上调长链非编码RNA SNHG12表达减轻过氧化氢诱导的神经细胞活性下降及细胞凋亡和过度自噬

Up-regulation of long non-coding RNA SNHG12 expression attenuates hydrogen peroxidation-induced decline in neuronal cell activity,apoptosis and excessive autophagy

背景:脊髓损伤后的继发性损伤被认为是其致残率高的主要因素。抑制氧化应激和细胞凋亡的病理级联反应,延缓疾病进程,是减轻脊髓损伤的一大治疗方向。长链非编码RNA SNHG12被认为与细胞的氧化还原稳态具有相关性,其能否调控过氧化氢过载引起的神经细胞氧化损伤目前仍未知。目的:探究长链非编码RNA SNHG12在过氧化氢诱导神经细胞HT22氧化损伤中的作用及机制。方法:(1)100μmol/L过氧化氢干预HT22细胞6 h,建立细胞氧化损伤模型。(2)通过转染提升HT22细胞中SNHG12表达水平,采用CCK-8、锥虫蓝染色、TUNEL染色和蛋白质免疫印迹实验评估SNHG12表达水平对HT22细胞活力、凋亡和自噬的影响。(3)采用荧光素酶实验验证SNHG12/miR-320a/SIRT5的靶向关系。(4)通过转染分别促进HT22细胞中miR-320a和SIRT5上调,采用逆转实验探讨miR-320a和SIRT5在SNHG12发挥保护作用中的调控作用。结果与结论:上调SHNG12表达可以减轻过氧化氢诱导的细胞活性下降、细胞凋亡和过度自噬。荧光素酶实验证实SHNG12/miR-320a/SIRT5三者之间存在结合关系。miR-320a过表达可以逆转上调SNHG12对以上各指标的影响,而SIRT5过表达可以逆转上调miR-320a对以上各指标的影响。结果表明:SNHG12通过调控miR-320a/SIRT5轴抑制HT22细胞自噬和凋亡,减缓细胞氧化损伤。

BACKGROUND:After spinal cord injury,secondary injury is considered to be a major factor in its high morbidity rate.Inhibiting the pathological cascade of oxidative stress and apoptosis and delaying the disease process are the primary therapeutic direction for alleviating spinal cord injury.The long non-coding RNA SNHG12 is thought to be related to cellular redox homeostasis,and its function on regulating the oxidative damage in neurons caused by H2O2 overload remains unknown.OBJECTIVE:To investigate the function and mechanism of long non-coding RNA SNHG12 in H2O2-induced oxidative damage of HT22 cells.METHODS:(1)The HT22 cells were exposed to 100μmol/L H2O2 for 6 hours to establish the cellular oxidative damage model.(2)The expression level of SNHG12 in HT22 cells was increased by transfection.The influences of SNHG12 expression on HT22 cell viability,apoptosis level,and autophagy were evaluated using CCK8 assay,Tryphenol blue staining,TUNEL staining,and western blot analysis.(3)Luciferase assay was used to verify the targeting relationship between SNHG12/miR-320a/SIRT5.(4)The upregulation of miR-320a and SIRT5 in HT22 cells was induced by transfection,respectively,and the regulation of miR-320a and SIRT5 in the protective role of SNHG12 was explored by reverse experiments.RESULTS AND CONCLUSION:SHNG12 overexpression could alleviate the decline in cell viability,apoptosis,and excessive autophagy caused by H2O2.Luciferase reporter assay confirmed the binding relationship among SHNG12/miR-320a/SIRT5.miR-320a overexpression could reverse the effects of elevated SNHG12 on the above aspects,whereas SIRT5 overexpression can reverse the effect of up-regulation of miR-320a on the above indicators.It is concluded that SNHG12 suppressed autophagy and apoptosis of HT22 cells and alleviated oxidative damage via regulating miR-320a/SIRT5 axis.