In Vitro and In Vivo Evaluation of Kai-Xin-San Polysaccharides: Antioxidant, Anti-Inflammatory, and Anti-Alzheimer's Disease Activities

Introduction: Alzheimer's disease (AD) brings a considerable burden to families and society. Kai-Xin-San (KXS) is a traditional Chinese medicine formula used to treat AD with a good curative effect. The existing literature and our previous work suggest that KXS polysaccharides (KXS-P) may play an important role in the anti-AD effect of KXS. However, there is limited research available on the KXS-P and its potential anti-AD activities. Objective: To investigate the in vitro antioxidant, acetylcholinesterase (AChE) inhibitory effects, and anti-inflammatory activities of KXS-P, as well as to evaluate its anti-AD effect in vivo. Methods: KXS-P was characterized using scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FT-IR), and high-performance liquid chromatography (HPLC). The in vitro antioxidant activity and AChE inhibitory effects were evaluated. The in vitro anti-inflammatory activity of KXS-P was assessed using LPS-stimulated RAW264.7 cells. The in vivo anti-AD effects of KXS-P were evaluated using a rat model induced by D-galactose and Aβ25-35. The pharmacodynamic experiments included general behavior, open field test, Morris water maze, laser Doppler flowmetry, histopathological analysis (Nissl and HE staining), enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry. Results: KXS-P exhibited stronger antioxidant activity than single herb polysaccharides. KXS-P showed significant AChE inhibitory effects. KXS-P significantly inhibited the release of NO, TNF-α, IL-1β, and IL-6 in LPS-stimulated RAW264.7 cells. KXS-P effectively alleviated symptoms in AD model rats. Open-field tests and water maze tests demonstrated that KXS-P improved cognitive, learning, and memory functions in AD model rats. Laser Doppler flowmetry showed that KXS-P had a limited effect on cerebral blood flow in AD model rats. Nissl staining and immunohistochemistry of rat hippocampal tissue indicated that KXS-P protected hippocampal neurons. HE staining of rat colon revealed that KXS-P alleviated inflammation induced by intestinal flora imbalance. Conclusion: KXS-P exhibited potent anti-oxidation, anti-inflammatory activities and AChE inhibitory effects in vitro, as well as anti-AD effects in vivo. The anti-AD mechanism may be related to antioxidant effects, AChE inhibition, anti-inflammatory properties, and neuroprotection. other: Running title: The anti-AD activity of KXS-P