Maternal Diets Deficient in Vitamin D Increase the Risk of Kyphosis in Offspring: A Novel Kyphotic Porcine Model

Abstract

Background: The purpose of this study was to explore the role of perinatal vitamin-D intake on the development and characterization of hyperkyphosis in a porcine model.

Methods: The spines of 16 pigs were assessed at 9, 13, and 17 weeks of age with radiography and at 17 weeks with computed tomography (CT), magnetic resonance imaging (MRI), histology, and bone-density testing. An additional 169 pigs exposed to 1 of 3 maternal dietary vitamin-D levels from conception through the entire lactation period were fed 1 of 4 nursery diets supplying different levels of vitamin D, calcium, and phosphorus. When the animals were 13 weeks of age, upright lateral spinal radiography was performed with use of a custom porcine lift and sagittal Cobb angles were measured in triplicate to determine the degree of kyphosis in each pig.

Results: The experimental animals had significantly greater kyphotic sagittal Cobb angles at all time points when compared with the control animals. These hyperkyphotic deformities demonstrated no significant differences in Hounsfield units, contained a slightly lower ash content (46.7% ± 1.1% compared with 50.9% ± 1.6%; p < 0.001), and demonstrated more physeal irregularities. Linear mixed model analysis of the measured kyphosis demonstrated that maternal diet had a greater effect on sagittal Cobb angle than did nursery diet and that postnatal supplementation did not completely eliminate the risk of hyperkyphosis.

Conclusions: Maternal diets deficient in vitamin D increased the development of hyperkyphosis in offspring in this model.

Clinical relevance: This study demonstrates that decreased maternal dietary vitamin-D intake during pregnancy increases the risk of spinal deformity in offspring. In addition, these data show the feasibility of generating a large-animal spinal-deformity model through dietary manipulation alone.

Fig. 1

Photograph of a dramatic example of the clinically apparent kyphotic deformity produced in a prior study via dietary manipulation alone at our institution. Of note, the porcine thoracic spine is typically less kyphotic than the human spine is.

Fig. 2

Photographs of the custom motorized lift used to hold the pigs in an upright position for radiography. The animal straddled the bar (left image) in the chute while the entire apparatus was elevated to lift the hooves off the ground and maintain the animal’s thorax in a horizontal but upright (quadruped) position (right image).

Fig. 3

Standing radiographs of a hyperkyphotic pig at 9 and 13 weeks of age.

Fig. 4

Predictions for control and kyphotic pigs over time. Control pigs maintained lower Cobb angles, while kyphotic pigs’ measurements increased. The pale dashed (teal and orange) shadowed lines represent the observed measurements, while the solid (teal and orange) lines represent the predicted curves over time.

Fig. 5

Midsagittal CT scans of the explanted control and kyphotic spines. The vertebral body fracture in the control spine was a result of the process used during euthanasia. Spines were positioned supine during this imaging.

Fig. 6

Midsagittal histologic evaluation of the apical and matched vertebrae of the control and kyphotic spines (magnification, ×1). In these slides, anterior is to the right, posterior is to the left, proximal is at the top, distal is at the bottom, and the vertebral disc is in the middle. Note that the anterior epiphysis of the kyphotic spine is less ossified and the cartilage is confluent with the physis compared with the control spines. H & E = hematoxylin and eosin.

Fig. 7

Trichrome-stained images centered on the anterior epiphysis of the control (Fig. 7-A) and the kyphotic (Fig. 7-B) vertebrae (magnification, ×2). In these images, the metaphyseal bone is at the bottom, the disc space is at the top, the epiphyseal (secondary) ossification center is at the left, and the fibers from the anulus are in the upper right. Note the relatively increased area of unossified cartilage of the anterior epiphysis in the kyphotic sample (Fig. 7-B). um = micrometer.

Fig. 8

Trichrome-stained histologic sections of the control (Figs. 8-A and 8-C) and kyphotic (Figs. 8-B and 8-D) vertebrae. The secondary center of ossification is at the top and the metaphyseal bone is at the bottom in these examples. Increased pleating of the physis in the kyphotic sample (Fig. 8-B; magnification, ×2) is clearly demonstrated when compared with the control (Fig. 8-A; magnification, ×2). Similarly, the regular columnar organization of the physeal chondrocytes (Fig. 8-C; magnification, ×10) becomes less organized in the kyphotic spine (Fig. 8-D; magnification, ×10).

Fig. 9

Graphical representation of the Cobb angles measured in this study. Values outside of the dashed lines indicate angles that are 2 standard deviations from the mean sagittal Cobb angle of the most supplemented diet. For the purposes of this study, we labeled values greater than the normal range as kyphotic.