Abnormal cholesterol biosynthesis as in Smith-Lemli-Opitz syndrome disrupts normal skeletal development in the rat

Smith-Lemli-Opitz syndrome (SLOS) in human infants is a common autosomal recessive malformation syndrome (estimated incidence, 1:20,000). It is characterized clinically by congenital anomalies, especially craniofacial and limb defects, and biochemically by a defect in 7-dehydrocholesterol-delta7-reductase activity (7DHC-reductase), the final enzyme in cholesterol biosynthesis. In previous studies, early administration of the 7DHC-reductase inhibitor AY9944 to pregnant rats resulted in a high frequency of holoprosencephaly, relevant to craniofacial anomalies of SLOS. In order to test the effect of AY9944 on limb development, we treated dams on gestation day 7 (GD7), which delays the biochemical defect to about GD13 to GD14. Sera were sampled on GD12, GD14, and GD21 and cholesterol and dehydrocholesterols (7DHC and 8DHC) were measured by gas-chromatography-mass spectrometry (GC-MS), as for the diagnosis of SLOS. GD21 fetuses were examined for gross malformations and skeletal development. In treated dams, the SLOS biochemical marker 7DHC accounted for one fourth and one third of total sterols, respectively, on GD12 and GD14, and cholesterolemia on these two gestation days was reduced by 50% and 43%, respectively, as compared with control values. This maternal metabolic defect was associated with decrease in fetal weight and delayed ossification. In addition, scapular malformations were observed in four fetuses from three litters. The malformations could have been caused by the same mechanism as holoprosencephaly after early treatment with AY9944. These cholesterol-deficiency-based malformations could have a common cause in the abnormal expression of Hedgehog or other developmental gene proteins, and may thus explain various congenital polymalformative syndromes in humans, including SLOS.