Quantification of epidermal histological changes induced by topical retinoids and CD271 in the rhino mouse model using a standardized image analysis technique.

The rhino mouse has been used as an experimental model to screen topically active comedolytic agents. Adult rhino mice were treated on the back once daily for 5 consecutive days per week during 3 weeks. Skin histological preparations were analyzed by image analysis techniques to quantify the number of epidermal comedones, comedo profile and epidermal thickness. Using both a negative (treated with acetone) and a positive (treated with Aberel gel 0.025%) control group of animals in all experiments conducted over a period of about 3 years, we defined the upper and lower limit of acceptability of the results. Topical treatment with an acetone solution of all-trans retinoic acid (0.01, 0.03, 0.1%) and 13-cis-retinoic acid (0.1%) induced comedolysis and a marked increase in epidermal thickness. Commercial preparations of all-trans retinoic acid (Aberel lotion, gel and cream, Retin A cream, Retacnyl cream) presented a similar comedolytic activity. However, the epidermal thickening was higher with Retin A and weaker with Retacnyl. CD271, a new modulator of cell differentiation, applied either in acetone solution (0.01, 0.1%) or in lotion, gel or cream formulations (0.1%) also demonstrated a marked activity (i.e. comedolysis and epidermal thickening). These data confirm that the rhino mouse model can be used to assay drugs applied either in solvent or in topical formulations. Activity in this model compares favorably with published clinical observations in the treatment of acne.

Hexadecane-induced skin hyperplasia in the hairless rat: time course of histological and biochemical events related to the synthesis of polyamines and DNA.

Several biochemical parameters including ornithine decarboxylase activity (ODC) and tissue polyamine levels were measured during the hexadecane-induced epidermal hyperplasia of hairless rat skin. Animals received three applications of 200 microliters pure n-hexadecane on day 1. ODC activity and polyamine levels (putrescine, spermidine and spermine) in the epidermis were significantly increased and reached maximum elevations at 12 h after the start of n-hexadecane treatment with DNA synthesis peaking at 24 h. Histological studies confirmed a significant cellular edema at 24 h after the beginning of the treatment followed at 48 h by an epidermal hyperplasia which was maximum at 72 h. These data support the view that ODC activation, increased biosynthesis of polyamines and DNA are early events in epidermal cell hyperproliferation.

Effects of antiinflammatory agents on edema and DNA synthesis induced by 12-O-tetradecanoylphorbol-13-acetate in the guinea pig.

Inflammation and hyperplasia are frequently associated in skin diseases. In order to verify this relationship, we studied the antagonistic effect of different classes of antiinflammatory agents on the inflammatory and hyperplasiogenic responses elicited by one topical application of 12-O-tetradecanoylphorbol-13-acetate (TPA) to the ear of the guinea-pig. Edema and DNA synthesis were chosen as relevant parameters. All antiinflammatory agents tested significantly inhibited DNA synthesis induced by TPA. Moreover, all compounds except quinacrine and phenylbutazone also inhibited edema formation. In conclusion, our results demonstrate that while edema and hyperplasia are frequently associated, this is not always the case.

Measurement of arachidonate and its metabolites extracted from human normal and malignant gastrointestinal tissues.

This is the first report of human gastrointestinal arachidonate and prostanoids measured quantitatively by gas chromatography-mass spectrometry (GC-MS) in extracts of human cancers and macroscopically normal tissues from the stomach and colon. There were microgram/g amounts of arachidonate, and the particularly high yield from the tumours may explain why they usually produce more prostaglandins than the normal tissues in which they arise. There was only a small conversion of the arachidonate into prostanoids. 6-Keto-PGF1 alpha was the most abundant metabolite measured, particularly in the tumour extracts, with smaller amounts of prostaglandins E2, F2 alpha and D2.

A rapid and simple test system for the evaluation of the inhibitory activity of retinoids on induced ornithine decarboxylase activity in the hairless rat epidermis.

In mouse skin, antiproliferative agents including retinoids inhibit induction of ornithine decarboxylase activity by a variety of hyperproliferative stimuli. In the hairless rat skin, ornithine decarboxylase activity was induced by ten successive strippings with cellotape and by topical application of 12-O-tetradecanoyl-phorbol-13-acetate. Topical application of all trans-retinoic acid (25 nmol/cm2) immediately after the tape stripping of the skin significantly inhibited the induction of ornithine decarboxylase activity at all time points measured. The inhibition by all trans-retinoic acid of ornithine decarboxylase induced by cellotape stripping was dose dependent as was found to be the case for arotinoid, retinol, Ro-10-1670, motretinid, 13-cis-retinoic acid, etretinate, and vitamin A. Oral administration of all trans-retinoic acid also inhibited the ornithine decarboxylase activity induced by cellotape stripping. We propose the assay of ornithine decarboxylase activity in the hairless rat epidermis after tape stripping for a rapid evaluation of new retinoids.

Lipoxygenase products of arachidonic acid in human inflamed skin.

Monohydroxy acids (HETEs) and leukotriene B4 (LTB4) metabolites of arachidonic acid were measured in skin of healthy volunteers after ultraviolet B irradiation, and in the uninvolved skin of psoriatics after topical dithranol application. Exudate was collected from suction bullae on control and inflamed abdominal skin, and analysed for 12-HETE and PGE2 by GC-MS and LTB4 by bioassay. 12-HETE and PGE2 were raised at 24 h but not at 72 h after u.v.B irradiation: control and 24 h values were 13.7 and 41.5 ng ml-1 (P less than 0.05, n = 6) for 12-HETE respectively, and 4.5 and 30.2 ng ml-1 (P less than 0.01, n = 6) for PGE2. Dithranol application raised PGE2 levels from 23.1 ng ml-1 in control exudate to 62 ng ml-1 (P less than 0.01, n = 6) at 24 h before declining to base levels at 72 h. However, 12-HETE was raised at 72 h (200 ng ml-1, P less than 0.01, n = 5) but not at 24 h (104 ng ml-1) compared to control levels (50 ng ml-1, n = 5). The levels of the LTB4 were low (less than 100 pg ml-1), and no significant increases were observed. Arachidonic acid in inflamed skin can be metabolised by the cyclo-oxygenase and lipoxygenase pathway. It is probable that the lipoxygenase product 12-HETE is involved in these inflammatory reactions.

Arachidonic acid and prostaglandin levels in dithranol erythema: time course study.

Arachidonic acid and prostaglandins were measured in uninvolved psoriatic skin before and after treatment for up to 24 h with dithranol. Prior to treatment, skin exudate contained 2744 ng ml-1 arachidonic acid and 26.4 ng ml-1 PGE2. After treatment with dithranol, the arachidonic acid concentration increased to a maximum of 5556 ng ml-1 at 48 h whilst PGE2 increased to 93.4 ng ml-1 at 12 h and then declined. The erythemal response was apparent at 6-12 h and maximal at 72 h. These results suggest that PGE2 mediates the early development of dithranol erythema.

Histologic changes associated with ultraviolet A--induced erythema in normal human skin.

We have examined the effects of a standardized, moderately erythemogenic dose of long-wave ultraviolet (UVA) radiation on normal human skin, with the use of an appropriately filtered solar simulator and sequential biopsy specimens processed as 1-micron Epon-embedded sections. Histologic changes were present immediately after irradiation and evolved slowly during the 48-hour study. The epidermis manifested slight intracellular and intercellular edema and progressive loss of Langerhans cells to approximately one-fifth control values. A dermal infiltrate of neutrophilic polymorphonuclear leukocytes was present in all postirradiation specimens and peaked at 3 hours. A perivascular lymphocytic infiltrate, moderate endothelial cell enlargement, mast cell hypogranulation, occasional massive venular dilation, and sparse red blood cell extravasation were also noted. Overall, our findings expand and quantify earlier impressions that, compared to UVB, UVA has a relatively greater histologic effect on the dermis than on the epidermis, depletes epidermal Langerhans cells, and recruits neutrophils into irradiated human skin.

Relative potencies of several non-steroidal anti-inflammatory drugs against soya bean 15-lipoxygenase.

The 15-lipoxygenase from soya bean has been claimed to represent a rapid assay procedure for drugs and chemical agents that inhibit other lipoxygenase products (Kingston, 1981). Although the exact role of products derived from skin lipoxygenase enzymes is not well established, many of these products, e.g. leukotriene B4, have inflammatory characteristics that warrant further examination (Camp, 1982). We have therefore started to evaluate anti-inflammatory drugs in vitro for their ability to inhibit soya bean 15-lipoxygenase, before appraising their effects in vivo in animal and human skin.

Increased concentrations of arachidonic acid, prostaglandins E2, D2, and 6-oxo-F1 alpha, and histamine in human skin following UVA irradiation.

The buttock skin of clinically normal human subjects was subjected to approximately 2.5 minimal erythema doses of ultraviolet A irradiation. Deep red erythema developed during irradiation, faded slightly within the next few hours, increased to maximum intensity between 9-15 h, and decreased gradually thereafter although still persisting strongly at 48 h. Suction blister exudates were obtained at 0, 5, 9, 15, 24, and 48 h after irradiation as well as suction blister exudates from a contralateral control site and assayed for arachidonic acid, prostaglandins D2 and E2, and the prostacyclin breakdown product 6-oxo-prostaglandin F1 alpha by gas chromatography-mass spectrometry, and for histamine by radioenzyme assay. Increased concentrations of arachidonic acid and prostaglandins D2, E2, and 6-oxo-prostaglandin F1 alpha were found maximally between 5-9 h after irradiation, preceding the phase of maximal erythema. Elevations of histamine concentration occurred 9-15 h after irradiation, preceding and coinciding with the phase of maximal erythema. At 24 h, still at the height of the erythemal response, all values had returned to near control levels. Hence increased concentrations of arachidonic acid and its products from the cyclooxygenase pathway, and of histamine, accompany the early stages up to 24 h. A causal role in production of the erythema seems likely for these substances although other mediators are almost certainly involved.