# Single-Step Extrusion Printing of Microgrooved Annulus Fibrosus Scaffolds via Patterned Nozzles

## TL;DR

A new 3D-printing method creates microgrooved scaffolds that guide cell alignment and support potential spinal disc repair.

## Contribution

A single-step extrusion-printing method with patterned nozzles to create microgrooved scaffolds that mimic the annulus fibrosus structure.

## Key findings

- Patterned nozzles enabled fabrication of PCL scaffolds with aligned microgrooves promoting cell organization.
- Cells cultured on patterned scaffolds aligned within grooves and deposited extracellular matrix in guided arrays.
- TGF-β3 supplementation upregulated outer AF markers, and patterned scaffolds increased TAGLN expression.

## Abstract

Intervertebral disk pathology, including disk herniation and degeneration, is a major contributor to chronic low back pain, and when conservative treatment fails, surgical management often involves discectomy-based procedures that leave residual annulus fibrosus (AF) defects associated with reherniation and progressive degeneration. These limitations have motivated interest in regenerative strategies using biomaterial scaffolds; however, reproducing the hierarchical, angle-ply architecture of the AF remains challenging. Here, we present a single-step extrusion-based 3D-printing approach to fabricate polycaprolactone (PCL) scaffolds with aligned microscale surface grooves that promote AF-like organization. Patterned nozzles with circumferential peaks generated uniaxial concave microgrooves (10–17 µm wide) directly during printing, enabling formation of multilamellar angle-ply constructs. Human bone marrow-derived mesenchymal stem cells cultured on patterned scaffolds aligned longitudinally within concave grooves, forming end-to-end arrays that guided extracellular matrix deposition. Gene expression analysis showed that topographical cues governed cellular organization without significantly altering gene expression profiles, while TGF-β3 supplementation upregulated outer AF-associated markers, including COL1, COL12, SFRP2, MKX, MCAM, and SCX. TAGLN expression increased specifically on patterned scaffolds in the absence of TGF-β3, indicating an association between microgroove-guided cellular organization and TAGLN expression, warranting further investigation into potential tension-related mechanisms. This novel single-step extrusion-printing approach leverages custom nozzle geometry to impart concave microgrooves, facilitating scalable fabrication of multilamellar angle-ply scaffolds that induce aligned cellular organization and support potential applications in annulus fibrosus repair, as well as mechanobiological studies of anisotropic musculoskeletal tissues.