Mern Demissew S, Beierfuβ Anja, Fontana Johann, Thomé Claudius, Hegewald Aldemar A
Department of Neurosurgery, Innsbruck Medical University, Innsbruck, Tirol, Austria.
Department of Neurosurgery, University Medical Center Mannheim, Heidelberg University, Mannheim, Baden-Württemberg, Germany.
PLoS One. 2014 May 7;9(5):e96870. doi: 10.1371/journal.pone.0096870. eCollection 2014.
Degenerative disc disease (DDD) of the cervical spine is common after middle age and can cause loss of disc height with painful nerve impingement, bone and joint inflammation. Despite the clinical importance of these problems, in current publications the pathology of cervical disc degeneration has been studied merely from a morphologic view point using magnetic resonance imaging (MRI), without addressing the issue of biological treatment approaches. So far a wide range of endogenously expressed bioactive factors in degenerative cervical disc cells has not yet been investigated, despite its importance for gene therapeutic approaches. Although degenerative lumbar disc cells have been targeted by different biological treatment approaches, the quantities of disc cells and the concentrations of gene therapeutic factors used in animal models differ extremely. These indicate lack of experimentally acquired data regarding disc cell proliferation and levels of target proteins. Therefore, we analysed proliferation and endogenous expression levels of anabolic, catabolic, ant-catabolic, inflammatory cytokines and matrix proteins of degenerative cervical disc cells in three-dimensional cultures. Preoperative MRI grading of cervical discs was used, then grade III and IV nucleus pulposus (NP) tissues were isolated from 15 patients, operated due to cervical disc herniation. NP cells were cultured for four weeks with low-glucose in collagen I scaffold. Their proliferation rates were analysed using 3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide. Their protein expression levels of 28 therapeutic targets were analysed using enzyme-linked immunosorbent assay. During progressive grades of degeneration NP cell proliferation rates were similar. Significantly decreased aggrecan and collagen II expressions (P<0.0001) were accompanied by accumulations of selective catabolic and inflammatory cytokines (disintegrin and metalloproteinase with thrombospondin motifs 4 and 5, matrix metalloproteinase 3, interleukin-1β, interleukin-1 receptor) combined with low expression of anti-catabolic factor (metalloproteinase inhibitor 3) (P<0.0001). This study might contribute to inhibit inflammatory catabolism of cervical discs.
颈椎退行性椎间盘疾病（DDD）在中年后较为常见，可导致椎间盘高度降低，并伴有疼痛性神经受压、骨骼和关节炎症。尽管这些问题具有临床重要性，但在当前的出版物中，颈椎间盘退变的病理学仅从形态学角度使用磁共振成像（MRI）进行了研究，而未涉及生物治疗方法的问题。到目前为止，尽管其对基因治疗方法很重要，但退变颈椎间盘细胞中多种内源性表达的生物活性因子尚未得到研究。虽然退变腰椎间盘细胞已成为不同生物治疗方法的靶点，但动物模型中使用的椎间盘细胞数量和基因治疗因子浓度差异极大。这些表明缺乏关于椎间盘细胞增殖和靶蛋白水平的实验性数据。因此，我们分析了三维培养中退变颈椎间盘细胞的合成代谢、分解代谢、抗分解代谢、炎性细胞因子和基质蛋白的增殖及内源性表达水平。采用术前颈椎间盘MRI分级，然后从15例因颈椎间盘突出症接受手术的患者中分离出III级和IV级髓核（NP）组织。NP细胞在I型胶原支架中用低糖培养4周。使用3-（4,5-二甲基噻唑-2）-2,5-二苯基四氮唑溴盐分析其增殖率。使用酶联免疫吸附测定法分析其28种治疗靶点的蛋白表达水平。在退变的进展阶段，NP细胞增殖率相似。聚集蛋白聚糖和II型胶原表达显著降低（P<0.0001），同时伴有选择性分解代谢和炎性细胞因子（含血小板反应蛋白基序的解聚素和金属蛋白酶4和5、基质金属蛋白酶3、白细胞介素-1β、白细胞介素-1受体）的积累，以及抗分解代谢因子（金属蛋白酶抑制剂3）的低表达（P<0.0001）。本研究可能有助于抑制颈椎间盘的炎性分解代谢。
