Abstract
With its traditional use in relieving insomnia and anxiety, our previous study has identified onjisaponin B from Radix Polygalae (RP), as a novel autophagic enhancer with potential neuroprotective effects. In current study, we have further identified a novel active fraction from RP, contains 17 major triterpenoid saponins including the onjisaponin B, by the combinational use of cell membrane chromatography (CMC) and ultra-performance liquid chromatography coupled to (quadrupole) time-of-flight mass spectrometry {UHPLC-(Q)TOF-MS}. By exhibiting more potent autophagic effect in cells, the active fraction enhances the clearance of mutant huntingtin, and reduces protein level and aggregation of α-synuclein in a higher extent when compared with onjisaponin B. Here, we have reported for the first time the new application of cell-based CMC and UHPLC-(Q)TOF-MS analysis in identifying new autophagy inducers with neuroprotective effects from Chinese medicinal herb. This result has provided novel insights into the possible pharmacological actions of the active components present in the newly identified active fraction of RP, which may help to improve the efficacy of the traditional way of prescribing RP, and also provide new standard for the quality control of decoction of RP or its medicinal products in the future.
Highlights
Neurodegenerative diseases such as Alzheimer’s disease (AD), Parkinson’s disease (PD) or Huntington’s disease (HD), are caused by the formation of inclusion bodies and protein aggregates, or the deposition of abnormal proteins in neuronal cells, which lead to degeneration and selective neuronal vulnerability in specific brain regions[1,2,3]
It is reported for the first time the use of PC-12 cells coupled cell membrane chromatography (CMC) model to identify active autophagic Chinese herbal medicine (CHM) components which bind on the cell membrane (Fig. 1a)
To further study the molecular mechanisms of the isolated bioactive Radix Polygalae (RP) fractions, we investigated the autophagic effect of onjisaponin B (5 μ M), RP (70–80% methanol fraction (MF)) (62.5 and 125 μ g/mL), RP (NF) (62.5 μ g/mL) and RP (TEE) (62.5 μ g/mL) in both autophagy related gene 7 (Atg7)-wild type or -knockout mouse embryonic fibroblasts (MEFs) (Fig. 6e,f), which are resistant to autophagy induction[38]
Summary
Neurodegenerative diseases such as Alzheimer’s disease (AD), Parkinson’s disease (PD) or Huntington’s disease (HD), are caused by the formation of inclusion bodies and protein aggregates, or the deposition of abnormal proteins in neuronal cells, which lead to degeneration and selective neuronal vulnerability in specific brain regions[1,2,3]. The human epidermal squamous cells (A431 cells) and human embryonic kidney (HEK 293 cells) coupled CMC model were used for screening of epidermal growth factor receptor (EGFRs) antagonists[28,29], and the human umbilical vein endothelial cell (HUVEC) coupled CMC model was applied for analyzing the competitive binding activity on the receptor of AGEs (RAGE)[30] To this end, we applied the CMC, ultra-performance liquid chromatography time-of-flight mass spectrometry (UHPLC-TOF-MS) and ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS) to identify the active fraction and components of RP, which are responsible for the autophagic and neuroprotective effects in PC-12 cells[28,29,30]. With a more potent autophagic and neuroprotective effect induced by the active methanol fraction of RP (70–80% MF) when compared with onjisaponin B, the identification of the active fraction may help to further explain the pharmacological and mechanistic action of RP, improve the efficacy of the traditional way of prescribing RP decoction as medication, and serve as a new standard for the quality control of RP
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