Department of Spinal Surgery, Shanghai East Hospital, Tongji University, School of Medicine, 150 Jimo Road, Shanghai 200120, China; Department of Joint and Bone Disease, Shanghai East Hospital, Tongji University, School of Medicine, 150 Jimo Road, Shanghai, 200120, China.
Department of Spinal Surgery, Shanghai East Hospital, Tongji University, School of Medicine, 150 Jimo Road, Shanghai 200120, China.
Biochem Biophys Res Commun. 2019 Aug 27;516(3):1026-1032. doi: 10.1016/j.bbrc.2017.03.111. Epub 2017 Mar 23.
Cartilage endplate (CEP) cell calcification and apoptosis play a vital role in the intervertebral disc degeneration (IVDD). Oxidative stress is a key factor in inducing programmed cell death and cartilage calcification. However, the cell death and calcification of cartilage endplate cells under oxidative stress have never been described. The present study investigated the apoptosis and calcification in the cartilage endplate cell under oxidative stress induced by HO to understand the underlying mechanism of IVDD. The cartilage endplate cells isolated from human lumbar discs were subjected to different concentrations of HO for various time periods. The cell viability was determined by CCK-8 assay, whereas Western blot, immunofluorescence, and Alcian blue, Alizarin red, and Von Kossa staining evaluated the apoptosis and calcification. The level of mitochondria-specific reactive oxygen species (ROS) was quantified with an oxygen radical-sensitive probe-MitoSOX. The potential signaling pathways were investigated by Western blot after the addition of N-acetyl-l-cysteine (NAC). We found that the oxidative stress induced by HO increased the apoptosis and subsequently the calcification in the cartilage endplate cells through the ROS/p38/ERK/p65 pathway. The apoptosis and the calcification of the cartilage endplate cells induced by HO can be abolished by NAC. These results suggested that regulating the apoptosis and the calcification in the cartilage endplate cells under oxidative stress should be advantageous for the survival of cells and might delay the process of disc degeneration.
软骨终板 (CEP) 细胞钙化和凋亡在椎间盘退变 (IVDD) 中起着至关重要的作用。氧化应激是诱导细胞程序性死亡和软骨钙化的关键因素。然而，氧化应激下软骨终板细胞的死亡和钙化从未被描述过。本研究通过 HO 诱导的氧化应激研究了软骨终板细胞的凋亡和钙化，以了解 IVDD 的潜在机制。从人腰椎间盘分离的软骨终板细胞用不同浓度的 HO 处理不同的时间段。通过 CCK-8 测定法测定细胞活力，而 Western blot、免疫荧光和茜素红、茜素蓝和 Von Kossa 染色评估凋亡和钙化。用氧自由基敏感探针 MitoSOX 定量测定线粒体特异性活性氧 (ROS) 水平。用 N-乙酰-l-半胱氨酸 (NAC) 处理后通过 Western blot 研究潜在的信号通路。我们发现 HO 诱导的氧化应激通过 ROS/p38/ERK/p65 通路增加软骨终板细胞的凋亡，随后增加钙化。HO 诱导的软骨终板细胞的凋亡和钙化可以被 NAC 消除。这些结果表明，调节氧化应激下软骨终板细胞的凋亡和钙化有利于细胞的存活，并可能延缓椎间盘退变的进程。
