Eomesodermin expression in CD4+T-cells associated with disease progression in amyotrophic lateral sclerosis.

AIM: To clarify the role of Eomesodermin (EOMES) to serve as a disease-relevant biomarker and the intracellular molecules underlying the immunophenotype shifting of CD4+T subsets in amyotrophic lateral sclerosis (ALS). METHODS: The derivation and validation cohorts included a total of 148 ALS patients and 101 healthy controls (HCs). Clinical data and peripheral blood were collected. T-cell subsets and the EOMES expression were quantified using multicolor flow cytometry. Serum neurofilament light chain (NFL) was measured. In 1-year longitudinal follow-ups, the ALSFRS-R scores and primary endpoint events were further recorded in the ALS patients of the validation cohort. RESULTS: In the derivation cohort, the CD4+EOMES+T-cell subsets were significantly increased (p < 0.001). EOMES+ subset was positively correlated with increased serum NFL levels in patients with onset longer than 12 months. In the validation cohort, the elevated CD4+EOMES+T-cell proportions and their association with NFL levels were also identified. The longitudinal study revealed that ALS patients with higher EOMES expression were associated with higher progression rates (p = .010) and worse prognosis (p = .003). CONCLUSIONS: We demonstrated that increased CD4+EOMES+T-cell subsets in ALS were associated with disease progression and poor prognosis. Identifying these associations may contribute to a better understanding of the immunopathological mechanism of ALS.

Potential Pleiotropic Genes and Shared Biological Pathways in Epilepsy and Depression Based on GWAS Summary Statistics.

Current epidemiological and experimental studies have indicated the overlapping genetic foundation of epilepsy and depression. However, the detailed pleiotropic genetic etiology and neurobiological pathways have not been well understood, and there are many variants with underestimated effect on the comorbidity of the two diseases. Utilizing genome-wide association study (GWAS) summary statistics of epilepsy (15,212 cases and 29,677 controls) and depression (170,756 cases and 329,443 controls) from large consortia, we assessed the integrated gene-based association with both diseases by Multimarker Analysis of Genomic Annotation (MAGMA) and Fisher's meta-analysis. On the one hand, shared genes with significantly altered transcripts in Gene Expression Omnibus (GEO) data sets were considered as possible pleiotropic genes. On the other hand, the pathway enrichment analysis was conducted based on the gene lists with nominal significance in the gene-based association test of each disease. We identified a total of two pleiotropic genes (CD3G and SLCO3A1) with gene expression analysis validated and interpreted twenty-five common biological process supported with literature mining. This study indicates the potentially shared genes associated with both epilepsy and depression based on gene expression, meta-data analysis, and pathway enrichment strategy along with traditional GWAS and provides insights into the possible intersecting pathways that were not previously reported.

Novel Intronic Mutations of TBK1 Promote Aberrant Splicing Modes in Amyotrophic Lateral Sclerosis.

TANK-binding kinase 1 (TBK1) has been identified as a causative gene of amyotrophic lateral sclerosis (ALS) in the Caucasian population in 2015. Here, we sequenced for TBK1 variants in a cohort of 15 familial ALS (fALS) and 275 sporadic ALS (sALS) of Chinese origin by targeted next-generation sequencing. We identified one likely benign missense variant (p. Ser398Pro), two missense variants of uncertain significance (p. Ile37Leu and p. Tyr677Asn), and two novel heterozygous variants in introns of TBK1, c.1522-3T > G and c.2066 + 4A > G. We performed splicing assays through minigene plasmids and RNA pull-down assay to determine that the two substitutions of nucleotides disrupted the binding of the important splicing regulator hnRNPA1 and promoted aberrant pre-mRNA splicing modes. The c.1522-3T > G variant promoted nearly 50.0% of abnormal transcripts (3 different types of insertions and deletions (indels) in junction of intron 13-exon 14) and the c.2066 + 4A > G variant inhibited about 75.0% inclusion of exon 19, both causing premature stop codon and producing TBK1 protein without CCD2. Immunofluorescence analysis showed that the expression of TBK1 with intronic variants was lower since less TBK1 distribution was observed in HEK293T cells. Both patients carrying TBK1 c.1522-3T > G and c.2066 + 4A > G variants developed a rapidly progressive ALS, with a survival of 31 and 10 months, respectively. The frequency of loss of function (LoF) variants in TBK1 was 0.73% in sALS in our cohort. We emphasize that intronic sequencing and pre-mRNA splicing analysis cannot be ignored to demonstrate the complex mutational spectrum and pathogenesis of ALS.

Phenotype of VCP Mutations in Chinese Amyotrophic Lateral Sclerosis Patients.

Mutations in the valosin-containing protein (VCP) gene have been linked to amyotrophic lateral sclerosis (ALS) in the Caucasian populations. However, the phenotype of VCP mutations in Chinese patients with (ALS) remains unclear. Targeted next-generation sequencing covered 28 ALS-related genes including the VCP gene was undertaken to screen in a Chinese cohort of 275 sporadic ALS cases and 15 familial ALS pedigrees. An extensive literature review was performed to identify all patients with ALS carrying VCP mutations previously reported. The clinical characteristics and genetic features of ALS patients with VCP mutations were reviewed. One known p.R155C mutation in the VCP gene was detected in two siblings from a familial ALS pedigree and two sporadic individuals. In addition, the same VCP p.R155C mutation was detected in an additional patient with ALS referred in 2021. Three patients with VCP p.R155C mutation presented with muscular weakness starting from proximal extremities to distal extremities. The other patient developed a phenotype of Paget's disease of bone in addition to the progressive muscular atrophy. We reported the first VCP mutation carrier manifesting ALS with Paget's disease of bone in the Chinese population. Our findings expand the phenotypic spectrum of the VCP mutations in Chinese patients with ALS and suggest that ALS patients with VCP p.R155C mutations tend to present with relatively young onset, symmetrical involvement of proximal muscles weakness of arms or legs, and then progressed to distal muscles of limbs.

Novel Variants in the FIG4 Gene Associated With Chinese Sporadic Amyotrophic Lateral Sclerosis With Slow Progression.

BACKGROUND AND PURPOSE: Mutations in the FIG4 gene have been linked to amyotrophic lateral sclerosis (ALS) type 11 in Caucasian populations. The purpose of this study was to identify FIG4 variants in a cohort of 15 familial ALS (FALS) indexes and 275 sporadic ALS (SALS) patients of Han Chinese origin. METHODS: All 23 exons of FIG4 were sequenced using targeted next-generation sequencing. An extensive literature review was performed to detect genotype-phenotype associations of FIG4 mutations. RESULTS: No FIG4 variants were identified in the FALS patients. One novel heterozygous missense variant (c.352G>T [p.D118Y]) and one novel heterozygous nonsense variant (c.2158G>T [p.E720X]) in FIG4 were identified in two SALS patients. The p.E720X variant is interpreted as likely pathogenic while the p.D118Y variant is a variant of uncertain significance. The patient carrying the p.E720X mutation developed lower-limb-onset slowly progressive ALS, and survived for 11.5 years. The patient harboring the FIG4 p.D118Y variant also presented with progressive ALS, with the score on the ALS Functional Rating Scale-Revised (ALSFRS-R) decreasing by 0.4 per month. The rate of decrease in the ALSFRS-R scores from symptom onset to diagnosis seemed to be lower in the patients carrying FIG4 variants than the no-FIG4-mutation ALS patients in this study. CONCLUSIONS: Our findings suggest that ALS patients carrying FIG4 mutations are not common in the Chinese population and are more likely to exhibit slow progression.

Novel TARDBP missense mutation caused familial amyotrophic lateral sclerosis with frontotemporal dementia and parkinsonism.

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder that predominately involves the motor neurons in the brain and spinal cord. The TARDBP gene, encoding TAR DNA-binding protein 43 (TDP-43) protein, has been identified as a major causative gene in ALS. In this study, we screened 275 SALS patients and 20 unrelated FALS probands for TARDBP mutations. We identified three TARDBP mutations in three SALS patients and two TARDBP mutations in two FALS probands, including a previously unreported mutation, p.K176I, in FALS patients consistent with frontotemporal dementia (FTD) and parkinsonism. The p.K176I mutation is the first mutation outside exon 6 of the TARDBP gene manifesting parkinsonism and the first TARDBP mutation manifesting parkinsonism identified in the Chinese population. Our results support that TARDBP mutations are one of the most common changes in both FALS and SALS in China. Patients with TARDBP mutations may have a broad phenotype spectrum of ALS, FTD, and parkinsonism. The TARDBP gene should be included in genetic screening for ALS with FTD, and/or parkinsonism.

Novel FUS mutation Y526F causing rapidly progressive familial amyotrophic lateral sclerosis.

FUS gene is one of the most common mutated genes in amyotrophic lateral sclerosis (ALS). We sequenced for FUS mutations in a cohort of 15 familial ALS and 275 sporadic ALS of Chinese origin. All 15 exons of the FUS gene were sequenced by targeted next-generation sequencing in a cohort of 15 familial ALS indexes and 275 sporadic ALS patients of Chinese origin. One novel p.Y526F mutation in FUS was detected in one familial ALS proband. Another novel FUS p.Q140R variant and two known FUS mutations (p.R495Efs*33 and p.R521C) were identified in four sporadic ALS cases. The frequency of FUS mutation in our cohort is 6.7% in familial ALS and 1.5% in sporadic ALS. The familial ALS proband carrying the FUS p.Y526F mutation presented with juvenile-onset lower limbs weakness and demonstrated an aggressive course, with respiratory muscles involvement 6 months after onset. The other patients in the family all had limbs weakness and died 1-2 years after disease onset. Our results strengthen that FUS mutations are the most frequent genetic causes of young-onset aggressive ALS. Genetic testing of the FUS gene should be performed in early-onset ALS patients especially those with a rapid progression.

Novel mutation in optineurin causing aggressive ALS+/-frontotemporal dementia.

OBJECTIVE: Mutations in optineurin (OPTN) have been identified in familial and sporadic amyotrophic lateral sclerosis (ALS). We screened a cohort of Chinese patients for mutations in optineurin. We also performed an extensive literatures review of all mutations in optineurin identified previously to detect genotype-phenotype associations. METHODS: All 16 exons of the OPTN gene in a cohort of 15 familial ALS indexes and 275 sporadic ALS patients of Chinese origin were sequenced by targeted next generation sequencing. RESULTS: Two known heterozygous missense mutations in the OPTN, c.1481T> G (p.L494W), and c.1546G> C (p.E516Q), as well as one novel heterozygous missense mutation c.1690G> C (p.D564H) were each detected in one sporadic ALS patient. The patient carrying the p.E516Q mutation developed clinical features of ALS-frontotemporal dementia (FTD) and the patient carrying the p.D564H mutation showed a phenotype of ALS. They both had an aggressive course, with a survival of 18 and 14 months respectively. Literature review showed that the clinical phenotypes in OPTN mutated ALS were not homogeneous, although some individuals showed a relatively slow progression and a long duration, some mutations carriers developed an aggressive progression and a short survival. INTERPRETATION: OPTN mutations contribute to ALS in Chinese population and account for 0.8% of sporadic ALS patients and 1.5% of familial ALS in the pooled Chinese ALS cohorts. Mutations in optineurin can cause aggressive ALS+/-FTD.

Novel insights into the effect of paroxetine administration in pilocarpine­induced chronic epileptic rats.

The aim of the present study was to investigate the role of paroxetine intervention in epilepsy, and its association with the expression of serotonin transporter (SERT) and hippocampal apoptosis. Thirty adult male Sprague Dawley rats were divided into control vehicle (n=6) and epileptic (n=24) groups. Status epilepticus (SE) was induced via systemic injection of pilocarpine, and seizure activity was monitored via video electroencephalogram. The epileptic group was then randomly divided into two groups; Four weeks following SE induction, paroxetine (5 mg/kg/day; SE + paroxetine group) or normal saline (SE group) was intraperitoneally injected for 4 weeks. Brain tissue was collected to evaluate apoptosis via terminal deoxynucleotidyl transferase dUTP nick­end labeling. SERT, B­cell lymphoma­2 (Bcl­2) and brain derived neurotropic factor (BDNF) expression levels were evaluated by western blotting, and miR­16 expression was evaluated by reverse transcription­quantitative polymerase chain reaction. Paroxetine did not affect the mortality of the pilocarpine­induced chronic epileptic rats. Spontaneous recurrent seizures (SSRs) were observed 7­28 days following SE induction. The frequency and stage of the SSRs were reduced by paroxetine administration. Apoptotic cells were observed in the epileptic hippocampus. Following paroxetine intervention, the staining intensity and number of apoptotic cells were significantly decreased. Expression levels of BDNF and Bcl­2 were lower in the SE group compared with the vehicle group. The former was not altered by paroxetine injection; however, the latter was increased. In the SE group, SERT expression was not altered in the raphe nucleus but was decreased in the hippocampus. Following paroxetine administration, SERT expression was decreased in the raphe nucleus and increased in the hippocampus. In the SE group, miR­16 expression was decreased in the raphe nucleus and increased in the hippocampus. Following paroxetine administration, miR­16 expression was not altered in the raphe nucleus but was reduced in the hippocampus. In conclusion, the seizures and hippocampal apoptosis observed in chronic epileptic rats were alleviated by paroxetine treatment. This effect may be associated with the reduced Bcl­2 and BDNF expression and the modulation of SERT expression. The alterations in miR­16 expression may provide a potential explanation for the modulation of apoptosis; however, further research is required to determine the complete underlying molecular mechanism.

DL-3-n-butylphthalide protects endothelial cells against advanced glycation end product-induced injury by attenuating oxidative stress and inflammation responses.

Endothelial dysfunction, regarded as a key step in the pathophysiological course of diabetic vascular complications, is initiated and deteriorated by advanced glycation end products (AGEs). DL-3-n-butylphthalide (DL-NBP) has been proven to have protective effects on neurons and vascular endothelial cells against ischemic and anoxic damage. The aim of the present study was to investigate whether NBP is able to attenuate AGE-induced endothelial dysfunction in vitro, and also elucidate the possible underlying mechanism. An injury model of human umbilical vein endothelial cells (HUVECs) induced by AGEs (200 µg/ml) was established. The results demonstrated that pretreatment with NBP (1-100 µM) significantly increased HUVEC viability and inhibited the apoptosis induced by AGEs. In addition, AGEs stimulated the expression levels of the receptor for AGEs protein and the downstream protein nuclear factor-κB in HUVECs, which were inhibited by pretreatment with NBP. Furthermore, it significantly reduced reactive oxygen species generation and the level of the inflammatory cytokines, intercellular cell adhesion molecule-1 and monocyte chemotactic protein-1, in HUVECs mediated by AGEs. The current findings indicated that NBP attenuated AGE-induced endothelial dysfunction by ameliorating inflammation and oxidative stress responses.

Genetic epidemiology of amyotrophic lateral sclerosis: a systematic review and meta-analysis.

BACKGROUND: Genetic studies have shown that C9orf72, SOD1, TARDBP and FUS are the most common mutated genes in amyotrophic lateral sclerosis (ALS). Here, we performed a meta-analysis to determine the mutation frequencies of these major ALS-related genes in patients with ALS. METHODS: We performed an extensive literature research to identify all original articles reporting frequencies of C9orf72, SOD1, TARDBP and FUS mutations in ALS. The mutation frequency and effect size of each study were combined. Possible sources of heterogeneity across studies were determined by meta-regression, sensitivity analysis and subgroup analysis. RESULTS: 111 studies were included in the meta-analysis. The overall pooled mutation frequencies of these major ALS-related genes were 47.7% in familial amyotrophic lateral sclerosis (FALS) and 5.2% in sporadic ALS (SALS). A significant difference was identified regarding the frequencies of mutations in major ALS genes between European and Asian patients. In European populations, the most common mutations were the C9orf72 repeat expansions (FALS 33.7%, SALS 5.1%), followed by SOD1 (FALS 14.8%, SALS 1.2%), TARDBP (FALS 4.2%, SALS 0.8%) and FUS mutations (FALS 2.8%, SALS 0.3%), while in Asian populations the most common mutations were SOD1 mutations (FALS 30.0%, SALS 1.5%), followed by FUS (FALS 6.4%, SALS 0.9%), C9orf72 (FALS 2.3%, SALS 0.3%) and TARDBP (FALS 1.5%, SALS 0.2%) mutations. CONCLUSIONS: These findings demonstrated that the genetic architecture of ALS in Asian populations is distinct from that in European populations, which need to be given appropriate consideration when performing genetic testing of patients with ALS.

Comparative Efficacy and Safety of Nine Anti-Platelet Therapies for Patients with Ischemic Stroke or Transient Ischemic Attack: a Mixed Treatment Comparisons.

Anti-platelet treatments, an effective anti-thrombotic therapy, are widely used in non-cardioembolic ischemic stroke or transient ischemic attack (TIA), including aspirin, cilostazol, clopidogrel, and other mono or dual therapies, while the optimal choice remains uncertain. All the literatures of 38 eligible randomized control trials were searched in PubMed, Embase, and China National Knowledge Internet (CNKI) without language limitation. And, nine anti-platelet therapies were assessed, including aspirin, clopidogrel, cilostazol, ticlopidine, triflusal, terutroban, sarpogrelate, dipyridamole plus aspirin, and clopidogrel plus aspirin. Additionally, we extract data of composite vascular events, major bleeding, ischemic stroke, intracranial hemorrhage, and all-cause death, as indicators of efficacy and safety. And among them, composite vascular events were the primary outcome. The binary outcomes were expressed as odds ratios (ORs) with corresponding 95 % confidence intervals (CIs). Both traditional meta-analysis and network meta-analysis were performed. Besides, for each outcome, the rank order was applied to reflect the superiority of every therapy compared with others, using the surface under the cumulative ranking curve (SUCRA). A cluster analysis was also conducted. Through the network meta-analysis, the synthesized data shows that cilostazol performed best on composite vascular events compared with placebo (OR = 0.62, 95 % CI 0.46-0.83) and aspirin (OR = 0.71, 95 % CI 0.53-0.95). In terms of ischemic stroke, clopidogrel plus aspirin seems the optimal, and it has significant difference between placebo (OR = 0.53, 95 % CI 0.35-0.74) and aspirin (OR = 0.75, 95 % CI 0.61-0.95). Meanwhile, cilostazol is also the first rank in major bleeding, especially when it is in contrast to aspirin (OR = 0.13, 95 % CI 0.02-0.70) and clopidogrel plus aspirin (OR = 0.09, 95 % CI 0.01-0.50). There is no significant difference among these nine treatments and placebo, as to all-cause death and intracranial hemorrhage. According to the cluster analysis, cilostazol can be the best choice with comprehensive assessment of composite vascular events, ischemic stroke and major bleeding. Based on this network meta-analysis, cilostazol was recommended as the optimal choice with good performance in both efficacy and safety for patient with ischemic stroke or TIA among nine anti-platelet therapies.

Toward precision medicine in amyotrophic lateral sclerosis.

Precision medicine is an innovative approach that uses emerging biomedical technologies to deliver optimally targeted and timed interventions, customized to the molecular drivers of an individual's disease. This approach is only just beginning to be considered for treating amyotrophic lateral sclerosis (ALS). The clinical and biological complexities of ALS have hindered development of effective therapeutic strategies. In this review we consider applying the key elements of precision medicine to ALS: phenotypic classification, comprehensive risk assessment, presymptomatic period detection, potential molecular pathways, disease model development, biomarker discovery and molecularly tailored interventions. Together, these would embody a precision medicine approach, which may provide strategies for optimal targeting and timing of efforts to prevent, stop or slow progression of ALS.

[Changes of left ventricular remodeling in hypertension patients with carotid atherosclerosis of phlegm-dampness syndrome].

OBJECTIVE: To study changes of left ventricular remodeling (LVR) in hypertension patients with carotid atherosclerosis (CAS) of phlegm-dampness syndrome (PDS). METHODS: Doppler ultrasonography data of CAS were observed in 223 hypertension patients with CAS (as the hypertension group, including 119 patients of the PDS group and 104 of the non-PDS group), 81 CAS patients with non-hypertension, and 19 non-hypertension non-CAS patients (as the control group). The difference in the degree of LVR was compared among the above groups. RESULTS: The left ventricular posterior wall thickness (LVPWT), inter ventricular septum thickness (IVS), E/A were higher in the hypertension group than in the non-hypertension group (P < 0.05). The left ventricular end-diastolic dimension (LVEDD), left ventricular end-systolic diameter (LVESD), stroke volume (SV) were higher in the soft plaque hypertension group and the soft plaque non-hypertension group than in the hard plaque group, the thickening intimal group, and the normal intimal group (P < 0.01 , P < 0.05). The LVEDD, LVESD, and SV were higher, and the ejection fraction (EF) was lower in the PDS hypertension group than in the non-PDS hypertension group (all P < 0.05). Of them, LVEDD, LVESD, and SV were higher in the soft plaque group than in the hard plaque group (P < 0.01), the thickening intimal group (P < 0.01) and the normal intimal group (P < 0.05). There was no statistical difference in PDS hypertension between the soft plaque group and the hard plaque group (P > 0.05). CONCLUSION: The hypertension patients with CAS of PDS might be correlated to LVR, and LVR was more obviously in the soft plaque patients.

Seizure-induced 5-HT release and chronic impairment of serotonergic function in rats.

We analyzed the dynamic concentration change of serotonin (5-HT) and its main metabolite 5-hydroxyindoleacetic acid (5-HIAA) within the epileptic hippocampus in rats. Seizure was induced by systemic injection of pilocarpine (320mg/kg, i.p.). Using electroencephalography (EEG) recordings, we found that primary seizure discharge was induced 30min after pilocarpine administration and that recurrent discharge peaked 14d after the onset of status epilepticus (SE). The extracellular fluid in the hippocampus was sampled by microdialysis from conscious animals at various time points before and after SE. The concentrations of 5-HT and 5-HIAA in the samples were measured by high-performance liquid chromatography and electrochemical detection (HPLC-ECD). Interestingly, 5-HT levels in the hippocampus were dramatically increased within the 30min following SE. This reversed to basal level by 4d after SE and continued to drop to 48% at 7d and 28% of basal level 14d after SE. Accordingly, a marked increase of 5-HIAA in the hippocampus appeared at 2d after SE, then gradually declined to levels below baseline. To identify serotonergic neurons in the raphe nuclei (a major source of 5-HT release in the brain), brain sections were immunostained for tryptophan hydroxylase (TPH). The number of TPH positive neurons and the intensity of TPH staining significantly decreased at 28d after SE. These data suggest that pilocarpine induces depletion of 5-HT in the hippocampus and significantly compromise serotonergic neurons in the raphe nuclei. The loss of serotonergic function may play a significant role in the pathophysiology of epilepsy.

[Analysis of CLCN1 gene mutations in 2 patients with myotonia congenita].

OBJECTIVE: To investigate chloride channel 1 (CLCN1) gene mutation and clinical features of 2 Chinese patients with myotonia congenita. METHODS: Clinical data of a patient from a family affected with myotonia congenita in addition with a sporadic patient from Fujian province were analyzed. Exons of CLCN1 gene were amplified and sequenced. RESULTS: The proband from the affected family was found to carry a c.1024G>A heterozygous missense mutation in exon 8, whilst the sporadic patient has carried a c.1292C>T heterozygous missense mutation in exon 11. CONCLUSION: Detection of CLCN1 gene mutation is an effective method for the diagnosis of myotonia congenita. Exon 8 of CLCN1 gene may be a mutational hotspot in Chinese patients with myotonia congenita.

[Effect of serotonin depletion on seizures learning-memory in pilocarpine-induced epileptic rats].

OBJECTIVE: To investigate the relationship between serotonin (5-HT) and epilepsy and the mechanism of learning-memory in pilocarpine (PILO)-induced epileptic rats after 5,7-dihydroxytryptamine (5,7-DHT) microinjection in median raphe nucleus. METHODS: Adult S D rats were randomly divided into 3 groups: PILO group, PILO+ 5,7-DHT group, vehicle control group; PILO group was divided into two groups by status epilepticus (SE): PILO + SE group and PILO - SE group. The rats' seizures and cortex electroencephalography (EEG) were observed by video EEG. The rats' spatial learning-memory was evaluated by Morris water maze. Finally, serotonergic neuron in raphe nuclei was observed by immunohistochemistry. RESULTS: After treatment of 5,7-DHT (PILO + 5,7-DHT group), the success rate, the mortality and the frequency of chronic spontaneous seizures in pilocarpine-induced epilepsy model were all improved. Compared with the control group, the number of serotonergic neuron in raphe nuclei was decrease in PILO + SE group (P < 0.05). Moreover, it's extremely decrease in PILO + 5,7-DHT group (P < 0.01). Compared with control group, the mean escape latency was prolonged, the times of crossing target was decreased and the retention time in target zone was shortened in PILO + SE group (P < 0.05), but there was no significant difference between PILO + SE group and PILO + 5,7-DHT group. CONCLUSION: Depletion of serotonin may facility the rats' epileptic seizures, but we could not interpret which may cause epileptic rats' cognitive deficit.

Ginsenoside Rb1 selectively inhibits the activity of L-type voltage-gated calcium channels in cultured rat hippocampal neurons.

AIM: To investigate the effect of ginsenoside Rb1 on voltage-gated calcium currents in cultured rat hippocampal neurons and the modulatory mechanism. METHODS: Cultured hippocampal neurons were prepared from Sprague Dawley rat embryos. Whole-cell configuration of the patch-clamp technique was used to record the voltage-gated calcium currents (VGCCs) from the hippocampal neurons,and the effect of Rb1 was examined. RESULTS: Rb1 (2-100 µmol/L) inhibited VGCCs in a concentration-dependent manner, and the current was mostly recovered upon wash-out. The specific L-type Ca(2+) channel inhibitor nifedipine (10 µmol/L) occluded Rb1-induced inhibition on VGCCs. Neither the selective N-type Ca(2+) channel blocker ω-conotoxin-GVIA (1 µmol/L), nor the selective P/Q-type Ca(2+) channel blocker ω-agatoxin IVA (30 nmol/L) diminished Rb1-sensitive VGCCs. Rb1 induced a leftward shift of the steady-state inactivation curve of I(Ca) to a negative potential without affecting its activation kinetics or reversal potential in the I-V curve. The inhibitory effect of Rb1 was neither abolished by the adenylyl cyclase activator forskolin (10 µmol/L), nor by the PKA inhibitor H-89 (10 µmol/L). CONCLUSION: Ginsenoside Rb1 selectively inhibits the activity of L-type voltage-gated calcium channels, without affecting the N-type or P/Q-type Ca(2+) channels in hippocampal neurons. cAMP-PKA signaling pathway is not involved in this effect.

[The effects of adipose-derived stem cells transplantation on the expression of TGF-ß1 in rat brain after cerebral ischemia].

AIM: To explore the mechanisms of adipose-derived stem cells (ADSCs) transplanting induced angiogenesis in rat brain after focal cerebral ischemia. METHODS: 108 male adult Sprague-Dawley rats were randomly assigned into 4 groups: sham-operated group, middle cerebral artery occlusion (MCAO) group, vehicle group and MCAO+ADSCs-treated group. Rat's focal cerebral ischemia model was established by right middle cerebral artery occlusion (MCAO) with modified Longa's method. ADSCs were pre-labeled by CFSE before the transplantation into the rat brain. At 1 d after MCAO, 30 µL cell suspension which contained 1×10(6); ADSCs was injected into the lateral ventricle of MCAO+ADSCs-treated rats, and the same dose of PBS was given to the rats of vehicle group as control. At 4 d, 7 d and 14 d after MCAO, rats were decapitated to detect the TGF-ß1 expression in the infarct area. RESULTS: The expression of TGF-ß1 in brain tissues in MCAO+ADSCs-treated group was significantly higher than MCAO group and vehicle group at 4 d, 7 d and 14 d after MCAO, respectively. After transplantation into MCAO rats, ADSCs could survive and express TGF-ß1 in the ischemic brain. CONCLUSION: These data suggest that ADSCs transplantation can promote revascularization in cerebral ischemic rats, partly by promoting TGF-ß1 expression in the brain.

Microglial phagocytosis induced by fibrillar ß-amyloid is attenuated by oligomeric ß-amyloid: implications for Alzheimer's disease.

BACKGROUND: Reactive microglia are associated with ß-amyloid (Aß) deposit and clearance in Alzhiemer's Disease (AD). Paradoxically, entocranial resident microglia fail to trigger an effective phagocytic response to clear Aß deposits although they mainly exist in an "activated" state. Oligomeric Aß (oAß), a recent target in the pathogenesis of AD, can induce more potent neurotoxicity when compared with fibrillar Aß (fAß). However, the role of the different Aß forms in microglial phagocytosis, induction of inflammation and oxidation, and subsequent regulation of phagocytic receptor system, remain unclear. RESULTS: We demonstrated that Aß(1-42) fibrils, not Aß(1-42) oligomers, increased the microglial phagocytosis. Intriguingly, the pretreatment of microglia with oAß(1-42) not only attenuated fAß(1-42)-triggered classical phagocytic response to fluorescent microspheres but also significantly inhibited phagocytosis of fluorescent labeled fAß(1-42). Compared with the fAß(1-42) treatment, the oAß(1-42) treatment resulted in a rapid and transient increase in interleukin 1ß (IL-1ß) level and produced higher levels of tumor necrosis factor-α (TNF-α), nitric oxide (NO), prostaglandin E2 (PGE2) and intracellular superoxide anion (SOA). The further results demonstrated that microglial phagocytosis was negatively correlated with inflammatory mediators in this process and that the capacity of phagocytosis in fAß(1-42)-induced microglia was decreased by IL-1ß, lippolysaccharide (LPS) and tert-butyl hydroperoxide (t-BHP). The decreased phagocytosis could be relieved by pyrrolidone dithiocarbamate (PDTC), a nuclear factor-κB (NF-κB) inhibitor, and N-acetyl-L-cysteine (NAC), a free radical scavenger. These results suggest that the oAß-impaired phagocytosis is mediated through inflammation and oxidative stress-mediated mechanism in microglial cells. Furthermore, oAß(1-42) stimulation reduced the mRNA expression of CD36, integrin ß1 (Itgb1), and Ig receptor FcγRIII, and significantly increased that of formyl peptide receptor 2 (FPR2) and scavenger receptor class B1 (SRB1), compared with the basal level. Interestingly, the pre-stimulation with oAß(1-42) or the inflammatory and oxidative milieu (IL-1ß, LPS or t-BHP) significantly downregulated the fAß(1-42)-induced mRNA over-expression of CD36, CD47 and Itgb1 receptors in microglial cells. CONCLUSION: These results imply that Aß oligomers induce a potent inflammatory response and subsequently disturb microglial phagocytosis and clearance of Aß fibrils, thereby contributing to an initial neurodegenerative characteristic of AD. Antiinflammatory and antioxidative therapies may indeed prove beneficial to delay the progression of AD.