半枝莲黄酮抑制复合Aβ所致大鼠皮层细胞NFT沉积及其调节机制

Scutellaria barbata flavonoids inhibits NFT aggregation and regulatory mechanism in rats induced by composited Aβ

目的:探讨半枝莲黄酮(SBF)对复合Aβ,即β淀粉样蛋白25-35(Aβ25-35)联合三氯化铝(AlCl_3)和重组人类转化生长因子-β1(RHTGF-β1)所致大鼠脑内神经元纤维缠结(NFT)沉积,tau蛋白磷酸化的影响及糖原合成激酶(GSK)3β和蛋白磷酸酶(PP)2A的调节机制。方法:雄性SD大鼠,脑室注射复合Aβ建立拟阿尔茨海默病(AD)大鼠记忆障碍模型,Morris水迷宫进行记忆障碍模型筛选,模型成功大鼠灌胃35、70和140 mg/kg的SBF和140 mg/kg的阳性对照药银杏叶黄酮(GLF),持续37 d。硝酸银法测定大鼠大脑皮层的NFT,Western blot法检测大鼠海马、皮层中总tau蛋白、Ser199和Ser214位点磷酸化的tau蛋白水平以及相关GSK3β和PP2A蛋白的表达水平。RT-PCR法检测海马、皮层中GSK3β和PP2A的mRNA水平。结果:大鼠脑室注射复合Aβ可以引起大鼠脑内NFT生成增加、Ser199和Ser214位点磷酸化tau蛋白、GSK3β蛋白和mRNA表达水平皆明显增加,PP2A的蛋白和mRNA表达水平明显降低。3种剂量的SBF灌胃37 d不同程度地逆转复合Aβ所致大鼠脑内上述异常改变。GLF也表现出与SBF相似的结果。结论:SBF能够抑制复合Aβ所致大鼠脑内NFT沉积,该作用可能是通过抑制GSK3β活性、增加PP2A活性从而降低tau蛋白磷酸化而实现的。

AIM：To investigate the effects of Scutellaria barbata flavonoids（ SBF） on neurofibrillary tangle（ NFT） aggregation,tau protein phosphorylation and the regulated mechanism of glycogen synthase kinase（ GSK） 3β and protein phosphatase（ PP） 2A in the rats induced by amyloid β protein 25-35（ Aβ25-35） in combination with AlCl3 and recombinant human transforming growth factor（ RHTGF）-β1（ composited Aβ）.METHODS：The male SD rats were used to establish the simulated Alzheimer disease（ AD） model by intracerebroventricular injection of composited Aβ.The Morris water maze was applied for screening the successful model rats with learning and memory deficits.The successful model rats were daily and orally administrated with SBF at doses of 35,70 and 140 mg/kg or positive control drug Ginkgo biloba leaves flavonoids（ GLF） at 140 mg/kg for 37 d.The silver nitrate staining was used to determine the cortical NFT.The protein levels of total tau,phosphorylated protein of tau at Ser199 and Ser214 sites,GSK3β and PP2A in hippocampus and cortex were determined by Western blot.The mRNA expression of GSK3β and PP2A in the hippocampus and cortex was detected by RT-PCR.RESULTS：Compared with sham group,the cell number of positive NFT with silver nitrate staining in modelrat cerebral cortex was significantly increased.The protein levels of phosphorylated tau protein at Ser199 and Ser214 sites,GSK3β in the hippocampus and cerebral cortex in the model rats dramatically elevated,and PP2A was marked decreased as compared with the sham group rats.Meanwhile,the mRNA expression of GSK-3β significantly increased but PP2A was decreased.However,these above abnormalities were differently attenuated by treating with SBF at different doses or GLF at140 mg/kg for 37 d.CONCLUSION：SBF suppresses the NFT aggregation by inhibition of the regulatory functions of GSK-3β and PP2A,thus reducing the phosphorylation of tau protein.