Species-dependent effects of adenosine receptor agonists on contractile responses of vas deferens to ATP.

1. Experiments were carried out to examine the postjunctional actions of adenosine receptor agonists on the smooth muscle of the vas deferens of the guinea-pig and rabbit. 2. Although they produced neither contraction nor relaxation by themselves, adenosine analogues enhanced contractions of the guinea-pig vas deferens induced by 10 microm ATP. The rank order of potency was N6-cyclopentyladenosine (CPA) > 5'-N-ethylcarboxamidoadenosine (NECA) > adenosine > CGS 21680. Dose-response curves for NECA were shifted to the right by the nonselective adenosine receptor antagonist 8(p-sulphophenyl)theophylline (8-SPT; 100 microM) and by the selective A1-receptor antagonist 1,3-dipropyl-8-cyclopentylxanthine (DPCPX; 1 mM). 3. In the rabbit vas deferens, contractions induced by ATP (1 mM) were inhibited rather than facilitated by NECA. Neither CPA, R(-)-N6-(2-phenyl isopropyl)-adenosine (R-PIA) nor CGS 21680 had any effect. 4. The results indicate that the smooth muscle of the guinea-pig vas deferens expresses facilitatory adenosine A1 receptors but not adenosine A2 receptors. In contrast, in rabbit there are postjunctional inhibitory adenosine A2A receptors but not adenosine A1 receptors.

Commercial sunscreen lotions prevent ultraviolet radiation-induced depletion of epidermal Langerhans cells in Skh-1 and C3H mice.

There is much controversy regarding the ability of sunscreens to prevent ultraviolet (UV)-induced immune suppression. Epidermal Langerhans cells (LC) play a key antigen-presenting role in the afferent limb of the immune system's response to antigens introduced through the skin. It has been suggested that depletion of LC in UV-exposed skin is a critical step toward the induction of immunosuppression by UV radiation. There are a number of disparate reports with inconsistent results concerning the ability of sunscreens to prevent UV-induced depletion of LC. The purpose of this study was to systematically evaluate the ability of sunscreens to prevent UV-induced LC depletion in mice. Epidermal sheets obtained from skin biopsies taken from mice exposed to UV radiation from Kodacel-filtered FS20 sunlamps, which do not emit UV power at wavelengths < 290 nm, were immunoperoxidase stained for LC using a rat monoclonal antibody against mouse Ia (major histocompatibility complex class II antigen). Time course and dose-response curves for LC depletion were generated for Skh-1 and C3H mice. Dose-response curves for acute UV exposure induced depletion of LC in Skh-1 and C3H mice were similar, but not identical. LC density in the skin of Skh-1 mice that received chronic UV exposure (3 days/week for 8 weeks) was reduced by 62% after 2 weeks of exposure, but returned to normal levels by 6 weeks. Five commercial sunscreen lotions with labeled sun protection factors (SPF) of 4, 8, 15, 30 and 45 were tested for their capacity to block UV-induced depletion of LC. LC were depleted approximately 75% in the skin of unprotected or placebo lotion treated Skh-1 mice exposed to UV given on two consecutive days. Conversely, LC depletion was prevented in similarly UV exposed Skh-1 mice protected with a SPF 30 sunscreen. In C3H mice the levels of protection against LC depletion provided by the five sunscreens were proportional to the level of protection predicted by their labeled SPF. Comparisons of dose-response curves showed that significantly higher doses of UV were required for LC depletion and induction of skin edema than for the induction of local suppression of contact hypersensitivity. Thus, at UV doses where sunscreens provide complete protection against immunosuppression of contact hypersensitivity, prevention of LC depletion and skin edema would be expected.

Percutaneous penetration of dimethylnitrosamine through human skin in vitro: application from cosmetic vehicles.

Human skin penetration of N-dimethylnitrosamine (DMN) from three vehicles has been determined in vitro. When applied as an infinite dose in isopropyl myristate (IPM, 1 microgram/microliter) the average total absorption over 48 hr was 2.6 +/- 1.2% of the applied dose (all data presented are expressed as means +/- standard errors). When applied as a finite dose in a representative oil-in-water emulsion vehicle the average total absorption over 48 hr was 4.0 +/- 0.3% of the applied dose. When applied as a finite dose in a representative shampoo vehicle for 10 min followed by rinsing (i.e. to represent in-use exposure conditions) the average total absorption over 48 hr was 1.1 +/- 0.1% of the applied dose. Approximately 72% of the DMN in the applied shampoo vehicle was removed by rinsing. There was considerable evaporative loss of DMN from the IPM and oil-in-water emulsion vehicles, such that absorption was complete within 3 hr of application. The overall data indicate that DMN can penetrate the skin rapidly but that in practice the amount actually available for penetration is significantly reduced by high permeant volatility. In contrast, application of N-nitrosodiethanolamine (NDELA) at a concentration of 1 microgram/microliter as an infinite dose generated an average total absorption over 48 hr of 23.6 +/- 6.4%, representing a total flux of 103.9 +/- 28.4 micrograms/cm2. In the case of NDELA, no evaporative loss was evident.

Percutaneous penetration of N-nitrosodiethanolamine through human skin (in vitro): comparison of finite and infinite dose applications from cosmetic vehicles.

N-Nitroso compounds (nitrosamines) have been detected at the parts per billion level in a wide variety of matrices including industrial chemicals, pharmaceuticals, and food. Although N-nitrosodiethanolamine (NDELA) may be detected as an impurity in some cosmetic products, studies on NDELA absorption through human skin have been limited. A study to determine the extent of NDELA absorption following topical application was therefore undertaken to assist in the proper assessment of risk following unintended exposure. NDELA absorption was measured in vitro through human cadaver skin using isopropyl myristate (IPM) and generic prototype personal-care formulations (sunscreen and shampoo) spiked with [14C]NDELA. When applied as a finite dose at a concentration of 0.06% or lower, NDELA absorption was found to be a linear function of concentration. Total absorption at 48 hr ranged from approximately 35 to 65% of the dose and was formulation dependent (IPM > shampoo > or = sunscreen). Absorption occurred relatively rapidly from all formulations and peak rates of absorption were seen within the first 5 hr from the IPM and shampoo formulations. When applied as an infinite dose, total NDELA absorption followed a different rank order (shampoo > or = IPM > sunscreen) and evidence of barrier damage was seen with the shampoo formulation.

The ultraviolet C energy emitted from FS lamps contributes significantly to the induction of human erythema and murine ear edema.

FS ultraviolet (UV) lamps are used extensively to study the biological effects of ultraviolet radiation. Using published data, we investigated the relative contributions of the UVC (250-289 nm), UVB (290-319 nm), and UVA (320-400 nm) portions of the FS lamp emission spectrum to the induction of human erythema and murine ear edema. Many investigators contend that the biological activity of the spectrum emitted from these lamps resides primarily in the UVB region, based on the proportions of the power emission spectrum in this region (53.6%) versus the small relative contribution of UVC (3.2%) and the low biological activity of UVA (43.2%). However, if the biological effectiveness spectrum of FS lamps is calculated by multiplying the power spectrum with different action spectra, the biological effects of UVC are readily observed. For example, 10.4% of the murine ear edema activity and 11.1% (Parrish) or 16.7% (McKinlay-Diffey) of the human erythemal activity is due to the energy emitted in the UVC region. Experimental determination of human erythema and murine ear edema demonstrated that, for an equal amount of energy delivered, radiation from the unfiltered lamps was more potent in causing these responses than radiation from filtered lamps, and the ratio of effectiveness could be predicted by the effectiveness spectra. Thus, the contribution of UVC emitted from FS lamps to biological effectiveness spectra should not be ignored.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of systemic cyclosporin A on a hairless mouse model of photoaging.

The mechanisms that cause skin wrinkling in response to chronic exposure to sunlight are unknown. We investigated the possibility that wrinkling of Skh-1 hairless mice is associated with an ultraviolet (UV) radiation-induced immunologic alteration. Exposing Skh-1 hairless mice to a regimen of nonerythemal UV-B (290-320 nm) radiation induced skin wrinkles after 6-7 weeks. Concomitant treatment with cyclosporin A decreased the time to the onset of wrinkles to approximately 4 weeks. Exposing HRS/J hairless mice or athymic nude mice to a similar nonerythemal UV-B radiation regimen for 10 weeks failed to induce skin wrinkles. Concomitant administration of cyclosporin A and UV-B radiation for 7 weeks to HRS/J hairless mice induced no skin wrinkles. Ultraviolet-B or UV-B plus cyclosporin A exposure caused increased immunohistochemical staining for Ia and F4/80 antigens in the upper dermis of tissue from Skh-1 mice, as compared to controls. Treating Skh-1 mice with UV-B radiation plus cyclosporin A was also associated with a large increase in the number of CD3+ cells in the dermis. These staining patterns were absent in similarly treated HRS/J hairless mice. Dermal mast cell numbers in Skh-1 mice were 2-3-fold higher than in HRS/J, athymic nude or NSA mice. Treatment with cyclosporin A increased Skh-1 dermal mast cell numbers approximately 2-fold but had no effect on the dermal mast cell numbers in HRS/J or NSA mice. Based on these findings we postulate that UV-B light and cyclosporin A exacerbate an immunological condition in Skh-1 mice, one consequence of which is manifested as skin wrinkles. Thus, the induction of skin wrinkles in this mouse strain may have no relevance to the wrinkles observed in human skin after chronic exposure to sunlight.

The hairless mouse model of photoaging: evaluation of the relationship between dermal elastin, collagen, skin thickness and wrinkles.

Quantitative and qualitative changes in dermal collagen and elastin occur in response to chronic ultraviolet (UV) irradiation. These changes have been implicated in the genesis of the wrinkling seen in chronically irradiated, or photoaged skin. We examined the relationship between wrinkle formation and changes in dermal structural protein content and type. Skh-1 hairless mice were irradiated with suberythemal doses of UV-B three times a week for up to 20 wk. Visible wrinkling was present after 6-7 wk of irradiation. Dermal elastic fiber content was quantified by color image analysis of paraffin-embedded tissue. There was no significant difference in dermal elastic fiber content between irradiated and age-matched control mice after either 10 or 20 wk of irradiation. The effect of UV-B irradiation on total dermal collagen content, ratio of collagen type III-type I, and extent of glycosylation and crosslinking of collagen was no different in irradiated and age-matched control mice after 10 wk of irradiation. Increased epidermal thickness was evident in frozen sections after 6 wk of irradiation, and the thickness increased with continued irradiation. Dermal thickening was evident after 10 wk of irradiation. Sufficient UV-B irradiation will eventually cause changes in dermal elastin and collagen content; however, wrinkle formation precedes such changes. A causal relationship between wrinkle formation and dermal structural protein content changes in Skh-1 hairless mice could not be established in this study.

Quantitative determination of murine dermal elastic fibers by color image analysis: comparison of three staining methods.

We compared three different staining methods to determine if the dermal elastic fiber content of the HRS/Skh-1 hairless mouse could be accurately measured by color image analysis. Comparisons were made among Kligman's modification of Luna's mast cell stain for elastin, Unna's orcein stain with or without potassium permanganate preoxidation, and Gomori's aldehyde fuchsin stain with potassium permanganate preoxidation. The color image analysis system could be used to identify and quantify murine dermal elastin fibers in sections stained by all three methods. Gomori's aldehyde fuchsin stain with preoxidation demonstrated twice the content of dermal elastic fibers demonstrated by either Kligman's modification of Luna's mast cell stain or Unna's orcein stain with or without preoxidation. Gomori's aldehyde fuchsin method with preoxidation should be considered the stain of choice for evaluating murine dermal elastic fiber content.

Numbers of murine dermal mast cells remain unchanged during chronic ultraviolet B irradiation.

Dermal mast cell numbers reportedly increase in response to chronic ultraviolet irradiation in both humans and in the HRS/Skh-1 mouse model of human photoaging. It has been hypothesized that these increased numbers of mast cells are responsible, at least in part, for the damage in this chronically irradiated or photoaged skin. However, few actual quantitative data have been reported to support this claim of increased dermal mast cell numbers caused by chronic ultraviolet irradiation. We sought to quantify the numbers of dermal mast cells in the skin of chronic ultraviolet-irradiated and control HRS/Skh-1 hairless mice. Dermal mast cells from irradiated and age-matched control mice were quantified by digital image analysis during a 20-week period of exposure to ultraviolet B (UVB) radiation. During the entire course of irradiation, there was no difference in the numbers of dermal mast cells between the irradiated and nonirradiated age-matched control mice. Visible physical evidence of the effects of chronic UVB irradiation, i.e., skin wrinkling, was evident after 6 weeks of treatment. The numbers of dermal mast cells in unirradiated age-matched NSA (CF-1) haired mice were three- to four-fold lower than those in either ultraviolet-exposed or unexposed HRS/Skh-1 mice. These findings indicate that dermal mast cell numbers in HRS/Skh-1 mice are not increased by chronic exposure to UVB radiation.

Teaching employees breast self examination. Factors that may influence practice.

Breast self examination is a widely known technique used for detection of breast lumps. Many women do not practice monthly breast self examination, although it has been proven that with early detection the prognosis of breast cancer can be improved greatly. When tend to be taught, or learn about, breast self examination at an early age. The educational techniques used during the instruction can influence the practice of BSE later on. Health professionals can make a difference in the practice of BSE by communicating the value of this detection technique. BSE is a viable breast cancer detection technique that employees can be taught easily in employee wellness programs by health professionals.

The in-vitro percutaneous absorption of glycerol trioleate through hairless mouse skin.

The chemical composition of the aqueous receptor fluid in one-chambered diffusion cells used for in-vitro percutaneous absorption studies has been shown to significantly affect the apparent extent of absorption of the triglyceride, glycerol trioleate. Using murine skin samples it was found that the addition of albumin to the receptor fluid resulted in an increase in the apparent extent of absorption, while the presence of the bacteriostatic agent thiomersal resulted in a decrease. The viability of the skin samples had no effect on absorption. It was determined that the chemical species in the receptor fluid was the free fatty acid. Albumin presumably bound the fatty acid, thereby creating a sink which enabled the fatty acid to partition from the skin into the receptor fluid.

Alkane-induced edema formation and cutaneous barrier dysfunction.

Certain mineral oils and hydrocarbons require repeated topical application to cause irritation. A structure activity relationship of pure n-alkanes was undertaken in a mouse ear edema model to investigate the mechanism of cumulative irritancy. Alkanes were applied twice daily over a 4-day period. Dodecane was found to be non-irritating, while tridecane elicited a response only at 96 h. Tetradecane was the strongest irritant with significant increases (p less than 0.05) in ear thickness observed at 48 h. Hexadecane, octadecane, and eicosane exhibited progressively decreasing activity. Permeability of the ears to hydrocortisone was monitored in vitro during tridecane- and tetradecane-induced irritation. Significant increases in permeability were observed 24 h before edema formation. A positive correlation was found between the extent of edema formation and enhancement of permeability. Loss of barrier function would result in increased cutaneous availability of the alkanes. Increased permeability prior to edema formation indicates that induction of barrier dysfunction may be a factor in the mechanism of alkane-induced irritation.

Recurring digital fibroma of childhood.

A recurrent digital fibroma of childhood is reported. This case illustrates difficulties in the management of these recurrent tumors. Despite tumor-free margins on the excised tumors, recurrence occurred at other sites. Recurrences or new primary lesions are reported in 75% of the cases. Because of the rare tendency of these lesions to regress and the high recurrence rate, an individualized approach based on lesion location and behavior is recommended.

Osseous choristoma of the tongue.

A 5-year-old black female, the youngest patient so far reported, presented with osseous choristoma of the tongue. The lesion was located on the dorsum of the posterior tongue at the circumvallate papillae area. The origin of the tumor is unknown; however, ossification of remnants of the branchial arches is accepted by most authorities. The tumor was pedunculated with no associated symptoms, and its excision was not followed by recurrence to date.

Propane and propylene formation during the microsomal metabolism of iproniazid and isopropylhydrazine.

Both iproniazid and isopropylhydrazine were metabolized to the hydrocarbon products, propane and propylene, with nearly identical Michaelis constants and rates. This reaction appeared to be catalyzed by microsomal cytochrome P-450. Isonicotinic acid, a product of iproniazid hydrolysis by various amidases, was produced in only very small quantities, suggesting that the other amidase product, isopropylhydrazine, may not be an obligatory intermediate in the pathway of hydrocarbon formation from iproniazid. Hydrocarbon formation from iproniazid was more sensitive to inhibition in vitro by bis-p-nitrophenylphosphate (used in vivo as an amidase inhibitor) than was isopropylhydrazine. Iproniazid must be directly metabolized by cytochrome P-450 to yield propane and propylene, presumably via an azo ester intermediate which could give rise to an isopropyl radical, the chemical species presumed to be responsible for the hepatoxicity apparent after administration of large doses of iproniazid in vivo.

Metabolic activation of the terminal N-methyl group of N-isopropyl-alpha-(2-methylhydrazino)-p-toluamide hydrochloride (procarbazine).

The NADPH-dependent microsomal metabolism of [14C]procarbazine, labeled on the terminal N-methyl group, resulted in the covalent binding of the drug to exogenously added DNA; this reaction was inhibited by metyrapone. Procarbazine metabolism was also shown to result in covalent binding of the methyl group of the drug to microsomal protein upon metabolism, but the extent of protein binding was at least an order of magnitude smaller than that seen with its primary oxidative metabolite. N-isopropyl-alpha-(2-methylazo)-p-toluamide. The characteristics of the reactions leading to the covalent binding of the N-methyl group of the azo derivative to microsomal protein and its metabolism to form the hydrocarbon, methane, possessed a number of similarities in the apparent kinetic parameters (Km and Vmax), induction, and inhibition patterns indicating a common pathway of metabolism to form a reactive intermediate and the involvement of cytochrome P-450. Reduced glutathione stimulated methane formation and inhibited covalent binding to protein. One azoxy derivative, N-isopropyl-alpha-(2-methyl-ONN-azoxy)-p-toluamide, was chemically unstable and its decomposition was shown to lead to covalent binding to microsomal protein. A diazene intermediate and a methyl radical are proposed to be intermediates in the formation of methane during the oxidative metabolism of the azo derivative of procarbazine and a common intermediate in the activation of procarbazine may result in both covalent binding to cellular macromolecules and methane production. In addition, chemical decomposition of the azoxy metabolites may also contribute to a small portion of the covalent binding, but not to methane formation.

Effect of cytosolic components on the metabolism of the hydrazide iproniazid.

The effects of thiols, such as glutathione (GSH), and the cytosolic glutathione S-transferases on the microsomal metabolism of the hydrazide iproniazid to hydrocarbon products were investigated. Thiol compounds stimulated propane production and depressed propylene production. Addition of preparations of cytosolic proteins to the microsomal reaction mixtures in the presence of GSH depressed production of propane by more than 80% and propylene by 50% compared to the GSH-mediated reaction. The purified glutathione S-transferases A and B were most potent in eliciting this effect; isozymes AA, C, and E had little or no effect on hydrocarbon production. Further, a mixture of these purified isozymes in the concentrations known to exist in cytosol affected hydrocarbon production in a manner similar to cytosol. Experiments performed with isolated hepatocytes and an inhibitor of these cytosolic enzymes further supported the involvement of these enzymes in altered hydrocarbon production. These isozymes were subsequently shown to catalyze the formation of a GSH conjugate, S-(2-propyl)glutathione. The decreases in hydrocarbon production by microsomes in the presence of the glutathione S-transferases and GSH were accompanied by production of slightly larger amounts of conjugate. These data indicate that the cytosolic glutathione S-transferases interact with an oxidative microsomal metabolite of iproniazid to enzymatically produce an S-(2-propyl)glutathione conjugate and thus prevent formation of a reactive species which would otherwise chemically decompose to yield hydrocarbons or to covalently bind to cellular macromolecules.

The interactions of hydrazine derivatives with rat-hepatic cytochrome P-450.

The ability of different classes of hydrazine derivatives to modify cytochrome P-450 function during turnover as judged by loss of absorbance at 416 nm, loss of CO-reactive cytochrome P-450, or destruction of haem has been studied. Addition of monosubstituted hydrazines to rat-liver microsomes caused considerable loss of CO-reactive cytochrome P-450 and haem destruction; monosubstituted hydrazides caused mainly loss of CO-reactive cytochrome P-450, most likely due to abortive complex formation. Metabolism of 1,1-disubstituted hydrazines by microsomal cytochrome P-450 resulted in loss of CO-reactive cytochrome P-450 only, with no haem destruction. The 1,2-disubstituted hydrazines and hydrazides, procarbazine and iproniazid, acted similarly to the monosubstituted hydrazines, while 1,2-dimethylhydrazine elicited no response, either in observable spectral changes or loss of CO-reactive cytochrome P-450. Synthetic diazene intermediates of phenylhydrazine and N-aminopiperidine reacted rapidly with microsomal cytochrome P-450 to form a spectral intermediate resembling the putative iron porphyrin-diazenyl complex. The decomposition of certain iron porphyrin-diazenyl derivatives apparently leads to destruction of the porphyrin prosthetic group, most likely due to haem alkylation.