Effects of Xixin Decoction （洗心汤） on the Diversity of Intestinal Flora and Levels of Aβ1-42，LPS，SAA，and ACH in Brain and Intestinal Tissues of Rapidly Aging Model Mice / 中医杂志

ObjectiveTo observe the possible mechanism of Xixin Decoction （洗心汤， XXD） in the prevention and treatment of Alzheimer 's disease（AD）. MethodsFifty rapid aging model mice （SAMP8） were randomly divided into model group， probiotic group， high-， moderate- and low-dose group of XXD， with 10 mice in each group. Another 10 homologous anti-rapid aging mice （SAMR1） were set as control group. After 10 weeks of feeding， the control group and the model group were given 10 ml·kg-1·d-1 of distilled water by gavage， while the probiotic group （0.39 g·kg-1·d-1）， the high-dose group of XXD （5.08 g·kg-1·d-1）， the moderate-dose group of XXD （2.54 g·kg-1·d-1）， and the low-dose group of XXD （1.27 g·kg-1·d-1） were given corresponding drugs or decoctions by gavage， once a day in all groups. After 10 weeks of intragastric administration， Morris water maze was used to detect the spatial learning and memory ability of mice in each group. HE staining was used to observe the pathological changes of hippocampal CA3 region and colon. The levels of β-amyloid 1-42 （Aβ1-42）， lipopolysaccharide （LPS）， serum amyloid A （SAA） and acetylcholine （ACH） in hippocampus and colon were detected by ELISA.The diversity of intestinal flora in mouse feces was detected by 16S rRNA sequencing. ResultsCompared to those in the control group， the levels of Aβ1-42，LPS， SAA increased， while the level of ACH decreased in the model group （P＜0.05 or P＜0.01）. Compared to those in the model group， the escape latency period of the probiotic group was significantly shortened on the 2nd and 5th days， while the escape latency period was shortened， and the residence time in the target platform quadrant increased in the high-dose XXD group during the 2nd to 5th days； the escape latency period was shortened significantly in the moderate-dose XXD group on the 5th day （P＜0.05 or P＜0.01）. Compared to those in the model group， the hippocampal neuron cells in the high- and moderate-dose XXD groups were arranged more closely， with decreased levels of SAA， Aβ1-42 and LPS， increased ACH level， Simpson and Shannon index （P＜0.05 or P＜0.01）； the arrangement of hippocampal neuron cells in the probiotic group and the low-dose XXD group was relatively loose； the proportions of Bacteroidetes and Prevotella were significantly reduced in the probiotic group and the high-dose XXD group， while that of Firmicutes and Lactobacillus significantly increased （P＜0.05 or P＜0.01）. Compared to those in the probiotic group and the high-dose XXD group， the number of goblet cells in the moderate-dose XXD group decreased， and the number of glands in the low-dose XXD group decreased with atrophy. The high-dose XXD group had decreased Aβ1-42 level in the hippocampus， increased ACH level in thehippocampus and colon tissue， and decreased SAA in the colon tissue than the moderate- and low-dose XXD groups （P＜0.05 or P＜0.01）； moreover， the SAA level in the hippocampus was significantly higher in the low-dose XXD group than the high- and moderate-dose groups （P＜0.01）. ConclusionXXD can improve the spatial learning and memory ability of SAMP8， reduce the production and deposition of LPS， SAA and Aβ1-42 in brain and intestine， and increase the content of ACH. The mechanism of its prevention and treatment of AD maybe related to regulating intestinal microecology， affecting flora diversity and improving inflammatory response.

Changes in Ileal Flora Induced by Qishengwan in Treatment of Alzheimer's Disease： An Exploration Based on "Interior-Exterior Relationship Between Heart and Small Intestine" / 中国实验方剂学杂志

ObjectiveTo explore the effect of Qishengwan on ileal flora during its treatment of Alzheimer's disease （AD） under the guidance of the theory of "interior-exterior relationship between heart and small intestine". MethodThe AD model was established by bilateral intraventricular injection of β-amyloid 1-42 （Aβ1-42）. The rats were then randomly divided into the blank group， sham-operated group， model group， low-， medium-， and high-dose （5.6， 11.2，22.4 g·kg-1·d-1） Qishengwan groups， and donepezil （0.46 mg·kg-1·d-1） group. After medication for 28 successive days， the spatial memory ability of rats was observed in water maze test， and the levels of Aβ1-42， nuclear transcription factor-κB （NF-κB）， tumor necrosis factor-α （TNF-α）， and interleukin-6 （IL-6） in the hippocampus were analyzed by enzyme-linked immunosorbent assay （ELISA）. Additionally， the contents of the ileum were collected and subjected to 16SrRNA-sequencing analysis for figuring out the changes in ileal flora. ResultCompared with the blank group and sham-operated group， the model group exhibited significantly reduced stay time in the target quadrant and number of target quadrant and platform crossings （P<0.05， P<0.01） and elevated Aβ1-42 content in the hippocampus （P<0.01） and central inflammatory factors NF-κB， TNF-α， and IL-6 （P<0.05， P<0.01）. Compared with the model group， Qishengwan at each dose significantly alleviated the impaired spatial memory function （P<0.05， P<0.01）， improved the deposition of Aβ1-42 in the hippocampus of rats （P<0.05， P<0.01）， and reduced the expression of central nervous system inflammatory factors （P<0.05， P<0.01）， thus exerting a good therapeutic effect on AD rats. The 16SrRNA-sequencing analysis results showed that the structure of the ileal flora in the model group was significantly separated from those in the blank group and sham-operated group. The abundance of Lachnospiraceae NK4A136 group was significantly increased （P<0.01）， while that of Escherichia-Shigella was reduced （P<0.05， P<0.01）. Qishengwan at each dose significantly changed the ileal flora structure and regulated the relative abundance of Lachnospiraceae NK4A136 group， Escherichia-Shigella， and Ruminococcaceae. ConclusionQishengwan has a positive therapeutic effect on AD. It can significantly enhance the memory and cognitive abilities in AD rats， which may be related to its regulation of the structure of rat ileal flora and the relative abundance of Lachnospiraceae NK4A136 group， Escherichia-Shigella， and Ruminococcaceae， the attenuation of the central neuroinflammatory response， and the reduction of central Aβ1-42 deposition.

Mechanism of Linggui Zhugantang in Repairing Blood-brain Barrier Injury of Alzheimer's Disease / 中国实验方剂学杂志

ObjectiveTo observe the effect of Linggui Zhugantang （LG） on the blood-brain barrier （BBB） model of Alzheimer's disease （AD） in vitro and to explore the mechanism of LG in repairing the BBB injury in AD. MethodA total of 50 male SPF rats were randomized into five groups： high-dose （4.8 g·kg-1）， medium-dose （2.4 g·kg-1）， and low-dose （1.2 g·kg-1） LG groups， western medicine （0.5 g·kg-1 donepezil hydrochloride） group， and normal group （normal saline of equivalent volume）. They received （ig） corresponding drugs twice a day for 7 d. Drug-containing serum was respectively collected from the abdominal aorta 1 h after the last administration. The BBB injury of AD in vitro was induced with the cell co-culture method， and 6 groups were designed： normal group， model group， high-， medium-， and low-dose LG groups， and western medicine group. The model group was added with 100 μL amyloid β1-42 （Aβ1-42， final concentration： 5 μmol·L-1）， and high-dose， medium-dose， and low-dose LG groups and the western medicine group were added with corresponding 10% drug-containing serum in addition to the 100 μL Aβ1-42 （final concentration： 5 μmol·L-1）. Cell survival rate was detected by methyl thiazolyl tetrazolium （MTT） assay， expression of BBB-related skeleton proteins （claudin-5， ZO-1， occludin）， matrix metalloproteinase-2 （MMP-2）， and matrix metalloproteinase-9 （MMP-9） by Western blot， and content of inflammatory factors interleukin-1β （IL-1β）， interleukin-6 （IL-6）， and tumor necrosis factor-α （TNF-α） by enzyme-linked immunosorbent assay （ELISA）. BBB Aβ transporter low-density lipoprotein receptor-related protein 1 （LRP-1） and advanced glycation end product receptor （RAGE） at different time points in high-dose， medium-dose， and low-dose LG groups were determined by Real-time PCR and Western blot. ResultCell survival rate of the model group was lower than that of the normal group （P<0.05） and the survival rates of the western medicine group and high-dose LG group was higher than that in the model group （P<0.05）. The skeleton proteins were down-regulated and MMP-2 and MMP-9 were up-regulated in the model group compared with those in the normal group （P<0.05）. The expression of skeleton proteins was higher （P<0.05） and that of MMP-2 and MMP-9 was lower （P<0.05） in the western medicine group and high-dose LG group than in the model group. Compared with the model group， only the medium-dose LG group showed the up-regulation （P<0.05） of claudin-5 （P<0.05） and the decrease （P<0.05） of MMP-2. IL-1β， IL-6， and TNF-α in the model group were up-regulated （P<0.05） compared with those in the normal group， and those inflammatory factors in the western medicine group and high-dose and medium-dose LG groups were lower （P<0.05） than those in the model group. LRP-1 expression was up-regulated and RAGE expression was down-regulated at 3 h compared with those at 0 h （P<0.05）， while the expression of the two became stable at 6， 12， 24， 36 h. At 3 h， LRP-1 expression was down-regulated and RAGE expression was up-regulated in model group compared with those in the normal group at 3 h （P<0.05）. Moreover， the LRP-1 content was higher and RAGE content was lower in the western medicine group and high-dose LG group than in the model group. ConclusionLG can repair the BBB injury in vitro by inhibiting the expression of inflammatory factors and MMP-2， MMP-9， promoting the expression of skeletal proteins， and regulating the balance of transporters.

Danggui Shaoyaosan Inhibits Neuroinflammation in AD Rats by Regulating NLRP3/Caspase-1 Pathway / 中国实验方剂学杂志

Objective:To investigate the neuroprotective effect of Danggui Shaoyaosan （DSS） in a rat model of amyloid-<italic>β</italic>-peptide_1-42 （A<italic>β</italic>_1-42）-induced Alzheimer's disease （AD） as well as its regulatory effect on NOD-like receptor protein 3 （NLRP3）/cysteinyl aspartate-specific protease-1 （Caspase-1） signaling pathway. Method:The AD animal model was established via intracerebral injection of A<italic>β</italic>_1-42 and treated with different concentrations of DSS after the division of rats into the sham operation group， model group， as well as the high-， medium-， and low-dose DSS groups. Morris water maze test was conducted to determine the learning and memory abilities of rats. The morphology and function of neurons were detected by hematoxylin-eosin （HE） staining and Golgi staining， followed by immunofluorescence co-localization of NLRP3 inflammasome activation. The mRNA expression levels of interleukin （IL）-1<italic>β</italic> and IL-18 were measured by Real-time polymerase chain reaction （Real-time PCR）， and the protein expression levels of NLRP3， Caspase-1， and IL-1<italic>β </italic>were assayed by Western blot. Result:Compared with the sham operation group， the model group exhibited significantly decreased learning and memory abilities （<italic>P</italic><0.01）， impaired neuronal morphology and function， up-regulated IL-1<italic>β</italic> and IL-18 mRNA expression， enhanced NLRP3 inflammasome activation， and elevated NLRP3， Caspase-1， and IL-1<italic>β</italic> protein expression （<italic>P</italic><0.01）. Compared with the model group， DSS at both medium and high doses remarkably improved the learning and memory abilities of AD rats （<italic>P</italic><0.05， <italic>P</italic><0.01）， restored neuronal morphology and function， down-regulated the mRNA expression levels of inflammatory factors IL-1<italic>β</italic> and IL-18， reduced the activation of NLRP3 inflammasomes， and lowered the protein expression levels of NLRP3， Caspase-1， and IL-1<italic>β</italic> （<italic>P</italic><0.01）. Conclusion:DSS inhibits inflammasome activation and neuroinflammatory response possibly by regulating the NLRP3/Caspase-1 signaling pathway， thus exerting the neuroprotective effect.

Abdominal acupoint thread embedding therapy based on "brain-intestinal connection" for mild-to-moderate Alzheimer's disease and its effects on serum levels of APP and Aβ / 中国针灸

OBJECTIVE@#To compare the clinical efficacy of abdominal acupoint thread embedding therapy based on "brain-intestinal connection" combined with donepezil hydrochloride tablets and oral donepezil hydrochloride tablets alone for mild-to-moderate Alzheimer's disease (AD) and observe its effects on amyloid precursor protein (APP) and β-amyloid protein@*METHODS@#Sixty patients with AD were randomly divided into an observation group (30 cases, 3 cases dropped off) and a control group (30 cases, 3 cases dropped off). The patients in the control group were treated with donepezil hydrochloride tablets (5 mg per day); based on the treatment in the control group, the patients in the observation group were treated with abdominal acupoint thread embedding therapy at Zhongwan (CV 12), Xiawan (CV 10), Huaroumen (ST 24), Wailing (ST 26), Daheng (SP 15), etc., once every 10 days. Both groups were treated for 2 months. The mini-mental state examination (MMSE), Alzheimer's disease assessment scale-cognitive subscale (ADAS-Cog), activity of daily living scale (ADL), neuropsychiatric inventory questionnaire (NPI) as well as the serum levels of APP and Aβ@*RESULTS@#After treatment, the MMSE scores in the two groups were higher than those before treatment (@*CONCLUSION@#The abdominal acupoint thread embedding therapy based on the theory of "brain-intestinal connection" combined with donepezil hydrochloride tablets can improve cognitive function, self-care ability of daily life and mental behavior, and reduce the serum levels of APP and Aβ

Effect of Huangjingwan on Antioxidant Functions and Aβ1-42 and APP Protein Expressions in Brain of Alzheimer' s Disease Rats / 中国实验方剂学杂志

Objective::To observe the effect of Huangjingwan (HW) on antioxidant functions and β-amyloid 1-42 (Aβ1-42) and amyloid precursor protein (APP) expressions in the brain of Alzheimer' s disease (AD) rats. Method::SD rats were randomly divided into normal control group, sham model control group, AD model group, and low, medium, high-dose (equivalent raw drug dose 1, 3, 9 g·kg-1·d-1) HW groups.The AD models were established through intraperitoneal injection with 1.25% D-galactose (120 mg·kg-1·d-1, 6 consecutive weeks) and then one-time right ventricular injection with Aβ1-42 (10 μg). Two weeks after modeling, the rats in each HW group received corresponding drugs through intragastric administration, once a day, while the rats in sham model control group, AD model group were given normal saline 1 mL through intragastric administration, once a day.Gastric perfusion lasted for 8 weeks.At the end of the experiment, learning and memory abilities of the rats were assessed by Platform Jumping Test.The changes of physical endurance in rats were tested by 10% weight swimming under load.The activities of superoxide dismutase (SOD), glutathione reductase (GR), glutathione peroxidase (GSH-Px) antioxidant enzymes and the contents of glutathione (GSH) and malondialdehyde (MDA) in rat brain tissue were detected by colorimetry.The changes of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), Aβ1-42 and APP protein in rat brain tissues were determined by enzyme linked immunosorbent assay (ELISA). Western blot analysis was used to measure the expression of APP protein in rat brain. Result::Compared with the normal control group, rats in AD model group showed an obvious dementia state, that is more lying and less movement, longer learning response time, significant increase in the number of learning and memory errors, significant attenuation in physical fitness, significant decrease in the activities of antioxidant enzymes (SOD, GR, GSH-Px) and anti-inflammatory factors GSH in brain, significant rise in the levels of inflammatory factors MDA, IL-1β and TNF-α and the content of Aβ1-42 protein, and significant reduction in the content of APP protein in brain (P<0.01). Low, medium and high-dose HW could ameliorate dementia symptoms in AD rats, improve the achievement of learning and memory, antagonize body weakness and increase physical fitness, promote SOD, GR, GSH-Px activities and anti-inflammatory factor GSH level in the brain, reduce the levels of MDA, IL-1β and TNF-α in the brain, decrease the level of Aβ1-42 and increase the level of APP protein in the brains of AD rats compared with the AD model group (P<0.05, P<0.01), besides, within the dose range of 1-9 g·kg-1·d-1, HW has a more obvious effect with the increase of dose. Conclusion::HW has the effects in preventing and treating AD, which is related to the HW' s mechanisms in enhancing the function of antioxidant system in brain, reducing neuroinflammatory reaction and deposition of Aβ1-42 induced by oxidative stress, and maintaining the expression level of APP protein.

Moxibustion improves learning-memory ability by promoting cellular autophagy and regulating autophagy-related proteins in hippocampus and cerebral cortex in APP/PS1 transgenic Alzheimer's disease mice / 针刺研究

OBJECTIVE: To observe the effect of moxibustion of acupoints of the Governor Vessel on the levels of cellular autophagy, β amyloid protein (Aβ) immunoactivity, and expression of LC3-Ⅰ， LC3-Ⅱ， p62 and p-P70S6K proteins in the hippocampal tissue of APPswe/PS1de9 (APP/PS1) double-transgenic Alzheimer's disease (AD) mice, so as to reveal its underlying mechanisms in improving AD. METHODS: APP/PS1 double-transgenic AD mice were randomly divided into AD model, moxibustion, autophagy-inducer (Rapamycin) and autophagy-inhibitor (3-MA)＋moxibustion groups (n＝10 in each group), and other 10 C57BL/6J male mice (the same age) were used as the normal control group. Herbal-cake (made of Chuanwu [Radix Aconiti Praeparata]) partitioned moxibustion was applied to "Baihui"(GV20), moxibustion was applied to "Fengfu"(GV16) and "Dazhui"(GV14), all for 20 min, once daily for 2 weeks, with one day's off between two weeks. For mice of the autophagy-inducer and 3-MA＋moxibustion groups, Rapamycin (2 mg•kg－1•d－1) and 3-MA (1.5 mg•kg－1•d－1) were separately administered by intraperitoneal injection for 2 weeks. The cognitive ability was examined by Morris water maze tests, and the ultrastructural changes (including autophagic lysosomes, etc.) of hippocampal neurons were observed by using transmission electron microscopy. The immunoactivity of cerebral cortex and hippocampal Amyloid β peptide 1-42 (Aβ1-42) was detected by immunohistochemistry, and the expression levels of hippocampal LC3-Ⅰ， LC3-Ⅱ， p62 and p-P70S6K proteins were detected by Western blot. RESULTS: After modeling, the escape latency of Morris water maze tasks was prolonged in the model group than in the normal control group (P<0.05) and obviously shortened in the moxibustion and autophagy-inducer groups (not the autophagy-inhibitor group) than in the model group (P<0.05). Results of transmission electron microscope showed deformed, irregular or atrophic neurons with rough and incomplete and fuzzy nuclear membrane, and decreased intracellular autophagosomes in the hippocampus in the model group, and partial irregular, atrophic neurons with more autophagic vesicles and lysosomes in the moxibustion group. The expression levels of Aβ1-42 in both cerebral cortex and hippocampus tissues, and LC3-Ⅰ， p62 and p-P70S6K proteins in the hippocampus were consi-derably up-regulated in the model group relevant to the normal control group (P<0.01), and evidently down-regulated in both moxibustion and autophagy-inducer groups (not the autophagy-inhibitor group) than in the model group (P<0.01), while that of hippocampal LC3-Ⅱ protein and LC3-Ⅱ/Ⅰ ratio levels were obviously down-regulated in the model group relevant to the normal control group (P<0.01), and significantly up-regulated in both moxibustion and autophagy-inducer groups (not the autophagy-inhibitor group) than in the model group (P<0.01)．. CONCLUSION: Moxibustion can improve the cognitive ability of APP/PS1 double-transgenic AD mice, which is associated with its effects in promoting hip-pocampal and cerebral cortex autophagy level, and down-regulating the expression levels of Aβ1-42, LC3-Ⅰ， p62 and p-P70S6K proteins in the hippocampus.

Effect of Hei Xiaoyaosan on Expressions of Aβ1-42, GSK-3β, NEP, IDE in Hippocampus Area of Alzheimer's Dementia Mice / 中国实验方剂学杂志

Objective: To observe the effect of Hei Xiaoyaosan on expressions of β-amyloid 1-42 peptide(Aβ1-42),glycogen synthase kinase-3β(GSK-3β),neprilysin(NEP),insulin-degrading enzyme(IDE) in the hippocampus area of Alzheimer's dementia mice. Method: After weighing, 42 APP/PSI bivalent transgenic mice were randomly divided into 4 groups:10 mice in the model group, 10 mice in the positive drug control group, 11 mice in the high-dose Hei Xiaoyaosan group, and 11 mice in the low-dose Hei Xiaoyaosan group; 10 wild C57BL/6J mice of the same age and strain were used for negative control group. Drugs were administered to mice by gavage once a day for 12 weeks. Then the behavior of all the mice were detected by Morris water maze, the morphological changes in hippocampal neurons were observed by hematoxylineosin(HE) staining, the expressions of Aβ1-42, GSK-3β, NEP and IDE proteins in hippocampus were detected by immunohistochemistry. Result: After 3 months of treatment, compared with negative control groups, the average escaping latency periods prolonged significantly, and the number of cross-platform was decreased significantly in model group (Pβ1-42 and GSK-3β proteins in model mice hippocampus were significantly increased (PPPβ1-42 and GSK-3β proteins in the hippocampus of drug groups were significantly decreased (PPPConclusion: Hei Xiaoyaosan can significantly improve the learning and memory abilities of AD mice, which may be related to the reduction of cognitive impairment in AD mice by regulating abnormal deposition and degradating Aβ in the hippocampus.

Effects of Exendin-4 on cognitive function and Nrf2 signaling pathway in rats with traumatic brain injury / 中华行为医学与脑科学杂志

Objective To explore the effect of Exendin-4 on dementia after traumatic brain injury (TBI) in rats and its related mechanism.Methods Thirty SD rats were randomly divided into sham group (n=10),TBI group(n=10) and Exendin-4 group(n=10).Cortical impact injury was used to construct the TBI model.Morris water maze test was used to test the memory function of rats one month after TBI.The beta-amyloid protein (Aβ1-42) and nuclear factor erythroid-2-related factor 2 (Nrf2) were detected by Western blot.Results One month after TBI compared with the sham group,the escape latency (EL) ((35.31 ± 13.23)s vs (8.79±9.71)s) was prolonged and the target quadrant stay time ((17.78±4.68)s vs (26.35± 5.83)s) was shortened,the number of crossing platforms ((1.40±1.75) vs (3.50±1.45)) decreased,the relative content of Aβ1-42 in hippocampus ((1.0140±0.0328) vs (0.4355±0.0152)) increased the relative content of tau protein ((0.8039±0.0251) vs (0.5170±0.0185)) increased,and Nrf2 expression levels ((0.3851±0.0188) vs (0.4901±± 0.0140)) decreased significantly,and the differences were statistically significant (t=5.110,3.625,4.068,16.010,9.208,4.474,all P＜0.01);Compared with TBI group,EL ((23.74±10.95) vs (35.31±13.23)) shortened,target quadrant dwell time ((24.28±5.37) vs (17.78±± 4.68)) shortened,the number of crossing platforms ((3.30±1.88) vs (1.40±1.75)) decreased,and the relative content of Aβ1-42 in hippocampus ((0.8370±0.0188) vs (1.0140±0.0328)) significantly decreased,the relative content of tau protein ((0.6693±0.0166) vs (0.8039±0.0251)) significantly decreased,and the expression level of Nrf2 ((0.4738 ± 0.0166) vs (0.3851 ± 0.0188)) significantly increased,and the differences were statistically significance (t=2.052,2.866,5.196,4.693,3.480,3.538,all P＜0.01).Conclusion Exendin-4 can significantly improve the learning and memory function of TBI rats,increase the expression of Nrf2,decrease the content of Aβ1-42 and tau in hippocampus,and improve the prognosis of neurological function of TBI rats.

Effect of Dl-3-n-butyphthalid on learning and memory deficiency and hippocampus synaptic long term potentiation in diabetic mice / 中华行为医学与脑科学杂志

Objective To study the effect and mechanism of Dl-3-n-butylphthalide(NBP) on mem-ory and learning ability in type 2 diabetes model db/db mice. Methods Sixteen male db/db mice were ran-domly divided into the NBP group and diabetes group,with 8 mice in each group. Eight male db/m mice of the same age were treated as the control group. NBP group was given NBP by gavage,while diabetes group and control group were given equal volume vegetable oil by gavage. After 6 weeks treatment,Morris water maze was used to examine the spatial memory and learning ability of the mice in each group. The changes of long-term potentiation ( LTP) in the hippocampus area of mice were detected by electrophysiological tech-niques. RT-PCR and Western blot were employed to measure expressions of VEGF,caspase-3,and Aβ1-42 in hippocampus of mice. Results Compared with the control group((21. 49±4. 41)s),average escape la-tency of day 5 in the diabetes group and NBP group were increased significantly ((51. 69±5. 45)s,(26. 92± 4. 76)s,t=9. 21,2. 35,both P<0. 05). Compared with the control group(7. 00±0. 60),the number of cross-ing the target platform in the diabetes group and NBP group were decreased significantly(2. 34± 0. 27), (4. 95±0. 54),t=-6. 56,-2. 49;both P<0. 05). Compared with the control group((255. 90±54. 24)%), LTP level in hippocampus of the diabetes group and NBP group were significantly decreased ( 130. 97 ± 14. 08)%,(176. 17 ± 18. 96)%,t=-4. 25,-2. 38; both P<0. 05). The relative expression of VEGF, caspase-3,and Aβ1-42 in the diabetes group and NBP group were significantly increased compared with those of the control group (t=4. 59,8. 42;7. 36,3. 85;3. 84,2. 11;all P<0. 05). Compared with the diabetes group,the escape latency of day 5,the number of crossing the target platform and LTP level were improved in the NBP group (t=-7. 25,4. 06,3. 25;all P<0. 05). The decreased expression of caspase-3,Aβ1-42 and increased expression of VEGF were reversed after NBP treatment in db/db mice (t=-4. 14,-2. 31,3. 42;all P<0. 05). Conclusion NBP might improve cognitive function and LTP by exerting anti-apoptosis effect through inhibiting the deposition of Aβ1-42 and upregulating VEGF expression in hippocampus of db/db mice.

Effect of astragaloside IV on damaged blood-brain barrier in vitro model induced by Aβ1-42 and its mechanism / 中草药

Objective To explore the effect of astragaloside IV on the damaged blood-brain barrier (BBB) in vitro model induced by amyloid-beta protein (Aβ1-42) and the underlying mechanism. Methods Firstly, the non-contact co-culture blood-brain barrier (BBB) model was established by Aβ1-42-induced mouse brain microvascular endothelial cell (bEnd.3) and primary rat astrocyte (As). Then the mice were divided into four groups: control, model (Aβ1-42 30 μmol/L), astragaloside IV low and high dose groups (Aβ1-42 30 μmol/L with Astragaloside IV 50 and 200 μmol/L). The effect of astragaloside IV on the vitality of bEnd.3 induced by Aβ1-42 was detected by methyl thiazolyl tetrazolium (MTT) assay. The permeability of BBB in vitro was determined by detecting the quantity of fluorescein sodium through BBB in various groups. In order to explore the mechanism of its protection, the apoptosis related proteins Caspase-3, cleaved Caspase-3 and tight junction proteins ZO-1, Claudin-5 and Occludin were detected by Western blotting. Results Compared with model group, the astragaloside IV groups improved the activity of bEnd.3 cells significantly (P < 0.001). The protective effect was positively correlated with the concentration of astragaloside IV. Astragaloside IV with low and high dose decreased the permeability of BBB model in vitro (P < 0.001). According to the results of Western blotting, the ratio of cleaved Caspase-3/Caspase-3 was significantly declined, and the expression levels of ZO-1, Claudin-5 and Occludin were significantly increased in astragaloside IV groups (P < 0.05). Conclusion Astragaloside IV may play a BBB protective role by inhibiting the apoptosis of bEnd.3 cells induced by fibrous Aβ1-42 and increasing the expression of tight junction proteins.

Effects of TGF-β1 on the expression and secretion of cytokines induced by Aβ in hippocampal neuron and microglia co-cultures / 中国应用生理学杂志

OBJECTIVE@#To investigate the neuroprotective effects of transforming growth factor beta 1(TGF-β1) on the expression and secretion of cytokines induced by Aβ in hippocampal neurons and microglial co-cultures.@*METHODS@#Hippocampal neurons and microglia obtained from SD rat were co-cultured. TGF-β1 was applied on day 5 after the neurons and microglia co-cultures were incubated at the concentrations of 5 or 20 ng/ml, Aβ was added 1 h following TGF-β1 application at a concentration of 5 μmol/L. They were incubated for 72 h and then assessed for further studies. Western blot analyses were employed to examine the expression of inducible nitric oxide synthase (iNOS); Real-time PCR and ELISA were used to detect the mRNA expression and secretion of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and insulin-like growth factor-1 (IGF-1).@*RESULTS@#In the hippocampal neuron-microglia co-cultures, Aβ induced upregulation of iNOS, TNF-α and IL-1β, downregulation of IGF-1. TGF-β1 pretreatment ameliorated the pro-inflammatory effects caused by Aβ.@*CONCLUSIONS@#TGF-β1 significantly inhibits the increase in inflammatory cytokines and the decrease in neurotrophic factor which are caused by Aβ-induced microglia activation.

Effects of Shenqi Yizhi Granules on PI3K/AKT Signaling Pathway in Aβ1-42 Bilateral Injection of AD Model Rats / 世界科学技术-中医药现代化

Objective: To investigate the effects of Shenqi Yizhi Granules on PI3K/AKT signaling pathway in Aβ1-42 bilateral hippocampal injection of AD model rats. Methods: A rat model of AD was established by bilateral hippocampus injection of Aβ1-42. Shenqi Yizhi granules were used for 60 days. Morris water maze was used to detect the learning and memory function of each rat. HE staining was used to observe the pathological changes of rat hippocampal CA3 area. Immunohistochemistry was used to detect the expression of PI3K and AKT protein in hippocampus. The relative expression levels of PI3K mRNA and AKT mRNA were detected by RT-PCR. Results: Compared with the blank group, the escape latency of the model group was significantly prolonged, and the number of entering the platform, the time and percentage of crossing the platform quadrant decreased significantly (P ＜ 0.05). At the same time, the vertebral cells in the hippocampal CA3 area were disordered and the neurons showed obvious lesions. PI3K/AKT showed significant inhibition at both protein and gene levels (P ＜ 0.05). After intervention with Shenqi Yizhi Granule, the escape latency of the model rats was significantly shortened, and the number of entering the platform and the time of crossing the platform quadrant were significantly increased (P ＜ 0.05). Conclusion: Shenqi Yizhi Granules can improve the cognitive function of Aβ1-42 bilateral hippocampus injection in AD rats, and its mechanism may be related to activation of PI3K/AKT signaling pathway in brain.

Effects of Angelica Polysaccharide on Learning and Memory Abilities and Aβ Metabolism in Model Rats with Alzheimer Disease / 中国中医药信息杂志

Objective To investigate effects of angelica polysaccharide on learning and memory abilities, Ach, ChAT, AChE, SOD, MDA in serum, APP and Aβ1-42 in hippocampus in model rats with Alzheimer disease (AD); To explore the mechanism of angelica polysaccharide for the treatment of AD. Methods Seventy SPF Wistar rats were selected for learning and memory ability by water maze. 10 rats were randomly selected (half female and half male) as sham-operation group, and the others were injected with Aβ25-35 by stereotatic techniques, copying AD model rats. 50 rats for learning and memory ability by water maze were successfully divided into model group, positive group, angelica polysaccharide low-, medium-, and high-dose groups, with 10 rats in each group. Rats in model group and sham-operation group were given normal saline for gavage, while rats in medication groups were given relevant medicine for gavage, 2 mL/(100 g?d), for 28 d. The learning and memory ability of rats in each group was tested by Morris water maze during 25-28 days, and the contents of Ach, ChAT, AChE, SOD, MDA in serum and APP and Aβ1-42 in hippocampus were determined. Results Compared with the sham-operation group, the escape latent period of model group was significantly prolonged in place navigation experiment; the target quadrant time was shortened; the latent time for the first time to reach the original escape platform was longer in spatial probe test; the residence time of crossing the original platform position and the target quadrant was shorter; the levels of Ach, the activity of ChAT and SOD in serum decreased; the levels of MDA, the activity of AChE in serum increased; the levels of APP and Aβ1-42 in hippocampus increased, with statistical significance (P<0.05, P<0.01). Compared with model group, the escape latent period of each medication group was shortened in different degrees after the intervention treatment; the residence time of target quadrant was prolonged; the latent time for the first time to reach the original escape platform was shortened; the number of cross platform increased; the levels of Ach, the activity of ChAT and SOD in serum increased; the levels of MDA and the activity of AChE in serum decreased; the levels of APP and Aβ1-42 in hippocampus significantly decreased, with statistical significance (P<0.05, P<0.01). Conclusion Angelica polysaccharide may effectively improve the learning and memory of ability of AD model rats to improve anti-free radical oxidation and promote Aβ metabolism and promote learning and memory ability of AD model rats, which have some preventive and therapeutic effects on AD.

Effects and mechanism of Anemarrhenae Rhizoma-Phellodendri Chinensis Cortex medicine effective parts to improve cognitive impairment in type 2 diabetic rats / 中草药

Objective To study the effects of Anemarrhenae Rhizoma-Phellodendri Chinensis Cortex medicine effective parts on phosphatidyl inositol 3 kinase (PI3K)/protein kinase B (Akt) signaling pathway and explore its mechanism of action to improve brain cognitive of type 2 diabetes rats. Methods A total of 50 male SPF SD rats, divided into five groups with 10 for each group, named as control group, model group, high/low dosage of Anemarrhenae Rhizoma-Phellodendri Chinensis Cortex medicine effective parts (ZBH and ZBL) group, and Ebixa (MHT) treatment group; The rats were immunized with ip of streptozotocin (STZ) jointly fed high-fat diet for three weeks. Then the rats were continuously gavaged with ZBH, ZBL, and MHT for 20 weeks, the serum Aβ1-42 levels were determinated in 8th and 16th week respectively, the changes of the cognitive impairment were analyzed, combined with Barnes maze and Morris water maze to detects the cognitive ability of each group rats; At the end of 20 weeks of administration, dissecting and preservatting the rat pancreatic tissue, the hippocampus to made into routine pathological sections and brain tissue pathology morphology inspection and PI3K, Akt, and Bcl-2 mRNA expression by fluorescence quantitative PCR method detection. Results After treatment for 8 weeks and 16 weeks, compared with control group, Aβ1-42 levels of model group were significantly increased (P < 0.05, 0.01); Administration 20 weeks, compared with control group, pathological section results showed normal hippocampus cells arranged in neat rows, morphological rules, and color is very even. DM models of hippocampal tissue cells arranged scattered through the administration significantly after repair. Compared with model group, ZBH and ZBL could effectively improve the form of neurons and cell arrangement of rat's hippocampus, repairing nerve injury; Compared with control group, PI3K and Akt mRNA expression of hippocampal tissue of model group rats decreased, Bcl-2 mRNA expression was abate (P < 0.05). After delivery, MHT and ZBH group could significantly improve the level of PI3K, Akt, and Bcl-2 mRNA expression (P < 0.05, 0.01). Conclusion Anemarrhenae Rhizoma-Phellodendri Chinensis Cortex medicine effective parts can significantly reduce the accumulation of Aβ1-42 protein, and decreased the expression of PI3K and Akt, indicating that the effects of improvement of cognitive impairment in type 2 diabetic rats may be achieved through regulation of PI3K/Akt pathway.

Protective effect of β-asarone on AD rat model induced by intracerebroventricular injection of Aβ₁₋₄₂ combined 2-VO and its mechanism / 中国中药杂志

This study was aimed to investigate the protective effect and mechanism of β-asarone on the animal model of Alzheimer's disease(AD) which was induced by intracerebroventricular injection of Aβ₁₋₄₂ combined cerebral ischemia. One hundred and five rats were randomly divided into seven groups including sham-operated group, AD model group, β-asarone 10 mg•kg⁻¹ group, β-asarone 20 mg•kg⁻¹ group, β-asarone 30 mg•kg⁻¹ group, donepezil group(0.75 mg•kg⁻¹) and Ginkgo biloba extract group(24 mg•kg⁻¹). Rats' learning and memory abilities, cerebric regional blood flow, pathological changes in hippocampal CA1 region, the expression level of HIF-1α and serum CAT, SOD and MDA level were detected 4 weeks later. The results showed that the application of intracerebroventricular injection of Aβ₁₋₄₂ joint 2-VO could lead to rats' dysfunction of learning and memory, decrease in regional cerebral blood flow. Neurons in CA1 region were arranged in disorder, and amyloid deposition was increased. The number of cerebral cortical cells expressing HIF-1α was increased as well. The level of serum CAT and SOD decreased, while level of serum MDA increased. However these symptoms were improved by 20 mg•kg⁻¹ and 30 mg•kg⁻¹ β-asarone. The results indicated that β-asarone could effectively relieve the symptoms of the AD model induced by intracerebroventricular injection of Aβ₁₋₄₂ combined cerebral ischemia, and the potential mechanism might be that it could attenuate damage of MDA to the body by improving the level of CAT and SOD, meanwhile the level of HIF-1α decreased as the decline of hyperoxide which might attenuate its damage to neuron, so it finally achieved alleviating Alzheimer's disease.

Protective Effect of Butylphthalide on Human Umbilical Vein Endothelial Cell Apoptosis Induced by Aβ1-42 and Its Mechanism / 中国药房

OBJECTIVE:To study the protective effect of butylphthalide on the apoptosis of human umbilical vein endothelial cells (HUVECs) induced by Aβ1-42 and its mechanism. METHODS:HUVECs were divided into normal control group,Aβ1-42 group,TAK242 group(10 nmol/L),DMSO group(1‰DMSO)and butylphthalide low-concentration,medium-concentration and high-concentration groups (40,80,160 μg/L). Except for normal control group and DMSO group,other groups were given 50 μmol/L Aβ1-42 to culture HUVECs for 24 h. TAK242 group,DMSO group and butylphthalide low-concentration,medium-concentra-tion and high-concentration groups were given relevant concentration of drugs for 30 min,with 3 holes for each concentration. The cell viability was determined by CCK-8 assay;cell apoptosis was observed by Hochest 33342/PI double staining;the cell apoptotic rate was detected by AnnexinⅤ-fluorescein isothiocyanate (FITC) flow cytometry;the protein expression of TLR-4 and COX-2 were determined by Western blot assay;the contents of IL-1 and TNF-α were detected by ELISA. RESULTS:Compared with nor-mal control group,cell viability of HUVECs were decreased in Aβ1-42 group;while apoptotic rate,protein expression of TLR4 and COX-2,the contents of IL-1 and TNF-α were increased. Compared with Aβ1-42 group,cell viability of HUVECs were increased in TAK242 group and butylphthalide low-concentration,medium-concentration and high-concentration groups;while apoptotic rate, protein expression of TLR4 and COX-2,the contents of IL-1 and TNF-α were decreased,with statistical significance(P<0.05 or P<0.01). CONCLUSIONS:Butylphthalide can reduce HUVECs apoptosis induced by Aβ1-42,which may be related with inhibiting the expression of TLR4,COX-2 and inflammatory factors.

Effects of Chrysophanol on Learning and Memory Impairment Induced by Sesflurane in Aged Rats / 医药导报

Objective To investigate the influence of inhalation anesthestic desfluran (Des) on the learning and memory abilities of rats and the protective role of chrysophanol.Methods Totally,50 male and 50 female rats,aging 24 months and weighing (500 ± 10) g,were randomly divided into five groups:normal control group,Des group,low-,mediumand high-dose Chr group (0.1,1.0 and 10.0 mg·kg-1),with 20 rats in each group.After anesthetization for 24 h,the Morris water maze was used to investigate the abilities of learning and memory of rats.The amount of Aβ1-42 was determined by ELISA assay,and the apoptosis of rat hippocampal neurons in five group was observed by TUNEL assay.Furthermore,the expression levels of Bcl-2,Bax and Caspase-3 were examined by Western blotting.The activity of acetylcholinesterase in each rats hippocampus was determined using iron trichloride chromogenic spectrophotometer colorimetric analysis method.Results Compared with the normal control group,the mean escape latency of the rats in Des group was significantly prolonged;the spatial exploring time (29.85 ± 4.51) s was reduced;the apoptotic rate of neurons (0.742 ± 0.052)％,the amount of Aβ1-42 peptide (9 618.72 ± 1 076.43) pmol· g-1,the expression levels of Caspase-3 (1.132 ± 0.217),and Bax (1.298 ± 0.209) were increased;the expression of Bcl-2 (0.318 ±0.038) were reduced;the activity of acetylcholinesterase (96.38 ±7.62) U·mL-1 was increased.Compared with the Des group,the rats in all Chr groups obtained shorter escape latency and longer spatial exploring time;the amount of Aβ1-42 peptide and the expression levels of Caspase-3 and Bax were down-regulated;the activity of acetylcholinesterase was reduced.In addition,chrysophanol improved the abilities of learning and memory of anesthetic rats in a dose-dependent manner.Conclusion Chrysophanol could improve the abilities of the learning and memory of rats after desflurane anesthesia,along with inhibition of Aβ deposition.

Effects of Dihuang Yinzi on RAGE/ROS/Apoptosis Pathway in SH-SY5Y Cells Induced by Aβ1-42 / 广州中医药大学学报

Objective To investigate the effects of Dihuang Yinzi (DY) on the receptor for advanced glycation end-products(RAGE)/reactive oxygen species(ROS)/apoptosis pathway in SH-SY5Y cells induced by amyloid-beta1-42 (Aβ1-42) oligomer. Methods Firstly, we adopted methyl thiazolyl tetrazolium(MTT) method to detect the cell vitality in fetal bovine serum (FBS) group, blank serum group, and low-, middle- and high- dose DY-containing serum groups, so as to confirm the optimal concentration and treatment time of DY-containing serum. Secondly, we applied MTT method to detect cell vitality and applied Annexin V/propidium iodide (PI) staining method to observe the apoptosis of SH-SY5Y cells treated with 0~20 μmol/L Aβ1-42 for 24 and 48 h, so as toconfirm the optimal concentration and treatment time of Aβ1-42 for establishing Alzheimer's disease (AD) model in vitro. Thirdly, MTT method was used for the detection of cell vitality, and Annexin V/PI staining method was used for detection of the apoptosis of SH-SY5Y cells in blank serum group, model group, western medicine control group and low-, middle-and high-dose DY-containing serum groups, and Dihydroethidium (DHE) method was used for the assay of ROS contents, so as to observe the effect of DY on the recovery of injured SH-SY5Y cells induced by Aβ1-42. Finally, we applied Western blot method to detect the expression level of RAGE in SH-SY5Y cells of blank group, model group and DY-containing serum group; after Aβ1-42-induced SH-SY5Y cells were transfected with RAGE gene, we adopted DHE staining method and Annexin V/PI staining method to detect ROS content and cell apoptotic rate in all of the above groups, so as to observe the effect of DY on SH-SY5Y cell apoptosis and RAGE expression. Results The cell vitalities were increased in low- and middle-dose DY-containing serum groups at 24 h (P < 0.05 or P < 0.01 compared with that in the blank serum group). The conditions for the establishment of AD model in vitro were as follows: the optimal concentration of Aβ1-42 was 5μmol/L, and the treatment time was 24 h. The cell vitalities were significantly enhanced, the cell apoptotic rate and ROS content were significantly lowered in Aβ1-42-induced SH-SY5Y cells of the medication groups(P <0.05 or P < 0.01 compared with those in the model group) , and the cell vitality was the highest and the cell apoptotic rate was the lowest in the middle-dose DY-containing serum group. The RAGE expression level was decreased in Aβ1-42-induced SH-SY5Y cells of the middle-dose DY-containing serum group(P < 0.05 compared with that in the model group) . ROS content and cell apoptotic rate were decreased in Aβ1-42-induced SH-SY5Y cells transfected with RAGE gene in the middle-dose DY-containing serum group (P<0.01). Conclusion DY may play an anti-oxidative role through inhibiting the production of ROS and cell apoptosis, thus to suppress RAGE protein and to achieve the preventive and therapeutic effect for AD.

Neuroprotective effect of LSS on Aβ1-42-induced BV-2 microglial cells through the inhibition of neuroinflammation / 中国药理学与毒理学杂志

OBJECTIVE To investigate the effect of neuroprotective effect of lychee seed saponins (LSS) in BV-2. METHODS Aβ1-42 induced BV-2 cells were incubated with LSS for 12 h, the content of the inflammatory factors such as IL-1β, TNF-α, COX-2 and iNOS in the supernatant of BV-2 cell were measured by ELISA. The detection of the mRNA levels and the protein expression of the inflammatory factors including IL-1β, TNF-α, COX-2 and iNOS using real-time PCR and Western blotting, respectively. RESULTS The level of IL-1β, COX-2 and iNOS significantly increased with the treatment of Aβ1-42, and 0.117 mg·L-1-0.469 mg·L-1 LSS can inhibit these increased level. CONCLUSION LSS conferred neuroprotection via inhibiting the inflammatory factors expression.