Nonclinical and human pharmacology of the potent and selective topical retinoic acid receptor‐γ agonist trifarotene

Acne is a chronic inflammatory skin disease, linked to changes in hormone levels. It mainly affects adolescents and young adults, and may lead to permanent scarring. 1.5 billion people aged between 15 and 45 suffer from some form of acne – mild, moderate or severe. Retinoic acid is one type of treatment for acne, and there are several different types, which are applied directly to the skin (topically). Trifarotene is a new type of retinoic acid which is able to act against only one particular retinoic acid receptor, and this study looked at whether it might be suitable for use in acne and other skin diseases, and potentially have an improved efficacy and safety profile compared with less selective retinoids (i.e. ones that target more than one retinoic acid receptor). Looking at the biological pathways of the drug, the authors suggest that it should be very effective in acne. Furthermore, trifarotene is expected to be rapidly eliminated in the blood stream, thereby potentially leading to fewer side effects and making it particularly useful for the treatment of large skin surface areas, such as the back and chest of acne patients. Based on these favourable characteristics of trifarotene, it is worth now investigating the clinical efficacy of this retinoid.     

Proteins of the extracellular matrix are sensitizers of photo-oxidative stress in human skin cells

Sensitized production of reactive oxygen species after photo-excitation of endogenous chromophores is thought to contribute to skin photo-oxidative stress. Here we present experimental evidence in support of a potential role of extracellular matrix proteins as skin photosensitizers. Human and bovine type I collagen and elastin sensitized of hydrogen peroxide generation upon irradiation with solar simulated light or ultraviolet A. Induction of intracellular oxidative stress by extracellular matrix-protein sensitization was demonstrated by flow cytometric analysis of fibroblasts preloaded with the intracellular redox dye dihydrorhodamine 123 and exposed to pre-irradiated type I collagen. Pre-irradiated collagen and elastin induced pronounced inhibition of proliferation in cultured keratinocytes and fibroblasts, which was reversed by antioxidant or catalase treatment and reproduced by exposure to concentrations of H2O2 formed during extracellular matrix-protein irradiation. In fibroblasts, chromosomal DNA damage as a consequence of collagen-sensitized H2O2 formation was demonstrated using a single cell electrophoresis assay. The enzymatic cross-links pyridinoline and desmosine were examined as candidate sensitizer chromophores contained in collagen and elastin, respectively. Pyridinoline, but not desmosine, sensitized light-driven H2O2 production and inhibition of fibroblast proliferation. Our results support the hypothesis that extracellular matrix proteins play a functional role in skin photoaging and carcinogenesis by sensitization of photo-oxidative damage.     

Cellular, immunologic and biochemical characterization of topical retinoic acid-treated human skin.

Histologic and clinical improvement of sun-exposed skin following topical treatment with retinoic acid has been reported. Daily application of retinoic acid typically results within 2-5 d in an erythematous scaling reaction, which lessens with continued usage. The cellular, immunologic, and biochemical basis of this retinoid reaction and its role in the repair of photodamaged skin are not known. To investigate the retinoid reaction in man, we have treated non-sun-exposed skin with 0.1% retinoic acid cream (Retin-A, Ortho Pharmaceutical Corporation, Raritan, NJ) under occlusion for 4 d to induce erythema and then examined changes in 1) histology, 2) expression of cell-surface molecules, 3) the enzymes and second messengers of the phospholipase C/protein kinase C signal-transduction system, 4) levels of eicosanoids, and 5) levels of interleukin-1 protein and mRNA. These parameters were chosen for measurement both because they are indicators of epidermal function and previous studies suggest they may be responsive to retinoic acid treatment. Epidermal cell growth as judged by increased epidermal thickness and mitotic figures was significantly increased in retinoic acid-treated skin compared to vehicle-treated controls. Increased numbers of CD4+ T cells accompanied by prominence of dermal dendrocytes in the papillary dermis and focal keratinocyte expression of intercellular adhesion molecule-1 were observed in retinoic acid-treated biopsies. Phosphoinositide-specific phospholipase C activity and 1,2-diacylglycerol content were also elevated in retinoic acid-treated epidermis. Protein kinase C activity was reduced by one third in both the soluble and membrane fraction, suggesting down-regulation. Surprisingly, in view of the inflammatory nature of the retinoid reaction, no increases were observed in arachidonic acid, its metabolites, interleukin-1 alpha, or interleukin-1 beta. To examine the specificity of the retinoid reaction, subjects were treated with the irritant sodium lauryl sulfate, under conditions that resulted in a reaction clinically similar to that observed with retinoic acid. The histologic alterations induced by sodium lauryl sulfate were found to be indistinguishable from those induced by retinoic acid. These data indicate that, although a wide range of cellular and molecular alterations occur in retinoic acid-treated skin, these changes may not be necessarily specific or unique for retinoic acid.     

Retinoic acid inhibits the production of collagenase by human epidermal keratinocytes

Lattices made of collagen and fibroblasts can be used as dermal equivalents to grow human keratinocytes in vitro. When these cultures are performed in a medium containing delipidized serum, the lattice is eventually degraded by the growing epithelium. The digestion of the dermal equivalent is due to the secretion of a collagenase by the keratinocytes. This degradation does not occur in cultures containing total serum or supplemented with retinoic acid. We show in this paper that retinoic acid inhibits the secretion of this keratinocyte collagenase in a dose-dependent manner. In the light of this result, the possible involvement of collagenase inhibition in the therapeutic effect of retinoic acid in skin disorders and skin aging must be considered.     

A comparison of dapsone with 13-cis retinoic acid in the treatment of nodular cystic acne

Forty male patients with nodular/cystic acne were randomly allocated to receive either 100mg dapsone daily, or 40 mg 13-cis retinoic acid, daily for a period of 16 weeks. The following parameters were assessed during Treatment and for a follow-up period of 20 weeks: lesion counts (pustules, nodules and cysts), photographic evaluation, patient visual analogue scale, sebum excretion rate and skin surface microflora. Seventeen patients from each group completed the study. Only marginal clinical benefit was observed with dapsone, which produced no significant effect either on sebum excretion rate, or on skin surface microflora. By contrast, patients on 13-cis retinoic acid showed significant reductions in sebum excretion rate and skin surface microflora, which were maximal at 16 weeks (P < 0.001 and P < 0.01, respectively), and significant improvements in all clinical parameters which were maintained at follow-up 5 months later. We conclude that dapsone is clearly inferior to 13-cis retinoic acid in the treatment of nodular cystic acne.     

Retinoic acid upregulates the plasminogen activator system in human epidermal keratinocytes

The activation of the proteolytic plasminogen activator system is important for the re-epithelialization of skin wounds. Keratinocytes synthesize and secrete the urokinase-type plasminogen activator, which binds to its specific receptor on keratinocytes. Receptor-bound urokinase-type plasminogen activator efficiently activates cell surface bound plasminogen. This results in pericellular proteolysis, which facilitates keratinocyte migration. Urokinase-type plasminogen activator activity is specifically controlled by plasminogen activator inhibitor-1 and -2. As retinoids have been reported to accelerate epithelialization of skin wounds in animal studies and clinical settings, we investigated the effects of all-trans retinoic acid on the plasminogen activator system in human epidermal keratinocytes. As tested in a chromogenic plasminogen activation assay, incubation with 10 microM all-trans retinoic acid caused a marked induction of cell-associated plasminogen activity after 24 h, and this induction was blocked by neutralizing anti-urokinase-type plasminogen activator antibodies, but not anti-tissue-type plasminogen activator antibodies. All-trans retinoic acid lead to a strong increase in urokinase-type plasminogen activator (enzyme-linked immunosorbent assay) and urokinase-type plasminogen activator receptor cell surface expression (flow cytometry) after 24 h. At this time-point, tissue-type plasminogen activator and plasminogen activator inhibitor-1 and -2 proteins were not or only slightly increased. Northern blot analyses revealed that all-trans retinoic acid caused an early and short-lived increase of plasminogen activator inhibitor-1, but a prolonged induction of urokinase-type plasminogen activator and urokinase-type plasminogen activator receptor mRNA levels. Collectively, these data suggest that all-trans retinoic acid activates the plasminogen activator system in human epidermal keratinocytes by differentially regulating activating and inhibiting components. The activation of the plasminogen activator system may be one mechanism by which all-trans retinoic acid exerts beneficial effects in cutaneous wound healing.     

Ultraviolet irradiation increases matrix metalloproteinase-8 protein in human skin in vivo

Humans express three distinct collagenases, MMP-1, MMP-8, and MMP-13, that initiate degradation of fibrillar type I collagen. We have previously reported that ultraviolet irradiation causes increased expression of MMP-1, but not MMP-13, in keratinocytes and fibroblasts in human skin in vivo. We report here that ultraviolet irradiation increases expression of MMP-8 in human skin in vivo. Western analysis revealed that levels of the full-length, 85 kDa proenzyme form of MMP-8 increased significantly within 8 h post ultraviolet irradiation (2 minimal erythema doses). Increased full-length MMP-8 protein was associated with infiltration into the skin of neutrophils, which are the major cell type that expresses MMP-8. Immunofluorescence revealed coexpression of MMP-8 and neutrophil elastase, a marker for neutrophils. Immunohistology demonstrated MMP-8 expression in neutrophils in the papillary dermis between 4 and 8 h post ultraviolet irradiation, and in the epidermis at 24 h post radiation. MMP-8 mRNA expression was not detected in nonirradiated or ultraviolet-irradiated human skin, indicating that increased MMP-8 following ultraviolet irradiation resulted from preexisting MMP-8 protein in infiltrating neutrophils. Pretreatment of skin with the glucocorticoid clobetasol, but not all-trans retinoic acid, significantly blocked ultraviolet-induced increases in MMP-8 protein levels, and neutrophil infiltration. In contrast, all-trans retinoic acid and clobetasol were equally effective in blocking ultraviolet induction of MMP-1 and degradation of collagen in human skin in vivo. Taken together, these data demonstrate that ultraviolet irradiation increases MMP-8 protein, which exists predominantly in a latent form within neutrophils, in human skin in vivo. Although ultraviolet irradiation induces both MMP-1 and MMP-8, ultraviolet-induced collagen degradation is initiated primarily by MMP-1, with little, if any, contribution by MMP-8.     

Carbon dioxide laser treatment promotes repair of the three-dimensional network of elastic fibres in rat skin

BACKGROUND: We have previously reported that ultraviolet (UV) B irradiation induces a loss of linearity in the three-dimensional structure of dermal elastic fibres, which results in the reduction of elastic properties of the skin and leads to wrinkle formation. We further reported that repair of wrinkles by all-trans retinoic acid is accompanied by recovery of the linearity of elastic fibres. Carbon dioxide (CO2) lasers are widely used for treating wrinkles in cosmetic surgery. OBJECTIVES: To perform CO2 laser treatment of wrinkles induced in rat skin by UVB irradiation and to evaluate changes in the three-dimensional structure of dermal elastic fibres during wrinkle repair. METHODS: Wrinkles were induced in the hind limb skin of Sprague-Dawley rats by UVB irradiation (130 mJ cm-2 three times weekly for 6 weeks), followed by CO2 laser treatment (11.3 J cm-2). The surface appearance of the skin was evaluated by replica observation 6 and 10 weeks after CO2 laser treatment followed by measurement of mechanical properties using a Cutometer. Subsequently, perfusion fixation and digestion with formic acid were performed and elastic fibres were observed by scanning electron microscopy (SEM). Image analysis of SEM micrographs was carried out to evaluate the linearity in the three-dimensional structure of elastic fibres. RESULTS: Six weeks after CO2 laser treatment, all parameters of skin mechanical properties in the UVB-irradiated group recovered to levels of the control non-irradiated group, accompanied by repair of wrinkles and a significant increase in linearity of the three-dimensional structure of elastic fibres. CONCLUSIONS: These findings indicate that CO2 laser treatment has a therapeutic potential to repair wrinkles to non-irradiated levels through recovery of the three-dimensional structure of elastic fibres.     

A short-term screening protocol, using fibrillin-1 as a reporter molecule, for photoaging repair agents

Photoaged skin is characterized by coarse and fine wrinkles. The mechanisms of wrinkle formation are undetermined, but appear to be due to changes within the matrix of the dermis and at the dermal-epidermal junction. Previous studies have identified marked reductions in procollagens I and III, collagen VII, and the fibrillin-rich microfibrillar apparatus in this area. Topically applied all-trans retinoic acid can repair photoaged dermal matrix, but this takes at least 6 mo of treatment. In this study, we have examined the abundance and distribution of fibrillin-1 prior to, and following, 192 wk of all-trans retinoic acid treatment. We have further developed a short-term protocol to determine the utility of potential repair agents, using fibrillin-1 as the marker for outcome. Individuals with clinically assessed severe photoaging were recruited to the study (n = 8). 0.025% all-trans retinoic acid, 5% sodium lauryl sulfate (irritant control), or vehicle were applied under occlusion to photoaged extensor forearm. A fourth control area was also occluded. After 96 h, punch biopsies were taken under local anesthesia and processed for either transmission electron microscopy or snap frozen. Frozen sections were prepared for immunohistochemistry and in situ hybridization immunohistochemistry. Electron microscopy revealed aberrant elastic fibers in the papillary dermis of photoaged forearm skin, with sparse microfibrillar apparatus and interstitial collagen. After application of 0.025% all-trans retinoic acid, there was increased deposition of both these dermal matrix components, with the aberrant elastic fibers no longer apparent. Significant increases (p < 0.05) were observed at the protein and mRNA levels for fibrillin-1 following all-trans retinoic acid and sodium lauryl sulfate treatments, with all-trans retinoic acid having a significantly greater effect than irritant control (p < 0.001); however, neither application had significant effect on the abundance of collagen VII or its mRNA. Investigation of collagen I synthesis revealed no difference following treatments. To ascertain the clinical relevance of using fibrillin-1 as a marker for photoaging, facial skin was biopsied at baseline and after long-term (192 wk) topical all-trans retinoic acid treatment (n = 5). Biopsies were wax-embedded and sections prepared for immunohistochemistry for fibrillin-1. Significant increases in the abundance of the microfibrillar apparatus was observed proximal to the dermal- epidermal junction (p < 0.001) following long-term all-trans retinoic acid application. This study indicates that all-trans retinoic acid can significantly affect fibrillin-1 content in photoaged skin. Furthermore, fibrillin-1 can be used as a "reporter" molecule in short-term protocols for testing the utility of topical agents in the repair of photoaged skin.     

Topical retinoic acid does not alter the vasoconstrictive properties of topical corticosteroids in humans

Dermo-epidermal atrophy is one of the main side effects of long-term treatment with topical corticosteroids. Retinoic acid may prevent and even reverse these effects in animals. Extension of this concept to therapy in humans implies that several studies have been performed; among others, it has to be established that treatment with topical retinoic acid does not interfere with the anti-inflammatory action of topical corticosteroids. The present study on the cutaneous vasoconstriction test comprised two different double-blind approaches: (i) vasoconstriction tests with betamethasone dipropionate (Diprolene) and clobetasone butyrate (Emovate) were carried out on skin that had previously been treated for 10 days with retinoic acid 0.01, 0.025 or 0.05% (or excipient); (ii) vasoconstriction tests with a combination of triamcinolone acetonide 0.1% and retinoic acid 0.025% were compared with triamcinolone acetonide 0.1% alone. Pretreatment for 10 days with retinoic acid did not alter the vasoconstriction induced by corticosteroids: no decrease or increase in the vasoconstriction score was observed, whether the skin had been previously treated with retinoic acid or with excipient. The vasoconstriction scores obtained with a combination of retinoic acid and triamcinolone acetonide were identical with those obtained with the steroid alone. This study indicates that retinoic acid does not inhibit the vasoconstriction induced by topical corticosteroids and suggests that the anti-inflammatory effect of the latter should be maintained in association with retinoic acid.     

Selective upregulation of fibroblast Fas ligand expression, and prolongation of Fas/Fas ligand-mediated skin allograft survival, by retinoic acid: the skin as a retinoide-inducible immune privilege site

Fas/Fas ligand-mediated lymphocyte apoptosis has been implicated in the suppression of immune responses and may cause immune privilege. Human corneas exhibit immune privilege and can be transplanted across allogeneic barriers without immunosuppressive therapy, perhaps, because corneal keratinocytes express Fas ligand. To characterize Fas and Fas ligand expression in skin, we examined expression by murine keratinocytes, dermal fibroblasts, melanocytes, and human umbilical endothelial cells. We also studied the regulation of Fas and Fas ligand in skin cells by retinoic acid, vitamin D3, and dexamethasone as well as various cytokines. Among the molecules and cells tested, retinoic acid selectively upregulated the expression of Fas ligand molecule by fibroblasts. Retinoic acid-induced Fas ligand+ fibroblasts killed Fas+ target cells, and this killing was blocked by anti-Fas ligand antibody. The function of Fas ligand on dermal fibroblasts in vivo was tested in a cutaneous allograft system. Histoincompatible BALB/C mouse (H-2d) donor skin was grafted on to allogeneic C57BL/6 mice (H-2b). Daily local injection of retinoic acid blocked inflammation and extended graft survival for more than 10 d. Injection of retinoic acid into Fas ligand mutated gld/gld donor skin did not prevent leukocyte infiltration into the allograft or prolong graft survival. These experiments indicate that, in skin, retinoic acid selectively increases Fas ligand expression by fibroblasts and that retinoic acid has potent Fas/Fas ligand-dependent immunosuppressive activity.     

Angiogenesis in skin aging and photoaging

Angiogenesis, the process of generating new blood vessels, is affected by various physiological and pathological conditions of skin. The skin aging process can be divided into intrinsic aging and photoaging. With aging, cutaneous blood vessels undergo pronounced alterations. A reduction of the cutaneous microvasculature has been observed in the skin of elderly individuals. Human skin is exposed daily to solar ultraviolet (UV) radiation, infrared rays and heat, and these stimuli are known to induce skin angiogenesis. Interestingly, although acute UV irradiation stimulates skin angiogenesis, cutaneous blood vessels are decreased in chronically photodamaged skin. The reason for the differential effects of acute and chronic UV exposure on skin angiogenesis remains to be elucidated. This review discusses the vascularization changes in intrinsically aged and photoaged human skin, the effects of UV irradiation, infrared rays and heat on skin angiogenesis, and the effects of topical retinoic acid treatment on UV-induced angiogenesis and cutaneous vascularity in aged and photoaged human skin. An understanding of the molecular mechanisms of aging- and photoaging-dependent changes of skin angiogenesis may provide us with new insights to prevent and treat the skin aging process.     

All-trans retinoic acid induces functional maturation of epidermal Langerhans cells and protects their accessory function from ultraviolet radiation

Abstract Retinoids provide some protection against ultraviolet radiation-induced skin damage. We have previously shown that topical all-trans retinoic acid prevents ultraviolet light from reducing the density of epidermal Langerhans cells in the epidermis but does not inhibit the development of immunosuppression to a locally applied contact sensitizer. We therefore investigated the ability of all-trans retinoic acid to modulate Langerhans cell induction of allogeneic T-cell proliferation in the mixed epidermal cell lymphocyte reaction. Langerhans cells isolated from all-trans retinoic acid-treated mice induced an enhanced mixed epidermal cell lymphocyte reaction. This is similar to Langerhans cells cultured with granulocyte-macrophage colony stimulating factor. Retinoic acid treatment also enhanced the allogeneic cell-stimulating capability of Langerhans cells isolated from ultraviolet-irradiated mice. Langerhans cells from all-trans relinoic acid-treated, ultraviolet-irradiated mice which were “matured” by 3 days in culture induced a larger mixed epidermal cell lymphocyte reaction than mice treated with solvent and ultraviolet irradiation. Thus all-trans retinoic acid treatment of mice causes Langerhans cell maturation and inhibits ultraviolet light from reducing their density or impairing their allogeneic cell-stimulating capacity. However, these mice remained immuno-suppresscd upon application of a contact sensitizer to irradiated or unirradiated skin. It is thus likely that, whereas all-trans relinoic acid protects local Langerhans cell numbers and function, it does not inhibit the production of an ultraviolet radiation-induced photoproduct which causes immunosuppression.     

UVB irradiation stimulates deposition of new elastic fibers by modified epithelial cells surrounding the hair follicles and sebaceous glands in mice

UVB irradiation stimulates the synthesis of elastin in the skin of humans and experimental animals. In this study we localized the site and the cells that are responsible for the synthesis of murine dermal elastic fibers. SKH-1 hairless mice were irradiated with UVB and the skin removed for light microscopy, electron microscopy, in situ hybridization, immunohistochemistry, and biochemical studies. In response to chronic low doses of UVB there was an initial moderate increase in tropoelastin mRNA in the papillary dermis. By contrast, there was a continuous marked elevation of collagen alpha1(I) message localizing to sites of inflammatory cell influx throughout the upper and lower dermis. After 25 wk of UV irradiation there was a 2-fold increase in skin elastin, yet total collagen remained unchanged. Serial desmosine analysis from en face sections indicated the increase in elastin content was due to dermal elastic fibers, an increase in the size and number of the dermal cysts, and an increase in subpanniculus elastic fibers. Elastin stains of en face sections suggested that the elastic fibers in the upper dermis were exclusively derived from cells lining the epithelial root sheath and sebaceous glands. In response to UV irradiation, the elastic fibers increased in number and size, wrapping around these structures and aligning in both directions as long fibers parallel to the body axis. Electron micrographs indicated that modified epithelial cells in close proximity to the flattened epithelial cells that encircled the root sheath and sebaceous glands were the source of the elastic fibers.     

Lack of enhancement of experimental photocarcinogenesis by topical retinoic acid

Summary A controversy exists regarding the ability of retinoic acid to enhance photocarcinogenesis. Divergent results have been obtained with albino hairless mice. We examined this issue with the lightly pigmented variety. We followed two designs: 1. ultraviolet light and topical retinoic acid were given concomitantly while the retinoic acid was continued for many weeks after stopping irradiation; 2. tumors were first induced by ultraviolet light and then treated topically with retinoic acid. In both studies, retinoic acid did not enhance photocarcinogenesis with regard to latent period, tumor yield or tumor progression. It appears that different treatment schedules and different varieties of mice can produce widely disparate results.     

Topical retinoic acid enhances, and a dark tan protects, from subedemal solar-simulated photocarcinogenesis

Studies into the effects of topical retinoic acid on photocarcinogenesis have yielded ambiguous findings. This may be due to different Experimental protocols and ultraviolet spectra. Retinoic acid is commonly used for a range of dermatologic conditions, and therefore it is important to resolve whether it affects skin tumor formation. To address this issue we used a protocol to mimic as closely as possible human use of retinoic acid. Two mouse strains were used: Skh:HR-1 (albino) and Skh:HR-2 (lightly pigmented). The pigmented mice more closely resemble Caucasian skin as they develop a light tan in response to ultraviolet radiation. This tan is greatly augmented by retinoic acid. As these are congenic mice, any differences can be attributed to the development of a tan. Mice were exposed to solar-simulated ultraviolet radiation, followed by treatment with 0.05% retinoic acid. This modeled exposure to sunlight during the day followed by retinoic acid treatment and a night-time period in the absence of sunlight. As it is recommended that ultraviolet exposure is minimized when using topical retinoic acid, the mice were only exposed to one-third of minimal edemal dose of ultraviolet radiation per day. This retinoic acid protocol augmented photocarcinogenesis. Retinoic acid decreased the latency period, reduced the probability that a mouse would survive without a tumor, and increased the number of tumors per mouse. All tumors induced were squamous cell carcinomas, and the skin between the tumors on mice treated with retinoic acid was found to contain carcinoma in situ upon histologic diagnosis. The light tan of the solvent-treated pigmented mice did not provide any protection, whereas the dark tan, which developed in Skh:HR-2 mice in response to retinoic acid, reduced photocarcinogenesis but did not overcome the augmenting effect of retinoic acid. Thus, using this experimental design, topical retinoic acid augmented photocarcinogenesis, and the ability to develop a dark but not light tan provided some, but limited, protection.     

Quantitative analysis of skin surface lipids using a sebumeter method. Random lipid levels on the skin surface during therapy with 13-cis-retinoic acid, minocycline hydrochloride and UVA rays

The Sebumeter method used in this study for quantitative analysis of skin surface lipids differs from previous techniques in simple handling and quick practicability and is therefore helpful in the clinical routine. Sebumetrical measurements carried out on healthy persons revealed symmetrical distribution of skin surface lipids. The highest levels were found on the forehead; young people generally showed higher values than older persons. During therapy with 13-cis retinoic acid on acne patients for 6 months (0.5 mg/kg daily), we found reduction of the random level in the first month, particularly in the forehead region. During therapy with minocycline on 3 acne patients for 3 months (2 X 50 mg daily), there was no variation of the random level observed. UVA irradiation on the face for 10 minutes daily (0.55 J/cm2/min) over a period of 3 weeks resulted in continuous reduction of the random level in most of the tested persons; a small part of them reacted by fluctuating values.     

Effects of retinoic acid on prostaglandin biosynthesis in guinea-pig skin.

Topical application of retinoic acid on guinea-pig skin resulted within 70 hours in erythema with a concomitant elevation of endogenous prostaglandin E2 (PGE2) in the treated areas of the skin. Prolonged daily treatment resulted in the development of severe scaly dermatoses and a corresponding decrease in the level of PGE2 in the skin. Examination of retinoic acid effects on the in vitro biosynthesis of PGE2 from arachidonic acid by extracts from guinea-pig skin and sheep vesicular gland demonstrated that retinoic acid inhibits prostaglandin synthesis in a concentration and time-dependent manner. These results indicate that retinoic acid may exert a regulatory role on prostaglandin biosynthesis in the skin.     

Differential modulation of transforming growth factor-beta 1 expression and mucin deposition by retinoic acid and sodium lauryl sulfate in human skin.

Immunohistochemical staining of skin sections with two polyclonal antibodies (anti-CC 1-30 and anti-LC 1-30), specific for transforming growth factor-beta 1, revealed increased extracellular and decreased intracellular expression of transforming growth factor-beta 1 in retinoic acid-treated, compared to vehicle-treated, skin. Transforming growth factor-beta 1 staining, with both antibodies, was most marked in the upper layers of the epidermis, although dermal staining was also evident. The modulation of transforming growth factor-beta 1 expression by retinoic acid occurred in the absence of any change in its mRNA level. Transforming growth factor-beta 1 protein, as detected by rabbit polyclonal antibody (anti-LC 50-75) and mRNA, were only minimally detected in either retinoic acid- or vehicle-treated skin. Similar changes in TGF-beta 1 and TGF-beta 2 immunoreactivity and mRNA levels, as observed in retinoic acid-treated skin, were observed in skin following topical application of the irritant sodium lauryl sulfate, indicating that the alterations induced by retinoic acid were not specific. In contrast, mucin deposition, which is induced by transforming growth factor-beta, was elevated in retinoic acid-treated but not sodium lauryl sulfate-treated skin. Cultured adult human keratinocytes also expressed predominantly transforming growth factor-beta 1 protein, as measured by ELISA, and mRNA. Treatment of keratinocytes with retinoic acid resulted in a 50% induction of transforming growth factor-beta 1 protein, without any detectable change in transforming growth factor-beta 2. These data demonstrate disassociation of modulation of transforming growth factor-beta 1 expression and mucin deposition by retinoic acid and sodium lauryl sulfate in human skin in vivo. Whereas alterations in transforming growth factor-beta 1 expression were observed in both retinoic acid- and sodium lauryl sulfate-treated skin, accumulation of mucin was specific to retinoic acid-treated skin.