Purpose <p>Paraspinal muscle degeneration and dysfunction are commonly observed in patients with adult spinal deformity, though their causative role remains unclear. This study aimed to evaluate the effect of unilateral glycerol-induced muscle degeneration on the development of spinal deformity. It was hypothesized that glycerol injections would increase fibrosis (collagen content) and lead to spinal curvature towards the left side.</p> Methods <p>Male and female mice were assigned to control (no injection), saline (sham), or glycerol-injection groups. Injections were administered to the right-side lumbar multifidus and erector spinae muscles at 4 timepoints, each spaced two weeks apart. Immediately post-sacrifice, spinal curvature (Cobb angle) and muscle density were assessed using micro-CT. Muscle biopsies were taken from both the left and right sides for histological analysis and passive mechanical testing.</p> Results <p>No significant differences in spinal curvature were observed in either the coronal or sagittal planes. Relative muscle density was significantly lower, indicating reduced muscle quality on the injected (right) side in the glycerol group compared to the saline and control groups. Histological staining showed evidence of degeneration and regeneration cycles in the glycerol-injected (right) muscles, with significantly greater collagen content (fibrosis) on the right side for both the multifidus (<i>p</i> = 0.0084) and erector spinae (<i>p</i> = 0.0074). Passive stiffness was also increased in the right erector spinae compared to the left.</p> Conclusion <p>Although the hypothesized spinal curvature did not develop in this study, micro-CT, histology, and passive mechanical testing confirm that intramuscular glycerol injections effectively induce localized degeneration of lumbar paraspinal muscles.</p>