Roles of SDF-1/CXCR4 axis in cartilage endplate stem cells mediated promotion of nucleus pulposus cells proliferation

Stem cells transplantation has shown considerable promise in intervertebral disc repair and low-back pain release. Cartilage endplate stem cells (CESCs) also showed potential for nucleus pulposus (NP) regeneration in a rabbit disc degeneration model, the precise mechanism was unclear. Here we investigated the effects of CESCs on NP cells (NPCs) proliferation and the mechanism in vitro. CESCs and NPCs were isolated from surgical specimens of degenerative human lumbar disc. NPCs were co-cultured with CESCs at a 1:1 ratio or cultured in CESCs conditioned medium (CESCs-CM). NPCs proliferation was evaluated by Ki-67 staining, CCK-8 assay and cell cycle analysis. Gene expressions were detected by qRT-PCR and activation of Akt and ERK1/2 was detected by western blot. CXCR4 antagonist AMD3100 was used to block SDF-1/CXCR4 axis. ERK1/2 and Akt inhibitors were used to block Akt and ERK1/2 activation. Results showed that NPCs proliferation was promoted by direct-contact co-culturing with CESCs as well as culturing in CESCs-CM. SDF-1 expression level in CESCs was significantly higher than that in NPCs, while CXCR4 was the opposite. Promotion of NPCs proliferation mediated by CESCs-CM could be partially attenuated by AMD3100. CESCs-CM activated both Akt and ERK1/2 in NPCs, while rhSDF-1 scarcely activated Akt but obviously activated ERK1/2. Akt and ERK1/2 inhibitors could partially inhibited CESCs-CM mediated promotion of NPCs proliferation and showed cumulative effect, while ERK1/2 inhibitor and AMD3100 could significantly abrogate SDF-1 mediated promotion of NPCs proliferation. Our results suggested that CESCs might promote NPCs proliferation in a paracrine pathway, which was partially mediated by SDF-1/CXCR4 axis via ERK1/2 signaling transduction pathway. (C) 2018 Published by Elsevier Inc.

干细胞移植在椎间盘修复和缓解腰痛方面显示出相当大的潜力。软骨终板干细胞（CESCs）在兔椎间盘退变模型中也显示出对髓核（NP）再生的潜力，但其确切机制尚不清楚。在此我们研究了CESCs对NP细胞（NPCs）增殖的影响及其体外机制。CESCs和NPCs从退变的人腰椎间盘手术标本中分离出来。NPCs与CESCs以1∶1的比例共培养，或在CESCs条件培养基（CESCs - CM）中培养。通过Ki - 67染色、CCK - 8检测和细胞周期分析评估NPCs的增殖。通过实时定量聚合酶链反应（qRT - PCR）检测基因表达，通过蛋白质印迹法（western blot）检测Akt和ERK1/2的激活。使用CXCR4拮抗剂AMD3100阻断SDF - 1/CXCR4轴。使用ERK1/2和Akt抑制剂阻断Akt和ERK1/2的激活。结果显示，与CESCs直接接触共培养以及在CESCs - CM中培养均可促进NPCs增殖。CESCs中SDF - 1的表达水平显著高于NPCs，而CXCR4则相反。AMD3100可部分减弱由CESCs - CM介导的NPCs增殖促进作用。CESCs - CM激活NPCs中的Akt和ERK1/2，而重组人SDF - 1（rhSDF - 1）几乎不激活Akt但明显激活ERK1/2。Akt和ERK1/2抑制剂可部分抑制由CESCs - CM介导的NPCs增殖促进作用，并显示出累积效应，而ERK1/2抑制剂和AMD3100可显著消除由SDF - 1介导的NPCs增殖促进作用。我们的结果表明，CESCs可能通过旁分泌途径促进NPCs增殖，该途径部分由SDF - 1/CXCR4轴通过ERK1/2信号转导通路介导。（C）2018年由爱思唯尔公司出版