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 nonimmunogenic upon implantation. The aim of this initial proof of technical feasibility study was to develop a decellularization protocol on bovine IVDs with endplates (EPs) and assess protocol performance before application of the protocol to human IVDs with attached EP and vertebral bone (VB). A decellularization protocol based on hypotonic low concentration sodium dodecyl sulfate (0.1% w/v) with proteinase inhibitors, freeze/thaw cycles, and nuclease and sonication treatments was applied to IVDs. Histological, biochemical, and biomechanical comparisons were made between cellular and decellularized tissue. Cell removal from bovine IVDs was demonstrated and total DNA levels of the decellularized 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% confidence level [CL]). These values were significantly lower than in cellular tissue. No significant difference in DNA levels between bovine cellular and decellularized 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 postdecellularization (2957.2 ± 340.8 N.mm⁻¹) (predecellularization: 2685.4 ± 263.1 N.mm⁻¹; n = 5, 95% CL), but the difference was within natural tissue variation. Total DNA levels for all decellularized 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 decellularized 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.

中文翻译：

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脱细胞椎间盘：退化人类椎间盘的潜在替代品

椎间盘 (IVD) 变性是背痛的主要原因。目前的手术干预有局限性。另一种方法是用天然生物支架代替退化的 IVD。从人类 IVD 中去除细胞成分应使它们在植入后无免疫原性。这项技术可行性研究的初步证明的目的是开发一个关于带有终板 (EP) 的牛 IVD 的脱细胞协议, 并在将该协议应用于具有附加 EP 和椎骨 (VB) 的人类 IVD 之前评估协议性能。基于低渗低浓度十二烷基硫酸钠 (0.1% w/v) 和蛋白酶抑制剂、冷冻/解冻循环以及核酸酶和超声处理的脱细胞方案应用于 IVD。组织学、生化、并在细胞组织和脱细胞组织之间进行了生物力学比较。证明了从牛 IVD 中去除细胞，脱细胞内纤维环 (iAF)、外纤维环 (oAF) 和 EP 的总 DNA 水平分别为 40.7 (±11.4)、25.9 (±3.8) 和 29.3 (±3.1) ng .mg分别为^-1干组织重量（n  = 6，±95% 置信水平 [CL]）。这些值明显低于细胞组织中的值。没有发现牛细胞和脱细胞髓核 (NP) 之间 DNA 水平的显着差异。糖胺聚糖 (GAG) 主要保留在 NP、iAF 和 oAF 中。循环压缩测试显示出足够的灵敏度来检测脱细胞后牛 IVD 刚度的增加 (2957.2 ± 340.8 N.mm ^-1 )（脱细胞前：2685.4 ± 263.1 N.mm ^-1；n  = 5，95 % CL），但差异在自然组织变异范围内。人类 IVD（NP、iAF、oAF、EP 和 VB）的所有脱细胞组织区域的总 DNA 水平低于 50 ng.mg ^-1干组织重量（范围：2 ng.mg ^-1，iAF 至 29 ng.mg ^-1，VB）并且组织保留了高水平的 GAG。现在需要进一步研究以评估脱细胞人类体外诊断体外和体内的生物相容性和再生潜力；然而，技术可行性的证据已经得到证明，IVD 样本中的骨骼保留将允许组织合并到受体脊柱中。