Expression Analysis of Susceptibility Genes for Ossification of the Posterior Longitudinal Ligament of the Cervical Spine in Human OPLL-related Tissues and a Spinal Hyperostotic Mouse (ttw/ttw)
Expression Analysis of Susceptibility Genes for Ossification of the Posterior Longitudinal Ligament of the Cervical Spine in Human OPLL-related Tissues and a Spinal Hyperostotic Mouse (ttw/ttw)
复制标题
DOI：
10.1097/brs.0000000000003648
复制
发表时间：
2020-11-15
期刊：
影响因子：
3
通讯作者：
Matsumine, Akihiko
中科院分区：
文献类型：
--
作者：
Nakajima, Hideaki;Watanabe, Shuji;Matsumine, Akihiko
研究设计。免疫组织化学和实时逆转录聚合酶链反应（RT-PCR）分析。目的。本研究的目的是分析全基因组关联研究中发现的人类后纵韧带骨化 (OPLL) 的五个易感基因（RSPO2、HAO1、CCDC91、RHPH9 和 STK38L）的表达。背景数据摘要。需要对五种易感基因进行详细的表达和功能研究，以帮助阐明 OPLL 的病因和发病机制。方法。对症状性 OPLL 颈前路减压期间收集的骨化韧带样本（n = 39 名患者）和对照非 OPLL 样本（n = 8 名患者）进行免疫染色、细胞培养和实时 RT-PCR。还对脊柱骨质增生小鼠 (ttw/ttw) (n = 25) 进行了免疫组织化学分析。样品切片进行 RSPO2、HAO1、CCDC91、RHPH9、STK38L、Runx2、Sox9 和 CD90 染色。还在经受循环拉伸应变的培养的人 OPLL 和非 OPLL 细胞中分析了 5 个易感基因的 mRNA 表达水平。结果。 RSPO2 和 Sox9 的免疫反应性在人 OPLL 组织和 ttw/ttw 小鼠的增殖软骨细胞中很明显。对培养的人 OPLL 细胞应用循环拉伸应变导致 RSPO2、HAO1 和 CCDC91 的 mRNA 水平增加。然而，人类 OPLL 相关样本中的表达存在个体差异。仅在 3 至 6 周龄的 ttw/ttw 小鼠中检测到 HAO1 阳性细胞，这些小鼠不同时表达 RSPO2 阳性样本。结论。在五个易感基因中，RSPO2、HAO1 和 CCDC91 可能是 OPLL 进展的影响因素。 RSPO2可能参与软骨内骨化，特别是在混合型或连续型OPLL中，HAO1可能是OPLL的启动因子，这在成熟的人类OPLL样本中很少见，而CCDC91可能与机械应力引起的骨化进展有关。这些发现为了解 OPLL 的发病机制和治疗靶点提供了重要见解。
Study Design. Immunohistochemical and real-time reverse transcription-polymerase chain reaction (RT-PCR) analysis.Objective. The aim of this study was to analyze the expression of five susceptibility genes (RSPO2, HAO1, CCDC91, RHPH9, and STK38L) for human ossification of the posterior longitudinal ligaments (OPLL) identified in a genome-wide association study.Summary of Background Data. Detailed expression and functional studies for the five susceptibility genes are needed to aid in clarification of the etiology and pathogenesis of OPLL.Methods. Immunostaining, cell culture, and real-time RT-PCR were performed on ossified ligament samples collected during anterior cervical decompression for symptomatic OPLL (n = 39 patients) and on control non-OPLL samples (n = 8 patients). Immunohistochemical analysis in spinal hyperostotic mice (ttw/ ttw) (n = 25) was also performed. The sample sections were stained for RSPO2, HAO1, CCDC91, RHPH9, STK38L, Runx2, Sox9, and CD90. The mRNA expression levels of the five susceptibility genes were also analyzed in cultured human OPLL and non-OPLL cells subjected to cyclic tensile strain.Results. Immunoreactivity for RSPO2 and Sox9 was evident in proliferating chondrocytes in human OPLL tissues and ttw/ttw mice. Application of cyclic tensile strain to cultured human OPLL cells resulted in increases in mRNA levels for RSPO2, HAO1, and CCDC91. However, individual differences in expression in human OPLL-related samples were seen. HAO1-positive cells were detected only in 3- to 6-week-old ttw/ttw mice that did not simultaneously express RSPO2-positive samples.Conclusion. Among the five susceptibility genes, RSPO2, HAO1, and CCDC91 might be contributory factors in progression of OPLL. RSPO2 may be involved in endochondral ossification, especially in mixed or continuous type OPLL, HAO1 may be an initiation factor for OPLL that is rarely seen in mature human OPLL samples, and CCDC91 may be associated with progression of ossification caused by mechanical stress. These findings provide important insights into the pathogenesis and therapeutic targets for OPLL.