Orthopaedic Surgery, Uijongbu St. Mary's Hospital, The Catholic University of Korea School of Medicine, Uijongbu, Korea.
Int Orthop. 2013 Dec;37(12):2507-14. doi: 10.1007/s00264-013-2037-8. Epub 2013 Aug 2.
Diabetes mellitus is associated with an increased risk of intervertebral disc degeneration (IDD). Reactive oxygen species (ROS), oxidative stressors, play a key role in autophagy of diabetes-associated diseases. Mitochondria are known to be the main source of endogenous ROS in most mammalian cell types. The authors therefore conducted the following study to evaluate the effects of high glucose concentrations on the induction of oxidative stress and autophagy through mitochondrial damage in rat notochordal cells.
Rat notochordal cells were isolated, cultured, and placed in either 10% fetal bovine serum (normal control) or 10% fetal bovine serum plus two different high glucose concentrations (0.1 M and 0.2 M) (experimental conditions) for one and three days, respectively. We identified and quantified the mitochondrial damage (mitochondrial transmembrane potential) and the generation of ROS and antioxidants (manganese superoxide dismutase [MnSOD] and catalase). We also investigated expressions and activities of autophagy markers (beclin-1, light chain3-I [LC3-I] and LC3-II, autophagy-related gene [Atg] 3, 5, 7, and 12).
An enhanced disruption of mitochondrial transmembrane potential, which indicates mitochondrial damage, was identified in rat notochordal cells treated with both high glucose concentrations. Both high glucose concentrations increased production of ROS by rat notochordal cells in a dose- and time-dependent manner. The two high glucose solutions also enhanced rat notochordal cells' compensatory expressions of MnSOD and catalase in a dose- and time-dependent manner. The proautophagic effects of high glucose concentrations were manifested in the form of enhanced rat notochordal cells' expressions of beclin-1, LC3-II, Atg3, 5, 7, and 12 in a dose- and time-dependent manner. The ratio of LC3-II/LC3-I expression was also increased in a dose- and time-dependent manner.
The findings from this study demonstrate that high glucose-induced oxidative stress promotes autophagy through mitochondrial damage of rat notochordal cells in a dose- and time-dependent manner. These results suggest that preventing the generation of oxidative stress might be a novel therapeutic target by which to prevent or to delay IDD in patients with diabetes mellitus.
糖尿病与椎间盘退变（IDD）的风险增加有关。活性氧（ROS）和氧化应激在糖尿病相关疾病的自噬中起关键作用。线粒体被认为是大多数哺乳动物细胞类型中内源性 ROS 的主要来源。因此，作者进行了以下研究，以评估高葡萄糖浓度通过大鼠脊索细胞中线粒体损伤对诱导氧化应激和自噬的影响。
分离、培养大鼠脊索细胞，分别置于 10%胎牛血清（正常对照）或 10%胎牛血清加两种不同高葡萄糖浓度（0.1 M 和 0.2 M）（实验条件）中 1 天和 3 天。我们鉴定和量化了线粒体损伤（线粒体跨膜电位）和 ROS 和抗氧化剂（锰超氧化物歧化酶[MnSOD]和过氧化氢酶）的产生。我们还研究了自噬标志物（beclin-1、轻链 3-I [LC3-I]和 LC3-II、自噬相关基因[Atg]3、5、7 和 12）的表达和活性。
用两种高葡萄糖浓度处理的大鼠脊索细胞，发现线粒体跨膜电位的破坏增强，表明线粒体损伤。两种高葡萄糖浓度均以剂量和时间依赖的方式增加了大鼠脊索细胞 ROS 的产生。两种高葡萄糖溶液还以剂量和时间依赖的方式增强了大鼠脊索细胞 MnSOD 和过氧化氢酶的代偿性表达。高葡萄糖浓度的促自噬作用表现为 beclin-1、LC3-II、Atg3、5、7 和 12 的表达在剂量和时间上均增强。LC3-II/LC3-I 表达的比值也呈剂量和时间依赖性增加。
本研究结果表明，高糖诱导的氧化应激通过大鼠脊索细胞中线粒体损伤以剂量和时间依赖的方式促进自噬。这些结果表明，预防氧化应激的产生可能是预防或延缓糖尿病患者 IDD 的新治疗靶点。
