亚急性砷中毒对大鼠脑氧化应激及Fos/Jun蛋白表达影响

Effect of sub-acute arsenic exposure on cerebral oxidative stress and Fos/Jun expression in rats

目的探讨亚急性砷中毒对大鼠脑神经氧化应激影响及机制。方法清洁级SD大鼠56只,随机分为4组,即对照组(蒸馏水)及低、中、高剂量染砷组(2、10、50 mg/L亚砷酸钠溶液),雌雄各半,每组14只,以自由饮水方式染砷30 d。用电感耦合等离子体发射光谱(ICP-AES)测定大鼠血砷、脑砷浓度,用5,5–二硫二硝基苯甲酸速率微板法测定脑组织还原型谷胱甘肽(GSH)含量,用微量定磷法测定脑组织中γ–谷氨酰半胱氨酸合成酶(γ-GCS)活性,免疫组化法测定脑海马和皮质区c-fos和c-jun蛋白表达。结果与对照组比较,各剂量染砷组大鼠血砷、脑砷浓度明显升高,差异均有统计学意义(P <0.05);脑海马区神经元肿胀率、变性率、坏死率高于对照组;与对照组比较,低、中、高剂量染砷组大鼠脑组织中γ-GCS活性[分别为(4.79±0.60)、(3.71±0.87)、(3.13±0.43) U/mgpro]、GSH含量[分别为(29.59±1.73)、(24.57±1.53)、(20.59±2.51)μmol/gpro]明显下降,差异均有统计学意义(均P <0.05);与对照组比较,低、中、高剂量染砷组大鼠海马和皮质c-fos阳性率[分别为(65.36±6.38)%、(71.51±6.05)%、(72.20±6.13)%和(67.81±7.47)%、(70.99±6.67)%、(70.77±4.79)%],c-jun阳性率[分别为(56.67±8.29)%、(65.65±5.70)%、(69.88±2.85)%和(59.46±6.05)%、(65.80±4.31)%、(67.51±3.84)%]明显升高,差异均有统计学意义(均P <0.05)。结论亚急性砷中毒导致脑组织c-fos和c-jun蛋白表达增加、γ-GCS活性下降、GSH含量减少、降低脑组织氧化应激能力,可能是亚急性砷暴露致中枢神经毒性作用机制之一。

Objective To investigate the effect and mechanism of sub-acute arsenic exposure on cerebral oxidative stress in rats. Methods Fifty-six Sprague-Dawley （SD） rats of clean grade were randomly divided into 4 groups （7 males and 7 females in each group）： a control group, low-, moderate, and high-dose groups （administered with drinking water containing 2.0, 10, and 50 mg/L sodium arsenite [NaAsO2] for 30 consecutive days）. Arsenic content in brain tissues and blood were detected with inductively coupled plasma atomic emission spectrometry （ICP-AES）; the level of reduced glutathione （GSH） in brain tissues was determined with 5, 5 ＇-dithiobis-[2-nitrobenzoic acid] （DTNB） method; the activity of gamma- glutamylcysteine （γ-GCS） in brain tissues was measured with microdetermination of phosphorus; and c-fos and c-jun expressions in brain tissues were determined with immunohistochemical method. Results Compared with those of the control group, the arsenic contents in blood and brain tissues were significantly higher in arsenic exposed groups （both P 〈 0.05）; the rate of swelling, denaturation, and necrosis of neurons in brains of arsenic exposed groups were all higher than those of the control group. In brain tissues of the rats exposed to low-, moderate-, and high-dose arsenic, significantly decreased activity ofy-GCS （4.79 ± 0.60, 3.71 ± 0.87, and 3.13 ±0.43 U/mgpro） and GSH level （29.59 ± 1.73, 24.57 ± 1.53, and 20.59 ± 2.51 μmol/gpro） were observed in comparison with those of the control rats （P 〈 0.05 for all）; whereas, both the c-fos expression in hippocampus （65.36 ± 6.38%, 71.51 ± 6.05%, and 72.20 ± 6.13%） and in cortex （67.81 ± 7.47%, 70.99 ±： 6.67, and 70.77 ± 4.79 %） and c-jun expression in hippocampus （56.67 ± 8.29%, 65.65 ± 5.70%, and 69.88 ± 2.85%） and in cortex （59.46 ± 6.05%, 65.80 ± 4.31%, and 67.51 ± 3.84%） increased significantly for the rats exposed to low-, moderate-, and high-dose arsenic compared to the control rats （P 〈 0.05 for all）. Conclusion In rats, sub-acute arsenic exposure results in overexpressions of c-los and c-jun protein and downregulations of gamma-GCS activity and GSH level in brain tissues, which may attenuate cerebral capacity of against oxidative stress and may partially explain the mechanisms of central neurotoxicity induced by subacute arsenic exposure.