目的:椎间盘退变(IVDD)是下背痛(LBP)的主要原因。β-抑制蛋白1(ARRB1)是一种多功能蛋白,可调节多种病理过程。本研究旨在探讨ARRB1在IVDD中的作用。
方法:测定IVDD大鼠髓核中ARRB1的表达。接下来,用含有shArrb1(LV-shArrb1)和过表达Arrb1(LV-oeArrb1)的慢病毒感染大鼠髓核细胞(NPC)。通过测量细胞凋亡来研究Arrb1在血清剥夺型NPCs中的作用,细胞外基质降解,和自噬通量。对于体内实验,将LV-oeArrb1慢病毒注射到IVDD大鼠的NP组织中,以评估Arrb1过表达对NP的影响。
结果:在IVDD大鼠的NP组织中,ARRB1和caspase-3表达增加,LC3II/LC3I蛋白表达比例上调。Arrb1敲低加重细胞外基质降解,细胞凋亡,以及血清剥夺条件下大鼠NPCs自噬通量的损害,而Arrb1过表达显著逆转了这些效应。ARRB1与Beclin1相互作用,而Arrb1敲低抑制了Beclin1-PIK3C3核心复合物的形成。自噬抑制剂3-甲基腺嘌呤(3-MA)抵消了血清剥夺型NPC中Arrb1过表达的保护作用。此外,Arrb1过表达抑制细胞凋亡和细胞外基质降解,促进NP中的自噬,并延缓了大鼠IVDD的发展。
结论:ARRB1通过上调自噬和改善IVDD进展,防止NPCs细胞外基质降解和凋亡,提出了治疗IVDD的创新策略。
OBJECTIVE: Intervertebral disc degeneration (IVDD) is the leading cause of lower back pain (LBP). β-arrestin 1 (ARRB1) is a multifunctional protein that regulates numerous pathological processes. The aim of this study was to investigate the role of ARRB1 in IVDD.
METHODS: The expression of ARRB1 in nucleus pulposus (NP) of rats with IVDD was assayed. Next, rat nucleus pulposus cells (NPCs) were infected with lentiviruses containing shArrb1 (LV-shArrb1) and overexpressing Arrb1 (LV-oeArrb1). The roles of Arrb1 in serum-deprived NPCs were investigated by measuring apoptosis, extracellular matrix degradation, and autophagic flux. For experiments in vivo, LV-oeArrb1 lentivirus was injected into the NP tissues of IVDD rats to evaluate the effects of Arrb1 overexpression on NP.
RESULTS: In the NP tissues of IVDD rats, ARRB1 and cleaved caspase-3 expression increased, and the ratio of LC3II/LC3I protein expression was upregulated. Arrb1 knockdown aggravated extracellular matrix degradation, cellular apoptosis, and impairment of autophagic flux in rat NPCs under serum-deprived conditions, whereas Arrb1 overexpression significantly reversed these effects. ARRB1 interacted with Beclin 1, and Arrb1 knockdown suppressed the formation of the Beclin1-PIK3C3 core complex. The autophagy inhibitor 3-methyladenine (3-MA) offset the protective effects of Arrb1 overexpression in serum-deprived NPCs. Furthermore, Arrb1 overexpression inhibited apoptosis and extracellular matrix degradation, promoted autophagy in NP, and delayed the development of IVDD in rats.
CONCLUSIONS: ARRB1 prevents extracellular matrix degradation and apoptosis of NPCs by upregulating autophagy and ameliorating IVDD progression, presenting an innovative strategy for the treatment of IVDD.