antigens are not polarized; lack of polarization is also evidenced by the distribution of actin. Differentiation markers appear but with a topography slightly different from that of epidermis in vivo; the 67-kD keratin does not appear in the first suprabasal layer as in vivo but above; involucrin, which appears in the granular layers in vivo appears as soon as the cells leave the basal layer.

Wound healing of human skin transplanted onto the nude mouse. I. An immunohistological study of the reepithelialization process.

Two months after transplantation of human skin onto the nude mouse, excisional wounds were made through the entire thickness of the skin, at the center of the graft, using a 2-mm punch. At various time intervals thereafter, ranging from 2 days to 9 weeks, the graft sites were harvested and processed for an immunohistological study. With a monoclonal antibody directed against HLA-ABC antigens, it was demonstrated that the healing epidermis is of human origin.

Localization of bullous pemphigoid antigen (BPA) in isolated human keratinocytes.

In early studies, the bullous pemphigoid antigen (BPA) has been localized extracellularly in the lamina lucida in the basement membrane zone. However, trypsin-dissociated basal cells can be tagged with bullous pemphigoid sera (BPS). By immunofluorescence, BPA appears located at the dermal pole of basal cells (BC).

Detection of distinct subpopulations of Langerhans cells by flow cytometry and sorting.

Flow cytometry was found to be a very appropriate tool for the study of Langerhans cells (LC), which represent a minor cell population (2-3%) of human epidermis, and allowed us to obtain new phenotypic, functional, and cell cycle data on these rare cells. The phenotypic analysis of cell surface antigens demonstrates the existence of two subpopulations of LC: the former is HLA-DR+ and OKT 6+ (about 90% of total HLA-DR+ cells) and the latter is HLA-DR+ and OKT 6- (about 10% of total HLA-DR+ cells). These subpopulations of LC are both able to stimulate the proliferation of peripheral blood lymphocytes (PBL) in the presence of keratinocytes i.

Biology of human epidermal Langerhans cells: cell cycle studies.

Langerhans cells (LC) are considered to play an important role in the initiation of the immune response in the skin. This study was performed to analyse the kinetics of LC in normal human epidermis. Using flow cytometry (FCM), we have applied three methods to estimate LC DNA distribution: FCM DNA measurement in LC-enriched suspensions (70-90% purity), FCM-correlated analysis of DNA and OKT6-positive cells in original epidermal cell suspensions, and staining of LC-enriched suspensions by the Feulgen technique on microscopic slides and counter labelling of contaminating keratinocytes with anti-keratin antiserum to eliminate them from the LC DNA estimation.

Epidermal Langerhans cells--a cycling cell population.

The limited number of Langerhans cells (LC) in human epidermis and the resultant technical difficulties have left open the question of LC kinetics. In the present study using flow cytometry (FCM) we have applied 3 methods to estimate LC-DNA distribution: (1) FCM-DNA measurement on highly enriched LC suspensions, (2) FCM-correlated analysis of DNA and OKT-6(+) cells in total epidermal cell suspensions, (3) LC-enriched suspensions (70-90%) were FACS (fluorescence-activated cell sorter) sorted on microscopic slides, and stained with the Feulgen technique, and DNA was measured densitometrically. In the latter method, contaminating keratinocytes were counterlabeled with antikeratin serum to eliminate them from LC-DNA estimation.