电离辐射诱导椎间盘纤维环细胞衰老的研究△

Senescence of annulus fibrosus cells induced by ionizing radiation

摘要:

[目的] 探讨电离辐射对大鼠纤维环细胞衰老及衰老相关分泌表型表达的影响。[方法] 选取6 只成熟SD 大鼠，取其腰尾段椎间盘纤维环细胞做体外细胞培养，将同等条件下培养的纤维环细胞随机分为两组，分别接受剂量为0 Gy、15 Gy 的X 线电离辐射，检测两组纤维环细胞的衰老情况以及细胞衰老相关分泌表型（senescence-associated secretory phenotype, SASP）的表达。[结果] 经15 Gy 电离辐射处理后，纤维环细胞在形态学上表现出典型的衰老特征，SA-β-Gal 测定显示纤维环细胞阳性染色百分比为77%。CCK-8 实验显示纤维环细胞9 d 内，未辐射组的OD 值逐渐增加（P＜0.05），而辐射组OD 值比较的差异无统计学意义（P＞0.05）。IF 及PCR 检测均显示，辐射组衰老相关基因，包括P16 [(5.2±0.3) vs (1.0±0.1), P＜0.001]、P21 [(12.9±0.5) vs (1.0±0.1), P＜0.001] 的mRNA 相对表达量均显著高于未辐射组。此外，辐射组SASP 的表达，包括胶原蛋白Col-1 [(1.6±0.1) vs (1.0±0.1), P=0.004]、Col-2 [(7.0±0.4) vs (1.0±0.1), P＜0.001]、白细胞介素（interleukin, IL） -6 [(7.9±0.2) vs (1.0±0.1), P＜0.001]、IL-8 [(1.8±0.1) vs (1.0±0.1), P＜0.001]、基质金属蛋白酶（matrix metalloproteinase, MMP） -1 [(39.2±0.6) vs (1.0±0.1), P＜0.001]、MMP-3 [(4.5±0.3) vs (1.0±0.1), P＜0.001] 的相对表达量均显著高于未辐射组。[结论] 电离辐射能诱导大鼠椎间盘纤维环细胞衰老，其衰老相关分泌表型的表达明显增加。

Abstract:

[Objective] To investigate the effects of ionizing radiation on aging and aging-related secretion phenotype expression of ratannulus fibrosus cells. [Methods] Six mature SD rats were selected to harvest lumbar intervertebral disc annulus fibrosus cells. The annu-lus fibrosus cells cultured under the same conditions were randomly divided into two groups, which received ionizing X-ray radiation indose of 0 Gy and 15 Gy respectively. The senescence-associated secretory phenotype (SASP) was assayed to determine the senescence ofannulus fibrosus cells. [Results] After being treated with 15 Gy ionizing radiation, the annulus fibrosus cells showed typical characteristicsof aging morphology, and the positive staining percentage of annulus fibrosus cells was 77% by SA-β-Gal assay. The CCK-8 test showedthat the OD value in the non-irradiated group increased gradually (P＜0.05), but there was no statistical significance in the comparison of OD values in the irradiated group (P＞0.05). Both IF and PCR tests showed that aging related genes in the radiation group, including P16[(5.2±0.3) vs (1.0±0.1), P＜0.001], P21 [(12.9±0.5) vs (1.0±0.1), P＜0.001] were significantly higher than those in the non-radiation group,whereas the expression of SASP in the radiation group including Col-1 [(1.6±0.1) vs (1.0±0.1), P=0.004], Col-2 [(7.0±0.4) vs (1.0±0.1), P＜0.001], IL-6 [(7.9±0.2) vs (1.0±0.1), P＜0.001], IL-8 [(1.8±0.1) vs (1.0±0.1), P＜0.001], matrix metalloproteinase (MMP)-1 [(39.2±0.6) vs(1.0±0.1), P＜0.001] and MMP-3 [(4.5±0.3) vs (1.0±0.1), P＜0.001] were significantly higher than those in the non-radiation group. [Conclu-sion] Ionizing radiation can induce senescence of rat intervertebral disc annulus cells, and the expression of senescence related secretionphenotype is significantly increased.