为了提高研究生的培养质量,系统地对研究生病原分子生物学技术进行优质课程建设和实践,包括师资队伍、课程目标、课程设计理念、课程内容、教学方法、教学手段、教学评价、教材建设、课程资源、课程管理、课程实践条件等,探索出提高研究生培养质量的优质课程模式.

The effects of deoxynivalenol in different dose including 100, 200, 500, 1000, and 1500 mu g/L on primary cranial osteoblasts from fetal mice were investigated. Fluorescence staining, flow cytometric measurement, 3-(4,5-dimethylthiazol 2-y1)-3,5-di-phenyltetrazolium bromide assay, quantitative PCR, and Western blot were used for the test. Mineralization and proliferation of osteoblasts decreased upon 100 mu g/L and higher deoxynivalenol treatment and apoptosis of osteoblasts was increased upon 500, 1000, and 1500 mu g/L deoxynivalenol treatment. Karyopyknosis, membrane breakage, and a decreased number of calcium nodes were also observed upon 500 mu g/L deoxynivalenol treatment. The mRNA and protein levels of B-cell lymphoma-2-associated X protein were upregulated, B-cell lymphoma-2 protein downregulated with increasing concentrations of deoxynivalenol treatment and their ratio increased. Deoxynivalenol induces apoptosis of osteoblasts, suggesting a mechanism by which deoxynivalenol can affect murine skeletal development.

目的 了解宝鸡市人民医院耐甲氧西林金黄色葡萄球菌的临床分布特点及对抗菌药物的耐药性,为临床合理使用抗生素和医院感染控制提供依据. 方法 回顾性分析该院2012-2015年耐甲氧西林金黄色葡萄球菌(methicillin resistant Staphylococcus aureus,MRSA)的临床分布及耐药性情况.采用VITEK-2 Compact全自动微生物鉴定药敏仪对可疑菌落进行鉴定和药敏检测,使用WHONET 5.6软件分析数据. 结果 2012-2015年共分离出MRSA菌株177株,MRSA菌株检出率为28.0％,男性患者MRSA检出率(33.1％)明显高于女性(21.2％),差异有统计学意义(x2=10.97,P=0.00).MRSA菌株主要源于痰液(占50.2％)和分泌物(占45.7％),二者MRSA菌株检出率分别为35.7％、26.0％;不同科室MR-SA主要以儿科(占33.9％)、皮肤科(占14.7％)为主,各科室MRSA检出率差异有统计学意义(x2=11.202,P=0.011);各年度MRSA检出率差异有统计学意义(x2=14.898,P=0.002),2012年最高(37.2％),2014年最低(20.3％).MASA对青霉素G、苯唑西林完全耐药,对红霉素(89.8％)、克林霉素(74.6％)耐药率较高,尚未发现对利奈唑胺、万古霉素、替考拉宁耐药. 结论 2012-2015年宝鸡市人民医院MRSA检出率28.0％,低于全国平均水平,不同标本、不同科室、不同年度MRSA检出率不同.MRSA对青霉素G、苯唑西林、红霉素、克林霉素常出现耐药.

Objective To understand the clinical distribution and drag resistance of Staphylococcus aureus,and to provide the basis for the rational use of antibiotics and the hospital control of the bacteria.Methods The clinical distribution and drug resistance of 632 strains of staphylococcus aureus from 2012 to 2015 in a hospital were retrospectively analyzed.VITEK-2 was used to identify the suspected colonies and the drug susceptibility of them.The statistics was conducted with WHONET 5.6 and SPSS17.0.Results A total of 6 777 strains of bacteria were isolated in the past 4 years,632 strains of them were Staphylococcus aureus,177 strains of them were Methicillin resistant Staphylococcus aureus,and 455 strains of them were Methicillin sensitive Staphylococcus aureus.The detection rate of Staphylococcus aureus,Methicillin resistant Staphylococcus aureus and Methicillin sensitive Staphylococcus aureus were lower than national average.The resistance rations of Staphylococcus aureus to Penicillin G,Erythromycin and Clindamycin were 94.5％,73.6％ and 60％,respectively,to Oxacillin,Amikacin,Rifampicin,Levofloxacin and Cotrimoxazole were less than 30％,to Linezolid,Vancomycin and Teicoplanin were all 0％.The resistance rations of methicillin resistant Staphylococcus aureus to Penicillin G,Oxacillin,Amikacin,Rifampin,Levofloxacin,Erythromycin and Clindamycin were more than that of Methicillin sensitive Staphylococcus aureus (P＜O.05).Conclusion The control of Staphylococcus aureus in the hospital is better than that in other hospitals,but the resistance ration is high.Multiple drug resistance and high resistance are found IN Methicillin resistant Staphylococcus aureus and Methicillin sensitive Staphylococcus aureus.

Objective To ascertain the molecular mechanism by which Helicobacter pylori CagA upregulates CIP2A.Methods After AGS and GES-1 cells were separately cultured to exponential phase,AGS or GES-1 cells were separately transfected with either the recombinant plasmid WT-cagA or the control vector plasmid pcDNA3.1.The cells were then treated with a signal inhibitor of B-Raf or JNK2 and transfected with the plasmid recombinant WT-cagA.The expression of CIP2A was detected with Western blotting,and the cell clonogenic potential was determined using a cell cloning assay.Cell migration was detected with a cell migration assay.Results AGS and GES-1 cells transfected with the plasmid WT-cagA had a higher level of CIP2A expression than that in the control.After treatment with a signal inhibitor of B-Raf or JNK2 and subsequent transfection with the plasmid WT-cagA,the level of CIP2A expression in the cells was decreased.The cell clonogenic potential and cell migration decreased below the cell clonogenic potential and cell migration noted in cells transfected with the plasmid WT-cagA alone.Conclusion H.pylori CagA upregulates the expression of the oncogene CIP2A through B-Raf and JNK2,affecting the cell clonogenic potential and cell migration.

目的:探讨用大米淘米水、牛奶治疗口腔溃疡的临床效果.方法:先用浓度为40%的冰醋酸对40只健康的昆明小鼠进行口腔溃疡模型制作,然后随机将这40只小鼠分成4组,分别为生理盐水组、自来水组、大米淘米水组和牛奶组,每组各有10只小鼠.为生理盐水组小鼠口服一定量的生理盐水,为自来水组小鼠口服一定量的自来水,为大米淘米水组小鼠口服一定量的大米淘米水,为牛奶组小鼠口服一定量的牛奶,然后观察其口腔溃疡的愈合情况.结果:与自来水组小鼠和生理盐水组小鼠相比,淘米水组小鼠和牛奶组小鼠经干预后其舌体黏膜恢复至湿润有光泽状态的时间和溃疡面愈合的时间均更短,差异有统计学意义(P<0.05).淘米水组小鼠和牛奶组小鼠经治疗其舌体黏膜恢复至湿润有光泽状态的时间及溃疡面愈合的时间相比差异无统计学意义(P>0.05).结论:用大米淘米水或牛奶治疗口腔溃疡的效果显著,均具有疗效好、见效快等优点.

ABSTRACT Coxsackievirus A10 (CVA10) is one of the major pathogens associated with hand, foot, and mouth disease (HFMD). CVA10 infection can cause herpangina and viral pneumonia, which can be complicated by severe neurological sequelae. The morbidity and mortality of CVA10-associated HFMD have been increasing in recent years, particularly in the pan-Pacific region. There are limited studies, however, on the pathogenesis and immunology of CVA10-associated HFMD infections, and few antiviral drugs or vaccines have been reported. In the present study, a cell-adapted CVA10 strain was employed to inoculate intramuscularly 5-day-old ICR mice, which developed significant clinical signs, including reduced mobility, lower weight gain, and quadriplegia, with significant pathology in the brain, hind limb skeletal muscles, and lungs of infected mice in the moribund state. The severity of illness was associated with abnormally high expression of the proinflammatory cytokine interleukin 6 (IL-6). Antiviral assays demonstrated that ribavirin and gamma interferon administration could significantly inhibit CVA10 replication both in vitro and in vivo . In addition, formaldehyde-inactivated CVA10 whole-virus vaccines induced immune responses in adult mice, and maternal neutralizing antibodies could be transmitted to neonatal mice, providing protection against CVA10 clinical strains. Furthermore, high-titer antisera were effective against CVA10 and could relieve early clinical symptoms and improve the survival rates of CVA10-challenged neonatal mice. In summary, we present a novel murine model to study CVA10 pathology that will be extremely useful in developing effective antivirals and vaccines to diminish the burden of HFMD-associated disease. IMPORTANCE Hand, foot, and mouth disease cases in infancy, arising from coxsackievirus A10 (CVA10) infections, are typically benign, resolving without any significant adverse events. Severe disease and fatalities, however, can occur in some children, necessitating the development of vaccines and antiviral therapies. The present study has established a newborn-mouse model of CVA10 that, importantly, recapitulates many aspects of human disease with respect to the neuropathology and skeletal muscle pathology. We found that high levels of the proinflammatory cytokine interleukin 6 correlated with disease severity and that ribavirin and gamma interferon could decrease viral titers in vitro and in vivo . Whole-virus vaccines produced immune responses in adult mice, and immunized mothers conferred protection on neonates against challenge from CVA10 clinical strains. Passive immunization with high-titer antisera could also improve survival rates in newborn animals.

Previous study has demonstrated that mitochondrial-dependent apoptotic pathway is involved in the nephroprotective effect of puerarin (PU) against lead-induced cytotoxicity in primary cultures of rat proximal tubular (rPT) cells. To further clarify how PU exerts its antiapoptotic effects, this study was designed to investigate the role of mitochondrial permeability transition (MPT) and subsequent apoptotic events in the process of PU against Pb-induced cytotoxicity in rPT cells. The results showed that Pb-mediated mitochondrial permeability transition pore (MPTP) opening together with mitochondrial cytochrome c release, activations of caspase-9 and caspase-3, and subsequent poly-ADP-ribose polymerase (PARP) cleavage can be effectively blocked by the addition of PU. Simultaneously, upregulation and downregulation of Bcl-2 and Bax with increased Bcl-2/Bax ratio due to PU administration further alleviated Pb-induced mitochondrial apoptosis. Moreover, PU can reverse Pb-induced ATP depletion by restoring mitochondrial fragmentation to affect ATP production and by regulating expression levels of ANT-1 and ANT-2 to improve ATP transport. In summary, PU produced a significant protection against Pb-induced mitochondrial apoptosis in rPT cells by inhibiting MPTP opening to ameliorate the mitochondrial dysfunction.

OBJECTIVE:To screen the differential gene expression and related pathways of fetal skeletal malformations induced by deoxynivalenol(DON) in mice,and to explore the mechanism of deformities induced by DON at the molecular level.METHODS:30 pregnant mice were randomly divided into 3 experimental groups and 2 control groups,6 in each group. DON injection was performed from days 7 to 10 of pregnancy by intraperitoneal injection into pregnant mice. At GD 18,all mice were killed under isoflurane anesthesia,and vertebral bone tissue of fetuses were collected. Total RNA of vertebral bone tissues was extracted and cDNA was obtained by RT-PCR. Using whole genome microarray technology,the gene expression profiles and the pathways of the rat vertebral bone tissue were studied. Analysis of the function of differentially expressed genes was performed by using software of gene ontology. Screening signaling pathways of differentially expressed genes was done by DAVID database. The results were verified by fluorescence quantitative PCR(qPCR) technology. RESULTS:Microarray analysis showed that 282 genes,including 148 down-regulated and 134 up-regulated genes,were abnormally expressed in fetal vertebral bones after maternal DON exposure. 153 pathways were related to the differentially expressed genes. The relative quantitative results ofqPCR were consistent w ith t he r esults o f g ene c hip t est i nFbn1,Co19a2,Papss2,Pax1,Runx2 a ndPthlh g enes.CONCLUSION: There were differential gene expressions in deformed fetal skeleton between deoxynivalenol-treated rats and normal rats, involving multiple signaling pathways. The differentially expressed genes and signaling pathways may be related to the molecular mechanisms of deformities induced by DON.

We compared the numbers of nucleotide substitutions occurring in the non-coding regions and coding regions of Ebola virus genomes and found that non-coding regions contain indispensable phylogenetic and evolutionary information. The omission of genetic data from non-coding regions can lead to unreliable phylogenies and inaccurate estimates of evolutionary parameters.

目的 监测医院中气载耐甲氧西林金黄葡萄球菌(MRSA)对临床常用抗生素的耐药性及相关耐药基因.方法 用FA-1型六级筛孔撞击式空气微生物采样器采集5所医院的大厅、ICU及病房等场所的空气,分离金黄葡萄球菌,采用头孢西丁纸片法检测MRSA,应用K-B法测定MRSA药敏情况,应用多重PCR扩增MRSA甲氧西林耐药基因mecA,氨基糖苷修饰酶基因 aacA-aphD,大环内酯类23s rRNA甲基化酶基因 ermA、ermC,四环素类核糖体保护蛋白基因tetM、tetK以及金黄葡萄球菌属特异性基因 16s rDNA.结果 共采集空气样品250份,分离气载金黄葡萄球菌219株,其中MRSA 88株.所有气载MRSA均对青霉素及头孢曲松耐药,对庆大霉素、红霉素及四环素耐药率＞90％,但均对万古霉及素替考拉宁敏感.所有MRSA菌株mecA基因和16s rDNA基因均阳性,vat(A)、vat(B)、vat(C)基因均阴性.96％以上MRSA菌株可同时检出氨基糖苷类、红霉素类和四环素类耐药基因,呈现多重耐药;携带aacA-aphD、erm(A)或(和)erm(C)、tetM或(和)tetK耐药基因的MRSA菌株总检出率分别为96.59％、100％和96.59％.结论 5所医院空气中均存在MRSA分布且呈多重耐药性,相关耐药基因检出率高.因此应加强对医院空气中致病菌的监测与消毒,预防和控制医院气源病原微生物感染的发生.

Objective To prepare Helicobacter pylori Omp18 protein and polyclonal antibodies against that protein Methods The Omp18 gene was amplified using PCR and cloned into the vector pMD-18T.It was then was cloned into the prokaryotic expression plasmid vector pTriEx(TM)-4.The vector was transformed into Escherichia coli JM109DE.Omp18 protein was produced and purified,and then rabbits were repeatedly administered the protein to produce polyclonal antibodies.Results The Omp18 gene was successfully cloned into the prokaryotic expression vector pTriEx(TM)-4 and the vector was successfully transformed into E.coli JM109DE.Omp18 protein was successfully produced,and purified Omp18 protein was administered to rabbits to yield polyclonal antibodies.Conclusion Omp18 protein was successfully expressed in prokaryotic cells,and polyclonal antibodies were successfully prepared in rabbits.This research will benefit study of the function and regulation of Omp18 and it will lay a foundation for detection of H.pylori and preparation of vaccines.

Objective To prepare Helicobacter pylori Omp18 protein and polyclonal antibodies against that protein Methods The Omp18 gene was amplified using PCR and cloned into the vector pMD-18T.It was then was cloned into the prokaryotic expression plasmid vector pTriEx(TM)-4.The vector was transformed into Escherichia coli JM109DE.Omp18 protein was produced and purified,and then rabbits were repeatedly administered the protein to produce polyclonal antibodies.Results The Omp18 gene was successfully cloned into the prokaryotic expression vector pTriEx(TM)-4 and the vector was successfully transformed into E.coli JM109DE.Omp18 protein was successfully produced,and purified Omp18 protein was administered to rabbits to yield polyclonal antibodies.Conclusion Omp18 protein was successfully expressed in prokaryotic cells,and polyclonal antibodies were successfully prepared in rabbits.This research will benefit study of the function and regulation of Omp18 and it will lay a foundation for detection of H.pylori and preparation of vaccines.

Objective The effects of Escherichia coli on the motility,ultrastructure,and apoptosis of human sperm in vitro were studied in order to identify the cause of male infertility and facilitate diagnosis and therapy for male infertility caused by pathogenic infections.Methods Semen samples were obtained from 30 healthy volunteers.Four groups of sperm suspensions were prepared: a normal sperm suspension serving as negative control(Group A),a sperm suspension mixed with suspended E.coli(Group B),a sperm suspension mixed with suspended E.coli treated with traditional Chinese medicine(Group C),and a sperm suspension mixed with suspended E.coli treated with furadantin(Group D).These samples were collected at 0,2,and 4 h.Motility parameters were analyzed using a computer-aided semen analysis(CASA) system.Changes in the ultrastructure of human sperm were observed with transmission electron microscopy.The rate of apoptosis of semen sperm was detected with flow cytometry combined with FITC-Annexin V/PI dual color fluorescence staining.Results Group B had significantly lower motility parameters,including straight line velocity(VSL),curve velocity(VCL),average path velocity(VAP),amplitude of lateral head displacement(ALH),and progressive motility,in comparison to the other groups at 4h(P0.01).There were no significant differences in the ultrastructure of sperm in the control group(A) and groups treated with medicine(C and D),while there were significant changes in the ultrastructure of sperm in Group B.In addition,the rate of sperm apoptosis increased significantly(P0.01) in Group B.Conclusion E.coli significantly inhibited sperm motility,changed the ultrastructure of sperm,and increased the rate of apoptosis of sperm in vitro.Therefore,E.coli infection may be one cause of male infertility.

IFN-γ plays an indirect anti-cancer role through the immune system but may have direct negative effects on cancer cells. It regulates the viability of gastric cancer cells, so we examined whether it affects their proliferation and how that might be brought about. We exposed AGS, HGC-27 and GES-1 gastric cancer cell lines to IFN-γ and found significantly reduced colony formation ability. Flow cytometry revealed no effect of IFN-γ on apoptosis of cell lines and no effect on cell aging as assessed by β-gal staining. Microarray assay revealed that IFN-γ changed the mRNA expression of genes related to the cell cycle and cell proliferation and migration, as well as chemokines and chemokine receptors, and immunity-related genes. Finally, flow cytometry revealed that IFN-γ arrested the cells in the G1/S phase. IFN-γ may slow proliferation of some gastric cancer cells by affecting the cell cycle to play a negative role in the development of gastric cancer.

基层医院检验科在日常工作中易忽略一些重要的问题,如未对每天的工作进行总结;未明确各种分析仪器是否经过较准;工作人员是否经过岗前培训等,尤其是没有形成文件存留,这与ISO15189体系(以下简称体系)不相符。该体系要求如下:(1)对人员的要求:每个工作人员必须建立详细的个人档案,必须进行岗前培训。(2)科室内使用的仪器:仪器应该经专业人员维护、修理,再由专人做质控校准,并定期检查,形成文件记录存档。(3)科室内的定期质控:每周质控的检验报告与临床回馈差错都应详细记录存档。该体系着重要求人员素质和质量控制。(4)实验室的设施与环境。

Objective To determine if gastric epithelial cells produce IFN-γ and the mechanism of its production.Methods AGS gastric epithelial cells were infected with H.pylori and the production of IFN-γ was detected using RT-PCR and ELISA.AGS gastric epithelial cells were transfected with CagA plasmid and tested for IFN-γ using ELISA.MEK / ERK,Src,P38,and NF-kB inhibitors were then added to the cells transfected with CagA plasmid and they were tested for IFN-γ using ELISA.Results When AGS gastric epithelial cells were infected with H.pylori,the cells produced IFN-γ.When the cells were transfected with CagA plasmid,the cells also produced IFN-γ.When MeK/ErK,P38 and NF-kB inhibitors were added to the cells transfected with CagA plasmid,the production of IFN-γ was inhibited.Conclusion H.pylori AGS gastric epithelial cells to produce IFN-γ.As the main virulence factor for H.pylori,CagA induces cells to produce IFN-γ,and that production depends on signal pathways involving MeK / ErK,P38,and NF-kB.

Deoxynivalenol (DON [vomitoxin]), one of trichothecene mycotoxins produced by the fungus Fusarium, is commonly detected in cereal foods across the world. DON induces diverse toxic effects in humans and animals, including emesis and diarrhea, anorexia, and immunotoxicity, and impaired maternal reproduction and fetal development. Recently, the teratogenic potential of DON has been shown and has received much attention. DON can cause various skeletal deformities in fetuses, but the underlying mechanisms have not yet been fully examined. In this study, fetal skeletal malformations in DON-treated maternal mice were thoroughly investigated using microarray assay. The results showed that DON administration caused various skeletal defects in fetuses, including misaligned or fused sternebrae and vertebrae, divided or fused ribs and polydactyly, hemivertebrae, short toes, and tail anomalies. Microarray analysis showed that 282 genes, including 148 downregulated and 134 upregulated genes, were abnormally expressed in fetal vertebral bones after maternal DON exposure. These identified genes can be classified into several categories: skeletal development, carcinogenesis, nervous disorders, sperm development and embryogenesis, and inflammation. Of these, 6 genes, mostly related to bone development, were intentionally selected for further validation using real-time reverse transcription-Polymerase Chain Reaction (RT-PCR). It was confirmed that the mRNA expression of 4 genes, i.e., fibrillin-1, Col9A2, 3′-phosphoadenosine 5′-phosphosulfate synthase 2, and Pax1, was upregulated significantly by DON administration, whereas that of 2 other genes, Runx2 and parathyroid hormone-like hormone, was downregulated significantly. Taken together, the results of our study suggest that altered expression of these 6 genes plays a critical role in fetal skeletal deformities induced by DON and thus they are worthy of further investigation.

PBL was used to teach sections 1-2 of students specializing in clinical medicine in the Class starting in 2008,and traditional teaching was used teach sections 3-4 of students specializing in clinical medicine in the same Class.Results on final exams and a questionnaire were used to evaluate the effectiveness of using PBL to teach Medical Microbiology.Students learning via PBL had significantly better exam results compared to students learning conventionally.The questionnaire indicated that PBL can improve the motivation and drive to learn,the ability to express one's self,independent thinking and independent learning,interpersonal communication skills,and ability to work in groups.PBL is clearly superior to traditional teaching.

研究生是我国创新体系中最重要的执行主体和推动社会发展的动力源泉。目前,我国研究生培养还不能完全适应创新社会发展的需要,所以迫切需要提高研究生培养的质量,培养出具有国际竞争力的高层次人才。结合普通医学院校的实际情况,在网络高度发达的今天,充分利用网络资源,为研究生成长营造良好的氛围,为研究生培养提供国际化的成长环境,有助于快速提高研究生的培养质量。