Histological dermal changes caused by preparation and application procedures in percutaneous dose toxicity studies in dogs, rabbits and rats.

We reevaluated histological slides of dorsal skin in control animals from past percutaneous dose toxicity studies using dogs, rabbits and rats to provide background data concerning histological changes related to preparation and application procedures and vehicles or embrocations of every variety. Acanthosis, dermal or perifollicular inflammatory cell infiltration in dogs; hyperkeratosis, acanthosis, dermal inflammatory cell infiltration or hemorrhage in rabbits; and acanthosis, dermal inflammatory cell infiltration, crust or foreign body granuloma in rats were present as procedure-related underlying histological changes in the control animals. Four mechanical acts, (1) rubbing with gauze to remove an administered substance for reapplication, (2) use of a taut bandage to avoid slipping from the application site, (3) peeling a patch off as a preparation procedure for reapplication, and (4) clipping or shaving, were considered to cause injury to the skin. The degree of influence of the various application procedures was found to be as follows: sham, lotion < cream < ointment and tape in dogs; untreated control, sham < lotion < tape and poultice in rabbits; and sham, sodium carboxymethylcellulose < olive oil and lotion < ointment and tape in rats. The degree of ointment influence on rabbits is equivocal.

Semi-quantitative immunohistochemical analysis of male rat-specific alpha2u-globulin accumulation for chemical toxicity evaluation.

We purified male rat urinary alpha(2u)-globulin, prepared the antibody in rabbits, and improved an immunohistochemical detection method using this antibody for male rat-specific alpha(2u)-globulin accumulation appearing as hyaline droplets in the kidneys. Our prepared antibody reacted specifically with alpha(2u)-globulin in both immunohistochemical and Western blotting analyses, furthermore, and the graded immuno-reactivities on the slide were well associated with computational image analyzing results. Using this method, we retrospectively analyzed the renal sections from the toxicity studies of 12 nephrotoxic chemicals, which had already been conducted under the Japanese Existing Chemicals Survey Program. We demonstrated that the hyaline droplets induced by treatment with 10 chemicals (1,4-dibromobenzene, dicyclopentadiene, 3,4-dimethylaniline, 1,4-dicyanobenzene, tetrahydrothiophene-1,1-dioxide, 1,3-dicyanobenzene, acenaphthene, 3,4-dichloro-1-butene, 3a,4,7,7a-tetrahydro-1H-indene and 3,5,5-trimethylhexan-1-ol) were directly associated with alpha(2u)-globulin accumulation. This immunohistochemical method is convenient for applying, even retrospectively, paraffin sections from general toxicity studies and could be useful for qualifying male rat-specific hyaline droplets consisting of alpha(2u)-globulin and renal risk in humans.

Effects of Various Antibiotics on the Excitability of Newt Ectodermal Cells in vitro: (amphibian embryos/epidermal action potential/cell culture/antibiotics).

With the addition of actinomycin D (AMD), cycloheximide (CH) or tunicamycin (TM) to a culture medium at 23°c, the epidermal action potentials (EAPs) of the cultured epidermal explants or monolayered cell colonies taken from newt embryos were examined for confirmation of their relationship to transcriptional and translational events. EAPs of ectodermal cells from the pregastrula were first recorded after 84 hr of monolayered cell culture in FCS-Steinberg's medium. The addition of 10 µg/ml CH or 1 µg/ml TM for 36 hr prevented the appearance of these potentials in cells precultured in FCS-Steinberg's medium for 60 hr. Ectodermal cells precultured in FCS-Steinberg's medium for 68 hr generated the EAPs after they were cultured in the same concentration of CH or TM for 28 hr. Thus, it appears that certain proteins and glycoproteins may be responsible for evoking the EAPs and that these molecules are synthesized at about 68 hr in culture following the pregastrula stage. Ectodermal cells precultured in FCS-Steinberg's medium for 48 hr generated EAPs after they were cultured in 0.5 µg/ml AMD for 72 hr. Cells precultured for 36 hr failed to produce the EAPs even when they were cultured in the same concentration of AMD for 84 hr.