Prevalidation of the Embryonic Stem Cell Test (EST)-A New In Vitro Embryotoxicity Test.

Pluripotent embryonic stem cells (ES cells) of the mouse (cell-line D3) can be maintained in the undifferentiated state in the presence of LIF (Leukaemia Inhibitory Factor). Upon withdrawal of LIF, these cells differentiate into various cell types under appropriate conditions. This property of ES cells allowed us to develop an in vitro embryotoxicity test, the Embryonic Stem Cell Test (EST; In Vitro Toxicology 1997, 10, 119-127), which does not require taking embryonic cells or tissues from pregnant animals.

The Performance of the Tissue Equivalent Assay using the Skin(2)(TM) ZK1200 Model in the COLIPA International Validation Study on Alternatives to the Draize Eye Irritation Test.

The tissue equivalent assay (TEA) (Osborne et al., 1995) was used to evaluate 55 mixed ingredients and formulations in the COLIPA International Validation Study on Alternatives to the Draize Rabbit Eye Irritation Test (Brantom et al., 1997).

Investigation of ingredient interactions in cosmetic formulations using isolated bovine corneas.

The purpose of this paper is to report on use of a modified bovine cornea opacity and permeability assay (BCOP) to test the effects of several cosmetic formulations on eye-derived tissue in vitro. The results from these studies suggest that a BCOP protocol using prolonged exposure and repeated treatments may be useful for screening the eye effects of cosmetic formulations. Further work will be required, however, before the model is ready for formal validation.

A summary report of the COLIPA international validation study on alternatives to the draize rabbit eye irritation test.

The principal goal of this study was to determine whether the results from a set of selected currently available alternative methods as used by cosmetics companies are valid for predicting the eye irritation potential of cosmetics formulations and ingredients and, as a consequence, could be valid replacements for the Draize eye irritation test. For the first time in a validation study, prediction models (PMs) that convert the in vitro data from an assay to a prediction of eye irritation were developed for each alternative method before the study began. The PM is an unequivocal description of the relationship between the in vitro and the in vivo data and allows an objective assessment of the reliability and relevance of the alternative methods.

New epidermal model for dermal irritancy testing.

An interlaboratory comparison of a new model of the human epidermis (EpiDerm) was conducted using a range of anionic and non-ionic surfactants and surfactant-containing final formulations. The toxicity of the materials was estimated by MTT conversion, using both concentration (EC(50)) and time (ET(50)) protocols. A range of 16 compounds was tested on different production lots of EpiDerm following storage periods of 1 and 2 days (after shipping) at MatTek and at two independent testing laboratories, Microbiological Associates (MA), USA and Scotland, UK.

Immunocytochemical localization of oxytocin in corpora lutea and luteinized cysts from anoestrous ewes stimulated with gonadotrophin-releasing hormone.

Anoestrous Romney Marsh ewes with or without progesterone pretreatment were injected with multiple low-doses of gonadotrophin-releasing hormone followed by a single, larger bolus. Blood samples were taken at twelve-hourly intervals for progesterone radioimmunoassay. Ewes were slaughtered on day 3 or 5 after the bolus injection, and the ovaries were collected for histology and immunocytochemical examination for oxytocin-immunoreactivity.

Heterogeneity in the luteal population following superovulation with pregnant mare serum gonadotrophin and human chorionic gonadotrophin in the sheep.

In order to investigate the development and possible heterogeneity in the luteal population following superovulation, anoestrous ewes were induced to ovulate using progestagen priming followed by injections of pregnant mare serum gonadotrophin (1000 IU) and hCG (1000 IU). Ovaries were recovered from ewes on each of days 2, 4, 6, 8, 10, 12 and 15, and the weight, progesterone content, 125I-labelled hCG binding and progesterone synthesis in vitro of the individual corpora lutea measured. The results obtained showed that plasma progesterone concentrations on the day of slaughter were significantly correlated with time (P less than 0.

Role of prostaglandin F-2 alpha and oxytocin in the regression of GnRH-induced abnormal corpora lutea in anoestrous ewes.

Anoestrous Romney Marsh ewes with (+P) and without (-P) progesterone pretreatment were induced to ovulate by multiple low-dose injection of GnRH followed by a bolus injection of GnRH. Luteal function was assessed by twice daily measurement of plasma progesterone. Animals were slaughtered on Days 3 or 5 after the end of GnRH treatment and CL and endometrium were recovered.

Effect of hysterectomy on the short life-cycle corpus luteum produced after GnRH-induced ovulation in the anoestrous ewe.

Anoestrous Romney Marsh ewes were treated with small-dose (250 ng) multiple injections of GnRH. Ewes in Groups 1 and 3 were hysterectomized 2 weeks before treatment, while those in Groups 2 and 4 were intact controls. Groups 1 and 2 were primed with progesterone (+P) and treated with 2 h injections of GnRH (250 ng) for 36 h, while Groups 3 and 4 were not pretreated (-P) but were given 2 h injections of GnRH (250 ng) for 18 h.

Function of abnormal corpora lutea in vitro after GnRH-induced ovulation in the anoestrous ewe.

Normal and abnormal corpora lutea were recovered from anoestrous Romney Marsh ewes on Days 3, 4, 5 and 6 after treatment with small-dose (250 ng) multiple injections of GnRH followed by a bolus injection (125 micrograms) with (+P) and without (-P) progesterone pretreatment and a study made of their characteristics in vitro. Plasma progesterone concentrations initially rose concurrently in all animals but abnormal luteal function occurred in 70% of the -P ewes and was defined on Day 5 when plasma progesterone concentrations declined relative to those in the +P ewes. All corpora lutea recovered on Days 3 and 4 appeared macroscopically similar and there were no significant differences between the +P and -P groups in terms of luteal weight, progesterone content and binding of 125I-labelled hCG on these days.

Function of abnormal corpora lutea in vivo after GnRH-induced ovulation in the anoestrous ewe.

Anoestrous Romney Marsh ewes with and without progesterone treatment (+P, -P) were treated with small-dose (250 ng) multiple injections of GnRH at 2-h intervals for 48 h. Animals were slaughtered on Days 4, 5, 7 and 11 after the end of GnRH treatment and luteal function was assessed by the measurement of daily plasma progesterone concentrations. In all animals which ovulated (29/32, 91%) peripheral progesterone concentrations rose to 0.

Progesterone pretreatment has a direct effect on GnRH-induced preovulatory follicles to determine their ability to develop into normal corpora lutea in anoestrous ewes.

In two experiments carried out during seasonal anoestrus, Romney Marsh ewes were treated with small-dose (250 ng) multiple injections of GnRH at 2-h intervals with and without progesterone pretreatment. In Exp. 1, 8/8 progesterone-primed ewes ovulated and produced functionally normal corpora lutea compared with 2/9 non-primed ewes.