Endogenous distribution of retinoids during normal development and teratogenesis in the mouse embryo

Abstract

We have analysed the endogenous retinoids present in whole mouse embryos from day 9 to day 14 of development and in individual components of the embryo at two stages, day 10.5 and day 13, by HPLC. We can only detect two retinoids, all-trans-RA (tRA) and all-transretinol (t-retinol), and t-retinol is 510-fold in excess over tRA. We cannot detect 9-cis-RA or any didehydroretinoids; thus mammalian embryos seem to differ in their retinoid content from other embryos such as chick, Xenopus, and fish. The levels of tRA do not change significantly over the 6 days of development analysed, whereas t-retinol rises sharply as the liver develops. Within the embryo, tRA is present at high levels in the developing spinal cord and at very low levels in the forebrain; indeed there is a gradient of endogenous tRA from the forebrain to the spinal cord. Other parts of the embryo had intermediate levels of tRA. When a teratogenic dose of RA was administered to day 10.5 embryos, the levels of tRA present in individual tissues of the embryo rose dramatically-from 175-fold to 1,400-fold-and the levels rose in all tissues not in any exclusive areas. We then determined which areas of the embryo were malformed by such a teratogenic dose. The lower jaw, palate, vertebrae, tail, and limbs were consistently abnormal, and since these areas received a dose of tRA no higher than any other it was concluded that cell-specific factors must determine the teratogenic response of these tissues. We then considered whether cellular retinoic acid-binding protein I or I1 (CRABP I or 11) played any role in this response by determining their relative levels in each of the tissues analysed. There was no correlation between the presence of CRABP I and 11 and the distribution of administered RA. Neither was there a clear correlation in detail between the presence of CRABP I and I1 and the sites of teratogenesis. We therefore conclude that other factors, for example, nuclear factors, must be responsible for the teratogenic response to RA. o 1995 Wiley-Liss, Inc.

Retinoid metabolism: a balancing act

“…Questions about other retinoid metabolites remain, however, including the didehydroretinoids found in chick embryos, and retro-retinoids, a class of hydroxylated retinoids whose biological activities are apparently not mediated by nuclear receptors [12][13][14] . Perhaps most enigmatic is the role of 9-cis retinoic acid, a high-affinity ligand for RXR, whose existence in vivo has been questioned because of difficulties in extracting it from tissues 2,15 . As exemplified by the work of Niederreither et al 1 , continuing efforts focusing on retinoid metabolism will provide important answers.…”

Retinoid activation of retinoic acid receptor but not retinoid X receptor is sufficient to rescue lethal defect in retinoic acid synthesis

“…Even though a targeted mutation of the RXR␣ AF-2 ligand-binding domain shows that this protein domain is important for mouse development (14), there is no evidence that AF-2 needs to bind 9-cis-RA to perform its function. Also, although all-trans-RA is easily detectable in many mammalian tissues (15)(16)(17)(18), detection of 9-cis-RA as originally reported (6) has not been confirmed. Nevertheless, in vitro studies pursuing the mechanism of retinoid signaling suggest that transcriptional activation by RXR homodimers depends on 9-cis-RA (13), and that RAR/RXR heterodimers may in some contexts depend solely on binding of all-trans-RA to the RAR partner, so-called RXR subordination (19); but in other contexts RXR subordination is overcome, and binding of 9-cis-RA to RXR is involved in the transcriptional mechanism (20).…”

A transcriptionally silent RXRα supports early embryonic morphogenesis and heart development

“…Assuming that a requirement of RXR␣ AF2 actually reflects the binding of an agonistic ligand (20), such a requirement raises the question of the possible existence and of the nature of a physiological RXR ligand(s) in vivo. However, because 9-cis RA is undetectable in rodent embryos (42,43) Detection of the Rxra-and Rxrb-null mutations and deletion of RXR␣ AF2-AD core (corresponding to helix 12 amino acids 503-521; referred to as Rxrb af2o ) were as described (18,28,45).…”