An endophenotype network strategy uncovers YangXue QingNao Wan suppresses Aβ deposition, improves mitochondrial dysfunction and glucose metabolism.

Background: Alzheimer's disease (AD), an escalating global health issue, lacks effective treatments due to its complex pathogenesis. YangXue QingNao Wan (YXQNW) is a China Food and Drug Administration (CFDA)- approved TCM formula that has been repurposed in clinical Phase II for the treatment of AD. Identifying YXQNW's active ingredients and their mechanisms is crucial for developing effective AD treatments.

Purpose: This study aims to elucidate the anti-AD effects of YXQNW and to explore its potential therapeutic mechanisms employing an endophenotype network strategy.

Methods: Herein we present an endophenotype network strategy that combines active ingredient identification in rat serum, network proximity prediction, metabolomics, and in vivo experimental validation in two animal models. Specially, utilizing UPLC-Q-TOF-MS/MS, active ingredients are identified in YXQNW to build a drug-target network. We applied network proximity to identify potential AD pathological mechanisms of YXQNW via integration of drug-target network, AD endophenotype gene sets, and human protein interactome, and validated related mechanisms in two animal models. In a d-galactose-induced senescent rat model, YXQNW was administered at varying doses for cognitive and neuronal assessments through behavioral tests, Nissl staining, and transmission electron microscopy (TEM). Metabolomic analysis with LC-MS revealed YXQNW's influence on brain metabolites, suggesting therapeutic pathways. Levels of key proteins and biochemicals were measured by WB and ELISA, providing insights into YXQNW's neuroprotective mechanisms. In addition, 5×FAD model mice were used and administered YXQNW by gavage for 14 days at two doses. Amyloid-β levels, transporter expression, and cerebral blood flow have been detected by MRI and biochemical assays.

Results: The network proximity analysis showed that the effect of YXQNW on AD was highly correlated with amyloid β, synaptic function, glucose metabolism and mitochondrial function. The results of metabolomics combined with in vivo experimental validation suggest that YXQNW has the potential to ameliorate glucose transport abnormalities in the brain by upregulating the expression of GLUT1 and GLUT3, while further enhancing glucose metabolism through increased O-GlcNAcylation and mitigating mitochondrial dysfunction via the AMPK/Sirt1 pathway, thereby improving d-galactose-induced cognitive deficits in rats. Additionally, YXQNW treatment significantly decreased Aβ_1-42 levels and enhanced cerebral blood flow (CBF) in the hippocampus of 5×FAD mice. while mechanistic findings indicated that YXQNW treatment increased the expression of ABCB1, an Aβ transporter, in 5×FAD model mice to promote the clearance of Aβ from the brain and alleviate AD-like symptoms.

Conclusions: This study reveals that YXQNW may mitigate AD by inhibiting Aβ deposition and ameliorating mitochondrial dysfunction and glucose metabolism, thus offering a promising therapeutic approach for AD.