Objective To explore the composition situation of the risk factors of patients with stroke in Huainan area. Methods 471 continuous registration and hospitalized patients with stroke in Huainan were selected,of which,362 cases were ischemic stroke,109 cases were hemorrhagic stroke,and the known risk factors were statistically analyzed. Results Of all patients,the incidence of hypertension was first(69. 3% was in ischemic stroke, 77. 9% was in hemorrhagic stroke) ,and hypertension was the most important risk factors. The composition of risk factors in stroke patients of different gender were different,and the smoking,drinking problem was more prominent in male patients (51.1% ,33.2%respectively)than that in female patients(5.1% ,2.0% respectively) (x2 =111-81,67.62, all P＜0.01) ;The incidence of diabetes, heart disease, atrial fibrillation was higher in female patients (37. 6% ,31. 9% ,19.3%respectively) than that in male patients (22.3% ,20.4% ,8.8% respectively) (x2 = 13. 12,8.09,11. 12,all P＜0.01). Hypertension,diabetes accounted for only 40.8% and 47.4% of patients taking regular medication. Atrial fibrillation was more common in patients with hemorrhagic stroke. Conclusion Controlling blood pressure and blood sugar was still important in the primary prevention and secondary prevention of stroke in the region. People of different gender should be targeted for individual health education, and promoting a healthy lifestyle should be paid attention, so as to the occurrence of stroke could be effectively prevented.

The carboxyl-terminal amino acids 272-299-truncated apoE4 (delta272-299) is the main fragments of apoE4 hydrolysate in neurons. The effects of truncated-ApoE4 (delta272-299) overexpression on tau phosphorylation in cultured N2a cells were investigated. The truncated-apoE4 (delta272-299) cDNA was subcloned into pEGFP-c3 to form recombinant pEGFP-T-apoE4. pEGFP-c3, pEGFP-T-apoE4 and pEGFP-apoE4 were transfected into N2a cells respectively by lipofectamine 2000 method. After 24--48 h, tau phosphorylation was detected by Western blot assay and glycogen synthase kinase-3 (GSK-3) activity by using GSK-3 activity assay. The results showed that the overexpression of both full length-apoE4 and truncated apoE4 fragments in N2a cells induced a dramatic increase in phosphorylation of tau at Ser202 sites and the activation of GSK-3 as compared with untransfected cells, most significantly in the cells transfected with pEGFP-T-apoE4 (P < 0.05). It was concluded that in vitro overexpression of truncated-ApoE4 (delta272-299) can result in tau hyperphosphorylation in N2a cells by activating GSK-3, suggesting truncated-ApoE4 (delta272-299) might contribute the pathogenesis of Alzheimer disease.

BACKGROUND: Puerarin functions to relieve the injury caused by ischemia reperfusion, improve the microcirculation, and inhibit the agglutination of platelet. But it is not clear yet that whether its protection on neurons relates to the apoptosis of cells.OBJECTIVE: To observe the protection of puerarin on neurons injured by mimic ischemia reperfusion in vitro.DESIGN: A random control study. SETTING: Laboratory of Biochemistry and Moleculobiology Department of Preclinical Medicine College of Tongji Medical College of Huazhong University of Science and Technology.PARTICIPANTS: The experiment was done on March 11, 2002. The cultured N2a cells of rat neuromablast were observed.METHODS: Ninety minutes after the cultured N2a cells were put into the 37 ℃ incubator with 0.05 CO2 and 0.95 N2(v/v), puerarin 0.5 mmol/L was added for cuiture for 24 hours. The methyl thiazolyl tetrazolium (MTT)method was used to observe the surviving ability of cells. The Annexin-V staining was adopted to exam the severity of apoptosis in the early stage and the supernate was collected to analyze the activity of lactate dehydrogenase (LDH) to reflect the permeability of cytomembrane. The immunoblotting method was applied to analyze the expression of caspase-3, at the same time, its activity was measured.MAIN OUTCOME MEASURES: The severity of cell apoptosis, the activity of LDH, the expression and activity of caspase-3.RESULTS: Puerarin could improve significantly the surviving rate of N2a cells 24 hours after reperfusion, decrease remarkably the activity of LDH in the culture fluid, lessen obviously the N2a apoptosis (P ＜ 0.01), at the same time, reduce tremendously the activity and expression of caspase-3 induced by ischemia reperfusion (P ＜ 0.01).CONCLUSION: Puerarin can protect nerves, inhibit obviously the N2a apoptosis induced by ischemia reperfusion. The mechanism is that it inhibits greatly the expression and activity of caspase-3.

BACKGROUND: Leptin is a kind of polypeptide hormone secreted by fatty tissue, previous studies have shown that leptin plays a certain role in the formation of atherosclerosis.OBJECTIVE: To investigate the effects of leptin on the expression of tumor necrosis factor-alpha (TNF-α) in RAW264.7 cells, and investigate the possible mechanism from the angle of the change of nuclear factor-κB (NF-κB) activity.DESIGN: A controlled observational experiment.SETTING: Department of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: The experiments were carried out in the Department of Biochemistry and Molecular Biology, Tongji Medical College, and the Department of General Surgery, Union Hospital affiliated to Huazhong University of Science and Technology between April 2005 and February 2006.The cultured RAW264.7 cells were divided into leptin treated groups treated with different concentration (12.5, 25, 50, 100 μg/L), I kappa B kinase inhibitor group and blank control group. Each group had 3 sub-wells,and the experiments were repeated for 3 times.METHODS: Mouse macrophage RAW264.7 cells were incubated into a 6-well plate at the density of 109 cells L-1, cultured in RPMI-1640 culture medium containing bovine serum of 0.1 in volume fraction. When the cells grew to 80%, serum-free culture medium Opti-MEM was changed to culture for another 24 hours, and then the cells were divided into leptin groups treated with different concentration (12.5, 25, 50, 100 μg/L), I kappa B kinase inhibitor group and blank control group. After the cells were incubated with leptin for 4 hours, the expression of TNF-α mRNA expression in RAW264.7 cells was detected with reverse transcription-polymerase chain reaction (RT-PCR). After the RAW264.7 cells were incubated with leptin for 1, 3, 6 and 9 hours, the expression of TNF-c protein expression was detected with double antibody sandwich enzyme-linked immunosorbent assay (ELISA). After the RAW264.7 cells were incubated with leptin for different times, the activity of NF-κB was detected analyzed with electrophoretic mobility shift assay. Another, the RAW264.7 cells were treated with or without 50 μmol/L leptin and/or 100 μmol/L PS1145 (I kappa B kinase specific inhibitor)divided into four groups: blank control group, I kappa B kinase specific inhibitor PS1145 (10 μmol/L) treated group, leptin (50 μmol/L) treated group, leptin (50 μmol/L) + PS1145 (10 μmol/L) group. Aftere incubated for 6 hours, the activity of NF-κB and expression TNF-α mRNA were detected respectively.MAIN OUTCOME MEASURES: ① Effect of leptin of different concentration on the expression of TNF-α mRNA and protein in RAW264.7cells; ② Effect of leptin of different concentration on the activity of NF-κB in RAW264.7 cells; ③ Influence of inhibition I kappa B kinase activity inhibition on expression of TNF-a induced by leptin in RAW264.7cells.RESULTS: ① After the RAW264.7 cells were treated with leptin of different concentration, the TNF-α mRNA level was elevated in a dose-dependent manner, and it reached the peak value emerged in the 50 μg/L leptin treated group. ② The expression of TNF-α protein increased in dose-dependent and time-dependent manners, and it reached the peak val ue at 6 hours in the 50 μg/L leptin treated group. ③ The activity of NF-κB was also positively correlated with the leptin concentration, and it was the highest value at 6 hours treated with 50 μg/L leptin (P ＜ 0.05). ④ I kappa B kinase activity inhibition only partially suppressed the leptin induced elevation of TNF-α expression induced by leptin.CONCLUSION: Leptin can increase the expression and secretion of TNF-α in RAW264.7 cells directly in both dose-dependent and time-dependent manners, and the mechanism may be correlated with the activated NF-κB induced by leptin. It may be one of the mechanisms of atherosclerosis induced by leptin.

BACKGROUND: The degree of neurofilament (NF) phosphorylation is closely correlated with the occurrence of Alzheimer disease (AD), and apolipoprotein E4 (apoE4) is a generally acknowledged liability factor for AD, but the effect and mechanism of apoE4 on the NF phosphorylation in neurons are not very clear. It has been reported that in the neurons cultured in vitro and in brain tissue of AD patients, the amino acid residues of apoE4 protein C terminal (272-299) could be truncated by hydrolysis,and produce truncated-apoE4 fragment. The latter interacts with the NF phosphorylation in neurofibrillary tangles (NFTs), which are the characteristic pathological changes of AD.OBJECTIVE: To observe the effect of truncated-apoE4 overexpression on the NF phosphorylation in the cultured neurons.DESIGN: A non-randomized controlled observation.SETTING: Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: The experiment was carried out in the Laboratory of Biochemistry and Molecular Biology, Tongji Medical College, Huazhong University of Science and Technology in December 2005. The mice neuroma cell strain N2a was provided by Dr. Xu.METHODS: The truncated-apoE4(△272-299) cDNA was subcloned into pEGFP-c3 to form pEGFP-T-apoE4 recombinant. Then pEGFP-c3, pEGFP-apoE4 and pEGFP-T-apoE4 were transfected into N2a cells by lipofectamine2000 respectively. NF phosphorylation was detected by Western blot assay. The activities of glycogen synthase kinase-3 (GSK-3) and cyclin-dependent kinase 5(CDK-5) were measured.MAIN OUTCOME MEASURES: The degree of NF phosphorylation and the activities of GSK-3 and CDK-5 were mainly observed.RESULTS: In the transfected groups, the contents of phosphorylated NF were significantly increased, the GSK-3 activities were significantly increased, which were the most significant in the pEGFP-T-apoE4 transfected group (P＜0.05), but the CDK-5 activities were not significantly different from that in th e control group (P＞0.05).CONCLUSION: These results indicate that in vitro overexpression of truncated-apoE4(△272-299) can lead to NF hyperphosphorylation by activating of GSK-3 but not CDK-5, which may be the underline mechanism of AD induced by truncated-apoE4(△272-299).

BACKGROUND: One of the key neuropathological changes in Alzheimer disease is that neurofibrils over phosphorylated cytoskeletal protein (such as r and neurofilaments) composed of entwist together, and the phosphorylation of τ protein can be catalyzed by cyclin dependent kinase 5 (CDK5),however whether the phosphorylation of neurofilaments can be catalyzed by CDK5, as well as its role in the pathogenesis of Alzheimer diseases is less acknowledged.OBJECTIVE: To explore the role of over-expression of intracellular CDK5 in the phosphorylation of neurofilamentsDESIGN: Randomized controlled study.SETTING: Biochemical and Molecular Biological Department of Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: This study was conduced at Biochemical and Molecular Biological Department of Tongji Medical College, Huazhong University of Science and Technology between February and May 2001. In vitro cultured rat neuroblastoma cell strain (N2a) was adopted as subjects.METHODS: In vitro cultured N2a cells were divided into 2 groups, namely transfection group and non-transfection group. In transfection group,CDK5 gene was transfected into N2a cell line by using liposome transfection technique so as to obtain N2a/CDK5 cell line stably expressing CDK5, immune-precipitation and enzyme activity assay was used to detect the CDK5 activity, meanwhile immunofluorescence technique and immuneblot assay was used to detect CDK5 expression and phosphorylation of neurofilaments.MAIN OUTCOME MEASURES: Phosphorylation of neurofilaments in both groups.RESULTS: In transfection group of N2a cell line, CDK5 expression increased presented by deep coloration of SMI31 antibody and weak coloration of SMI32 antibody, implying hyper-phosphorylation of neurofilaments. Meanwhile, the activity of CDK5 was 3.5 times higher than that in non-transfection group.CONCLUSION: Intracellular over-expressison of CDK5 would lead to hyperactivity of CDK5 and hyper-phosphorylation of neurofilaments, however the hyper-phosphorylation of neurofilamentsmight invlove in the pathological development of AD.

Forty cases of type Ⅱ diabetes mellitus were treated by puncturing point Neiguan (PC 6), and the effect on their cardiac vegetative nerve functions were observed at 20 min, 40 min and 60 min after acupuncture respectively. The findings showed that all heart rate variables improved remarkably at the three time periods after acupuncture treatment, with significant differences (P＜0.01); but there was no significant difference in the curative effects among the three time periods.

OBJECTIVE: To standardize the management on drug use in primary hospitals in Beijing Dongcheng district so as to enhance the efficiency of drug administration.METHODS: A "Drug Use Monitoring Information Platform in Primary Hospital" comprising management information system on drug use,management system on information of drug wholesalers,drug monitoring management information system was established and its problems in the try-out were analyzed.RESULTS & CONCLUSIONS: The "Drug Use Monitoring Information Platform in Primary Hospital" still faced with the problems of legal basis,psychological concern,and construction of standard basic drug information database etc.However,its operation standardized the drug purchasing channel and management on drug use in hospitals of Beijing Dongcheng district and enhanced the drug administration level and efficiency,which thus deserves to be popularized throughout the country.

ObjectiveTo investigate the effects of using transforming growth factor-β3 (TGF-β3) combined with dental pulp stem cells (DPSCs) on the regeneration of rabbit facial nerve.MethodsSixteen New Zealand adult rabbits were selected randomly.These rabbits were divided into three groups (TGF-β3 and DPSCs treated group,TGF-β3 control group and PBS control group.treatment group (silicone guidance channel with collagen protein sponge were filled with TGF-β3 and DPSCs),TGF-β3 control group (silicone guidance channel with collagen protein sponge were filled with TGF-β3) and PBS control group(silicone guidance channel with collagen protein sponge were filled with PBS).After 3 months,a series of examinations were performed,including gross morphology,histological staining,neuroelectrophysiological tset.ResultsIn the experimental group,the diameter of the regenerating nerve and near distal nerve stem were almost the same.There were no formation of neuroma.The adventitial angiogenesis was rich and with tough texture.3 months after operation,in the experimental group,the facial nerve membrane integrity,nerve fibers arranged in neat rows,form a more complete,myelin swelling.The total number of regeneration of nerve fibers in the experimental group were more than of control groups,statistical analysis was significant (P＜0.05).Diameter of regenerating nerve fibers in the experimental group were greater than that of control groups,statistical analysis was significant (P＜0.05).The ultra-thin section showed that the regenerated fibers in the treatment group were mainly myelinated never fiber.The layer structure of myelin sheath was clear,and there were rich organells axoplasma.The neuroelectrophysiological examinations revealed that the latency of nerve and muscle action conduction in the treatment group was shorter than that of the control groups,the treatment group:(1.96±0.32) ms,the TGF-β3 control group:(2.35±0.41) ms,the PBS control group:(3.42±0.55) ms.The wave amplitude of nerve and muscle action conduction in the treatment group was obviously higher than that of control groups,the treatment group:(11.06±3.25) mV,the TGF-β3 control group:(8.40± 1.68) mV,the PBS control group:(4.62±0.77) mV.ConclusionThe combination of TGF-β3 and DPSCs can improve the effects on the repair of facial nerve injury.

In this study, we studied the effect of glycogen synthase kinase-3 (GSK-3) overactivation on neurofilament phosphorylation in cultured cells. After N2a cells were treated with the specific inhibitor (wortmannin) of phosphoinositol-3 kinase (PI-3K) or treated with wortmannin and the specific inhibitor (LiCl) of glycogen synthase kinase-3 (GSK-3), GSK-3 activity and neurofilament phosphorylation were detected by using GSK-3 activity assay, Western blots and immunofluoresence. Our results showed that after treatment of N2a cells with wortmannin for 1 h, overactivation of GSK-3 caused a reduced staining with antibody SMI32 and an enhanced staining with antibody SMI31. When N2a cells were treated with wortmannin and LiCl, the activity of GSK-3 was reduced substantially. At the same time, the phosphorylation of neurofilament was also reduced. The study demonstrated that overactivation of GSK-3 induced hyperphosphorylation of neurofilament and suggested that in vitro overactivation of GSK-3 resulted in neurofilament hyperphosphorylation and this may be the underlying mechanism for Alzheimer's disease.