Anomalies of Cartilaginous and Ossified Axial Skeleton in Rat Fetuses Treated with Cyclophosphamide: Type, Frequency, and Stage Specificity

The purpose of the present study was to assess the type, frequency and stage specificity of axial skeletal anomalies induced by cyclophosphamide (CP). Female Crj: Wistar rats were mated with male rats (vaginal sperm = gestation day 0, GDO). They were then injected intravenously with 5 mg/kg of CP in a saline solution on GD7, 8, 9, 10, 11, 12, 13, or 14, or with saline on GD8 for controls. Dams were sacrificed on GD20. Fetuses were stained with alcian blue and alizarin red S. Eighteen types of axial skeletal anomaly were detected, e. g., shifted ventral lamina, cervical rib, cleft arch of the atlas, axis fused to atlas, sternebral misalignment, bipartite sternum, fused ribs, change in pre-sacral vertebral number, supernumerary rib, misdirected transverse process, anteriorly-shifted spinous process, short spinous process, fused vertebral bodies, cartilaginous short rib at Th.7, short or absent rib at Th.12 or 13, dumbbell-shaped vertebral body, wavy rib, and rib cartilage not attached to the sternal cartilage. The present experimental study indicates that: 1) observations of cartilaginous skeletons can provide more information than observations of ossified skeletons; 2) the pattern of CP-induced anomalies along the anterior–posterior axis may depend on the sequence of development progression among structures, spinous process, vertebral arch, vertebral body, rib/transverse process; and 3) in more cranial vertebrae, anteriorly-shifted anomalies, such as anteriorly-shifted ventral lamina and anteriorly-shifted spinous processes, rather than posteriorly-shifted anomalies, may be the primary indicators for detecting developmental toxicity, since CP injection increased the frequencies of these anomalies.