RANK/RANKL/OPG system in the intervertebral disc
RANK/RANKL/OPG system in the intervertebral disc
复制标题
椎间盘内的 RANK/RANKL/OPG 系统
作者：
Norihiko Takegami;K. Akeda;J. Yamada;Tomohiko Sano;K. Murata;Jenny Huang;K. Masuda;A. Sudo
研究背景NF-κB配体受体激活因子（receptor activator of NF-κB ligand，RANKL）是TNF配体超家族的成员之一，参与骨代谢的调节。RANK/RANKL/骨保护素（OPG）系统各组分在椎间盘（IVD）中的表达尚未详细研究。本研究的目的是研究RANK/RANKL/OPG系统的表达，并评估RANKL在大鼠IVD.MethodsSprague-Dawley，12周龄，雄性大鼠的基质代谢的功能在这项研究中使用。从胸腰椎椎间盘分离纤维环（AF）、髓核（NP）和软骨终板（CEP）细胞进行单层培养。使用实时聚合酶链反应（PCR）和免疫组织化学分析（培养细胞和IVD组织）检测大鼠IVD中RANK/RANKL/OPG的表达。为了检测白细胞介素-1 β（IL-1β）刺激对RANK、RANKL和OPG mRNA水平的影响，将细胞与重组人IL-1β（rhIL-1β）一起培养。为了评价RANKL对分解代谢因子（IL-1β、基质金属蛋白酶-3（MMP-3）和MMP-13）mRNA表达的影响，在存在或不存在rhIL-1β的情况下与RANKL一起培养细胞。RANK、RANKL和OPG的mRNA表达水平在AF、NP和CEP细胞中均有明确的表达。对RANK、RANKL和OPG的免疫反应性分布于三种类型细胞的细胞膜和/或细胞质中。经rhIL-1β处理后，3种细胞RANKL mRNA水平均显著上调。在无rhIL-1β的情况下，RANKL处理对AF、NP和CEP细胞的分解代谢因子的mRNA表达无显著影响。然而，在存在rhIL-1β的情况下，RANKL刺激显著上调表达。有更多的RANK/RANKL/OPG阳性细胞在人类椎间盘组织中的退变的晚期阶段相比，在早期stage.ConclusionsOur研究表明，RANK/RANKL/OPG系统的可能性，可能发挥了一部分在椎间盘退变的过程中。
BackgroundThe receptor activator of NF-κB ligand (RANKL), a member of the TNF ligand superfamily, is known to regulate bone metabolism. The expression of each component of the RANK/RANKL/osteoprotegerin (OPG) system in the intervertebral disc (IVD) has not been examined in detail. The purposes of this study were to examine the expression of the RANK/RANKL/OPG system and to evaluate the function of RANKL in the matrix metabolism of the rat IVD.MethodsSprague-Dawley, 12-week-old, male rats were used in this study. Anulus fibrosus (AF), nucleus pulposus (NP) and cartilaginous endplate (CEP) cells isolated from dissected thoracolumbar discs were monolayer-cultured. RANK/RANKL/OPG expression in rat IVDs was examined using real-time polymerase chain reaction (PCR) and immunohistochemical analysis (cultured cells and IVD tissues). To examine the effect of interleukin-1β (IL-1β) stimulation on the mRNA levels of RANK, RANKL and OPG, the cells were cultured with or without recombinant human IL-1β (rhIL-1β). To evaluate the effect of RANKL on the mRNA expression of catabolic factors (IL-1β, matrix metalloproteinase-3 (MMP-3) and MMP-13), the cells were cultured with RANKL in the presence or absence of rhIL-1β. The immunohistochemical expression of this system was also evaluated using human IVD tissues with different grades of degeneration.ResultsmRNA expression levels of RANK, RANKL, and OPG were clearly identified in AF, NP and CEP cells. Immunoreactivity to RANK, RANKL and OPG was distributed in the cell membranes and/or cytoplasm of the three types of cells. The mRNA level of RANKL was significantly upregulated by treatment with rhIL-1β of the three types of cells. Treatment with RANKL without rhIL-1β did not induce significant effects on the mRNA expression of catabolic factors by AF, NP and CEP cells. However, the expression was significantly upregulated by stimulation with RANKL in the presence of rhIL-1β. There was a general trend for more RANK/RANKL/OPG-positive cells in human IVD tissues in an advanced stage of degeneration compared to an early stage.ConclusionsOur study showed the possibility that the RANK/RANKL/OPG system may play a part in the process of intervertebral disc degeneration.