地塞米松对体外培养胎鼠大脑皮质神经元胞浆Dynein重链和Dynactin表达的影响

Effect of dexamethasone on the expression of Dynein heavy chain and Dynactin in the cytoplasm of fetal rat cerebral cortical neurons cultured in vitro

目的研究地塞米松(dexamethasone,DEX)对体外培养胎鼠大脑皮质神经元胞浆动力蛋白Dynein重链(Dynein heavy chain,DHC)和Dynactin表达的影响。方法体外培养原代胎鼠大脑皮质神经元细胞,制作DEX干预细胞模型。依据DEX终浓度不同,分为对照组(未施加DEX)、0.1μmol/L组和1.0μmol/L组。分别于干预1 d、3 d及7 d时,应用实时荧光定量PCR研究DEX对DHC和Dynactin mRNA表达的影响;应用Western blot法研究DEX对DHC和Dynactin蛋白表达的影响。结果各时间点各组之间DHC mRNA和Dynactin mRNA比较差异均无统计学意义(P>0.05)。DEX干预后7 d,1.0μmol/L组胎鼠大脑皮质神经元DHC蛋白随时间推移表达至高峰,且明显高于对照组和0.1μmol/L组(P<0.05)。DEX干预后,对照组、0.1μmol/L组在3 d和7 d时体外培养神经元Dynactin蛋白表达均高于1 d时(P<0.05);对照组7 d时神经元Dynactin蛋白表达高于3 d时(P<0.05);0.1μmol/L组7 d时神经元Dynactin蛋白表达低于3 d时(P<0.05)。在DEX干预后3 d及7 d时,0.1μmol/L组及1.0μmol/L组胎鼠大脑皮质神经元Dynactin蛋白表达均明显低于对照组(P<0.05);且7 d时,1.0μmol/L组Dynactin蛋白表达低于0.1μmol/L组(P<0.05)。结论 DEX影响体外培养发育中胎鼠大脑皮质神经元DHC和Dynactin蛋白表达,并可能存在浓度依赖性及时间依赖性。[中国当代儿科杂志,2021,23 (6):639-644]

Objective To study the effect of dexamethasone(DEX) on the expression of Dynein heavy chain(DHC) and Dynactin in the cytoplasm of fetal rat cerebral cortical neurons cultured in vitro. Methods Primary cerebral cortical neurons of fetal rats were cultured in vitro and were used to establish a cellular model of DEX intervention.According to the final concentration of DEX, the neurons were divided into three groups: control(without DEX),0.1 μmol/L DEX, and 1.0 μmol/L DEX. On days 1, 3, and 7 after intervention, the quantitative PCR was used to observe the effect of DEX on the m RNA expression of DHC and Dynactin. The Western blot was used to observe the effect of DEX on the protein expression of DHC and Dynactin. Results There was no significant difference in the mRNA expression levels of DHC and Dynactin among the three groups at all time points(P>0.05). On day 7 after DEX intervention, the protein expression of DHC in the 1.0 μmol/L DEX group gradually increased and reached the peak over time, which was significantly higher than that in the control and 0.1 μmol/L DEX groups(P<0.05). The control and0.1 μmol/L DEX groups had a significant increase in the protein expression of Dynactin from day 1 to days 3 and 7 after DEX intervention(P<0.05). The control group had a significant increase in the protein expression of Dynactin from day3 to day 7 after intervention(P<0.05), while the 0.1 μmol/L DEX group had a significant reduction in the protein expression of Dynactin from day 3 to day 7 after intervention(P<0.05). On days 3 and 7 after DEX intervention, the0.1 μmol/L DEX and 1.0 μmol/L DEX groups had a significantly lower protein expression level of Dynactin in the cerebral cortical neurons than the control group(P<0.05). On day 7 after DEX intervention, the 1.0 μmol/L DEX group had a significantly lower protein expression level of Dynactin than the 0.1 μmol/L DEX group(P<0.05). Conclusions DEX affects the protein expression of DHC and Dynactin in the fetal rat cerebral cortical neurons cultured in vitro,possibly in a concentration-and time-dependent manner.[Chinese Journal of Contemporary Pediatrics, 2021, 23(6): 639-644]