Objective: To study the protective effect and effect of SphK1 overexpression on the injury of nerve cells induced by acrylamide. Methods: ACR with 99% purity was prepared into 1.25 mmol/L and 2.5 mmol/L solutions. SH-SY5Y cells were divided into control group (NC group) , experimental group and SphK1 activator group. The experimental group was given ACR solution with final concentration of 1.25 mmol/L and 2.5 mmol/L respectively for 24 h. In the SphK1 activator group, on the basis of the exposure concentration of the experimental group, the SphK1 specific activator (12-) phorbol tetradecanoate (-13-) acetate (PMA) solution[prepared by dimethyl sulfoxide (DMSO) , the final concentration was 100 nmol/l], and other treatments were the same as the experimental group. Control group (NC group) added PMA solution into normal cells. Western blot was used to detect the expression of SphK1 protein; CCK-8 was used to detect the proliferation of SH-SY5Y cells; hoechst33342 method was used to observe the morphological changes of nerve cells; flow cytometry was used to analyze the apoptosis of cells. Results: Compared with NC group, the expression of SphK1 protein in the experimental group and the SphK1 activator group was significantly lower (P<0.05) . Compared with the experimental group, the expression of SphK1 protein in each concentration of SphK1 activator group was increased, and the difference was statistically significant (P<0.05) . In addition to 1.25 mmol/L SphK1 activator group, compared with NC group, the relative growth survival rate of experimental group and 2.5 mmol/L SphK1 activator group were lower, the difference was statistically significant (P<0.05) . Compared with the experimental group, the relative survival rate of cells in the SphK1 activator group was higher, and the difference was statistically significant (P<0.05) . With the increase of exposure concentration, the cells in the experimental group showed the morphological characteristics of early apoptosis at ACR 1.25 mmol/L and late apoptosis at ACR 2.5 mmol/L. Compared with NC group, the apoptosis rate of experimental group and SphK1 activator group at ACR 2.5 mmol/L was significantly different (P<0.05) ; compared with experimental group, the apoptosis rate of SphK1 activator group at ACR 2.5 mmol/L was lower, the difference was statistically significant (P<0.05) . Conclusion: The SphK1 excessive expression plays the protective function to the nerve cell damage caused by acrylamide.
目的： 研究鞘氨醇激酶1（SphK1）过表达在丙烯酰胺（ACR）致神经细胞损伤中的保护作用以及影响。 方法： 将纯度为99%的ACR用生长液制备成浓度为1.25、2.5 mmol/L溶液；将人神经母细胞瘤（SH-SY5Y）细胞分为对照组（NC组）、实验组和SphK1激活剂组。NC组加入（12-）十四酸佛波酯（-13-）乙酸盐（PMA）溶液[SphK1特异性激活剂，二甲基亚砜（DMSO）配制，终浓度为100 nmol/L]。实验组给予终浓度分别为1.25和2.5 mmol/L的ACR溶液，染毒24 h。SphK1激活剂组在实验组染毒浓度的基础上，每个染毒浓度分别加入PMA溶液，其他处理与实验组一致。各组采用免疫印迹（Western blot）法检测SphK1蛋白的表达含量；CCK-8检测SH-SY5Y细胞的增殖活性；Hoechst33342法观察神经细胞形态学改变；流式细胞术分析细胞的凋亡。 结果： 与NC组比较，实验组和SphK1激活剂组细胞SphK1蛋白表达均降低，差异有统计学意义（P<0.05）。与实验组比较，SphK1激活剂组细胞在1.25和2.5 mmol/L浓度的SphK1蛋白表达均增高，差异均有统计学意义（P<0.05）。与NC组比较，实验组和2.5 mmol/L浓度SphK1激活剂组的细胞相对生长存活率均更低，差异有统计学意义（P<0.05）。与实验组比较，SphK1激活剂组细胞相对生长存活率均更高，差异有统计学意义（P<0.05）。随着染毒剂量的增加，实验组细胞在ACR 1.25 mmol/L浓度时呈现出早期凋亡、在ACR 2.5 mmol/L浓度时呈现出晚期凋亡的形态学特征。与NC组比较，实验组和SphK1激活剂组在ACR 2.5 mmol/L浓度时凋亡率差异均有统计学意义（P<0.05）；与实验组比较，SphK1激活剂组在ACR 2.5 mmol/L浓度时细胞凋亡率更低，差异有统计学意义（P<0.05）。 结论： SphK1过表达可以对丙烯酰胺引起的神经细胞损伤起到保护作用。.
Keywords: Acrylamide; Nerve damage; Protection; Ssphingosine kinase 1.