Low back pain (LBP) is often associated with the degeneration of human intervertebral discs (IVDs). However, the pain-inducing mechanism in degenerating discs remains to be elucidated. Here, we identified a subtype of locally residing human nucleus pulposus cells (NPCs), generated by certain conditions in degenerating discs, that was associated with the onset of discogenic back pain. Single-cell transcriptomic analysis of human tissues showed a strong correlation between a specific cell subtype and the pain condition associated with the human degenerated disc, suggesting that they are pain-triggering. The application of IVD degeneration-associated exogenous stimuli to healthy NPCs in vitro recreated a pain-associated phenotype. These stimulated NPCs activated functional human iPSC-derived sensory neuron responses in an in vitro organ-chip model. Injection of stimulated NPCs into the healthy rat IVD induced local inflammatory responses and increased cold sensitivity and mechanical hypersensitivity. Our findings reveal a previously uncharacterized pain-inducing mechanism mediated by NPCs in degenerating IVDs. These findings could aid in the development of NPC-targeted therapeutic strategies for the clinically unmet need to attenuate discogenic LBP.

Background: In vitro studies using nucleus pulposus (NP) cells are commonly used to investigate disc cell biology and pathogenesis, or to aid in the development of new therapies. However, lab‐to‐lab variability jeopardizes the much‐needed progress in the field. Here, an international group of spine scientists collaborated to standardize extraction and expansion techniques for NP cells to reduce variability, improve comparability between labs and improve utilization of funding and resources.Methods: The most commonly applied methods for NP cell extraction, expansion, and re‐differentiation were identified using a questionnaire to research groups worldwide. NP cell extraction methods from rat, rabbit, pig, dog, cow, and human NP tissue were experimentally assessed. Expansion and re‐differentiation media and techniques were also investigated.Results: Recommended protocols are provided for extraction, expansion, and re‐differentiation of NP cells from common species utilized for NP cell culture.Conclusions: This international, multilab and multispecies study identified cell extraction methods for greater cell yield and fewer gene expression changes by applying species‐specific pronase usage, 60–100 U/ml collagenase for shorter durations. Recommendations for NP cell expansion, passage number, and many factors driving successful cell culture in different species are also addressed to support harmonization, rigor, and cross‐lab comparisons on NP cells worldwide.

Introduction: Intervertebral disc (IVD) degeneration is often associated with low back pain and radiating leg pain. The purpose of this study is to develop a reproducible and standardized preclinical model of painful lumbar IVD degeneration by evaluation of structural and behavioral changes in response to IVD injury with increasing needle sizes. This model can be used to develop new therapies for IVD degeneration.Methods: Forty‐five female Sprague Dawley rats underwent anterior lumbar disc needle puncture at levels L4‐5 and L5‐6 under fluoroscopic guidance. Animals were randomly assigned to four different experimental groups: needle sizes of 18 Gauge (G), 21G, 23G, and sham control. To monitor the progression of IVD degeneration and pain, the following methods were employed: μMRI, qRT‐PCR, histology, and biobehavioral analysis.Results: T1‐ and T2‐weighted μMRI analysis showed a correlation between the degree of IVD degeneration and needle diameter, with the most severe degeneration in the 18G group. mRNA expression of markers for IVD degeneration markers were dysregulated in the 18G and 21G groups, while pro‐nociceptive markers were increased in the 18G group only. Hematoxylin and Eosin (H&E) and Alcian Blue/Picrosirius Red staining confirmed the most pronounced IVD degeneration in the 18G group. Randall‐Selitto and von Frey tests showed increased hindpaw sensitivity in the 18G group.Conclusion: Our findings demonstrate that anterior disc injury with an 18G needle creates severe IVD degeneration and mechanical hypersensitivity, while the 21G needle results in moderate degeneration with no increased pain sensitivity. Therefore, needle sizes should be selected depending on the desired phenotype for the pre‐clinical model.

Chronic lower back pain (LBP) is the leading cause of disability worldwide. Due to its close relationship with intervertebral disc (IVD) degeneration (IVDD), research has historically focused more on understanding the mechanism behind IVDD while clinical efforts prioritize pain management. More recently, there has been a shift toward understanding LBP as a distinct pathological entity. This review synthesizes current knowledge on discogenic LBP, combining known pathophysiology, molecular mechanisms, risk factors, diagnostic challenges, and available experimental models. IVDD is a complex, multifactorial process involving biochemical, mechanical, and inflammatory changes within the disc, leading to structural breakdown and potential discogenic pain. Key mechanisms include extracellular matrix degradation, upregulation of inflammatory mediators, immune cell infiltration, and aberrant nerve and vascular ingrowth. However, not all cases of IVDD result in LBP, highlighting the need for further investigation into the cellular, molecular, and biomechanical factors contributing to symptom development. Current diagnostic tools and experimental models for studying discogenic LBP remain limited, impeding the development of targeted treatments. Existing therapies primarily focus on symptom management rather than addressing underlying disease mechanisms.