Micro-computed tomographic evaluation of fetal skeletal changes induced by all-trans-retinoic acid in rats and rabbits

• Published in: Birth defects research. Part B. Developmental and reproductive toxicology 2010-10, Vol.89 (5), p.408-417
• Main Authors: Wise, L. David, Xue, Dahai, Winkelmann, Christopher T.
• Format: Article
• Language: English
• Subjects: alizarin red staining; all-trans-retinoic acid; Animals; Anthraquinones; Bone and Bones - abnormalities; Bone and Bones - embryology; Bone Development - drug effects; Bone mineral content; boric acid; Cesarean section; Computed tomography; Congenital defects; developmental toxicity; Female; Fetal Development - drug effects; Fetus - abnormalities; Fetus - drug effects; Fetuses; Hydroxyurea; imaging; Male; micro-Computed Tomography; micro-CT; Pregnancy; Rabbits; Rats; Rats, Sprague-Dawley; Retinoic acid; Rib; skeletal evaluation; Skull; Spine; Staining and Labeling; Supernumerary; Toxicity Tests; Tretinoin - adverse effects; Vertebrae; Viscera; X-Ray Microtomography - methods
• ISSN: 1542-9733; 1542-9741; 1542-9741
• DOI: 10.1002/bdrb.20267

BACKGROUND: Our laboratory has been conducting positive control studies to evaluate the utility of micro‐computed tomography (micro‐CT) for qualitative evaluation of fetal skeletal morphology. All‐trans‐retinoic acid (atRA) was used to produce a different spectrum of defects compared to our previous studies with boric acid and hydroxyurea. METHODS: Groups of five mated Crl:CD(SD) female rats each were administered vehicle or atRA (2.5–50 mg/kg) on GD 10, and groups of four mated Dutch Belted rabbits each were dosed with vehicle or atRA (6.25–25 mg/kg) on GD 9. Cesarean sections were performed on GD 21 and 28, respectively. Following external examination the viscera were removed and fetuses scanned in a micro‐CT imaging system. Fetuses were subsequently stained with alizarin red. Skeletal morphology was evaluated by each method without the knowledge of treatment group. Total bone mineral content (BMC) of each fetus was quantitated using the micro‐CT images. RESULTS: In rats there were dose‐related increases in the incidence of extra lumbar vertebra and non‐dose‐related increases in supernumerary ribs at all dose levels. There were decreases in mean number of ossified sacrocaudal vertebra at ≥5 mg/kg, and increases in skull bone malformations at ≥10 mg/kg. Rabbits were less sensitive on a mg/kg basis since skeletal malformations and a decrease in mean number of ossified sacrocaudal vertebra were observed only in the 25‐mg/kg group. Micro‐CT evaluation detected essentially the same incidence of skeletal abnormalities as seen in alizarin red‐stained rat and rabbit fetuses. BMC analysis showed a trend toward slight decreases in atRA‐treated rats, but no notable changes in rabbits. CONCLUSIONS: These results add support to our previous work that demonstrates that micro‐CT imaging can effectively assess rat and rabbit fetal skeletal morphology. Birth Defects Res (Part B) 89:408–417, 2010. © 2010 Wiley‐Liss, Inc.