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ObjectiveTo investigate the effect of Danzhi Xiaoyaosan on the phosphorylation of tau protein and different sites of glycogen synthase kinase-3β （GSK-3β） and phosphoseryl/suanyl phosphate protein phosphatase 2A （PP2A） in the hippocampus of rats with Alzheimer's disease （AD） and its mechanism. MethodThe rat model of AD was established by injecting okadaic acid into the bilateral hippocampus of 90 male Wistar rats in SPF grades. The rats with successful modeling were selected and randomly divided into model group， aricept group （0.5 mg·kg-1）， and Danzhi Xiaoyaosan high， medium， and low groups （17.55， 8.77， and 4.38 g·kg-1）， and then gavaged for 42 d， once a day. Morris water maze was used to detect the learning and memory ability of rats， Nissl's staining was used to observe the morphological structure of neurons in the hippocampus， and Real-time polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression levels of tau protein， GSK-3β， and PP2A. Western blot was used to determine the protein expression levels of tau protein， GSK-3β， and PP2A. ResultAs compared with the control group， the learning and memory abilities of the rats in the model group were significantly decreased （P<0.01）， and the hippocampal CA3 region cells had abnormal structure， disorderly arrangement， and decreased number. The expression levels of GSK-3β mRNA， GSK-3β， p-GSK-3β-Tyr216， p-PP2A， and p-tau were increased in the model group as compared with the control group （P<0.01）， and those of p-GSK-3β-Ser9 and PP2A decreased significantly （P<0.01）. As compared with the model group， the learning and memory ability of the Aricept group and the Danzhi Xiaoyaosan groups were improved （P<0.05， P<0.01）， and the cell morphology and the number of hippocampal CA3 regions were better. The mRNA expression levels of PP2A and tau in the Aricept group were significantly up-regulated （P<0.05）， the mRNA expression level of GSK-3β was significantly down-regulated （P<0.01）， and the protein expression levels of GSK-3β， p-GSK-3β-Tyr216， and p-PP2A were down-regulated （P<0.05， P<0.01）， and the protein expression level of PP2A was significantly up-regulated （P<0.01）. As compared with the model group， the mRNA expression level of PP2A in the high-dose Danzhi Xiaoyaosan group was significantly up-regulated （P<0.01）， and that of GSK-3β was significantly down-regulated （P<0.01）， whereas the protein expression levels of p-PP2A， p-GSK-3β-Tyr216， and p-tau were down-regulated （P<0.05， P<0.01）， and the protein expression level of PP2A was significantly up-regulated （P<0.01）. As compared with the model group， the mRNA expression level of GSK-3β was significantly down-regulated in the medium-dose Danzhi Xiaoyaosan group （P<0.01）， the protein expression levels of GSK-3β， p-GSK-3β-Tyr216， and p-tau were down-regulated （P<0.05， P<0.01）， and the protein expression level of PP2A was significantly up-regulated （P<0.01）. As compared with the model group， the mRNA expression level of PP2A was significantly up-regulated in the low-dose Danzhi Xiaoyaosan group （P<0.01）， and that of GSK-3β was significantly down-regulated （P<0.01）， whereas the protein expression levels of GSK-3β and p-GSK-3β-Tyr216 were down-regulated （P<0.05， P<0.01）， and those of p-GSK-3β-Ser9 and PP2A were significantly up-regulated （P<0.01）. ConclusionDanzhi Xiaoyaosan can improve the learning and memory ability of rats with AD， and its mechanism may be related to the regulation of the activities of GSK-3β and PP2A protein-related sites and the phosphorylation of tau protein.
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Alzheimer's disease （AD） is a neurological disease highly related to age， which is the main cause of senile dementia and the most common disease leading to the loss of daily living ability of the elderly. AD brings heavy mental burden and economic pressure to patients， families， and society. Traditional Chinese medicine （TCM） ascribes AD to category of "dementia"， believing that the treatment should start from kidney because kidney deficiency is the root cause. Combined with the physiological and pathological characteristics of liver， this paper proposed that liver-kidney homology was an important idea for the prevention and treatment of AD. The main pathological manifestations of AD were amyloid β-protein （Aβ） deposition and neurofibrillary tangles （NFT）， and the pathogenesis was complex. A growing number of studies showed that immune inflammation played an important role in the pathogenesis of AD. The important target of treating AD was the regulation of neuro-immune inflammation through the nuclear factor kappa B （NF-κB）/NOD-like receptor thermal protein domain associated protein 3 （NLRP3）/Caspase-1/interleukin-1β （IL-1β） signaling pathway. Based on the idea of liver-kidney homology， this paper selected the representative formula Hei Xiaoyaosan to explore its effect on the prevention and treatment of AD and the mechanism from the perspective of regulating NF-κB/NLRP3/Caspase-1/IL-1β signaling pathway and inhibiting neuro-immune inflammation， expecting to further promote the in-depth study on the prevention and treatment of AD， and provide references for the prevention and treatment of AD by TCM.
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Objective To explore the effects of ferulic acid on gastric cancer cell line MGC-803 proliferation;To discuss the mechanism of apoptosis induced by ferulic acid. Methods Ferulic acid with a variety of concentrations was used to treat gastric cancer cell line MGC-803 for 24, 48, 72 h;MTT experiment was used to detect the growth of gastric cancer cells. After gastric cancer cell line MGC-803 were treated with ferulic acid with a variety of concentrations (0, 5, 7.5, 10 mg/mL) for 48 h, all gastric cancer cells were collected and stained by Annexin V-FITC/PI for the detection of apoptosis by flow cytometry. RT-qPCR and Western-blot methods were used to detect mRNA and protein levels of Caspase-3, Caspase-9, Bax, Bcl-2 and Xiap. Results Compared with the control group, the OD value of cell line MGC-803 treated by ferulic acid with a variety of concentrations (5, 7.5, 10, 12.5 mg/mL) decreased significantly;the anti-tumor effect of ferulic acid was obvious;IC50 of 24, 48, and 72 hours after treated by ferulic acid was 12.93, 9.73 and 5.52 mg/mL respectively. Cell lineMGC-803 treated by ferulic acid with a variety of concentrations (5, 7.5, 10 mg/mL) after 48 h could induce the apoptosis of MGC-803 cells, up-regulate mRNA and protein levels of Caspase-3, Caspase-9, and Bax, down-regulate mRNA and protein levels of Bcl-2 and Xiap. Conclusion Ferulic acid can inhibit the proliferation of MGC-803 cells effectively and induce the apoptosis of MGC-803 cells, which mechanism is related to mitochondria apoptosis pathway and the down-regulation of Xiap expression.
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Objective To explore the mechanisms and the significance of compatibility of Heixiaoyao Powder by observing the effect of Heixiaoyao Powder on mice model of senile dementia induced by scopolamine. Methods Kunming mice were randomly divided into five groups including control group, model group, Heixiaoyao Powder group, Xiaoyao powder group and streamlined formula group, with 10 mice of each group. Scopolamine hydrobromide intraperitoneal injection was adopted to make memory impairment in mice model, and followed by drugs treatment with corresponding decoction of the crude drug 2.314 g /(kg·d). Control group and model group were given normal saline orally once a day, all groups were treated for 10 days continuously. After 6 and 8 days of the administration, water maze test, stair jumping test and darkness avoidance test were conducted to test behavior proficiency, and the activity of acetylcholinesterase (AchE) activity and choline acetyltransferase (ChAT) in brain tissue were tested also. Results Compared with model group, Heixiaoyao Powder group, Xiaoyao Powder group and streamlined formula group could significantly reduce the error number and shorten latency in the water maze test, decrease the time of electrical shocks and prolong the incubation period of time in both stair jumping test and darkness avoidance test, and could increase ChAT activity and lower AchE activity in brain tissue of mice (P<0.01 or P<0.05), especially the effects of Heixiaoyao Powder group. Conclusions Heixiaoyao Powder and its analogous formula have the function of preventing and treating senile dementia.