Individual Collagen Fibril Thickening and Stiffening of Annulus Fibrosus in Degenerative Intervertebral Disc

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Study Design.In vitro study using rat intervertebral discs (IVDs).Objective.To explore the alteration of annulus fibrosus collagen fibrils after loading on IVD and to investigate the degeneration pathogenesis at the nanoscale.Summary of Background Data.Abnormal loading can lead to IVD degeneration, but the precise mechanism has been hitherto elusive, especially at the nanoscale.Methods.A rat IVD loading model was used, which combined bending of the tail by 40° with compressive loading of 1.8, 4.5, and 7.2 N of the rat tail using an external fixation device. The structure and the elastic modulus of individual collagen fibrils within IVD Co8-Co9 was examined 2 weeks after loading at the nanoscale using atomic force microscopy.Results.Significant fibril disorder and a decrease in cell number within the annulus fibrosus after loading was observed at the microscale as judged by hematoxylin/eosin staining, suggesting initiation of rupture of the structure and degradation of the IVD. The annulus fibrosus collagen fibrils underwent a change in diameter and elastic modulus from 170 ± 18 to 310 ± 24 nm (P < 0.001) and 0.86 ± 0.12 to 1.27 ± 0.30 GPa (P = 0.003), respectively when measured on the concave side after a loading of 7.2 N. Thus the loading process resulted in a thickening and stiffening of collagen fibrils with a difference between the inner and outer layers.Conclusion.The results of the present study indicated that abnormal loading was not only associated with disorder at the microscale, but also alteration of the collagen fibrils at the nanoscale, possibly leading to changes in the mechanical and physiological environment around the cells of the annulus fibrosus.Level of Evidence: N/A

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