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Purpose: Vertebral compression fracture is a common complication of spinal stereotactic body radiation therapy. Development of an in vivo model is crucial to fully understand how focal radiation treatment affects vertebral integrity and biology at various dose fractionation regimens. We present a clinically relevant animal model to analyze the effects of localized, high-dose radiation on vertebral microstructure and mechanical integrity. Using this model, we test the hypothesis that fractionation of radiation dosing can reduce focused radiation therapy's harmful effects on the spine.Methods and Materials: The L5 vertebra of New Zealand white rabbits was treated with either a 24-Gy single dose of focused radiation or 3 fractionated 8-Gy doses over 3 consecutive days via the Small Animal Radiation Research Platform. Nonirradiated rabbits were used as controls. Rabbits were euthanized 6 months after irradiation, and their lumbar vertebrae were harvested for radiologic, histologic, and biomechanical testing.Results: Localized single-dose radiation led to decreased vertebral bone volume and trabecular number and a subsequent increase in trabecular spacing and thickness at L5. Hypofractionation of the radiation dose similarly led to reduced trabecular number and increased trabecular spacing and thickness, yet it preserved normalized bone volume. Single-dose irradiated vertebrae displayed lower fracture loads and stiffness compared with those receiving hypofractionated irradiation and with controls. The hypofractionated and control groups exhibited similar fracture load and stiffness. For all vertebral samples, bone volume, trabecular number, and trabecular spacing were correlated with fracture loads and Young's modulus (P < .05). Hypocellularity was observed in the bone marrow of both irradiated groups, but osteogenic features were conserved in only the hypofractionated group.Conclusions: Single-dose focal irradiation showed greater detrimental effects than hypofractionation on the microarchitectural, cellular, and biomechanical characteristics of irradiated vertebral bodies. Correlation between radiologic measurements and biomechanical properties supported the reliability of this animal model of radiation-induced vertebral compression fracture, a finding that can be applied to future studies of preventative measures. [ABSTRACT FROM AUTHOR]