Study On The Relationship Between Alzheimer’s Disease And The Syndrome Of Spleen Deficiency And Fluid Retention Based On Lingguizhugantang’s Intervention Of Aβ-Induced Inflammatory Injury

Alzheimer’s disease, (AD) is a kind of acquired, comprehensive and progressive degenerative disease in the central nervous system, characterized by memory impairment as initial cognitive dysfunction, daily life self-care ability decrease and mental behavior disorder. With the sharply accelerated global population aging process, AD has become a focus for medical research community in recent years. The pathogenesis of AD is complex. Inflammatory damage, as one of its important pathogenesis, has shown a very close relationship with the syndrome of spleen deficiency and fluid retention. Our experiment team applied Lingguizhugantang (LGZGT) in preliminary studies, a classic formula from the Treatise on Febrile Disease, and studied its possible mechanism of treating AD from molecular biology level, which turned out to be very good support for the previous relationship hypothesis. However, in addition to the previous proven treatment targets, still it remains unknown whether LGZGT can intervene beta-amyloid transportation in the model rat brain, reduce NF-kappa B signaling activation, protect brain cells from inflammation induced damages and block the vicious cycle of inflammation in the brain injury. If this hypothesis is true, we may be able to interpret LGZGT’s efficacy from in vivo experiment level, so to provide support of the treatment of AD from the perspective of spleen deficiency and fluid retention, fully back up the relationship hypothesis of Aβ Deposition-Inflammatory Injury-Spleen Deficiency and Fluid Retention, better illustrate the nature of spleen deficiency and fluid retention syndrome in AD, and further enrich the connotation of AD pathology.OBJECTIVESTo observe LGZGT’s effect in the treatment of AD from in vivo experiment level with the classic formula for invigorating the spleen and eliminating the fluid from the Treatise on Febrile Disease and to explore its possible mechanism; to illustrate the nature of spleen deficiency and fluid retention syndrome in AD, and to further enrich the connotation of AD pathology based on the relationship hypothesis of AD-Aβ Deposition-Inflammatory Injury-Spleen Deficiency and Fluid Retention.METHODSAD rat models were created and later they were treated with LGZGT for 3weeks. The experiment mainly includes five parts.1. Aβ induced AD rat models were established and evaluated. Bilateral intraventricular injection of Aβ1-42 in rats so as to establish AD models. Results of navigation experiment comparisons before and with Morris water maze, together with hematoxylin-eosin staining results were observed, which ensured the success of establishing AD rat models.2. LGZGT’s effect on model rats’learning and memory ability was evaluated. After first-time Morris water maze, rats with natural disability in swimming and memory were deleted; normal and healthy rats were randomly assigned into groups, modelling and given proper tretment.3 weeks later, their learning and memory ability in Morris water maze navigation experiment were compared to determine LGZGT’s effect on them.3. LGZGT’s effect on the hippocampus and cortex cell morphology of model rats was evaluated. After hematoxylin eosin staining, the morphology of hippocampus and cortex neurons, astrocytes, microglia and other indicators of each rat were compared to find out LGZGT’s intervention on AD model rats.4. LGZGT’s effect on expressions of IL-6, IL-1β and TNF-α at the hippocampus and cortex of model rats were evaluated. With immunohistochemical experiments, expressions of IL-6, IL-1β and TNF-ain each group were calculated so as to figure out mechanism of LGZGT treating inflammation in AD.5. LGZGT’s effect on RAGE and NF-κB expressions at the hippocampus and cortex of model rats were evaluated. With western-blot experiments, expressions of RAGE and NF-κB in each group were compared so as to figure out anti-inflammatory mechanism of LGZGT in AD. RESULTS1. Aβ induced AD rat models were successfully established. The average incubation time and swimming distance in Morris water maze navigation experiment before and after modelling showed significant difference; microscopically, after hematoxylin eosin staining, hippocampus and cortex injury pathological characteristics in AD have been observed, suggesting the success of AD modelling in rats.2. The second experiment showed the average incubation time in Morris water maze navigation experiment after LGZGT treatment was reduced significantly.3. The third experiment showed that after hematoxylin eosin staining, hippocampus and cortex injury pathological characteristics of neurons, astrocytes and microglia in model rats were improved significantly.4. The fourth immunohistochemical experiment demonstrated significant declines of IL-6, IL-1β and TNF-a expression at the hippocampus and cortex of model rats.5. The fifth experiment of western-blot demonstrated significant declines of RAGE and NF-κB expressions at the hippocampus and cortex of each model rats.CONCLUSIONS1. AD rat models can be well established via bilateral intraventricular injection of AP1.42 in rats.2. LGZGT can shorten the average incubation time and swimming distance in Morris water maze navigation experiment, suggesting its effect of improving rats’learning and memory abilities.3. LGZGT can effectively improve the pathological characteristics of neurons, astrocytes and microglia at hippocampus and cortex injury in model rats.4. LGZGT can significantly decrease IL-6, IL-1β and TNF-α expression at the hippocampus and cortex of model rats, indicating its good effects of suppressing inflammation in AD.5. LGZGT can significantly cut down RAGE and NF-κB expressions at the hippocampus and cortex of each model rats, suggesting it can prevent and treat AD through regulating NF-κB signaling pathway.6. The treatment of AD with LGZGT is practical in theories and has been well supported by in vivo experiments.