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目的检测肿瘤坏死因子-α(TNF-α)在后纵韧带骨化中的表达及对体外培养后纵韧带细胞生物学性状的影响。方法切取手术后纵韧带骨化患者和非后纵韧带骨化患者非骨化部位的脊柱韧带,进行体外培养建立8个后纵韧带骨化细胞系(OL1~OL8)和7个非后纵韧带骨化细胞系(NL1~NL7)。检测体外培养条件下各细胞系的形态学特征及成骨特性。采用免疫组织化学法检测TNF-α在正常及退变后纵韧带组织中的表达;分析不同浓度的重组人TNF-α(rhTNF-α)对两类细胞系分化方向的作用。结果①只有4个OL细胞系表达成骨细胞表型,其余细胞系均表达成纤维细胞表型;②免疫组织化学染色显示,TNF-α在退变及成熟的骨化区均有表达,在正常及软骨化后纵韧带组织中无表达;③rhTNF-α对各细胞系的作用不同,它可增强各细胞系的碱性磷酸酶(ALP)活性、DNA及胶原合成,能诱导所有OL细胞系分泌骨钙素(OC),只有2个NL细胞系分泌较低水平OC。结论后纵韧带骨化患者脊柱韧带细胞体外培养可表达成骨细胞表型,TNF-α在后纵韧带骨化发生中起着重要作用。
Objective To investigate the expression of tumor necrosis factor-α (TNF-α) in the ossification of the posterior longitudinal ligament and its effect on the biological character of the ligamentum chongllus in vitro. Methods The posterior longitudinal ligament ossification and non-ossification site non-ossification spinal ligaments of the patients with ossification of the posterior longitudinal ligament were dissected. Eight posterior longitudinal ligament ossifying cell lines (OL1 ~ OL8) and seven non-posterior longitudinal ligations Ligament ossification cell line (NL1 ~ NL7). The morphological and osteogenic characteristics of each cell line were detected under in vitro culture conditions. The expression of TNF-α in normal and degenerated posterior longitudinal ligament tissues was detected by immunohistochemistry. The effect of different concentrations of recombinant human TNF-α (rhTNF-α) on the differentiation of the two cell lines was analyzed. Results ① Only 4 OL cell lines expressed osteoblast phenotype, and the other cell lines expressed fibroblast phenotype; ② Immunohistochemical staining showed that TNF-α was expressed in degenerative and mature ossification area, The expression of rhTNF-α in each cell line was different, and it could enhance alkaline phosphatase (ALP) activity, DNA and collagen synthesis in all cell lines and induce all OL cell lines Secretion of osteocalcin (OC), only two of the NL cell lines secreted lower levels of OC. CONCLUSION: Spinal ligament cells from ossification of the posterior longitudinal ligament can express osteoblast phenotype in vitro. TNF-α plays an important role in the development of ossification of the posterior longitudinal ligament.