Multiscale mechanical heterogeneity and structural gradients in the annulus fibrosus–endplate interface in the spine characterized using AFM nanomechanical testing

The intervertebral disc (IVD)–endplate–vertebra junction forms a structurally heterogeneous soft–hard tissue interface involved in spinal load transmission, yet its microscale structural–mechanical heterogeneity remains incompletely understood. Here, we employ atomic force microscopy (AFM)-based nanoindentation to quantify spatial variations in apparent elastic modulus across the annulus fibrosus (AF) and endplate in a healthy, young porcine spine model. High-resolution morphological mapping reveals a radial transition from loosely organized fibrillar networks in medial AF to densely packed lamellar bundles in lateral regions. AFM nanomechanical mapping indicates lamellar- and fibril-scale heterogeneity, with localized stiffness variations associated with fiber orientation, aggregation, and geometric effects. Feature-based analysis further suggests regional differences in apparent mechanical response between fibrillar and non-fibrillar structures within the medial AF. AFM measurements acquired in the AF, cartilaginous endplate (CEP), and bony endplate (BEP) regions indicated heterogeneous apparent elastic modulus distributions under the present measurement conditions. Together, these observations highlight spatial variations in structure and local mechanical response across the AF–endplate–vertebra interface under the present measurement conditions.