PGC-1α在海人酸(KA)诱导培养大鼠海马神经元氧化应激损伤中的作用

The role of PGC-1α on kainic acid (KA) induced oxidative stress and neuronal injury in cultured rat hippocampal neuron

目的探讨过氧化物酶体增殖物激活受体γ辅激活子(peroxisome proliferator-activated receptor-γcoactivator,PGC)-1α在海人酸(kainic acid,KA)诱导的培养大鼠海马神经元氧化应激及神经损伤中的作用。方法离体培养大鼠海马神经元,采用电穿孔基因转染技术分别转染PGC-1αshRNA质粒和空载体质粒。培养7天后,用KA刺激上述海马神经元24h,检测转染空载体质粒海马神经元(空白对照组,n=10)、KA刺激的转染PGC-1αshRNA质粒(实验组,n=11)和KA刺激的空载体质粒海马神经元(KA对照组,n=10)的谷胱甘肽(glutathione,GSH)含量、超氧化物歧化酶(superoxidase dismutase,SOD)和乳酸脱氢酶(lactic dehydrogenase,LDH)活性变化。FJB染色检测KA刺激24h后实验组和KA对照组培养大鼠海马神经元神经损伤情况。结果KA刺激24h,培养的大鼠海马神经元GSH含量和SOD活性分别为(5.74±0.54)μmol.L-1.μg-1 protein和(10.57±1.25)U/mg protein,较空白对照组显著降低[(9.38±0.72)μmol.L-1.μg-1 protein,P<0.01;(23.12±3.23)U/mg protein,P<0.01];LDH活性为(2.21±0.18)mmol NADH,H+/min/100mg protein,较空白对照组海马神经元显著增高[(1.29±0.09)mmol NADH,H+/min/100mg protein,P<0.01]。转染PGC-1αshRNA质粒组,KA刺激24h海马神经元GSH含量和SOD活性分别为(3.37±0.42)μmol.L-1.μg-1protein和(4.54±0.91)U/mg protein,较KA对照组海马神经元明显降低(P均<0.05);LDH活性为(2.74±0.19)mol/NADH+/min/mg protein,较KA对照组海马神经元显著增加(P<0.01)。FJB染色显示转染PGC-1αshRNA质粒组,KA刺激24h培养大鼠海马神经元损伤较KA对照组明显增加(P<0.05)。结论 PGC-1α基因下调加重KA刺激后培养大鼠海马神经元损伤,可能与其加重氧化应激损伤有关。

Objective To explore the role of peroxisome proliferator-activated receptor- γcoaetivator- 1α（PGC-1α） on kainic acid （KA） induced oxidative stress and neuronal injury in cultured rat hippocampal neuron. Methods Hippocampal neurons from SD rat were cultured and transferred with PGC-1α shRNA or vector （control） by the method of electroporation-based gene transfection. After 7 days, cultured hippocampal neurons were stimulated with KA for 24 hours. In vehicl control group （n = 10） ,experiment group （n = 11） and KA control group （n = 10） ,the content of glutathione （GSH） and activities of superoxidase dismutase （SOD） and lactic dehydrogenase （LDH） were measured, and neuronal injury was observed by FJB staining. Results The content of GSH and activity of SOD in KA control group [（5. 74 ± 0. 54） μmol·L^-1·μg^-1 protein, （10. 57 ± 1. 25） U/mg protein respectively] were less than those in vehicle control group [（9.38 ± 0.72）μmol·L^-1·μg^-1 protein, P〈0.01, （23.12 ± 3.23） U/rag protein, P〈0.01, respectively]. The activity of LDH was significantly elevated in KA control group [（2.21 ± 0.18） mmol NADH, H^＋/min/100 mg protein] compared with vehicle control group I-（1.29±0.09） mrnol NADH, H^＋/min/100 rng protein, P〈0. 01]. PGC-la shRNA transfer reduced the content of GSH and activity of SOD [（3.37 ± 0. 42） μmol·L^-1·μg^-1 protein, P〈0.05, （4.54 ± 0.91） U/rag protein, P〈0.05 respectively] and increased the activity of LDH [（2.74 ± 0. 19） mol/NADH^＋/rain/rag protein, P〈0. 01] further. FJB staining indicated that neuronal injury in experiment group was more severe than that in KA control group （P〈0.05）. Conclusions Down-regulation of PGC-1α aggravates KA induced neuronal injury in cultured rat hippocampal neuron,which may be due to exacerbation of neuronal oxidative stress.