Li Li, Al-Jallad Hadil, Sun Aiwei, Georgiopoulos Miltiadis, Bokhari Rakan, Ouellet Jean, Jarzem Peter, Cherif Hosni, Haglund Lisbet
Department of Surgery, Division of Orthopaedics McGill University Montreal Quebec Canada.
The McGill Scoliosis and Spine Group, McGill University Health Centre Montreal Quebec Canada.
JOR Spine. 2024 Nov 5;7(4):e70007. doi: 10.1002/jsp2.70007. eCollection 2024 Dec.
Extracellular vesicles (EVs) function as biomarkers and are crucial in cell communication and regulation, with therapeutic potential for intervertebral disc (IVD)-related low back pain (LBP). EV cargo is often affected by tissue health, which may affect the therapeutic potential. There is currently limited knowledge of how the cargo of IVD cell-derived EVs varies with tissue health and how differences in proteomic profile affect the predicted biological functions.
Our study purified EVs from human IVD cell conditioned media by size-exclusion chromatography. Nanoparticle tracking analysis was conducted to measure EV size and concentration. Transmission electron microscopy and Western blot were performed to examine EV structure and markers. Tandem mass tag-mass spectrometry was conducted to determine protein cargo.
Most EVs were exosomes and intermediate microvesicles with an increasing amount linked to disease progression. Of the proteins detected, 88.6% were shared across the non-degenerate, mildly-degenerate, and degenerate samples. GO and KEGG analyses revealed that cargo from the mildly-degenerate samples was the most distinct, with the proteins in high abundance strongly associated with extracellular matrix (ECM) organization and structure. Shared proteins, highly expressed in the non-degenerate and degenerate samples, showed strong associations with cell adhesion, ECM-receptor interaction, and vesicle-mediated transport, respectively.
Our findings indicate that EVs from IVD cells from tissue with different degrees of degeneration share a majority of the cargo proteins. However, the level of expression differs with degeneration grade. Cargo from the mildly-degenerate samples exhibits the most differences. A better understanding of changes in EV cargo in the degenerative process may provide novel information related to molecular mechanisms underlying IVD degeneration and suggest new potential treatment modalities for IVD-related LBP.
细胞外囊泡（EVs）作为生物标志物发挥作用，在细胞通讯和调节中至关重要，对椎间盘（IVD）相关的下腰痛（LBP）具有治疗潜力。EVs的货物通常受组织健康状况影响，这可能会影响其治疗潜力。目前对于IVD细胞衍生的EVs的货物如何随组织健康状况变化以及蛋白质组学特征的差异如何影响预测的生物学功能了解有限。
我们的研究通过尺寸排阻色谱法从人IVD细胞条件培养基中纯化EVs。进行纳米颗粒跟踪分析以测量EVs的大小和浓度。采用透射电子显微镜和蛋白质印迹法检测EVs的结构和标志物。进行串联质谱标签-质谱分析以确定蛋白质货物。
大多数EVs是外泌体和中间微囊泡，其数量随着疾病进展而增加。在检测到的蛋白质中，88.6%在非退变、轻度退变和退变样本中是共有的。基因本体（GO）和京都基因与基因组百科全书（KEGG）分析表明，轻度退变样本的货物最为独特，其中高丰度蛋白质与细胞外基质（ECM）组织和结构密切相关。在非退变和退变样本中高表达的共有蛋白质分别与细胞粘附、ECM-受体相互作用和囊泡介导的运输密切相关。
我们的研究结果表明，来自不同退变程度组织的IVD细胞的EVs共享大部分货物蛋白质。然而，表达水平随退变等级而不同。轻度退变样本的货物表现出最大差异。更好地了解退变过程中EVs货物的变化可能为IVD退变的分子机制提供新信息，并为IVD相关的LBP提出新的潜在治疗方式。
