交泰丸入脑成分通过PTEN/AKT/NF-κB通路抑制LPS诱导BV2细胞炎症的机制研究  

Brain-penetrating components of Jiaotai Pills inhibit LPS-induced inflammation in BV2 cells via the PTEN/AKT/NF-κB signaling pathway

作　　者：陈佩瑶 戴国梁 王艳军 黄茜 王一然 邵雪文 居文政 CHEN Peiyao;DAI Guoliang;WANG Yanjun;HUANG Qian;WANG Yiran;SHAO Xuewen;JU Wenzheng(Department of Clinical Pharmacology,Jiangsu Province Hospital of Chinese Medicine,Affiliated Hospital of Nanjing University of Chinese Medicine,Jiangsu Nanjing 210029,China)

机构地区：[1]南京中医药大学附属医院/江苏省中医院临床药理科,江苏南京210029

出　　处：《中国医院药学杂志》2025年第6期614-621,共8页Chinese Journal of Hospital Pharmacy

基　　金：江苏省自然科学基金面上项目资助(编号:BK20211394);江苏省中药临床药理医学重点实验室项目(编号:ZDXYS202209);江苏省研究生科研与实践创新计划项目(编号:KYCX24_2339)。

摘　　要：目的:基于超高效液相色谱与四极杆-静电场轨道阱-线性离子阱三合一式质谱仪(UHPLC-Orbitrap-MS)、分子对接及分子生物学技术探讨交泰丸入脑成分抑制小鼠小胶质细胞(BV2)炎症的作用机制。方法:采用UHPLC-Orbitrap-MS技术分析交泰丸入脑成分,并结合分子对接技术筛选关键入脑成分。进一步构建脂多糖(lipopolysaccharides,LPS)诱导的BV2细胞炎症模型,并给予小檗碱(berberine,BBR)、木兰花碱(magnoflorine,MAG)干预处理。采用RT(q)-PCR检测细胞内肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interlenkin-1β,IL-1β)、IL-6 mRNA表达水平并使用ELISA检测细胞上清液中TNF-α、IL-1β、IL-6蛋白水平;倒置显微镜观察各组细胞形态;Western blot检测各组细胞PTEN/AKT/NF-κB通路关键蛋白的表达。结果:鉴定出交泰丸入脑成分13个,均为生物碱类化合物。RT-PCR、ELISA结果显示,BBR、MAG均能显著降低BV2细胞炎症模型中TNF-α、IL-1β、IL-6 mRNA及蛋白表达水平。同时,BBR与MAG能改善BV2细胞炎症模型的细胞形态。此外,与正常组相比,模型组中p-PTEN/PTEN、p-AKT/AKT、p-p65/p65蛋白水平显著提高,而BBR、MAG组中上述蛋白的表达均得到显著抑制。结论:交泰丸入脑成分BBR、MAG可能通过抑制PTEN/AKT/NF-κB信号通路,进而抑制细胞炎性因子的表达并改善细胞形态,发挥抑制BV2细胞炎症的作用。OBJECTIVE To investigate the potential of brain-penetrating components of Jiaotai Pills in inhibiting the inflammation in mouse microglial cells BV2 via ultra-high-performance liquid chromatography combined with an Orbitrap mass spectrometer(UHPLC-Orbitrap MS),molecular docking,and molecular biology.METHODS UHPLC-Orbitrap-MS was used to analyze the brain-penetrating components of Jiaotai Pills,and key components were identified using molecular docking.An in vitro inflammatory model was established in lipopolysaccharides(LPS)-induced BV2 cells,followed by treatment with berberine(BBR)and magnoflorine(MAG).The quantitative reverse-transcription polymerase chain reaction(qRT-PCR)was utilized to measure the mRNA levels of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),and IL-6 in cells.Enzyme-linked immunosorbent assay(ELISA)was utilized to measure the contents of TNF-α,IL-1β,and IL-6 in cell supernatants.Cell morphology was observed under an inverted microscope.Western blot analysis was performed to assess the protein expressions of key proteins in the phosphatase and tensin homolog deleted on chromosome 10(PTEN)/protein kinase B(AKT)/nuclear factor kappaB(NF-κB)signaling pathway.RESULTS A total of 13 brain-penetrating components of Jiaotai Pills were identified,all of them were alkaloid compounds.RT-PCR and ELISA results showed that BBR and MAG significantly decreased the levels of TNF-α,IL-1β,and IL-6 in LPS-induced BV2 cells.Meanwhile,BBR and MAG improved inflammatory morphology in LPS-induced BV2 cells.Furthermore,compared with the normal group,LPS-induced BV2 cells presented significantly elevated protein levels of p-PTEN/PTEN,p-AKT/AKT,and p-p65/p65,which were markedly suppressed by BBR and MAG treatment.CONCLUSION The brain-penetrating components BBR and MAG from Jiaotai Pills may exert anti-inflammatory effects on LPS-induced BV2 cells by reducing the expressions of inflammatory cytokines and improving cell morphology via inhibiting the PTEN/AKT/NF-κB signaling pathway.

关 键 词：交泰丸 超高效液相色谱与四极杆-静电场轨道阱-线性离子阱三合一式质谱仪 BV2细胞 炎症反应 第10号染色体磷酸酶和张力蛋白同源物 

分 类 号：R282[医药卫生—中药学] R966[医药卫生—中医学]