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目的探讨丙烯酰胺(acrylamide,ACR)染毒对突触素Ⅰ(SynapsinⅠ)上游丝裂原活化蛋白激酶(MAPK)、环磷酸腺苷(PKA)和Ⅱ型钙离子/钙调素依赖蛋白激酶(Ⅱ型Ca~(2+)/CaM激酶)表达的影响。方法MTT法检测不同剂量(0、25、50、75、100、125、150、200μg/ml),不同时间(24、48h)ACR染毒对成熟的NB-1细胞的毒作用;western-blot法检测突触素Ⅰ(SynapsinⅠ)的蛋白表达、SynapsinⅠ上游MAPK、PKA和Ⅱ型Ca~(2+)/CAM蛋白激酶的表达。结果 ACR对诱导成熟的NB-1具有明显的细胞毒性,细胞相对存活率随染毒剂量增加而逐渐降低,50μg/ml和100μg/ml ACR组SynapsinⅠ蛋白表达较对照组显著降低。100μg/ml ACR组MAPK蛋白表达显著降低,对照组、25和50μg/ml ACR染毒组与100μg/ml ACR组之间比较差异有统计学意义(P<0.05)。25μg/ml ACR组PKA蛋白表达显著降低,对照组、50和100μg/ml ACR染毒组与25μg/ml ACR组间差异有统计学意义(P<0.05)。25、50和100μg/ml组Ⅱ型Ca~(2+)/CaM激酶蛋白表达较对照显著增加(P<0.05),100μg/ml分别与25μg/ml与50μg/ml剂量组之间差异有统计学意义(P<0.05)。结论 ACR对诱导成熟的NB-1细胞具有明显的细胞毒性,ACR染毒可致SynapsinⅠ蛋白表达降低,且同时影响SynapsinⅠ上游的磷酸蛋白激酶MAPK、PKA和Ⅱ型Ca~(2+)/CaM的蛋白表达,可能是造成突触损伤的机制之一。
Objective To investigate the effect of acrylamide (ACR) on MAPK, PKA and Ⅱ (Ca2 + / calmodulin-dependent protein kinase) in synapsin Ⅰ Ⅱ type Ca ~ (2 +) / CaM kinase expression. Methods MTT assay was used to detect the cytotoxicity of ACR at different doses (0,25,50,75,100,125,150,200μg / ml) and different time (24,48 hours) on mature NB-1 cells. Western-blot The protein expressions of Synapsin Ⅰ, MAPK, PKA and type Ⅱ Ca ~ (2 +) / CAM protein kinases in the upstream of Synapsin Ⅰ were detected. Results ACR had obvious cytotoxicity on mature NB-1 cells. The relative survival rate of ACR decreased gradually with the increase of exposure dose. The protein expression of Synapsin I in 50μg / ml and 100μg / ml ACR group was significantly lower than that in control group. The MAPK protein expression in 100μg / ml ACR group was significantly lower than that in ACR group and 100μg / ml ACR group (P <0.05). The protein expression of PKA in 25μg / ml ACR group was significantly lower than that in ACR group (50μg / ml and 100μg / ml) and 25μg / ml ACR group (p <0.05). The expression of Ca ~ (2 +) / CaM kinase Ⅱ in groupⅡwas significantly increased compared with control (P <0.05) at 25, 50 and 100μg / ml, and there was statistical difference between 100μg / ml and 25μg / ml and 50μg / ml Significance (P <0.05). Conclusion ACR has obvious cytotoxicity on mature NB-1 cells. ACR could decrease the expression of Synapsin Ⅰ protein and affect MAPK, PKA and Ⅱ Ca 2+ / CaM of synapse Ⅰ Protein expression may be one of the mechanisms that cause synaptic damage.