Objective To investigate carotid atherosclerosis and its relevant risk factors of ischemic cerebral apoplexy in high-risk population and to explore new modes of behavioral influencing factors based on health management platform.Methods Using the carotid ultrasound screening of personnel undergoing physical examination,the differences of the related risk factors were studied among 1 152 cases of carotid atherosclerosis with differ ent gender and different age and different disease periods.Results High-risk population between 40-49 years wasmainly focused on mild lesions,and the numbers of serious lesions increased significantly among people older than 70;male patients numbered 51 were significantly much more than the females numbered 21 (P＜0.01)during mild lesion period,there was no statistical significance between male numbered 36 and female numbered 29 during serious lesion period (P＞0.05).Setting mild and serious period respectively as the dependent variables,and hypertension,overweight,dyslipidemia,diabetes mellitus,smnoking,physical activity and atrial fibrillation as the independent variables to conduct logistic regression analysis,it showed that mild lesions were related to overweight,hypertension and dyslipidemia.The risk factors of serious lesion period included hypertension,overweight,diabetes mellitus,dyslipidemia,atrial fibrillation,smoking and lack of physical activity.The female detection rates of hypertension and dyslipidemia in female patients,which were 83.8％ and 44.1％ respectively,were higher than those in male patients,which were 67.6％ and 31.0％ respectively.The detection rates of smoking and arterial fibrillation in female patients,which were 0.7％ and 0.3％ respectively,were lower than those in male patients,which were 20.7％ and 1.0％ respectively.Conclusions Early detection of carotid atherosclerosis lesions and its relevant risk factors as soon as possible,and early intervention influencing factors to mild lesions to prevent the atherosclerosis from getting worse.A new model of zero level prevention can be developed to control of ischemic stroke based on health management platform.

Grass carp reovirus (GCRV), a disaster agent to aquatic animals, belongs to Genus Aquareovirus of family Reoviridea. Sequence analysis revealed GCRV genome segment 8 (s8) was 1296 bp nucleotides in length encoding an inner capsid protein VP6 of about 43kDa. To obtain in vitro non-fusion expression of a GCRV VP6 protein containing a molecular of fluorescence reporter, the recombinant baculovirus, which contained the GCRVs8 and eGFP (enhanced green fluorescence protein)genes, was constructed by using the Bac-to-Bac insect expression system. In this study, the whole GCRVs8 and eGFP genes, amplified by PCR, were constructed into a pFastBacDual vector under polyhedron (PH) and p10 promoters, respectively. The constructed dual recombinant plasmid (pFbDGCRVs8/eGFP) was transformed into DH10Bac cells to obtain recombinant Bacmid (AcGCRVs8/eGFP) by transposition. Finally, the recombinant bacluovirus (vAcGCRVs8/eGFP) was obtained from transfected Sf9 insect cells. The green fluorescence that was expressed by transfected Sf9 cells was initially observed 3 days post transfection, and gradually enhanced and extended around 5days culture in P1(Passage1) stock. The stable high level expression of recombinant protein was observed in P2 and subsequent passage budding virus (BV) stock. Additionally, PCR amplification from P1 and amplified P2 BV stock further confirmed the validity of the dual-recombinant baculovirus. Our results provide a foundation for expression and assembly of the GCRV structural protein in vitro.

Objective To establish an easier and better method for primary culture of rabbit fibroblast-like synoviocytes(FLS) by modified tissue cell culture.Method Healthy rabbit joint synovial layers were cut into small pieces and siphoned off water attached on the tissue with sterile filter paper and then cultured.The growth status and morphology were observed.The growth curve of the 3rd passage cells was measured by CCK-8 assay, and the expression of vimentin was detected by immunocytochemistry.Results The FLS cultured in vitro exhibited a spindle-shaped appearance and could rapidly expand.The cell growth curve was typical of S type, and the cells highly expressed vimentin.Conclusions Primary culture of rabbit FLS by the improved explant culture method is successfully established.The method is simple and highly efficient.It provideds a new method for isolation of FLS.

In order to synthetically train students to do scientific researches independently,inspire their enthusiasm and go-aheadism about study,and improve the quality of experimental teaching,we have been exploring to update experimental content and reform experimental system for many years,and have commenced a number of "Series Experiments".The setup of"Series Experiments" which means several separate experiments are organized together by their internal relations has already showed us a favorable effect.

Objective To clone CYP3A4 gene and to construct CYP3A4 recombinant mammalian expression vectors which are transferred into K562 cells for expression,and detect the anti-tumor effect of CYP3A4 combined with Cyclophosphamide (CPA) in vitro.Methods Mammalian expression vector containing CYP3A4 gene cloned from human hepatocytes by RT-PCR were constructed and transferred into K562 cells via liposome. The expression of CYP3A4 was detected by RT-PCR and Western blot respectively. MTT detected the anti-tumor effect of CYP3A4 recombinant mammalian expression vectors combined with CPA.Results We cloned CYP3A4 gene and constructed the recombinant mammalian expression vectors pcDNA3, 1/myc-His A (+)-CYP3A4 successfully. Both RT-PCR and Western blot showed significantly higher mRNA and protein expression of CYP3A4 in gene-transfected group than in empty vector-transfected controls and in empty cells controls. The IC_(50) values were 1. 090 16 mmol/L in gene-transfected group, which were significantly lower than in other two groups and could be reduced and increased by the revulsant and inhibitor respectively.Conclusions CYP3A4 recombinant mammalian expression vectors had anti-tumor effect cooperating with CPA, and the effect could be increased and reduced by the revulsant and inhibitor of CYP3A4 respectively.

Grass carp reovirus (GCRV), a double stranded RNA virus that infects aquatic animals, often with disastrous effects, belongs to the genus Aquareovirus and family Reoviridea. Similar to other reoviruses, genome replication of GCRV in infected cells occurs in cytoplasmic inclusion bodies, also called viral factories. Sequences analysis revealed the nonstructural protein NS80, encoded by GCRV segment 4, has a high similarity with uNS in MRV(Mammalian orthoreoviruses), which may be associated with viral factory formation. To understand the function of the uNS80 protein in virus replication, the initial expression and identification of the immunogenicity of the GCRV NS80 protein inclusion forming-related region (335.742) was investigated in this study. It is shown that the over-expressed fusion protein was produced by inducing with IPTG at 28oC. In addition, serum specific rabbit antibody was obtained by using super purified recombinant NS80(335.742) protein as antigen. Moreover, the expressed protein was able to bind to anti-his-tag monoclonal antibody (mouse) and NS80(335-742) specific rabbit antibody. Further western blot analysis indicates that the antiserum could detect NS80 or NS80C protein expression in GCRV infected cells. This data provides a foundation for further investigation of the role of NS80 in viral inclusion formation and virion assembly.

Sequences analysis revealed Grass carp reovirus (GCRV) s10 was 909 nucleotides coding a 34 kDa protein denoted as VP7, which was determined to be a viral outer capsid protein (OCP). To obtain expressed OCP in vitro, a full length VP7 gene was produced by RT-PCR amplification, and the amplified fragment was cloned into T7 promoted prokaryotic expression vector pRSET. The recombinant plasmid,which was named as pR/GCRV-VP7,was then transformed into E.coli BL21 host cells. The data indicated that the expressed recombinant was in frame with the N-terminal fusion peptide. The over-expressed fusion protein was produced by inducing with IPTG, and its molecular weight was about 37kDa, which was consistent with its predicted size. In addition, the fusion protein was produced in the form of the inclusion body with their yield remaining steady at more than 60% of total bacterial protein. Moreover,the expressed protein was able to bind immunologically to anti-his-tag monoclonal antibody (mouse) and anti-GCRV serum (rabbit). This work provides a research basis for further structure and function studies of GCRV during entry into cells.

A retrospective analysis was carried out in 36 patients with blunt diaphragmatic rupture during March 1991 and March 2006. Twenty-two diagnoses were confirmed preoperatively, and the rest 14were confirmed perioperatively. Three patients underwent surgical treatment after no response to conservative therapy. Thoracotomy was performed in 21 patients, laparotomy in 9, thoracotomy plus laparotomy in 5 and combined thoraco-laparotomy in 1. Most diaphragmatic rupture may be caused by blunt collision and could lead to thoracoabdominal injury. In spite of high mortality rate, the condition is usually under diagnosed. Definite diagnosis and timely operation are important to increase survival rate and reduce mortality.

To investigate the relationship between plasma level of homocysteine(Hcy) and the methylenetetrahydrofolate reductase ( MTHFR ) gene polyroorphism with non-alcoholic fatty liver in patients with type 2 diabetes mellitus (T2DM). Methods In a case-control study, plasma levels of Hcy, folic acid (FA), vitamin B12 (VitB12), glycosylated hemoglobin Alc (HbAlc), fasting blood glucose (FBG), total cholesterol and triglyceride were measured in 159 T2DM patients with and without non-alcoholic fatty liver ( NAFL), as well as 52 normal controls. Mutation of the C677T of MTHFR gene was determined by polymerase chain reaction-restricted fragment length polymorphism (PCR-RFLP) for all of them. Results Patients of T2DM both without NAFL (96 case) and with NAFL had higher prevalence of hyperhomocysteinemia (Hhcy) (49% and 21%, respectively ) than normal controls did (4 cases, 8% ) (P<0.05), while patients of T2DM with NAFL had higher prevalence of Hhcy than those without it did (P <0. 05). Plasma level of Hey positively correlated to genotype frequency of the MTHFR gene, plasma 0levels of HbAlc and FBG in patients of T2DM, with coefficients of correlation of 0.248, 0.423 and 0.242, respectively (P < 0.05). Results of multiple logistic regression analysis showed that course of the disease, body mass index, plasma levels of FBG and Hcy all were independent risk factors for non-alcoholic fatty liver in patients with T2DM. Conclusions Hhey was an independent risk for non-alcoholic fatty liver and plasma level of Hey was influenced by frequency of the TT genotype of the MTHFR gene, plasma levels of FA and VitB12, as well as metabolic disturbance in patients with T2DM.

Moyamoya disease (MMD) is a chronic and progressive cerebrovascular disease which is characterized by the bilateral internal carotid artery ends and (or) stenosis or occlusion of anterior cerebral artery and middle cerebral artery initial segments,compensatory proliferation of small blood vessels in the skull base and formation of abnormal vascular network.Its etiology and pathogenesis remains unclear.The present studies speculate that MMD may be a polygenic disease,inflammation,immune response,abnormal cytokine secretion,endothelial progenitor cell change and nitric oxide level change are associated with the occurrence and development of MMD.This article reviews the advances in research on the genetics and pathophysiological mechanism of MMD.