Pathological Changes In Hippocampus Research Articles

Folic acid and S-adenosylmethionine reverse Homocysteine-induced Alzheimer’s disease-like pathological changes in rat hippocampus by modulating PS1 and PP2A methylation levels

BackgroundAbnormally elevated homocysteine (Hcy) is recognized as a biomarker and risk factor for Alzheimer’s disease (AD). However, the underlying mechanisms by which Hcy affects AD are still unclear. ObjectivesThis study aimed to elucidate the effects and mechanisms by which Hcy affects AD-like pathological changes in the hippocampus through in vivo and in vitro experiments, and to investigate whether folic acid (FA) and S-adenosylmethionine (SAM) supplementation could improve neurodegenerative injuries. MethodsIn vitro experiments hippocampal neurons of rat were treated with Hcy, FA or SAM for 24 h; while the hyperhomocysteinemia (HHcy) in Wistar rats was established by intraperitoneal injection of Hcy, and FA was added to feed. The expression of β-amyloid (Aβ), phosphorylated tau protein, presenilin 1 (PS1) at the protein level and the activity of protein phosphatase 2A (PP2A) were detected, the immunopositive cells for Aβ and phosphorylated tau protein in the rat hippocampus were also evaluated by immunohistochemical staining. ResultsFA and SAM significantly repressed Hcy-induced AD-like pathological changes in the hippocampus, including the increased tau protein phosphorylation at Ser214, Ser396 and the expression of Aβ42. In addition, Hcy-induced PS1 expression increased at the protein level and PP2A activity decreased, while FA and SAM were able to retard that. ConclusionsThe increase in PS1 expression and decrease in PP2A activity may be the mechanisms underlying the Hcy-induced AD-like pathology. FA and SAM significantly repressed the Hcy-induced neurodegenerative injury by modulating PS1 and PP2A methylation levels.

Salidroside prevents gestational hypertension-induced impairment of offspring learning and memory via Wnt/Skp2 pathway

BackgroundSalidroside (Sal) has been found to protect against multiple impairments caused by diabetes, and we designed this study to investigate the effect of Sal on gestational hypertension (GHP)-induced impairment of offspring learning and memory. MethodsWe established a GHP rat model by intraperitoneal injection of NG-nitro-L-arginine methyl ester (L-NAME), and treated with Sal by daily gavage. We used Morris Water Maze test to evaluate the learning and memory ability of offspring rats. HE staining was used to measured the pathological changes in hippocampus of offspring. Immunohistochemistry, cellular immunofluorescence and western blot were used to detect the protein expression. ResultsThe learning and memory abilities of GHP offspring rats were significantly lower than those of normal rat offspring, while Sal treatment could significantly improve the learning and memory abilities of GHP offspring rats. HE staining did not reveal pathological differences in the hippocampus of normal rats, GHP rats and Sal-treated GHP offspring rats. However, Sal treatment can significantly increase the expression of Wnt1 and Skp2 protein, and decrease the expression of P27kiwf and P21waf1 protein in the hippocampus of GHP offspring rats. In vitro, Sal significantly promoted the proliferation and differentiation on neural stem cell, while Wnt1 knockdown could reverse these promotions by Sal. In the hippocampus of GHP offspring rats, Sal treatment significantly increased the expression of Tuj1, SOX2, Ki67 and DCX protein. ConclusionSalidroside significantly improves the learning and memory impairment of offspring caused by GHP, and its mechanism may be related to the fact that Salidroside promotes the proliferation and differentiation of neural stem cells by activating the Wnt1/Skp2 signaling pathway.

Electroacupuncture improves cognitive dysfunction in rats with Alzheimer's disease by regulating microglial cells.

To observe the effect of electroacupuncture (EA) on microglia (MG), Janus kinase-2 (JAK2) and signal transducer and activator of transcription-3 (STAT3) in hippocampal CA1 region of Alzheimer's di-sease (AD) rats, so as to explore its mechanisms in the treatment of AD. Thirty-six male SD rats were randomly divided into sham operation, model and EA groups, with 12 rats in each group. The AD rat model was established by intraperitoneal injection of D-galactose combined with intrahippocampal injection of aggregated Aβ25-35. The rats in the EA group were given EA (2 Hz/20 Hz, 2 mA) at "Baihui"(GV20) and"Shenting"(GV24) for 30 min, once daily, 6 days a week for 4 weeks. Morris water maze test was used to detect the learning and memory ability and spatial exploration ability of rats. HE staining was used to observe the pathological changes of hippocampus. The ultrastructure of hippocampal neurons was observed by transmission electron microscopy. The positive expression of MG marker io-nized calcium adaptor protein (Iba-1) in hippocampus was observed by immunofluorescence staining. The expression levels of serum inflammatory factor interferon-γ (IFN-γ) and transforming growth factor beta 1 (TGF-β1) were detected by ELISA. The mRNA expression levels of JAK2, STAT3, inducible nitric oxide synthase (iNOS) and arginase-1 (Arg-1) in hippocampal CA1 region were detected by real-time quantitative PCR. The protein and phosphorylation levels of JAK2 and STAT3 in hippocampal CA1 region were detected by Western blot. Compared with the sham operation group, the escape latency of the model group was significantly prolonged (P<0.01), and the number of crossing the original platform was significantly reduced (P<0.01), the positive expression of Iba-1 in CA1 region, the content of serum IFN-γ, the relative mRNA expressions of JAK2, STAT3 and iNOS, and the protein and phosphorylation levels of JAK2 and STAT3 were significantly increased (P<0.01), while the content of serum TGF-β1 and the relative expression of Arg-1 mRNA were significantly decreased (P<0.01). Compared with the model group, the escape latency of rats in the EA group was significantly shortened (P<0.01), the number of crossing the original platform was significantly increased (P<0.01), the positive expression of Iba1, the content of serum IFN-γ, the mRNA expressions of JAK2, STAT3 and iNOS, and the protein and phosphorylation levels of JAK2 and STAT3 were significantly decreased (P<0.05, P<0.01), while the content of serum TGF-β1 and the expression of Arg-1 mRNA were significantly increased (P<0.01). Moreover, pathological and ultrastructural observation showed a reduction in the number of hippocampal neurons, changement of nuclear morphology, dilation of intercellular space, and decreased number of mitochondria in the model group；these situations were relatively milder in the EA group. EA can improve the learning and memory function of AD rats, which may be associated with its functions in decreasing MG activities, and inhibiting the JAK2 / STAT3 signaling pathway in the hippocampus.

Downregulating NHE-1 decreases the apoptosis of hippocampal cells in epileptic model rats based on the NHE-1/calpain1 pathway

Seizure is associated with pathological changes of hippocampus, but the mechanism by which hippocampal neuronal apoptosis promotes epilepsy is unclear. Our previous study showed that the expression of NHE-1 was increased in epileptic model rats. Therefore, this study further explores the effect of NHE-1 on hippocampal cells apoptosis and seizure in lithium chloride-pilocarpine epileptic model rats. First, we established a lithium chloride-pilocarpine induced epileptic rat model and detected the expression of NHE-1, calpain1 and apoptosis in the hippocampus. Then, we further down-regulated NHE-1 to observe the expression of calpain1 and apoptosis in the hippocampus, as well as its effect on seizures in rats. We found that the expression of NHE-1 and calpain1 and apoptosis in the hippocampus was significant increased in the model group. After down-regulating NHE-1, the expression of calpain1 was decreased, and hippocampal cell apoptosis was alleviated. In addition, down-regulation of NHE-1 reduced the frequency and duration of seizures in epileptic rats. Therefore, hippocampal NHE-1 overexpression is closely related to the development of neuronal apoptosis in a rat model of epilepsy, and downregulating NHE-1 expression can reduce cell apoptosis. Moreover, the NHE-1/calpain1 signaling pathway may be an important mechanism leading to hippocampal cell apoptosis.

Echinacoside exerts antidepressant-like effects through enhancing BDNF-CREB pathway and inhibiting neuroinflammation via regulating microglia M1/M2 polarization and JAK1/STAT3 pathway.

The present study was performed to investigate the antidepressant effect of echinacoside (ECH) using chronic unpredictable mild stress (CUMS) induced depression mice and lipopolysaccharide (LPS)-stimulated N9 microglial cells. CUMS treatment was performed on C57BL/6 mice for 28days, followed by gavaging with different doses of echinacoside (15 and 60mg/kg) for 21 consecutive days. Sucrose preference test (SPT), open field test (OFT), tail suspension test (TST), and forced swimming test (FST) were measured to assess the effects of echinacoside on CUMS-Induced Depressive-Like Behaviors. After that, the pathological changes of hippocampus were determined by Hematoxylin and eosin (HE) staining and Nissl staining; the neurotransmitters, pro-inflammatory cytokines and indoleamine 2,3-dioxygenase (IDO) levels, and the hypothalamic-pituitary-adrenal (HPA) axis activity were determined by enzyme linked immunosorbent assay (ELISA); Iba 1were evaluated by Immunofluorescence assay; Key protein expression levels of CREB/BDNF signal pathway were measured by western blotting. Subsequently, N9 cells were stimulated with 1μg/ml LPS to induce N9 microglia activation, and were treated with 5-20μM of echinacoside for 24h. After that, the levels of NO, interleukin (IL)-1β, IL-6, tumor necrosis factor alpha (TNF-α), IL-4, IL-10, and transforming growth factor beta (TGF-β) in N9 cell culture supernatants were measured by enzyme-linked immunosorbent assay (ELISA) kits; morphology and Iba 1 expression level were observed by high-content screening assay; the M1 markers of CD11b, CD86 and M2 markers of CD206 were analyzed by imaging flow cytometry. Results show that treatment with echinacoside reversed CUMS-increased immobility time in OFT, TST, FST and reversed CUMS-reduced sucrose preference in SPT. In addition, echinacoside reduced the levels of pro-inflammatory cytokines and Iba 1. Moreover, echinacoside significantly increased p-CREB/CREB ratio and BDNF level in hippocampus. Furthermore, echinacoside reduced the secretion of inflammatory factors and inhibited microglia M1 polarization in N9 cells. In conclusion, echinacoside may be beneficial for the treatment of depression diseases through regulating the microglia balance by inhibiting the polarization of microglia to M1 phenotype, and improving hippocampal neurogenesis by the CREB-BDNF signaling pathway.

Effects of electroacupuncture on the expression of β-amyloid and autophagy-related proteins in hippocampal cells of mice with Alzheimer's disease

To investigate the effect of electroacupuncture (EA) on the expression of β-amyloid (Aβ) and autophagy-related proteins in hippocampal cells of Alzheimer's disease (AD) model mice, so as to explore its underlying mechanisms. Eighteen male APP/PS1 transgenic mice (6 months old) were randomly divided into model and EA groups, with 9 mice in each group. Nine male C57BL/6 wild-type mice of the same age were chosen as the normal group. Mice in the EA group were treated with acupuncture on "Baihui" (GV20) and EA (1 Hz/50 Hz, 1 mA) on bilateral "Yongquan" (KI1), once every other day, 20 min each time for a total of 21 times. After the interventions, the spatial learning and memory ability were observed by Morris water maze test. The autophagy-related pathological changes in hippocampus were observed by transmission electron microscopy. The expressions of microtublue associated protein 1 light chain 3 (LC3) and Aβ in hippocampus were observed by immunofluorescence and the expression levels of LC3 and p62 proteins were detected by Western blot. Compared with the normal group, the escape latency was prolonged (P<0.01), the residence time in the original quadrant platform was shor-tened (P<0.05), the positive expressions of LC3 and Aβ, the expression levels of LC3Ⅱ and p62 proteins, and the ratio of LC3Ⅱ/LC3Ⅰ proteins in hippocampus were increased (P<0.01, P<0.05) in the model group. Compared with the model group, the escape latency was shortened (P<0.05), the residence time in the original quadrant platform was prolonged (P<0.05), the positive expressions of LC3 and Aβ, the expression levels of LC3Ⅱ and p62 proteins, and the ratio of LC3Ⅱ/LC3Ⅰ proteins in hippocampus were decreased (P<0.05) in the EA group. The transmission electron microscopy showed that the structure of neurons was normal in the normal group, a large number of autolysosomes and autophagosomes existed in hip-pocampal nerve cells in the model group, and only a small number of autophagosomes were observed in the EA group. EA can reduce the expression levels of autophagy-related proteins LC3 and p62 in APP/PS1 transgenic mice, improve the hip-pocampal autophagy state, reduce intracellular Aβ aggregation, and thus improve the learning and memory ability.

Prenatal exposure to titanium dioxide nanoparticles induces persistent neurobehavioral impairments in maternal mice that is associated with microbiota-gut-brain axis.

Gestational exposure to titanium dioxide nanoparticles (TiO2NPs) has been widely reported to have deleterious effects on the brain functions of offspring. However, little attention has been paid to the neurotoxic effects of TiO2NPs on maternal body after parturition. The pregnant mice were orally administrated with TiO2NPs at 150mg/kg from gestational day 8-21. The potential effects of TiO2NPs on the neurobehaviors were evaluated at postnatal day 60. The gut microbiota, morphological alterations of intestine and brain, and other indicators that involved in gut-brain axis were all assessed to investigate the underlying mechanisms. The results demonstrated that exposure to TiO2NPs during pregnancy caused the persistent neurobehavioral impairments of maternal mice after delivery for 60 days, mainly including behavioural changes, pathological changes in hippocampus, cortex and intestine. Our data also showed that persistent dysfunction and tissue injuries were probably associated with the disruption of gut-brain axis, manifested by the shift in the composition of gut microbial community, alteration of Sstr1, inhibition of enteric neurons and reduction of diamine oxidase contents in maternal mice. These findings provide a novel insight that regulation of gut microecology may be an alternative strategy for the protection against the neurotoxicity of TiO2NPs in pregnant women.

FKN/NR Signaling Pathway Regulates Hippocampal Inflammatory Responses: the Survival of Hippocampal Neurons in Diabetic Rats with Chronic Unpredictable Mild Stress.

Aim To investigate the mechanism via which FKN/CX3CR1 signaling abnormalities mediate N-methyl-D-aspartic acid receptor (NMDA) overexcitation-induced hippocampal neuronal injury in diabetic rats complicated with depression (DD). Methods Sixty rats were randomly divided into 5 groups. The depression-like behaviors of the rats were evaluated by open field test and Morris water maze. The pathological changes of hippocampus in DD rats were observed by HE staining. The blood levels of inflammatory factors (IL-1β, TNF-α, and IL-6) and neurotransmitters (D-serine and glutamic acid) were determined by enzyme-linked immunosorbent assay (ELISA). The expressions of BDNF, A1 receptor (A1R), A2 receptor (A2R), A3 receptor (A3R), calmodulin dependent kinase II (CaMKII), CX3CR1, CX3CL1 (FKN), NR2A, and NR2B proteins were detected by immunohistochemistry and Western-blotting. Results Compared with the normal control group, blood glucose level increased significantly and body weight decreased in T2DM group and T2DMC group. In addition, the number of spontaneous activities significantly decreased and the capability of learning and memory was attenuated in T2DMC group and Chronic Stress group. The blood levels of IL-1β, TNF-α, IL-6, glutamate (Glu), and D-serine significantly increased in each model group. After intervention with CX3CR1 antibody, the expressions of BDNF, CaMK II, A1R, and A3R increased and those of A2R, CX3CR1, FKN, NR2A, and NR2B decreased. Conclusion In the diabetic state, the binding of FKN to CX3CR1 increases, which regulates a variety of adenosine receptors. When it exerts its effect on neurons, the overactivation of NR results in neuronal injury and causes depression.

Proteomic Analysis Reveals the Vital Role of Synaptic Plasticity in the Pathogenesis of Temporal Lobe Epilepsy.

Temporal lobe epilepsy (TLE) is a chronic neurological disorder that is often resistant to antiepileptic drugs. The pathogenesis of TLE is extremely complicated and remains elusive. Understanding the molecular mechanisms underlying TLE is crucial for its diagnosis and treatment. In the present study, a lithium-pilocarpine-induced TLE model was employed to reveal the pathological changes of hippocampus in rats. Hippocampal samples were taken for proteomic analysis at 2 weeks after the onset of spontaneous seizure (a chronic stage of epileptogenesis). Isobaric tag for relative and absolute quantization (iTRAQ) coupled with liquid chromatography-tandem mass spectrometry (LC–MS/MS) technique was applied for proteomic analysis of hippocampus. A total of 4173 proteins were identified from the hippocampi of epileptic rats and its control, of which 27 differentially expressed proteins (DEPs) were obtained with a fold change > 1.5 and P < 0.05. Bioinformatics analysis indicated 27 DEPs were mainly enriched in “regulation of synaptic plasticity and structure” and “calmodulin-dependent protein kinase activity,” which implicate synaptic remodeling may play a vital role in the pathogenesis of TLE. Consequently, the synaptic plasticity-related proteins and synaptic structure were investigated to verify it. It has been demonstrated that CaMKII-α, CaMKII-β, and GFAP were significant upregulated coincidently with proteomic analysis in the hippocampus of TLE rats. Moreover, the increased dendritic spines and hippocampal sclerosis further proved that synaptic plasticity involves in the development of TLE. The present study may help to understand the molecular mechanisms underlying epileptogenesis and provide a basis for further studies on synaptic plasticity in TLE.

Galangin alleviates learning and memory impairments in APP/PS1 double- transgenic mice by regulating Akt/MEF2D/Beclin-1 signaling pathway

The study aimed to investigate the effects of galangin on learning and memory impairments and Akt/MEF2 D/Beclin-1 signaling pathway in APP/PS1 double-transgenic mice. The mice in this experiment were divided into the normal group, model group, low-(25 mg·kg~(-1)), medium-(50 mg·kg~(-1)), and high-dose(100 mg·kg~(-1)) galangin groups, donepezil(3 mg·kg~(-1)) group, Akt inhibitor(25 mg·kg~(-1)) group, and autophagy inhibitor(30 mg·kg~(-1)) group, with ten in each group, and administered with the corresponding drugs for 30 successive days. On the 24 th day of medication, the water maze and dark avoidance tests were performed. The levels of p-tau, β-amyloid peptide 1-42(Aβ_(42)), acetylcholinesterase(AChE), β-site amyloid precursor protein cleaving enzyme 1(BACE1), and amyloid precursor protein(APP) in hippocampus were detected by ELISA, the Beclin-1 mRNA expression by RT-PCR, the expression of Aβ_(42) and glial fibrillary acidic protein(GFAP) by immunohistochemistry, and the expression of myocyte enhancer factor 2 D(MEF2 D) by immunofluorescence assay. The pathological changes in hippocampus were observed after HE staining, and the expression of Akt, MEF2 D, and Beclin-1 in hippocampus were assayed by Western blot. These results showed that compared with the normal group, the model group exhibited prolonged swimming time, increased number of errors and electric shocks, up-regulated p-tau, Aβ_(42), APP, AChE, BACE1, GFAP, and Beclin-1, shortened incubation period, decreased p-Akt and MEF2 D, and obvious hippocampal injury. Compared with the model group, donepezil and galangin shortened the swimming time, reduced the number of errors and electric shocks, down-regulated the expression of p-tau, Aβ_(42), APP, AChE, BACE1, GFAP, and Beclin-1, prolonged the incubation period, up-regulated p-Akt and MEF2 D, and improved the pathological changes in hippocampus. Compared with the autophagy inhibitor group, galangin prolonged the swimming time, elevated the number of errors and electric shocks, enhanced the expression of p-tau, Aβ_(42), APP, AChE, BACE1, GFAP, and Beclin-1, shortened the incubation period, and diminished the expression of p-Akt and MEF2 D. In conclusion, galangin improves the learning and memory impairments and hippocampal neuron injury of APP/PS1 mice, which may be related to its regulation of Akt/MEF2 D/Beclin-1 signaling pathway.

Influence of miR-1 on Nerve Cell Apoptosis in Rats with Cerebral Stroke via Regulating ERK Signaling Pathway.

To explore the effect of miR-1 on neuronal apoptosis in rats with stroke through the ERK signaling pathway. Methods. Forty male rats (180-220 g) were selected and randomly divided into the sham, model, miR-1 inhibitor, and miR-1 mimic groups (10 rats per group) by average body weight. Cerebral ischemia/reperfusion (I/R) models were established using a modified middle cerebral artery wire thrombosis (MCAO) method in rats in the model group, miR-1 inhibitor group, and miR-1 mimic group. After the successful model establishment, the miR-1inhibitor group and miR-1 mimic group were intravenously injected with miR-1 inhibitor and miR-1 mimic, respectively, once a day for 3 days. The sham and model groups were given the same dose of normal saline. TTC staining was applied to detect the cerebral infarct size and calculate the infarct volume. Histopathological changes in the hippocampus of rat brains were observed by HE staining. Flow cytometry was used to detect neuronal apoptosis in rat brains. The mRNA expressions of miR-1, ERK1/2, Bcl-2, and Bax in rat brain tissues were determined by QRT PCR, and the protein levels of ERK1/2, Bcl-2, Bax, and caspase-3 were determined by Western blot analysis. Results. Compared with the sham group, the neurological impairment score, cerebral infarct size, and volume of rats in the model group were significantly increased (p < 0.05). Compared with the model group, the neurological impairment score, cerebral infarct size, and volume were significantly increased in the miR-1 mimic group and significantly decreased in the miR-1 inhibitor group (p < 0.05). In the model group, the hippocampal tissue of rats had malaligned cells, neuron cell atrophy became smaller, the intercellular spaces became larger, and vacuoles appeared. Compared with the model group, the miR-1 inhibitor group could effectively alleviate the pathological changes in the hippocampus, and the miR-1 mimic group could significantly add to the pathological changes in the rat hippocampus. Compared with the sham group, the mRNA expression of miR-1 and Bax in the brain of model rats increased significantly (p < 0.05), and the mRNA expression of ERK1/2 decreased significantly; Compared with the model group, the miR-1 and Bax mRNA expressions in the brain tissues of rats in the miR-1 inhibitor group were significantly decreased, the ERK1/2 and bcl-2 mRNA expressions were significantly increased, and the miR-1 and Bax mRNA expressions in the brain tissues of rats in the miR-1 inhibitor group were significantly decreased, and the Bcl-2 mRNA expression was significantly increased (p < 0.05). Compared with the sham group, neuronal apoptosis was increased in the brain tissues of rats in the model group and miR-1 mimic group. Compared with the model group, neuronal apoptosis was decreased in the brain tissues of rats in the miR-1 inhibitor group. Compared with the sham group, the ERK1/2 proteins in the model group were significantly decreased, the Bcl-2, Bax, and caspase-3 proteins were significantly increased, and the ERK1/2, Bcl-2, Bax, and caspase-3 proteins in the miR-1 inhibitor group and miR-1 mimic group were significantly increased. Compared with the model group, the protein levels of ERK1/2 and Bcl-2 in the miR-1 inhibitor group were significantly increased, the proteins of Bax and caspase-3 were significantly decreased, and the protein levels of ERK1/2 and Bcl-2 in the miR-1 inhibitor group were significantly increased (p < 0.05). Conclusions. miR-1 can interfere with neuronal apoptosis in rats with stroke through the ERK signaling pathway.

Effect of acupuncture on hippocampal inflammation in rats with intrauterine distress induced hypoxic-ischemic brain injury

To observe the effects of acupuncture on the behavior, serum inflammatory cytokines, hippocampal Nod-like receptor protein 3(NLRP3) inflammasome-related proteins and microglia activation in rats with intrauterine distress induced ischemia-hypoxic brain injury (HIBD), so as to explore the mechanism of acupuncture underlying treatment of HIBD by regulating hippocampal inflammation. The HIBD model was established by clipping bilateral uterine arteries for 10 min and delaying caesarean delivery. HIBD rats were randomly divided into model group and acupuncture group according to body weight and litter factors, with 12 rats in each group. Twelve rats delivered normally were regarded as the normal group. Acupuncture was applied to bilateral "Benshen"(GB13) for 10 min, once a day for 14 consecutive days. H.E. staining was used to observe the pathological changes of hippocampus. The three chamber sociability test was used to determine the rat's social ability. The serum IL-1β and IL-18 levels, the hippocampal expressions of NLRP3, apoptosis associated speck like protein containing CARD(ASC) and cysteinyl aspartate specific proteinase-1(Caspase-1), andionized calcium bindingadaptor molecule-1(IBA-1) were detected by ELISA，Western blot and Immunofluorescence, separately. Compared with the normal group, The hippocampal neurons in the model group are arranged loosely, and the cells are solidified and shrunk and the whole cells are deeply stained. The time of exploring the stranger rat in the model group was significantly reduced (P<0.01). The contents of IL-1β and IL-18, the expressions of NLRP3, ASC, Caspase-1 and IBA-1 of the model group were higher than those of the normal group (P<0.01, P<0.05, P<0.001). Following acupuncture intervention, the cell structure of hippocampal neurons was improved, and there were fewer cases of cell solidification, atrophy, and deep staining. In comparison with the model group, the time of exploring stranger rat in the acupuncture group was significantly increased (P<0.01). The contents of IL-1β and IL-18, the expressions of NLRP3, ASC and Caspase-1, and IBA-1 of the acupuncture group were significantly down-regulated than those of the model group (P<0.01, P<0.05). Acupuncture can effectively improve the social behavior deficits of HIBD rats，which may be related to inhibition of the expression of inflammatory response proteins NLRP3, ASC and Caspase-1 in the hippocampus, decrease the activation of microglia in the hippocampus, and reduce serum IL-1β and IL-18, thereby alleviating inflammation.

Effects of Moringa flavone on cognitive function and neuropathological indexes in diabetic encephalopathy rats

To investigate the effect of Moringa flavone on cognitive impairment and neuropathological indexes in diabetic encephalopathy (DE) rats. Sixty male Sprague-Dawley (SD) rats were divided into control group, model group, positive drug group, Moringa low-dose and high-dose groups according to the random number table method, with 10 rats in each group. Diabetic rat model was established by intraperitoneal injection of 25 mg/kg streptozotocin (STZ) after continuous feeding of high fat and high sugar diet for one week. Blood was collected from the tail vein after 72 hours, the mean value of twice random blood glucose was ≥ 16.67 mmol/L, and the continuous positive urine glucose showed that the diabetes model was successfully prepared. The control group was fed with conventional feed. After successful model establishment, the rats in the Moringa low and high dose groups were given 4.0 g/kg and 8.0 g/kg Moringa extract (Moringa flavone) by gavage everyday, the rats in the positive drug group were given piracetam 0.48 g/kg, and the rats in the model group and control group were given the same amount of normal saline once a day for 30 days. Morris water maze was used to evaluate the cognitive impairment of the rats. The hippocampus of the rats was harvested 12 hours after the last administration, and the advanced glycation end product receptor (RAGE) and nuclear factor-κB (NF-κB) were detected by immunohistochemistry. The contents of acetylcholinesterase (AChE), advanced glycation end product (AGE) and choline acetyl transferase (ChAT) were detected by enzyme linked immunosorbent assay (ELISA). Compared with the control group, the escape latency and the exploration distance in model group were extended, target quadrant stay time was shortened, the levels of AChE and AGE in brain tissue were significantly increased, and ChAT level was significantly decreased. Morris water maze experiment showed that compared with the model group, in the Moringa low and high dose groups from the 3rd day, the escape latency (s: 35.07±7.21, 33.14±5.35 vs. 43.09±9.83, both P < 0.05) and the exploration distance (m: 8.32±4.23, 8.10±4.97 vs. 13.02±3.67) were significantly shortened (both P < 0.05). The target qauadrant stay time was extended (s: 35.12±3.12, 41.53±8.37 vs. 23.15±4.89, both P < 0.01). The results of ELISA showed that compared with the model group, the levels of AChE and AGE in brain tissue of the Moringa low and high dose groups were significantly decreased [AChE (U/L): 180.22±12.03, 142.67±20.56 vs. 205.27±25.14, AGE (μg/L): 439.10±25.19, 428.27±19.14 vs. 501.28±21.53, all P < 0.05], and the levels of ChAT were significantly increased (U/L: 51.95±5.27, 53.13±5.04 vs. 37.91±5.10, both P < 0.01). There were no significant differences in AChE, AGE or ChAT between the Moringa low and high dose groups. The results of immunohistochemistry showed that the number of RAGE and NF-κB positive cells in DG area of hippocampus increased significantly, and the average gray values of RAGE and NF-κB decreased significantly. Compared with the model group, the RAGE and NF-κB positive cells in the Moringa low and high dose groups were significantly reduced, and the average gray values of RAGE and NF-κB in hippocampus were significantly increased [RAGE (gray value): 110.46±10.04, 117.76±8.64 vs. 92.19±8.76, NF-κB (gray value): 109.40±8.93, 116.59±7.26 vs. 90.74±13.27, all P < 0.05]. There were no significant differences in the expressions of RAGE or NF-κB between the Moringa low and high dose groups. Moringa flavonoids could obviously improve the cognitive dysfunction and memory ability of DE model rats, improve the pathological changes of hippocampus, and have a certain protective effect on brain.

Carnosic acid alleviates depression-like behaviors on chronic mild stressed mice via PPAR-γ-dependent regulation of ADPN/FGF9 pathway.

The pathway of adiponectin (ADPN)/fibroblast growth factor 9 (FGF9) was recently thought as a key role in the development of depression. ADPN is crucially regulated by peroxisome proliferator-activated receptor-gamma (PPAR-γ). Natural material carnosic acid (CA) has been applied for therapeutics of mental disorders. To evaluate the antidepressive effect of CA in stress-treated mice and define whether its effects is involved in the regulation of ADPN/FGF9 pathway. In vivo study, the levels of ADPN and FGF9 in both serum and hippocampus tissues, the expressions of ADPN receptor 2 (AdipoR2) in hippocampus and PPAR-γ in abdominal adipose, as well as the pathological changes of hippocampus were determined in 28-day period of chronic unpredictable mild stress (CUMS)-induced depression model of male ICR (Institute of Cancer Research) mice or adipo-/- mice. In vitro study, the level of ADPN and the mRNA expressions of both ADPN and PPAR-γ were determined in mouse 3T3-L1 preadipocytes. In vivo study, treatment with CA (50 or 100 mg/kg per day) for 21 days markedly suppressed depressive-like behaviors, the elevating levels of FGF9 and decreasing levels of ADPN in both serum and hippocampus tissues, the downregulating protein and mRNA expressions of AdipoR2 in hippocampus and PPAR-γ in abdominal adipose, as well as the pathological injury of hippocampus induced by CUMS in male ICR mice. The antidepressive effects of CA were markedly attenuated in male CUMS-treated adipo-/- mice. In vitro study, incubation with CA (3-30 μmol/L) for 24 h could concentration-dependently upregulate the mRNA expressions of both PPAR-γ and ADPN as well as increase the level of ADPN. The experiments using PPAR-γ-specific inhibitor GW9662 and transient transfection with mutated PPAR-γ-binding site promotor constructs showed that the activation of PPAR-γ mediated CA-induced ADPN expression in adipocytes. CA could significantly improve stress-induced depressive disorder, which may be related to regulating the dysfunction of ADPN-FGF9 pathway via activating PPAR-γ in adipocytes.

Dexmedetomidine alleviates sevoflurane-induced neurotoxicity via mitophagy signaling.

Dexmedetomidine, a class of α2-adrenergic agonist, was reported to exert a neuroprotective effect on sevoflurane-induced neurotoxicity. However, the specific mechanisms have not been fully clarified yet. The aim of our study is to uncover the role of dexmedetomidine in sevoflurane-induced neurotoxicity.The rats pretreated with dexmedetomidine and/or Rapamycin 3-Methyladenine were housed in a box containing 30% O2, 68% N2 and 2% sevoflurane for 4 h for anesthesia. 24 h after drug injection, Morris water maze test was used to evaluate rats' learning and memory ability. Hematoxylin & eosin (H&E) staining was adopted to analyze the pathological changes of hippocampus. TUNEL assay was performed to measure cell apoptosis in hippocampus. Immunofluorescent assay was utilized to detect HSP60 level. The protein levels of LC3I, LC3II, Beclin-1, CypD, VDAC1 and Tom20 were examined by western blot. 5 weeks after drug injection, Morris water maze test was used to evaluate rats' learning and memory ability again. Dexmedetomidine alleviated sevoflurane-induced nerve injury and the impairment of learning and memory abilities. Additionally, dexmedetomidine inhibited sevoflurane-induced cell apoptosis in hippocampus. In mechanism, dexmedetomidine activated mitophagy to mitigate neurotoxicity by enhancing LC3II/LC3I ratio, HSP60, Beclin-1, CypD, VDAC1 and Tom20 protein levels in hippocampus.Dexmedetomidine alleviates sevoflurane-induced neurotoxicity via mitophagy signaling, offering a potential strategy for sevoflurane-induced neurotoxicity treatment.

MiR-126 suppresses neuronal apoptosis in rats after cardiopulmonary resuscitation via regulating p38MAPK

In this study, we aimed to evaluate the effect of microRNA-126 (miR-126) on neuronal apoptosis in cardiopulmonary resuscitation rats and to explore the related molecular mechanism. The expression of miR-126 in brain tissues of rats after cardiopulmonary resuscitation was measured by real-time fluorescence quantitative polymerase chain reaction (RT-qPCR). The basic parameters of cardiopulmonary resuscitation were recorded by miR-126 mimic injection in rats after cardiopulmonary resuscitation. Hematoxylin-eosin staining was used to observe the pathological changes of hippocampus. Immunohistochemistry was used to observe the expression of p38 and caspase-3 protein. Furthermore, the expression of p38 mitogen-activated protein kinase (p38MAPK), Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase 1/2 (ERK1/2) in rat hippocampus was detected by RT-qPCR and Western blot. In order to confirm whether miR-126 takes part in the p38MAPK pathway in the hippocampus of rats after cardiopulmonary resuscitation, the p38MAPK pathway inhibitor (SB203580) and activator (anisomycin) were added. The results showed overexpression of miR-126 could significantly increase the neurological function score and improve the pathological morphology of hippocampus in rats after cardiopulmonary resuscitation. miR-126 overexpression also could reduce the neuronal apoptosis, p38, and caspase-3 expression in the hippocampus. Moreover, the p38MAPK and JNK expression was downregulated and ERK1/2 expression was upregulated after miR-126 mimic injection (p < 0.05). The results of inhibition of p38MAPK pathway were consistent with those of overexpression of miR-126 (p > 0.05). This study indicated miR-126 could significantly reduce neuronal apoptosis of hippocampus in rats after cardiopulmonary resuscitation, which might be involved in the regulation of p38MAPK pathway.

High mobility group box 1 antibody represses autophagy and alleviates hippocampus damage in pilocarpine-induced mouse epilepsy model

As a neurological disorder, epilepsy has affected over 65 million people all over the world because of the unforeseeable seizures it might cause. However, in-depth understandings of the pathogenesis of epilepsy and effective treatments for the disease are still lacked. Recent discoveries suggest that autophagy, as an endogenous self-cleansing pathway in mammals, might be involved in the onset of epilepsy. Our study assumes that a non-histone DNA binding protein, high mobility group box-1 (HMGB1), formerly considered as a crucial inflammatory factor, may mediate the autophagy of neurons in epileptic mouse brain. To verify this hypothesis, pilocarpine induced epilepsy mouse model was constructed. The mice were treated with HMGB1 antibody for 4 weeks after the initial epileptic seizure. Behavioral test results suggested a recovery of learning ability and memory in epileptic mice when treated with HMGB1 antibody. Pathological changes in hippocampus were inspected under microscopes and hippocampus damages caused by seizures in mouse with epilepsy such as increased intracellular space were alleviated by HMGB1 antibody treatment. Moreover, the expressions of the proteins involved in autophagy pathways were detected by immunofluorescence staining and western blot. microtubule-associated protein 1A/1B-light chain 3 (LC3), Beclin 1, autophagy protein-5 (ATG5), and ATG7 levels were significantly decreased by HMGB1 antibody while the level of p62 was increased. TdT-mediated dUTP Nick-End Labeling (TUNEL) illustrated that cell apoptosis induced by seizures in hippocampus was mitigated by HMGB1 antibody. In conclusion, we propose that HMGB1 may induce increased autophagy in epilepsy mouse model.

The protective effect of Lonicera japonica polysaccharide on mice with depression by inhibiting NLRP3 inflammasome.

Depression is associated with the abnormal activation of the human inflammatory response system, which is a life-threatening disorder affecting millions of people of all ages around the world. The purposes of the present study were to examine the effect of Lonicera japonica polysaccharide (LJP), a polysaccharide extracted from Lonicera japonica Thunb., on depressed mice treated with the unpredictable emotional stress stimulation. The components of LJP after extraction were detected by HPLC. Depression model is established through chronic unpredictable stimulation, and the depression behavior was assessed by mouse behavioral assessment, including open field, elevated plus maze, tail suspension, forced swim. Pathological changes in hippocampus of mice were observed by HE and toluidine blue staining. Protein expression of NLRP3 inflammasome pathway was detected by WB. It was showed that LJP mainly consisted of 8.7% GalA, 8.2% Rha, 16.2% Gal, 19.5% Ara, 26.9% Glc and 20.5% Man, with the average molecular weight approximately ranging from 1 to 1,000 kDa, which could significantly reduce the time in the open arms and the immobility time of the depressed mice in behavioral tests, and the expressions of NLRP3, IL-1β, and caspase-1 in the hippocampus of depressed mice were significantly upregulated. LJP exhibited a strong protective effect on mice with depression by inhibiting NLRP3 inflammasome. The results will help to understand the potential use of Lonicera japonica Thun polysaccharide in pharmaceutical and food applications better.

Seizure characteristics and hippocampus pathological alterations in two rat models of temporal lobe epilepsy induced by hippocampus and amygdala injection of kainic acid

Objective To compare the behavioral manifestations, electroencephalogram(EEG) monitoring and hippocampal pathological changes in two rat epilepsy models induced by kainic acid (KA) injected into hippocampus and amygdala respectively. Methods Male adult Sprague-Dawley rats(n=24) were randomly divided into hippocampus model group (n=9), amygdala model group (n=9) and control group (n=6). Two epilepsy models were established by kainic acid (0.6 μg, 1.0 μg/μl) stereotactically injected into hippocampus CA3 region (hippocampus model) or amygdala (amygdala model). The status epilepticus (SE) of rats was observed by behavioral manifestations and EEG monitoring. Following successful modeling, those rats were randomly divided into 3 groups, including 1-day (acute phase), 7-day (latent phase) and 30-day (chronic phase) post SE groups (n=3 in each group). Rats in the control group were injected with an equal volume of saline in the hippocampal CA3 or amygdala. Immunohistochemistry was used for observing the pathological changes of neuron (NeuN), astrocyte (GFAP) and microglia (Iba1) in the rat hippocampus. Results Behavioral and EEG monitoring showed that rats from both models had typical seizure behaviors and EEG characteristics in the acute and chronic phases. However, there were different seize types and onset time between the 2 models in the acute phase. The hippocampus model group had partial seizures at 63.33±4.41 min post KA injection with intermittent generalized tonic-clonic seizures. Multi-phase spikes were recorded in acute phase. Amygdala model group had severe generalized tonic-clonic seizures at 28.67±3.48 min post KA injection, and the main cortical EEG form was sharp wave rhythm. Immunohistochemical staining revealed gradual hippocampal neuron death from acute to chronic phases in both models, which resulted in serious neuronal loss. Meanwhile, the aggravation of astrocyte proliferation and accumulation of microglia occurred in hippocampus in both models. Remarkably, compared with hippocampus model, the amygdala model in chronic phase had more neuronal loss in CA1 region (10.83±1.52 vs. 22.43±5.16, P<0.01) and CA4 region (12.87±2.13 vs. 25.81±4.60, P<0.05), more astrocyte proliferation in CA1 region (61.20±7.33 vs. 14.65±0.12, P<0.01) and CA4 region (76.73±5.40 vs. 43.01±1.35, P<0.01) as well as more widespread accumulation of microglia in CA1 region (13.70±3.88 vs. 1.08±0.01, P<0.01). Conclusions Our results have indicated that both rat epilepsy models simulated human temporal lobe epilepsy. The differences in the behavior, EEG performances and particularly pathological changes of hippocampus between two models should be taken into consideration in the future study. Key words: Epilepsy, temporal lobe; Hippocampus; Amygdala; Models, animal; Rats; Kainic acid

Study on Urine Metabolic Profile of Aβ25–35-Induced Alzheimer's Disease Using UHPLC-Q-TOF-MS

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder, with no effective method for its treatment so far. The pathogenesis of AD has been reported, but the endogenous metabolic profile and disease-related biomarkers are still not clear. To better understand AD, an AD model induced by injecting β-amyloid 25–35 (Aβ 25–35) solution into bilateral hippocampus was developed on Sprague–Dawley rats. After 8 weeks of modeling, the impairment of spatial learning and memory ability in AD rats were assessed by Morris water maze task. Hematoxylin and eosin staining and immunohistochemistry were used to investigate the pathological changes of hippocampus. The neurotransmitter concentrations in the hippocampus were measured using UHPLC-TQ-MS. Urinary metabolomics based on UHPLC-Q-TOF-MS was established to delineate the alterations of endogenous metabolites in AD rats. The results showed that compared with healthy control rats, AD rats suffered from cognitive dysfunction, hippocampus damage, Aβ formation and tau phosphorylation at 8 weeks after surgery, suggesting that the Aβ25–35-induced AD model was successfully established. In addition, the levels of γ-aminobutyric acid, acetylcholine, glycine, norepinephrine, serotonin, taurine and dopamine decreased and glutamate and aspartic acid increased in hippocampal tissue of AD rats. 45 altered metabolites mainly involved in 8 metabolic pathways were identified as the endogenous biomarkers of AD. According to the analysis of the biological significance of metabolic profiles, the pathogenesis of AD was mainly due to gut microbiome dysbiosis, inhibition of energy metabolism, oxidative stress injury and loss of neuronal protective substances.