海口市"三级甲等医院"医务人员人格特质调查研究

目的研究试图通过对海口市三甲医院医务人员人格特质的测量与分析,在与全国常模进行对比的情况下,得出海口市三甲医院医务人员人格特质的基本描述,建立医务人员人格特质的常模,为最终科学地建立海南省医学人才人格特质考评体系奠定必要的工作基础,也为海南省医学人才的培养和利用提供一项可行性的管理模式.方法采取分层整群随机抽样方法对海口市五家三甲医院医务人员人格特质进行调查,使用国际著名的人格问卷-加利福尼亚心理调查表CPI作为工具,对海口市三甲医院医务人员人格特质进行调查.结果被调查的海口市三甲医院588名医务人员人格类型分布以Beta型为主,各型分布情况为：Alpha型171人,占29.1%,Beta型302人,占51.4%,Gamma型59人,占10.0%,Delta型56人,占9.5%;在V3自我实现水平上海口市三甲医院医务人员有较高得分,除1例处于第二水平和4例处于第三水平外,其余583例都分别处于第四至第七水平;海口市三甲医院医务人员在20个分量表的测试中,在CS、IN、EM、SC、GI、WB、CM、TO、AI、AC、IE、PY、FX等13个分量表的平均得分上明显高于全国常模组（P〈0.01）,而在DO、SY、SP、SA、SO 5个分量表平均得分上明显低于全国常模组（P〈0.01）.结论海口市三甲医院医务人员各分量表平均得分大部分高于全国常模水平,尤其是在测量成就潜能方面的3个分量表AC、AI、IE平均得分全部明显高于全国常模水平（P〈0.01）,表明海口市三甲医院医务人员较常模有更强烈的成就动机,对智力活动与知识成就有较强的追求,在工作中能更有效地发挥智能,注重办事效率,见识广博,比较适合在激励自由和个人首创精神的场所独立地开展工作,并且能够专心于别人看来较为枯燥的工作.在测量社会价值内化程度的7个分量表中有5个（SC、GI、CM、WB、TO）高于全国常模水平.     

不同骨移植材料修复骨缺损的初步实验研究

目的对比观察不同骨移植材料对兔桡骨节段性骨缺损的修复效果。方法48只新西兰大白兔采用桡骨15mm节段性骨缺损模型,随机分为4组,每组12只,A组植入深冻异体骨复合自体骨髓,B组植入深冻异体骨,C组植入自体骨,D组植入新鲜同种异体骨。术后不同阶段分别行组织学、透射电镜及X线检测。结果四组骨缺损均有成骨现象发生,骨生成、骨连接情况A、C组优于B组,B组优于D组;A、C组细胞增生活跃、核呈分裂相、胞质丰富、核膜光整、细胞器丰富,同比均优于B、D组;骨缺损愈合时间A、C组为8-10周,B组为12周,D组骨缺损在术后12周仍未愈合。结论提示兔深冻异体骨复合自体骨髓具有良好的组织相容性及成骨作用,其取代自体骨植骨修复骨缺损具有可能性。     

企业员工应对方式与心理健康之间的关系

目的：验证不同应对方式对心理健康的单独与共同影响。方法：对4745名被试用分层回归的方法分析不同应对方式对心理健康的单独影响和交互作用。结论：单独使用“消极应对”的应对方式,对心理健康水平存在不利影响;而单独使用“积极应对”的应对方式能在一定程度上改善心理健康水平。“消极应对”与“积极应对”两种应对方式结合使用能显著改善心理健康水平,比单独使用这两种应对方式对心理健康更有利。     

胶原基真皮再生支架的微结构控制

综述了影响胶原基真皮再生支架微结构的各种因素及其控制方法。胶原基真皮支架的生物活性和修复创面的能力受到诸如除胶原外的其它主要材料 (第二组分 )、支架的孔径和孔隙率、支架厚度、生物活性因子以及交联度等多方面因素的综合影响。从材料学、生物学和医学的角度综合地应用物理、化学和生物学的手段探索各种影响因素的控制方法是组织工程皮肤研究的重点。     

NMDA受体在β-淀粉样蛋白引起海马神经元突触蛋白改变中的作用

为了研究NMDA受体活性对Aβ引发的海马神经元突触蛋白表达变化的影响,本文运用免疫细胞化学方法检测不同浓度NMDA受体激动剂以及拮抗剂对Aβ诱导的海马神经元突触蛋白变化的影响。结果显示:NMDA可浓度依赖性地缓解Aβ25-35引起的突触蛋白synaptophysin与PSD-95的减少。抑制突触内NMDA受体,NMDA缓解Aβ减少突触蛋白的作用减弱;抑制突触外NMDA受体,对抗Aβ的作用无显著变化。本研究结果提示NMDA受体活性改变影响Aβ诱导的突触蛋白减少,突触内NMDA受体激活可对抗Aβ的毒性作用。突触内NMDA受体活性减弱可能在谷氨酸兴奋毒性中发挥作用。     

降钙素基因的化学合成与克隆

本文报道用化学法全合成降钙素的基因。基因全长113碱基对。采用大肠杆菌偏爱密码子。整个基因分为6个片段合成。用T~4DNA连接酶一次连接成功,序列分析证明合成的基因序列与设计的一致。     

用BA－ELISA斑点法检测痢疾杆菌免疫小鼠GALT中的特异性抗体分泌细胞（ASC）

改进的ＢＡ－ＥＬＩＳＰＯＴ法使免疫酶斑更为清晰，保存时间延长。用此法检测了痢疾杆菌福氏２ａ经口及腹腔免疫后，小鼠派伊尔氏（ＰＰ）淋巴结、肠系膜淋巴结（ＭＬＮ）及脾脏（ＳＰＬ）中特异性ＩｇＡ、ＩｇＧ、ＩｇＭ抗体分泌细胞（ＡｎｔｉｂｏｄｙＳｅｃｒｅｔｉｎｇｃｅｌｌ，ＡＳＣ）的动态变化，得到有规律的结果：两种免疫途径均能在ＰＰ及ＭＬＮ中诱导出３类特异ＡＳＣ的显著升高，但口服导致的升高其持续时间较腹腔途径为短。此外，腹腔途径还能诱导ＳＰＬ中３种ＡＳＣ升高。     

Commercial preparations of lipoteichoic acid contain endotoxin that contributes to activation of mouse macrophages in vitro

Lipoteichoic acids (LTA), cell wall components of gram-positive bacteria, have been reported to induce various inflammatory mediators and to play a key role in gram-positive-microbe-mediated septic shock. In a large number of these studies, investigators used commercially available LTA purified from a variety of gram-positive bacteria, including Staphylococcus aureus, Bacillus subtilis, and Streptococcus sanguis. We report here that, although these commercially available LTA could be readily shown to stimulate production of nitric oxide (NO) in RAW 264.7 mouse macrophages, the activity was dramatically inhibited by polymyxin B, a relatively specific inhibitor of endotoxin biological activity. One-step purification of the commercially available S. aureus LTA using hydrophobic interaction chromatography resulted in two well-separated peak fractions, one highly enriched for LTA and a second highly enriched for endotoxin. The LTA-enriched fractions did not induce production of NO in RAW 264.7 macrophages, although they caused a dose-dependent induction of NO in the presence of low concentrations of gamma interferon (IFN-gamma) (which by itself induced little NO), regardless of the presence of polymyxin B. In contrast, the endotoxin-enriched fractions by themselves inhibited in high levels of NO in RAW 264.7 macrophages but activity was almost completely inhibited in the presence of polymyxin B. Consistent with these findings, our data also indicate that commercial LTA preparations from S. aureus, B. subtilis, and S. sanguis were not able to induce NO from lipopolysaccharide-hyporesponsive C3H/HeJ mouse peritoneal macrophages, but in the presence of IFN-gamma, these LTA preparations were able to induce relatively high levels of NO from C3H/HeJ macrophages. These results indicate that commercially available LTA can contain contaminating and potentially significant levels of endotoxin that can be expected to contribute to the putative macrophage-stimulating effects of LTA as assessed by NO production. The fact that the purified LTA, by itself, was not able to induce significant levels of NO secretion in RAW 264.7 macrophages supports the conclusion that caution in attributing high-level biological activity to this microbial cell wall constituent should be exercised.     

大鼠肝抑素纯化及其生物活性的检测

用ＳｅｐｈａｄｅｃＧ－５凝胶过滤层析法，进一步纯化具肝抑素生物活性的大鼠肝蛋白质粗提品，以分离的大鼠再生肝的肝细胞为靶细胞，体外检测各洗脱峰浓缩物对肝细胞增殖的制率结果证明，Ｅ峰浓缩物的抑制作用最强，其活性比为粗提品的２０倍，ＳＤＳ聚丙烯酰胺电泳图及蛋白质迁移率测定表明，该浓缩物的主要成分为分子量１３．５ｋＤ的多肽。本研究对大鼠肝抑素做了初步纯化，验证了该物质在肝再生中起重要调控作用的生物效应。     

A HIT-trapping strategy for rapid generation of reversible and conditional alleles using a universal donor

Targeted mutagenesis in model organisms is key for gene functional annotation and biomedical research. Despite technological advances in gene editing by the CRISPR-Cas9 systems, rapid and efficient introduction of site-directed mutations remains a challenge in large animal models. Here, we developed a robust and flexible insertional mutagenesis strategy, homology-independent targeted trapping (HIT-trapping), which is generic and can efficiently target-trap an endogenous gene of interest independent of homology arm and embryonic stem cells. Further optimization and equipping the HIT-trap donor with a site-specific DNA inversion mechanism enabled one-step generation of reversible and conditional alleles in a single experiment. As a proof of concept, we successfully created mutant alleles for 21 disease-related genes in primary porcine fibroblasts with an average knock-in frequency of 53.2%, a great improvement over previous approaches. The versatile HIT-trapping strategy presented here is expected to simplify the targeted generation of mutant alleles and facilitate large-scale mutagenesis in large mammals such as pigs.

Following the completion of animal genome sequencing projects, rapid and efficient mutagenesis strategies are needed for analyzing gene function and for creating human disease models. Gene trapping is a high-throughput mutagenesis strategy whereby random vector insertion can be achieved across the mouse genome. A typical gene-trap vector contains a promoter-less reporter/selection gene flanked by an upstream splice acceptor (SA) and a downstream poly(A) signal. Upon insertion into an intron of a gene, the vector both inactivates the trapped gene and enables the gene-specific expression of a reporter gene (Gossler et al. 1989; Stanford et al. 2001). To date, gene-trapping approaches have been successfully applied toward large-scale mutagenesis in mouse embryonic stem cells (mESCs) and generation of gene knockout mice (Skarnes et al. 2004). The main drawback of random gene trapping is that gene-trap alleles are not specifically engineered to target genes of interest in advance. Therefore, methods to streamline the introduction of predesigned, site-specific modifications into the genome by homologous recombination would represent a significant technological advance. Previously, a hybrid approach combining gene targeting and gene trapping (targeted trapping) enabled mutation of expressed genes in mESCs with high efficiency, using a gene-trap construct flanked by homologous sequences of the target locus (Friedel et al. 2005). Also, homologous recombination is commonly used for creating conditional alleles, which is essential to avoid embryonic lethality and to study the stage- and tissue-specific functions of genes (Branda and Dymecki 2004). However, both standard gene trapping and targeted trapping are only suitable for genes expressed in embryonic stem (ES) cells. Furthermore, construction of targeting donor vectors with homology arms is labor intensive and costly, and the low efficiency of homologous recombination is also a rate-limiting step for gene targeting in mammalian genomes.Recently, by taking advantage of precise genomic double-strand breaks (DSBs) created by the clustered regularly interspaced short palindromic repeats (CRISPR)-associated protein 9 (Cas9) system (Ran et al. 2013; Doudna and Charpentier 2014; Hsu et al. 2014), homology-directed repair (HDR) efficiency was substantially enhanced (Porteus and Carroll 2005), and even donors with short homology arms (Orlando et al. 2010) or single-stranded DNA oligonucleotides (Chen et al. 2011; Quadros et al. 2017) were found to be compatible with site-specific integration. However, each targeting donor for HDR still needs to be customized with gene-specific homology sequences. Because of the lack of ES cells for certain animals such as pigs, sheep, and cattle, the genome must be edited either in a zygote embryo or in a somatic cell for somatic cell nuclear transfer (SCNT) (Reddy et al. 2020). It is still not feasible to achieve large-scale insertional mutagenesis including conditional knockouts in these important species with random gene trapping or HDR-based methods. Also, the problem of genetic mosaicism in embryo editing remains unresolved (Mehravar et al. 2019), prompting a need for technological advances to accelerate genetic modification in somatic cells.Alternatively, the generally more efficient nonhomologous end joining (NHEJ) pathway has been exploited for site-specific insertion of exogenous DNA by simultaneous cleavage of both donor plasmid and genome using programmable nucleases (Cristea et al. 2013; Maresca et al. 2013; Brown et al. 2016; Suzuki et al. 2016; Sawatsubashi et al. 2018). In contrast to HDR-based strategies, NHEJ-mediated insertions do not require gene-specific homology arms, enabling diverse sites to be targeted with a universal donor vector. Therefore, we speculated that a gene-trap cassette could be inserted into a specific locus easily through this mechanism in any cell type.Here, by combining NHEJ-mediated knock-in and gene trapping, we developed a strategy for targeted mutagenesis, especially in somatic cells with low HDR activity, referred to as HIT-trapping. By using a universal donor, this strategy allows us to (1) create null alleles, (2) produce a fluorescent reporter signal that could potentially allow cells with null alleles to be identified very quickly, and (3) produce reversible and conditional alleles that would be very helpful to have in most animal models but are often cumbersome to create.