Niu Tong, Jin Liuzhong, Niu Shizhen, Gong Cunqi, Wang Hui
Department of Spine Surgery, Jining No. 1 People's Hospital, Jining, China.
School of Clinical Medicine, Jining Medical University, Jining, China.
Cell Physiol Biochem. 2018;50(2):460-472. doi: 10.1159/000494159. Epub 2018 Oct 11.
BACKGROUND/AIMS: Currently, scientists attempt to improve outcome of spinal cord injury (SCI) via reducing secondary injury during SCI. Oxidative stress is critical for pathophysiology of secondary damage, thus we mainly focused on the anti-oxidant effects of Lycium barbarum polysaccharides (LBPs) on PC-12 and SH-SY5Y cells as well as the underlying mechanisms.
Oxidative stress was induced by H2O2 stimulation. Effects of LBPs on cell viability, apoptosis, and expression of proteins associated with apoptosis and autophagy in H2O2-induced cells were assessed by CCK-8 assay, flow cytometry assay and Western blot analysis, respectively. Then, expression of miR-194 was determined by qRT-PCR. Expression of miR-194 was dysregulated, and whether LBPs affected H2O2-treated cells through modulating miR-194 was verified. The expression of key kinases in the PI3K/AKT pathway and the intracellular levels of ROS and NO were testified by Western blot analysis and flow cytometry with fluorescent probes.
H2O2-induced decrease of cell viability and increases of apoptosis and autophagy in PC-12 cells were mitigated by LBPs treatment. Next, we found that miR-194 expression was both down-regulated by LBPs treatment in PC-12 and SH-SY5Y cells. More experiments consolidated that influence of LBPs on H2O2-treated cells was reversed by miR-194 overexpression while was augmented by miR-194 inhibition. LBPs elevated the phosphorylated levels of PI3K and AKT and reduced levels of ROS and NO through miR-194.
LBPs alleviated H2O2-induced decrease of cell viability, and increase of apoptosis and autophagy through down-regulating miR-194. Moreover, LBPs activated the PI3K/AKT pathway and reduced oxidative stress through miR-194.
背景/目的：目前，科学家们试图通过减少脊髓损伤（SCI）期间的继发性损伤来改善其预后。氧化应激对继发性损伤的病理生理学至关重要，因此我们主要关注枸杞多糖（LBPs）对PC-12和SH-SY5Y细胞的抗氧化作用及其潜在机制。
通过H2O2刺激诱导氧化应激。分别采用CCK-8法、流式细胞术和蛋白质印迹分析评估LBPs对H2O2诱导细胞的细胞活力、凋亡以及与凋亡和自噬相关蛋白质表达的影响。然后，通过qRT-PCR测定miR-194 的表达。验证了miR-194表达失调，以及LBPs是否通过调节miR-194影响H2O2处理的细胞。通过蛋白质印迹分析和荧光探针流式细胞术验证PI3K/AKT通路中关键激酶的表达以及细胞内ROS和NO的水平。
LBPs处理减轻了H2O2诱导的PC-12细胞活力下降以及凋亡和自噬增加。接下来，我们发现LBPs处理在PC-12和SH-SY5Y细胞中均下调了miR-194的表达。更多实验证实，miR-194过表达可逆转LBPs对H2O2处理细胞的影响，而miR-194抑制则增强了这种影响。LBPs通过miR-194提高了PI3K和AKT的磷酸化水平，并降低了ROS和NO的水平。
LBPs通过下调miR-194减轻了H2O2诱导的细胞活力下降以及凋亡和自噬增加。此外，LBPs通过miR-194激活PI3K/AKT通路并降低氧化应激。
