Analysis of limb anomalies induced in vitro by vitamin A (retinol) in mice

Abstract

Forelimb buds of day 11 ICR-JCL mouse embryos were cultured on liquid medium consisting of 90% Eagle's MEM and 10% fetal calf serum. Experimental medium contained 10 iu/ml vitamin A alcohol (retinol). In controls four of five metacarpals chondrified and the epidermis began to keratinize after 3-4 days of culturing. In experimental explants many pycnotic cells were observed in the peripheral mesenchyme in the hand plate, especially in pre- and postaxial regions. Only 2-3 metacarpals chondrified, and keratinization was inhibited in the experimental explants. Uptake of labeled sulfate was suppresssed by vitamin A. Excess vitamin A is thus thought (1) to act directly on limb buds, and thereby to induce limb anomalies, (2) to induce cell death in the mesenchyme, (3) to suppres the formation of chondroitin sulfate, and (4) to inhibit keratinization.

Effects of Retinoic Acid on the Development of the Facial Skeleton in Hamsters: Early Changes Involving Cranial Neural Crest Cells

“…The ultrastructural changes which nccur- red in the cranial neural crest cells generally involved the expansion and ultimate break down of cellular and subcellular membranes and resembled changes reported to result from vitamin A exposure in other tissues both in vivo [Morriss, 1973;Theodosis and Fraser, 1978;Taylor, 1979;Geelen et al, 1980] and in vitro [Glauert et al, 1963;Dingle and Lucy, 1965b;Daniel et al, 1966;Nakamura, 1975;Morriss and Steele, 1977], These changes may relate to the abil ity of vitamin A to insert into biological membranes [Glauert et al, 1963;Bangham et al, 1964;Lucy and Dingle, 1964;Dingle and Lucy, 1965a, b], Glauert et al [1963] have suggested that by doing so vitamin A may cause expansion of membranous struc tures thereby altering their permeability. This could be expected to alter fluid trans port across these barriers causing dilatation of organelles as well as swelling and even tual lysis of the cells.…”

Caspase‐3 mediates retinoid‐induced apoptosis in the organogenesis‐stage mouse limb

“…All animal studies were fully compliant with the guidelines of The Canadian Council on Animal Care. All‐trans retinol acetate (RA, vitamin A) (Sigma, St. Louis, MO), dissolved in 100% ethanol, was added to the designated cultures with a final percentage of ethanol of no more than 10 −4 M. Ethanol was not added to controls, as these concentrations have previously been shown not to affect the explants substantially (Nakamura, 1975). The cultures were terminated at the specified times.…”

Teratological research using in vitro systems. II. Rodent limb bud culture system.

“…To answer some of these questions, the pattern of changes was carefully compared in the in vitro growth and differentiation of the limb bud after exposure to a compound in vitro (57,72) or at the same stage in utero (25,50). Results indicated that vitamin A probably acts directly on the limb bud to produce characteristic malformations and that suppression of the formation of chondroitin sulfate may be partly responsible for the morphological alterations.…”