BDNF乙酰化修饰参与毛蕊花糖苷对Aβ_(25-35)致PC12细胞损伤的保护作用

Involvement of BDNF acetylation modification in the protective effect of verbascoside on the damage to PC12 cells by Aβ_(25-35)

目的:通过观察肉苁蓉中毛蕊花糖苷(OC1)对大鼠肾上腺嗜铬瘤细胞(PC12)增殖的影响,探讨OC1对脑源性神经营养因子(BDNF)乙酰化修饰参与的β-淀粉样蛋白(Aβ_(25-35))诱导PC12细胞损伤的影响。方法:取Aβ_(25-35)20μmol·L-1分别与OC1 50,150μg·m L-1和人参皂苷(GSrd)20μg·m L-1共同作用48 h后,流式细胞术检测细胞凋亡情况;Hoechst33258染色观察细胞形态的变化。酶联免疫吸附实验检测组蛋白乙酰化酶(HATs)、组蛋白去乙酰化酶2(HDAC2)和BDNF的含量变化。Western Blotting检测HATs腺病毒E1A相关蛋白p300和HDAC2的表达水平变化。染色质免疫共沉淀(CHIP)检测BDNF基因exon IV组蛋白H3乙酰化(Ac-H3)水平的变化和HDAC2含量变化。结果:给予药物干预后,凋亡情况出现不同程度的好转。OC1和GSrd作用48 h后,HATs表达增加,p300/CBP表达增加,HDAC2表达下降,BDNF水平提高。BDNF基因exon IV组蛋白H3乙酰化水平升高,HDAC2水平降低。结论:OC1对Aβ_(25-35)暴露PC12细胞有明显的保护作用,其机制可能是Aβ_(25-35)通过引起HDAC2依赖的组蛋白H3乙酰化的降低诱导了持续BDNF基因转录的抑制,从而导致了PC12细胞的损伤。OC1可能通过抑制HDAC2的表达来活化BDNF基因转录从而起到保护PC12细胞、刺激PC12细胞再生的作用。

Objective： To investigate the protective effect of verbascoside（ OC1） on β-amyloid protein（ Aβ_（25-35））-induced rat adrenalpheochromocytoma（ PC12） injury,and study the effect of histone acetylation on Brain-derived neurotrophic factor（ BDNF） gene transcription. Methods： Aβ_（25-35） of 20 μmol·L^-1 was incubated with OC1 of 50,150 μg·mL^（-1） and ginsenoside（ GSrd） of 20 μg·mL^（-1） for 48 h,and the cell apoptosis was detected by flow cytometry. Hoechst33258 staining was used to observe the changes in cell morphology. Enzyme linked immunosorbent assay was adopted to detect changes in contents of histone acetyltransferases（ HATs）,histone deacetylase 2（ HDAC2） and BDNF. Western Blotting was used to detect changes in expression levels of HATsadenovirus E1 A-associated protein p300 and HDAC2. Chromatin immunoprecipitation（ CHIP） was used to detect changes in H3 acetylation（ Ac-H3） levels in BDNF gene exon IV and changes in HDAC2 content. Results： After the drug intervention,the apoptosis showed different degrees of improvement. After 48 h of OC1 and GSrd treatment,HATs expression increased,p300/CBP expression increased,HDAC2 expression decreased,and BDNF levels increased. The BDNF gene exon IV histone H3 acetylation level increased,HDAC2 levels decreased.Conclusion： OC1 has a significant protective effect on Aβ_（25-35） exposure to PC12 cells. The mechanism may be that Aβ_（25-35） induces sustained inhibition of BDNF gene transcription by causing a decrease in HDAC2-dependent histone H3 acetylation,resulting in PC12 cell damage. OC1 can activate BDNF gene transcription by inhibiting the expression of HDAC2 to protect PC12 cells and stimulate the regeneration of PC12 cells.