雷公藤红素抑制Cd2+诱导的神经毒性

Celastrol inhibits neurotoxicity induced by Cd2+

摘要:

镉(cadmium,Cd)是环境中常见的一种重金属，Cd²⁺可以通过穿透血脑屏障，产生神经毒性，从而诱发各种神经退行性疾病，雷公藤红素是雷公藤的一种有效成分，具有抗癌、抗炎等一系列药理作用，本文探究雷公藤红素对Cd²⁺诱导的相应神经毒性的影响作用。通过细胞增殖实验、细胞膜完整性实验、细胞形态实验探索了Cd²⁺对小胶质细胞HMC3活力的影响；通过一氧化氮(NO)检测实验、脂质过氧化(malondialdehyde,MDA)检测实验、蛋白免疫印迹实验分析了Cd²⁺的神经毒性以及雷公藤红素对Cd²⁺诱导的相应神经毒性的影响。结果表明：与对照组相比，当Cd²⁺浓度达到40 μmol/L时，对HMC3细胞增殖抑制率为(57.17±8.23)%(P<0.01,n=5)，继续增大Cd²⁺浓度，细胞活性将进一步降低；当Cd²⁺浓度达到40 μmol/L以上时，HMC3的细胞膜明显受到破坏，并且破坏作用与浓度呈剂量依赖性关系；随着Cd²⁺浓度的增加，细胞形态开始变化，贴壁效果变差。Cd²⁺使HMC3细胞释放的NO量显著增加，而雷公藤红素能够有效地抑制Cd²⁺诱导的HMC3细胞NO的释放；Cd²⁺使HMC3细胞脂质过氧化水平显著增加，加入10^–7 mol/L雷公藤红素后，MDA的释放量显著减少；Cd²⁺会使p-PI3K蛋白含量增加，而加入了雷公藤红素(10^–7、10^–6 mol/L)后，p-PI3K蛋白和p-AKT蛋白的激活均被抑制，从而抑制了细胞凋亡。综上所述，雷公藤红素能够抑制Cd²⁺诱导的小胶质细胞毒性，从而起到神经保护作用。

Abstract:

Cadmium (Cd) is a common heavy metal in the environment. Cd²⁺ may penetrate the blood-brain barrier and produce neurotoxicity, thus inducing various neurodegenerative diseases. Celastrol is an effective component of Tripterygium wilfordii Hook. F., which has many pharmacological effects such as anti-cancer and anti-inflammatory. Here we explored the effect of celastrol on the corresponding neurotoxicity induced by Cd²⁺. Cell proliferation test, cell membrane integrity test, and cell morphology were observed to analyze the effect of Cd²⁺ on the viability of HMC3. The neurotoxicity of Cd²⁺ and the effect of celastrol on the corresponding neurotoxicity induced by Cd²⁺ were analyzed by nitric oxide (NO) test, lipid peroxidation (MDA) test, and Western blotting. When the concentration of Cd²⁺ reached 40 μmol/L, the inhibition rate of HMC3 cell proliferation was (57.17±8.23)% (P<0.01, n=5), compared with the control group. The cell activity continued to reduce when the Cd²⁺ concentration further increased. When the concentration of Cd²⁺ was higher than 40 μmol/L, the cell membrane of HMC3 was significantly damaged, and the damage was dose-dependent. Upon increasing the Cd²⁺ concentration, the cell morphology began to change and the adhesion also became worse. Cd²⁺ significantly increased the amount of NO released by HMC3 cells, while celastrol effectively inhibited the NO release of HMC3 cells induced by Cd²⁺. Cd²⁺ greatly increased the release of MDA in HMC3 cells, and the level of MDA decreased rapidly upon the addition of 10^-7 mol/L celastrol. Cd²⁺ increased the expression of p-PI3K protein, and the levels of p-PI3K protein and p-AKT protein were inhibited by the addition of celastrol (10^‒7 mol/L, 10^‒6 mol/L), thus preventing cell apoptosis. In conclusion, celastrol inhibits Cd²⁺ induced microglial cytotoxicity and plays a neuroprotective role.