Examination of a rodent limb bud micromass assay as a prescreen for developmental toxicity

Abstract

The mouse limb bud micromass assay is one of many short-term tests proposed as preliminary screens for potential developmental toxicity. Previous efforts to validate this assay have used too few "nonteratogens." The purpose of this study was to examine additional compounds, most of which, based on the literature, were perceived to have low potential for developmental toxicity in vivo. In addition, a method of data analysis was sought that would identify selective developmental toxins in the micromass assay, i.e., those that are effective at dosages not maternally toxic. The concentration of each of 23 compounds that produced a 50% inhibition (IC50) of radiolabeled thymidine (T) and sulfate (S) incorporation was determined and used to calculate a T/S ratio. The T/S ratio may be a useful measure of developmental hazard, since T incorporation measures toxicity toward a general cell function (DNA synthesis) and S incorporation measures mainly toxicity toward a developmentally specific cell activity (chondroitin sulfate synthesis). All compounds tested produced T/S ratios of less than 2.0. Since 22 of these 23 compounds are classified as "nonteratogens" or nonselective developmental toxins in vivo, a low T/S ratio in this in vitro assay system may be capable of discriminating potential for developmental hazard in vivo.

Extracellular matrix environment influences chondrogenic pattern formation in limb bud micromass culture: Experimental verification of theoretical models

“…Micromass culture of limb bud mesenchyme cells has been used as an assay system for studying the differentiation of chondrocytes (Wise et al, 1990) or the mechanisms of pattern formation (Leonard et al, 1991), and we chose this culture as an experimental system to verify theoretical models for several reasons. First, in micromass culture, some cells remain fibroblastic while others differentiate into chondrocytes, and the distribution of the chondrogenic cells forms a ''whorl''-like pattern (Cottrill et al, 1987), probably because this culture retains the periodic nature of the pattern in vivo (Newman, 1996).…”

Evaluation of Embryotoxic Potential of Olaquindox and Vitamin A in Micromass Culture and in Rats

“…Using Alcian blue, a stain specific for cartilage proteoglycans, the degree of chondrogenesis can be visualized in the micromass cultures as well as quantified by extraction of the stain and spectrophotometric determination of its absorbance. Several studies have shown a fairly good correlation between inhibition of chondrogenesis in vitro and teratogenic activity in vivo (Flint and Orton, 1984 ; Guntakatta et al , 1984 ; Wise et al , 1990 ; Aulthouse and Hitt, 1994) .…”

Numeric Estimates of Teratogenic Severity from Embryo–Fetal Developmental Toxicity Studies

“…Several lists of reference compounds for screening assays have been put forward (Smith et al., ; Brown, ), and individual studies have utilized additional reference compounds (e.g., Wise et al., ; Brannen et al., ). The most recent list proposed by Daston and colleagues (, ) has made an important improvement by associating an exposure metric with the developmental outcome.…”