Decellularised intervertebral discs: A potential replacement for degenerate human discs.

Intervertebral disc (IVD) degeneration is a major cause of back pain. Current surgical interventions have limitations. An alternative approach is to replace degenerated IVDs with a natural biological scaffold. The removal of cellular components from human IVDs should render them non-immunogenic upon implantation. The aim of this initial proof of technical feasibility study was to develop a decellularisation protocol on bovine IVDs with endplates (EP) and assess protocol performance before application of the protocol to human IVDs with attached EP and vertebral bone (VB). A decellularisation protocol based on hypotonic low concentration sodium dodecyl sulphate (0.1 % w/v) with proteinase inhibitors, freeze/thaw cycles, nuclease and sonication treatments was applied to IVDs. Histological, biochemical and biomechanical comparisons were made between cellular and decellularised tissue. Cell removal from bovine IVDs was demonstrated and total DNA levels of the decellularised inner annulus fibrosus (iAF), outer annulus fibrosus (oAF) and EP were 40.7 (± 11.4), 25.9 (± 3.8) and 29.3 (± 3.1) ng.mg-1 dry tissue weight respectively (n = 6, ± 95 % CL). These values were significantly lower than in cellular tissue. No significant difference in DNA levels between bovine cellular and decellularised nucleus pulposus (NP) was found. Glycosaminoglycans (GAGs), were largely retained in the NP, iAF and oAF. Cyclic compression testing showed sufficient sensitivity to detect an increase in stiffness of bovine IVD post decellularisation (2957.2 ± 340.8 N.mm-1) (pre decellularisation: 2685.4 ± 263.1 N.mm-1; n = 5, 95 % CL) but the difference was within natural tissue variation. Total DNA levels for all the decellularised tissue regions of human IVDs (NP, iAF, oAF, EP and VB) were below 50 ng.mg-1 dry tissue weight (range: 2 ng.mg-1, iAF to 29 ng.mg-1, VB) and the tissue retained high levels of GAGs. Further studies to assess the biocompatibility and regenerative potential of decellularised human IVDs in vitro and in vivo are now required, however, proof of technical feasibility has been demonstrated and the retention of bone in the IVD samples would allow incorporation of the tissue into the recipient spine.