SphK2在氧糖剥夺/复糖复氧模型中对BV-2细胞损伤的影响  

Effect of SphK2 on BV-2 cell injury in oxygen glucose deprivation/reoxygenation models

作　　者：张佳乐 李家任 张雅轩 李超然 乔婧 姚璐 牛愉捷 郝肖琼 ZHANG Jiale;LI Jiaren;ZHANG Yaxuan;LI Chaoran;QIAO Jing;YAO Lu;NIU Yujie;HAO Xiaoqiong(School of Basic Medicine and Forensic Medicine,Baotou Medical College,Baotou 014040,China;The 2020 Graduate Student,Baotou Medical College)

机构地区：[1]包头医学院基础医学与法医学院,内蒙古包头014040 [2]包头医学院

出　　处：《包头医学院学报》2024年第9期24-28,共5页Journal of Baotou Medical College

基　　金：包头医学院科研基金项目青苗计划(BYJJ-QM201905);包头医学院博士科研启动基金(BSJJ201806);包头医学院2021年度花蕾计划项目(HLBTMC202247)。

摘　　要：目的:观察BV-2小胶质细胞在氧糖剥夺/复糖复氧(OGD/R)中鞘氨醇激酶2(SphK2)的表达情况,并探讨SphK2是否参与了OGD/R致BV-2小胶质细胞的损伤过程。方法:采用小鼠BV-2小胶质细胞制备OGD/R模型,选取氧糖剥夺时间为2 h,复氧复糖时间为0 h,3 h,6 h,12 h,24 h。(1)设对照组,OGD 2 h组,OGD 2 h/R 3 h组,OGD 2 h/R 6 h组,OGD 2 h/R 12 h组,OGD 2 h/R 24 h组,观察细胞形态变化。细胞经OGD/R处理后CCK-8法检测细胞存活率。(2)设对照组,OGD 2 h/R 12 h组,OGD 2 h/R 12 h+ABC294640(SphK2特异抑制剂)组,用q-PCR及Western blot测定SphK2的基因及蛋白表达变化。结果:与对照组相比,随着OGD/R时间的延长,BV-2活力逐渐下降(P<0.001),细胞形态发生了不同程度的改变,且有不同程度的损伤;通过观察细胞生存情况确定OGD2 h/R12 h为最适宜的OGD/R时间,与对照组相比,经OGD/R处理后BV-2细胞中SphK2 mRNA表达水平显著升高(P<0.05),加入ABC294640显著抑制其表达升高(P<0.001);与对照组相比,经OGD/R处理后BV-2细胞中SphK2蛋白表达水平显著升高(P<0.001),加入ABC294640后,SphK2蛋白表达明显降低(P<0.001)。结论:OGD/R损伤后的小胶质细胞可以被SphK2激活,在其损伤过程中发挥着调控作用。Objective:To investigate the expression of sphingosine kinase 2(SphK2)in BV-2 microglia during oxygen glucose deprivation/reoxygenation(OGD/R),and to investigate whether SphK2 is involved in the damage process of BV-2 microglia induced by OGD/R.Methods:The OGD/R model was established using mouse BV-2 microglial cells.The oxygen glucose deprivation time was 2 hours,and the reoxygenation and rehydration time was 0,3,6,12,and 24 hours.(1)The control group,OGD2h group,OGD2h/R3h group,OGD2h/R6h group,OGD2h/R12h group and OGD2h/R24h group were set up to observe the changes of cell morphology.Cell viability was detected by CCK-8 method after OGD/R treatment;(2)The control group,OGD2h/R12h group and OGD2h/R12h+ABC294640(SphK2 specific inhibitor)group were set up,and the expression of SphK2 gene and protein was determined by q-PCR and Western blot.Results:Compared with the control group,with the prolongation of OGD/R time,the activity of BV-2 gradually decreased(P<0.001),and the cell morphology changed to varying degrees,with varying degrees of injury By observing cell survival,OGD2h/R12h was determined as the most suitable OGD/R time.Compared with the control group,the expression level of Sphk2 mRNA in BV-2 cells treated with OGD/R was significantly increased(P<0.05),and the addition of ABC294640 significantly inhibited the increase in its expression(P<0.001).Compared with the control group,the expression level of SphK2 protein in BV-2 cells treated with OGD/R was significantly increased(P<0.001),while the expression of SphK2 protein was significantly decreased after adding ABC294640(P<0.001).Conclusion:SphK2 activates microglia after OGD/R injury and plays a regulatory role in the process of OGD/R injury.

关 键 词：鞘氨醇激酶2 氧糖剥夺/复糖复氧 小胶质细胞 缺血性脑卒中 

分 类 号：R743.3[医药卫生—神经病学与精神病学]