双孔钾离子通道TASK-1在腰椎软骨终板退变中的作用及其表达调控研究

Lumbar cartilage endplate degeneration is closely associated with the process of the lumbar intervetebral disc degeneration, and the chondrocytes play a pivotal role in maintaining the physiological function of the cartilage endplate. It is reported that potassium channels may invlove the process of cartilage degeneration in huamn osteoarthritis via regulation of chondrocyte celluar membrane potential, but the mechanism remains largely unkonwn. With our efforts on understanding the molecular mechanisms of lumbar cartilage endplate degeneration, we identidied downregulation of of a tandem pore potassium, i.e., TASK-1 in degenerated lumbar cartilage endplate. We further successfully generated an lumbar cartilage endplate degenration model in rabbit by intradiscal injection of IL-1β. And the injection of IL-1β resulted in an decreased expression of TASK-1. IL-1β-NF-kB is reportedly an important signaling pathway inducing cartilage degeneration, and NF-kB can regulate the expression level of several potassium channels. We planned to investigate the expression levels of TASK-1 in degenerated and healthy human lumbar cartilage endplate and its role in maintaining the chondrocyte resting membrane potential by molecular biological and electrophysiological techniques. We then planned to determine the role of TASK-1 in cartilage endplate degneration via gene silencing and overexpression and to validate the protective role of overexpression of TASK-1 in animal model. Furthermore, we proposed to investigate the regulatory effect of IL-1β on the expression of TASK-1 and the possible regulatory mechanism through IL-1β-NF-kB signaling pathway. Our study may probably provide a theoretical foundation for targeted biological therapy for the treatment of lumbar cartilage endplate degeneration.

腰椎软骨终板退变与腰椎间盘退变密切相关，而终板软骨细胞是维持终板生理功能的关键。钾离子通道在软骨细胞中通过调节细胞膜电位间接影响其基质代谢。文献报道钾通道可能参与了人骨关节炎软骨退变进程，但具体机制不明。我们前期研究发现人退变腰椎软骨终板中双孔钾通道TASK-1表达降低；IL-1β作用可致兔软骨终板退变并使TASK-1表达下降。而IL-1β-NF-kB是软骨退变中的重要信号通路，NF-kB参与多种钾通道表达的调节。本研究拟采用分子生物学、电生理学等方法在大样本人腰椎软骨终板标本中明确TASK-1的表达及其在稳定软骨细胞静息膜电位中的贡献，通过基因沉默和过表达评估TASK-1在软骨终板退变中的作用，并在动物模型中验证过表达对软骨终板退变的保护作用。在此基础上，探讨IL-1β对TASK-1的表达调控作用并通过NF-kB通路分析调控的可能机制，为开发以TASK-1为靶点的生物治疗提供理论依据。

本课题发现在人正常的腰椎软骨终板细胞中存在双孔钾离子通道TASK-1的表达，退变的腰椎软骨终板细胞中TASK-1的表达显著降低。电生理研究发现双孔钾离子通道在稳定人腰椎软骨终板细胞静息膜电位中起主要作用，亦提示其可能在调节腰椎软骨终板退变中有重要意义。TASK-1表达抑制和过表达的体外试验提示TASK-1参与了腰椎软骨终板退变的调节，包括影响I型胶原、II型胶原、aggrecan等的表达。动物体内实验进一步证实了TASK-1的过表达对于腰椎软骨终板的退变具有保护作用。机制研究发现IL-1β可下调人正常腰椎软骨终板细胞中TASK-1的表达水平，进而调节细胞外基质、基质降解酶的表达和细胞增殖活性。IL-1β刺激TASK-1表达下调通过NF-kB信号通路发挥作用。. 本课题首次发现了双孔钾离子通道TASK-1在调节腰椎软骨终板退变中起重要作用。IL-1β可通过IL-1β-NF-kB信号通路调节TASK-1的表达，进而调节软骨终板细胞基质的表达。本研究从分子生物学层面阐述了腰椎软骨终板退变的机制，为开发以TASK-1离子通道为靶点的腰椎软骨终板退变的生物治疗方案提供了理论依据。