Staphylococcal enterotoxin B causes proliferation of sensory C-fibers and subsequent enhancement of neurogenic inflammation in rat skin.

BACKGROUND: Staphylococcal enterotoxin B (SEB) may be associated with the exacerbation of atopic dermatitis. We investigated whether SEB causes proliferation of sensory C-fibers and subsequent enhancement of plasma leakage induced by sensorineural stimulation in rat skin. METHODS: SEB was applied intracutaneously to the abdomen of preweaning and adult rats. Evans blue dye leakage into the skin induced by topical 10% formalin was measured as an index of neurogenic skin vascular permeability. Local expression of substance P, tachykinin NK1 receptors, and nerve growth factor was assessed immunohistochemically. In addition, we assessed the effects of topical tacrolimus on these skin responses induced by SEB. RESULTS: Increased neurogenic skin plasma leakage was seen 7 days after SEB treatment in 2 different age groups. Innervation of substance P-immunoreactive nerves and expression of tachykinin NK1 receptors and nerve growth factor were also promoted by SEB, peaking at 7 days, 7 days, and 56 h after SEB treatment, respectively. Tacrolimus markedly inhibited these skin changes. CONCLUSIONS: SEB increased the innervation of sensory C-fibers and tachykinin NK1 receptors in rat skin, probably because of upregulated production of neurotrophins, including nerve growth factor, leading to enhancement of neurogenic skin inflammation. T cell activation induced by SEB may initiate these changes.

Identification of select glucocorticoids as Smoothened agonists: potential utility for regenerative medicine.

Regenerative medicine holds the promise of replacing damaged tissues largely by stem cell activation. Hedgehog signaling through the plasma membrane receptor Smoothened (Smo) is an important process for regulating stem cell proliferation. The development of Hedgehog-related therapies has been impeded by a lack of US Food and Drug Administration (FDA)-approved Smo agonists. Using a high-content screen with cells expressing Smo receptors and a beta-arrestin2-GFP reporter, we identified four FDA-approved drugs, halcinonide, fluticasone, clobetasol, and fluocinonide, as Smo agonists that activate Hedgehog signaling. These drugs demonstrated an ability to bind Smo, promote Smo internalization, activate Gli, and stimulate the proliferation of primary neuronal precursor cells alone and synergistically in the presence of Sonic Hedgehog protein. Halcinonide, fluticasone, clobetasol, and fluocinonide provide an unprecedented opportunity to develop unique clinical strategies to treat Hedgehog-dependent illnesses.

Glucocorticoids decrease tissue mast cell number by reducing the production of the c-kit ligand, stem cell factor, by resident cells: in vitro and in vivo evidence in murine systems.

The local delivery of glucocorticoids to tissues significantly decreases mast cell number. This pharmacologic effect of glucocorticoids is believed to be one of the mechanisms by which glucocorticoids regulate allergic inflammation. To determine the mechanism by which glucocorticoids are able to exert this effect, we first applied the glucocorticoid fluocinonide to mouse dermis and observed that the decrease in mast cell number was associated with an increase in mast cell apoptosis. This did not appear to be due to a direct effect of the glucocorticoid on mast cells, as the addition of 0.01-1.0 microM of the glucocorticoid dexamethasone into stem cell factor (SCF)-dependent mast cell cultures did not enhance mast cell death. However, addition of dexamethasone to cultured fibroblasts did result in a downregulation of SCF mRNA and a significant decrease in SCF protein production. Similarly, immunohistochemistry performed on fluocinonide-treated mouse dermis revealed a decrease in immunoreactive SCF. Administration of SCF at sites of fluocinonide administration to the dermis abolished the mast cell-depleting effect of this glucocorticoid. Thus, glucocorticoids decrease tissue mast cell number by downregulating tissue SCF production required for the survival of local mast cells. This observation may be applicable to the design of improved strategies to treat mast cell-mediated disorders.

Topical application of a corticosteroid destabilizes the host-parasite relationship in an experimental model of the oral carrier state of Candida albicans.

Using an experimental model in the mouse we have shown that both local and central lines of defense, involving CD4+ T cells, participate in a dynamic interaction to maintain a long-term carrier state of Candida albicans in the oral cavity. We have tested the impact of a predisposing factor to oral candidiasis in the form of a topical application of a corticosteroid (Topsyn gel) to the oral mucosa for 75 mice twice a day for a 20-day period. Very rapidly after the treatment was initiated, i.e. on day 4, the residual population of Candida increased up to 40-fold and by day 21, the population was 400-fold that of the carrier state. The resident population of intraepithelial CD4+ T cells in the oral mucosa virtually disappeared during the treatment. A topical corticosteroid application also resulted in a massive depletion of T cells in the lymph nodes and in the transient abrogation of the DTH reaction to Candida antigens. On cessation of treatment, normal levels of both Candida and intraepithelial CD4+ T cells were also quickly restored. These results suggest that resistance to superficial invasion by Candida is linked to the presence of an oral mucosal line of defense and that topical application of corticosteroids may dramatically shift the host-parasite relationship in favor of Candida.

[Silastic gel and elastomer in the cicatrization of wounds in the rabbit]. / Le gel et l'élastomère de Silastic dans la cicatrisation des plaies chez le lapin.

The aim of this experimental study was to examine two new wound healing adjuvants. Four experimental wounds were created on the back of rabbits to compare the kinetics of wound healing after applying the standard treatment, Silastic gel, and elastomer with and without compounds known to act on wound healing. We observed a tendency towards more rapid healing in the gel groups (13.7 days), the elastomer group (17.3 days) and the elastomer with active compound group (15.3 days) compared with the control group (17.3 days) and the standard treatment group (17.8 days). The differences were not however statistically different. Elastomer is the better carrier for active wound healing compounds because it has better stability over time. Silicon dressings are interesting for wound healing because they give satisfactory results in terms of healing time and in terms of redressing and patient comfort.

Intrinsic potencies of novel thiol ester corticosteroids RS-85095 and RS-21314 as compared with clobetasol 17-propionate and fluocinonide.

The intrinsic potencies of two novel topical thiol ester corticosteroids, RS-85095 and RS-21314, were compared with the high potency corticosteroids clobetasol 17-propionate and fluocinonide, using human vasoconstriction assays. In these assays, four or five concentrations (0.03 to 3 mg/L) of the corticosteroids in 95% ethanol (alcohol, USP) were applied to predetermined sites on the forearm of volunteers and were occluded following evaporation of the alcohol. The responses were scored in terms of the presence or absence of vasoconstriction and the degree of vasoconstrictive intensity. No statistically significant difference was found in the intrinsic potencies of RS-85095, RS-21314, and clobetasol 17-propionate, and all three corticosteroids were significantly more potent than fluocinonide.

Effects of retinoids on phorbol ester-stimulated epidermal DNA synthesis and hyperplasia in hairless mice.

We investigated the effects of the retinoids, all-trans retinoic acid (t-RA), 13-cis retinoic acid, etretinate, and arotinoid ethyl ester, on 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced DNA synthesis, and epidermal hyperplasia in hairless mouse skin. Topical application of these retinoids produced dose-dependent inhibition of the TPA-induced epidermal DNA synthesis as measured by [3H]thymidine incorporation at 15 h after TPA application. However, this inhibition was only transient and did not affect the corresponding increase in epidermal cell layers measured at 40 or 70 h after TPA application. Fluocinonide also inhibited the epidermal DNA synthesis and failed to block TPA-induced epidermal hyperplasia. However, fluocinonide did effectively suppress the inflammation caused by TPA. In this paper we have shown that the suppression of TPA-stimulated DNA synthesis is a general property of topically applied retinoids. The biologic significance of a temporary suppression of TPA-stimulated epidermal DNA synthesis by the retinoids and fluocinonide is not understood at this time.

Direct effects of glucocorticosteroids on epidermal Langerhans cells.

To determine the direct effects of glucocorticosteroids on epidermal Langerhans cells (LC), we treated isolated LC with dexamethasone (DEX) in vitro, and investigated Ia expression by LC using immunofluorescence microscopy and FACS analysis. We found that DEX directly decreased the number of Ia+ LC in a dose- and time-dependent manner. Pulse incubation with DEX also inhibited the immunostimulatory function of LC in vitro. FACS analysis demonstrated that LC detected in DEX-treated culture expressed a similar amount of Ia antigen and Fc receptor on the cell surface as LC cultured with the solvent control, suggesting that LC may be composed of a heterogeneous population in terms of sensitivity to DEX, and DEX may completely abolish the expression of surface molecules on a subpopulation of LC or may be cytolytic to this sensitive population.

Adrenocortical suppression by topically applied corticosteroids in healthy dogs.

Three corticosteroid products (triamcinolone acetonide, fluocinonide, betamethasone valerate) and a control product composed of water, petrolatum, mineral oil, cetyl alcohol, steryl alcohol, sodium lauryl sulfate, cholesterol, and methylparaben each were applied topically to healthy dogs (5 dogs/product) once daily for 5 consecutive days. Plasma concentrations of immunoreactive adrenocorticotropic hormone (iACTH) and cortisol were determined before 1 microgram of ACTH/kg of body weight was given intravenously (pre-ACTH values) and cortisol was again measured 60 minutes after ACTH was given (post-ACTH values). Cortisol and iACTH concentrations were determined in each dog before, during, and after administration of the corticosteroid products. All 3 corticosteroids caused prompt and sustained pituitary-adrenocortical suppression. Compared with control applications, the application of corticosteroids resulted in significant reduction of plasma cortisol and iACTH concentrations by day 2 of treatment, and the lower concentrations continued to day 5. One week after the last application of the corticosteroids, plasma iACTH concentrations in the corticosteroid-treated dogs had returned to the range of values for the control dogs; however, pre- and post-ACTH cortisol concentrations remained suppressed in all corticosteroid-treated dogs. Two weeks after the last treatment, the pre-ACTH plasma cortisol concentrations of corticosteroid-treated dogs returned to those of the control dogs, but the post-ACTH plasma cortisol concentrations remained suppressed. By 3 weeks after the last treatment, post-ACTH plasma cortisol concentrations of dogs treated with triamcinolone acetonide had returned to the range of values for the control dogs, but remained suppressed in the other 2 groups of dogs. All indices of pituitary-adrenocortical activity were within the control range by 4 weeks after the last treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

In vivo effects of glucocorticoids on serum corticosterone levels in rats.

In vivo effects of glucocorticoids on rat serum corticosterone level were investigated by using a simple and convenient HPLC-UV detection method. Serum corticosterone level was decreased by a single injection of hydrocortisone, methylprednisolone, betamethasone, dexamethasone or fluocinonide.

[Effect of difluprednate on adrenocortical and gonadal function].

The effects of difluprednate on the deposition of liver glycogen, the inhibition of adrenocortical function, the estrogenic, progestational and androgenic activities, and the excretion of electrolytes were investigated by comparing them with those of fluocinonide. The following results were obtained: 1) the deposition of liver glycogen was remarkably increased by subcutaneous administration of these two glucocorticoids, and in the dose of 0.1 mg/kg, the effect of difluprednate (35.1-fold the control value) was larger than that of fluocinonide (19.4-fold the control value) in mice. 2) the administration of difluprednate and fluocinonide greatly induced the decrease in the corticosterone concentration in the rat serum and adrenal gland (0.1 and 1 mg/kg, s.c.). 3) the estrogenic, progestational and androgenic activities were not recognized by administration of difluprednate in rats. 4) the two glucocorticoids induced an increase in the electrolytes excretion (especially K+) and the urine volume. 5) by the repeated injection of difluprednate (1.0 mg/kg) and fluocinonide (0.1 mg/kg), decrease of the body weight was observed in all of the experimental animals. In these experiments, it was recognized that the glucocortical action of difluprednate was similar or more potent in comparison with the action of fluocinonide and that the systemic effects of fluocinonide such as body weight loss was larger than that of difluprednate.

Adrenal suppression with high-potency corticosteroid ointment formulations in normal subjects.

In this 14-day, double-blind, in-clinic study, 24 healthy male volunteers were assigned at random to one of four treatment groups to compare the effects of a new formulation of 0.05% diflorasone diacetate ointment in a vehicle of propylene glycol (PG) with the effects of ointments of 0.05% fluocinonide, 0.05% clobetasol propionate, or the vehicle for diflorasone diacetate PG. The medication was applied to 75% of each subject's total body surface once a day for six consecutive days. During treatment and four days before and four days after treatment, various indicators of adrenal suppression were measured. A reduction in plasma cortisol levels was seen in several patients in each treatment group during the pretreatment period (days 1-4). Plasma cortisol continued to decrease during treatment and tended to return to pretreatment levels after cessation of therapy. There were no statistically significant differences between treatment groups. The lowering of plasma cortisol values in all groups was attributed to the applied medications as well as to the volunteers' change in daily routine and environment.

Applications of glucocorticosteroids. The effects of twice-daily vs once-every-other-day applications on mouse epidermal cell DNA synthesis.

To determine a schedule for continuous suppression of epidermal cell DNA synthesis, 0.05% fluocinonide ointment was applied either twice daily or once on alternate days to hairless mouse skin for 174 hours. Suppression occurred similarly for both regimens for 78 hours followed by an increase in DNA synthesis despite continued application of fluocinonide. The systemic effect was less marked with the alternate-day schedule. The effect of a single application of the fluocinonide ointment was also studied over 174 hours. Substantial inhibition of DNA synthesis occurred but for a shorter period followed by an increase and then return to normal values. Experiments were performed where second applications were made at various time intervals after the first. These did not appear to affect events until 192 hours had elapsed, when a second application caused profound inhibition of DNA synthesis once more.

Epidermal thinning: evaluation of commercial corticosteroids.

Epidermal thinning of mouse tail skin was compared for commercial preparations of clobetasol propionate (Dermovate), clobetasone butyrate (Eumovate), fluocinonide (Metosyn), and hydrocortisone butyrate (Locoid). The thickness measurements were ranked with those for hydrocortisone (1%), betamethasone valerate (Betnovate), triamcinolone acetonide (Ledercort), fluocinolone acetonide (Synlar), and prednisolone stearoylglycolate (Sinistrone) obtained in a previous study (Spearman and Jarrett, 1975). All steroids caused epidermal thinning, except clobetasone butyrate. Some cream and ointment vehicles were also assayed. Epidermal thickening was caused by the cream and ointment vehicles used for Eumovate and also by the cream employed for Locoid formulation.

The influence of synthetic corticoids on the isolated vas deferens of the rat.

The influence of corticosteroids on the tone of isolated smooth muscle (vas deferens of the rat) is reported. Corticoids alone are without effect on the muscle tone, but they potentiate the constrictor effect of catecholamines. Fluorinated synthetic steroids exhibit a more pronounced effect than non-halogenated ones.

Effects of antiinflammatory agents on mouse skin tumor promotion, epidermal DNA synthesis, phorbol ester-induced cellular proliferation, and production of plasminogen activator.

The antinflammatory ateroids fluocinoine acetonide, fluocinonide, and fluclorolone acetonide were found to be very effectiveinhibitory agents of mouse skin tumor promotion. These steroids also drastically inhibited epidermal DNA synthesis and epidermal cellular proliferation induced by a phorbal ester tumor promoter. In addition, these compounds were potent inhibitors, of plasminogen activator production in tumor cell cultures. The clinically used non-steroidal antiinflammatory agents oxyphenbutazone, indomethacin, and Seclazone also inhibite tumor promotion but were much less effective. Although these agents are useful against inflammatory disorders in general when given p.o., in our studies they had little effect on inflammation and epidermal cellular proliferation induced by a phorbol ester tumor promoter when given topically. The afore mentioned nonsteroidal antiinflammatory agents also had little effect on epidermal DNA synthesis. Oxyphenbutazone and indomethacin were less potent inhibitors of plasminogen activator production in tumor cells than were the antiinflammatory steroids, and Seclazone produced a negligible inhibition. There is, therefore, a general correlation in the potencies of a series of steroidal antiinflammatory agents for inhibition of tumor promotion and their ability to inhibit plasminogen activator production by tumor cell cultures and epidermal DNA synthesis.

Tachyphylaxis to topically applied steroids.

Acute tolerance to the vasoconstrictive effect of topically applied glucocorticoids in humans has recently been described. Similar acute tolerance to the antiproliferative properties of topically applied steroids in proliferating and normal hairless mouse epidermis is now shown to occur. Thus, at first, topically applied steroids cause profound inhibition of DNA synthesis and mitosis in the epidermis, but with continued treatment, DNA synthesis and mitosis recover and the tissue becomes insensitive to further stimulation. This suggests that there is an optimum approach to therapy with topically applied steroids that might reduce the incidence of side effects and improve therapeutic efficacy.

Acute tolerance to effects of topical glucocorticosteroids.

Tachyphylaxis (acute tolerance) to two important actions of topical glucocorticosteroids is demonstrated. In humans, topically active glucocorticoids applied to the skin cause vasoconstriction initially but this response fades with successive applications. In hairless mouse epidermis, inhibition of DNA synthesis and mitosis is caused by one application of a glucocorticoid but with repeated administration of the drug this effect is lost. The degree of inhibition of cell division appears to be similar 30 and 54 h after one application of steroid to that following three or five applications during such a time period. These findings suggest that an optimum therapeutic approach to the use of these agents exists.