Shandong Institute of Orthopaedics and Traumatology, Medical Research Center, Affiliated Hospital of Qingdao University, Qingdao, 266003, China.
Department of Spinal Surgery, Affiliated Hospital of Qingdao University, Qingdao, 266003, China.
Osteoarthritis Cartilage. 2021 Sep;29(9):1324-1334. doi: 10.1016/j.joca.2021.06.010. Epub 2021 Jul 6.
Nucleus pulposus (NP) plays a central role in disc degeneration pathogenesis, however, as a heterogeneous tissue, cell subsets in NP and their corresponding biological process in intervertebral disc degeneration (IVDD) are unreported.
Nucleus pulposus were isolated from normal control and IVDD, and then subjected to single-cell RNA sequencing (scRNA-seq). Unsupervised clustering of the cells based on the gene expression profiles using the Seurat package and passed to tSNE for clustering visualization. Rat model of disc degeneration was built to validate the pathways identified by scRNA-Seq.
Seven chondrocyte subsets were revealed in NP based on differential gene expression, among which 4 subsets (C1-C4) were reported for the first time. Furthermore, GO and KEGG analyses discovered that ferroptosis pathways were enriched. Rat model of disc degeneration was built (n = 6/group, control vs. model) to validate the pathways identified by scRNA-Seq. Iron levels of NP were significantly higher in model group than control group (means 0.712 vs. 0.248, respectively, mg/gpro, p = 0.0026), and the levels of Heme Oxygenase 1 (HO-1) were also elevated in model group (means 14.33 vs. 5.16 IOD, respectively, p = 0.0002). However, the levels of ferritin light chain (FTL) were significantly decreased in model group compared to control group (means 26.17 vs. 9.00 FTL cell number, respectively, p = 0.0011).
Novel chondrocyte subsets in nucleus pulposus were discovered through scRNA-Seq, which provided novel insight to understand the pathological change during the development of IVDD. Ferroptosis participated in disc degeneration pathogenesis and it might serve as a new target for intervening IVDD.
髓核（NP）在椎间盘退变发病机制中起核心作用，然而，作为一种异质性组织，NP 中的细胞亚群及其在椎间盘退变（IVDD）中的相应生物学过程尚未报道。
从正常对照和 IVDD 中分离 NP，然后进行单细胞 RNA 测序（scRNA-seq）。使用 Seurat 包根据基因表达谱对细胞进行无监督聚类，然后传递给 tSNE 进行聚类可视化。构建大鼠椎间盘退变模型以验证 scRNA-Seq 鉴定的途径。
根据差异基因表达，在 NP 中揭示了 7 个软骨细胞亚群，其中 4 个亚群（C1-C4）为首次报道。此外，GO 和 KEGG 分析发现铁死亡途径富集。构建大鼠椎间盘退变模型（n=6/组，对照组与模型组）以验证 scRNA-Seq 鉴定的途径。模型组 NP 的铁含量明显高于对照组（分别为 0.712 和 0.248，mg/gpro，p=0.0026），模型组血红素加氧酶 1（HO-1）水平也升高（分别为 14.33 和 5.16 IOD，p=0.0002）。然而，与对照组相比，模型组铁蛋白轻链（FTL）水平显著降低（分别为 26.17 和 9.00 FTL 细胞数，p=0.0011）。
通过 scRNA-Seq 发现了 NP 中的新型软骨细胞亚群，为理解 IVDD 发展过程中的病理变化提供了新的见解。铁死亡参与椎间盘退变发病机制，可能成为干预 IVDD 的新靶点。
