自噬相关基因Beclin1和LC3在颈椎后纵韧带骨化组织中的表达及其与成骨因子的相关性分析

Expression of autophagy-related genes Beclin1 and LC3 in ossified tissues of posterior longitudinal ligament of cervical spine and correlation analysis with osteogenic factors

中文关键词:  后纵韧带  颈椎  自噬  成骨因子  相关性

英文关键词:Posterior longitudinal ligament  Cervical spine  Autophagy  Osteogenic factor  Correlation

中文摘要:

  【摘要】 目的：探讨自噬相关基因Beclin1和LC3在颈椎后纵韧带骨化组织中的表达水平及其与成骨因子的相关性。方法：收集2020年10月～2021年5月经手术切除的颈椎后纵韧带骨化（ossification of posterior longitudinal ligament，OPLL）患者的18例后纵韧带组织标本（OPLL组）和15例非骨化的颈椎后纵韧带组织标本（非OPLL组）。苏木精-伊红（hematoxylin-eosin，HE）染色观察颈椎后纵韧带组织形态学改变；Von Kossa染色观察颈椎后纵韧带钙盐沉积情况；通过免疫组织化学染色及实时定量聚合酶链反应（RT-qPCR）技术分别测定两组标本中Beclin1、微管相关蛋白轻链（microtubule associated protein light chain 3，LC3）和Runt-相关转录因子-2 (runt-related transcription factor 2，RUNX2)，骨形态发生蛋白-2（bone morphogenetic protein-2，BMP-2）及Osterix的mRNA及蛋白的表达水平；采用Pearson相关性分析行Beclin1、LC3与RUNX2、BMP2 及Osterix的关系研究。结果：与非OPLL组相比，OPLL组细胞形态较大且形状不规则，细胞核较明显；Von Kossa染色可见非OPLL组无明显钙盐沉积，而OPLL组可见棕黑色钙盐并聚积成片状或团状；Beclin1、LC3、RUNX2、BMP2 及Osterix的蛋白及mRNA在OPLL组中的表达水平显著高于非OPLL组（P<0.05）；Beclin1 mRNA表达水平与BMP2、RUNX2和Osterix具有显著相关性（P<0.05，r＞0.5），而LC3与成骨因子不具备相关性。结论：自噬相关基因Beclin1和LC3在颈椎OPLL组织中显著高表达，Beclin1与后纵韧带成骨密切相关。

英文摘要:

  【Abstract】 Objectives: To investigate the expression levels of autophagy-related genes Beclin1 and LC3 in ossified tissues of posterior longitudinal ligament in cervical spine and their correlations with osteogenic factors. Methods: 18 posterior longitudinal ligament tissue specimens from cervical ossification of posterior longitudinal ligament(OPLL) patients underwent OPLL surgical removal from October 2020 to May 2021(OPLL group) and 15 tissue specimens of the posterior longitudinal ligament of the cervical spine that were not ossified(non-OPLL group) were collected. The morphological changes of posterior longitudinal ligament tissues were observed by hematoxylin-eosin(HE) staining; the deposition of calcium salts was observed by Von Kossa staining; The mRNA and protein expression levels of Beclin1, microtubule associated protein light chain 3(LC3), and the osteogenic factors runt-related transcription factor 2(RUNX2), bone morphogenetic protein-2(BMP2), and Osterix in the specimens of the two groups were determined with immunohistochemical staining and RT-qPCR respectively; Pearson correlation analysis was used to explore the relationship between Beclin1, LC3, RUNX2, BMP2, and Osterix. Results: Compared with the non-OPLL group, the cell morphology in the OPLL group was larger and irregular in shape, and the nuclei were more obvious; Von Kossa staining showed that there was no obvious calcium salt deposition in the non-OPLL group, whereas brownish-black calcium salts could be seen in the OPLL group, which were aggregated into flakes or clusters; The expression levels of Beclin1, LC3, RUNX2, BMP2 and Osterix proteins and mRNA in the OPLL group were higher than those in the non-OPLL group(P<0.05); Beclin1 mRNA expression level was significantly correlated with BMP2, RUNX2 and Osterix(P<0.05, r>0.5), while LC3 was not correlated with osteogenic factors. Conclusions: The autophagy-related genes Beclin1 and LC3 are significantly overexpressed in ossified tissues of posterior logitudinal ligament of cervical spine, and Beclin1 is closely related to posterior longitudinal ligament osteogenesis.