壳聚糖体内外抗幽门螺杆菌的实验研究

目的 研究壳聚糖体内外抗幽门螺杆菌（Hp）作用。方法 （1）采用打孔法检测了不同浓度、pH值、脱乙酰度壳聚糖及羧甲基壳聚糖在体外对蜘的抑菌作用。（2）建立BALB／c小鼠脚感染的动物模型后，随机分为8组：①对照组；②PPI组；③AM组；④AM＋PH组；⑤壳聚糖组；⑥壳聚糖＋PPI组；⑦壳聚糖＋AM组；⑧壳聚糖＋AM＋PPI组。分别给予上述药物每日2次灌胃，共2周。停药后4周，处死小鼠，无菌条件下取胃黏膜进行砷定量培养和石蜡包埋切片，进行Giemsa染色。结果 （1）壳聚糖和羧甲基壳聚糖在体外对3株邱标准菌株具有普遍的抑菌作用；在pH6-4范围内，随pH值降低，抗菌作用增强，差异有统计学意义（P〈0．01），最佳pH值为4；70％、88．5％脱乙酰度壳聚糖及羧甲基壳聚糖的抗伽作用差异有统计学意义（P〈0．05），抑菌强度依次为DD70壳聚糖、DD88．5，壳聚糖和羧甲基壳聚糖；在1％-5％浓度范围内羧甲基壳聚糖抗伽作用差异无统计学意义；在0．5％-2％浓度范围内70％、88．5％脱乙酰度壳聚糖抗伽作用差异也无统计学意义（P〉0．05）。（2）对饰感染的小鼠，以上8组的坤清除率分别为O％、O％、41．7％、58．3％、58．3％、66．7％、83．3％、91．7％，其中③-⑧组的坤清除率显著高于①和②组（P〈0．05），⑧组的坤清除率还显著高于③组（P〈0．05）。坤定植密度研究发现，各组之间坤定植密度差异有统计学意义（P〈0．001），饰定植密度在③一⑧组显著低于①和②组（P〈0．05）；⑥-⑧组显著低于③组（P〈0．05）；⑧组显著低于④组（P〈0．05）。结论 壳聚糖和其衍生物对坤有普遍的抑菌作用；壳聚糖及其衍生物的抗饰作用受多种因素影响，其中pH值对壳聚糖抗菌作用的影响最为明显，在pH值6—4范围内，壳聚糖的抗坤活性随着pH值的下降而显著增强；壳聚糖在体内有抗坤作用，并与AM有协同作用。     

新型聚己内酯／聚醚嵌段共聚物的合成及其释放蛋白药物的研究

目的合成一种含有活泼环氧基和羟端基的聚己内酯（PCL）／聚醚嵌段共聚物，这个新聚合物具有更理想的亲水性，并可通过与环氧基团的反应进一步衍生化。方法采用双官能团的环氧基化合物作为交联剂，通过化学偶联的方法合成嵌段共聚物，用^1H核磁共振（^1H—NMR）、热分析和接触角测定法表征其结构和性能。通过多官能环氧基化合物将肝素偶联到嵌段共聚物的端羟基上，用讯标记的肝素定量测定肝素的结合量和结合稳定性。采用溶液浇注和压膜方法制备含牛血清蛋白（BSA）的共聚物膜，37℃恒温摇床中进行体外模拟释放，用分光光度法（595nm波长）测定BSA的释放量。结果^1H—NMR测定证明共聚物中环氧基团（EO）对CL单元的比例与原始配方的计算值相符，热分析曲线表明嵌段共聚物有两个吸热峰，证明它具有微观相分离结构。接触角测定表明，随着共聚物中亲水组分的增加其亲水性增加。选择聚醚的类型和比例可以调控其亲水性，嵌段共聚物上化学偶联肝素的量可达到15％，具有明显高于物理混合的结合稳定性和很好的抗凝血活性。共聚物中亲水性聚醚组分的存在促进蛋白质的释放，而且通过改变聚醚片段的组成可以对蛋白质的释放速率进行调控。结论本研究提出了一种合成预设片段长度的聚酯-聚醚嵌段共聚物的新方法，并提供了一类新的肝素化的释放蛋白和多肽类药物的控释新材料。     

广东地区人禽流感H5N1毒株的核蛋白基因变异和进化

目的通过对人禽流感H5N1毒株核蛋白（NP）基因序列的变异分析，揭示毒株NP基因变异与进化。方法检测广东地区人禽流感H5N1毒株NP基因核苷酸序列，同时检索全球人禽流感H5N1毒株NP基因序列，采用DNAStar5．0软件对检索的人禽流感H5N1毒株NP基因核苷酸序列进行比对和分析；并结合流行病学资料对变异毒株进行进化速度分析。结果1997--2006年45株毒株胛基因核苷酸序列同源性分成3类，1997--1998年毒株为第1类，2004--2005年毒株为第2类，2003年毒株和2006年毒株为第3类；NP基因35个氨基酸位点全部置换，占7．03％（35／498）；2003—2006年H5N1毒株通过氨基酸第430位（1〈430T）位点的置换，增加一个糖基化位点（NGT430-432；GD—01—06毒株第370位发生N3加S变异；增加一个糖基化位点（NES368.370）。同义变异中，NP基因Ks（同义变异速度）值为2．03×10^-5～2．55×10^-5核苷酸／d，Ka（错义变异速度）值为1．58×10^-6～3．10×10^-6核苷酸／d，检验发现进化无明显选择性压力存在。结论1997--1998年毒株、2004--2005年毒株、2003年毒株和2006年毒株NP基因核苷酸有差异；2003--2006年人禽流感H5N1毒株NP基因增加一个糖蛋白位点、GD-01—06毒株再增加一个糖蛋白位点可能改变毒株抗原性。人禽流感H5N1毒株在自然界变异频繁，随着其自然进化，H5N1毒株具有人．人传播能力的概率较大。     

氧化应激条件下CD200对MG炎性反应的调控作用

目的探讨氧化应激条件下CD200如何参与调控MG（d＇胶质细胞）激活后的炎性反应。方法对原代培养的MG进行氧化应激处理，并利用急性分离的皮质神经元进行干预。“对照组”和“H2O2组”分别采用生理盐水和H2O2（100μmol／L）进行处理，“H2O2＋CD200组”在H2O2（100μmol／L）处理之前30min采用急性分离的神经元（约含1×10^6细胞）进行预处理。于处理后第0．5h、1h、2h、3h和4h时，采用Western blotting法对MG内p-p38 MAPK和IκB-α的蛋白表达量进行检测；以及第0h和72h时，采用ELISA法对培养介质内的TNF-α和IL-1β的含量进行检测。结果p-p38 MAPK和IκB—α的蛋白表达：“对照组”各时间点均无明显变化；“H2O2组”和“H2O2＋CD200组”0．5b时表达量亦基本相同；而3h时“H2O2组”p-p38 MAPK蛋白表达量明显高于“H2O2＋CD200组”；4h时“H2O2组”IκB-α的蛋白表达量明显低于“H2O2＋CD200组”。TNF-α和IL-1β含量：各组培养介质内0h时均未检出含有TNF-α和IL-1β；第72h时两细胞因子含量高低次序相同，即“对照组”〈“H2O2＋CD200”〈“H2O2组”。结论氧化应激条件下，CD200分子通过抑制MG胞质内p38 MARK的磷酸化，进而抑制了NF—κB的激活和炎性因子表达水平。     

内蒙古锡盟蒙族群体VWA、FGA、PENTA E基因座的多态性研究

目的 了解内蒙古锡盟地区蒙族人群VWA、FGA、PENTAE3个STR位点的遗传多态性分布并与汉族群体作对照，获得相应多态位点的群体遗传学数据。方法 采用多重PCR扩增技术和聚丙烯酰胺凝胶电泳方法，对263名蒙族个体进行调查。结果 VWA、FGA、PENTAE3个位点分别检出9、10、22个等位基因，多态性分布均符合Hardy—Weinberg平衡定律。杂合度(H)分别为0．8115、0．8487、0．9146，多态信息量(PIC)分别为0．7825、0．8298、0．9064。结论 上述3个位点均具有较高的杂合度和多态信息量，是较理想的遗传标记系统。同时表明蒙汉群体基因频率分布在VWA位点无显著性差异，而在FGA、PENTAE位点存在极显著性差异，为人类遗传学和民族学研究提供了依据。     

单纯疱疹病毒单克隆抗体靶抗原的鉴定及其编码基因的定位

用放射免疫沉淀的方法，鉴定单纯疱疹病毒(HSV)型共同性单克隆抗体(McAb)1D10的靶抗原是一分子量约为60，000道尔顿的糖蛋白，另6株McAbs 1A12．Mad-2、2D11．CM—D3．2A8及1C4的靶抗原的分子量在105，000-130，000之间。用Mcabs与HSV-1／HSV-2型间重组株作物理图谱的方法．定位出这些McAbs靶抗原的基因。McAb1 D10的靶抗原的编码基因定位于短单一序列(Us)上，在0．912-0.976遗传单位之间，这一区域与gD鳊码基因重叠，故认为是gD。另6株McAbs的靶抗原的蝙码基固定位于长单一序列(UL)上，在0．573-0．690遗传单位之间，这一区域与gC鳊码基因重叠，故认为是gC，其中，Mad-2和OM—D3分别为HSV-1和HSV-2型特异性，所以其靶抗原分别为gC-1和gC-2。ELISA阻断试验结果表明4株型共同性MeAbs 1A12．2D11．2A8和1C4，抗gC上4个独立的抗原位点。     

相同来源HIV-1 Env、Gag基因序列变异和宿主基因多态性与疾病进展关系的分析

目的探讨HIV—1基因序列变异和宿主基因多态性与疾病进展的关系。方法PCR方法从外周血细胞中扩增HIV—1 Env、Gag区片段和测序，分析序列变异、糖基化，超突变等指标。RFLP法确定宿主基因多态性。结果未治疗组中，env基因区PCR和克隆序列平均离散率分别为0．1和0．06，差异有统计学意义，治疗组内差异没有统计学意义。V3环顶端序列在未治疗和治疗组中均以GPGQ比例最大（61．5％和39％），治疗组出现稀有多肽序列如GPGH、GQGR、GLGR、12位I／V和21位Y／H变异与疾病快速进展相关。Env区段上进展较快速组（RRP）比典型进展组（TP）的糖基化程度高（平均值分别为14．56和13．20个），差异有统计学意义。Env区段上RRP比TP组GA取代百分率和绝对数平均值都高（8．7％／6．9％和10．1／7．6），差异也有统计学意义。TP组中SDF1—3’A和CCR2V62I基因频率均高于RRP组，但差异没有统计学意义。CX3CR1 V249I／M280T与疾病快速进展没有显著相关性。结论V3区序列主要位点的氨基酸变异、Env区段糖基化程度、GA取代与疾病进展相关。SDF1—3’A、CCR2V641和CX3CR1 2491／280M与疾病进展均无显著相关性。     

高碳酸血症对丙泊酚／瑞芬太尼静脉全麻下熵指数监测的影响

目的观察丙泊酚／瑞芬太尼全身麻醉下行腹腔镜手术时高碳酸血症对患者麻醉深度及熵指数监测的影响。方法22例行妇科腹腔镜手术通过积极增加分钟通气量仍未能纠正高碳酸血症，需降低气腹压力进一步处理的病人，观察30min后按呼气末二氧化碳分压（PetCO2）进行分组，8例病人PetCO2波动于51-55mmHg，归为I组；6例病人波动于46～50mmHg，归为Ⅱ组；8例病人波动于40～45mmHg，归为Ⅲ组。分别记录麻醉前（T1）、切皮后（T2）、CO2气腹30min（T3）、降低CO2腹压30min后（T4）、停止麻醉时（T5）、拔除气管导管时（拍）的RE、SE、MBP、HR值，并记录停止麻醉至拔除气管导管的时间（T5—6）。结果I组病人T4和T5的RE、SE值均低于Ⅲ组（P〈0．05）；H组RE、SE值各时间点与其他两组差异无统计学意义。I组病人T4和T5RE、SE值低于T2和T3（P〈0．05）；I组病人意识恢复时间T5—6长于Ⅱ组和Ⅲ组（P〈0．05）。结论应用丙泊酚／瑞芬太尼在妇科腹腔镜手术时，CO2气腹造成的高碳酸血症可使熵指数下降和意识恢复时间延长，在麻醉中应予以重视。     

Surface properties and hemocompatibility of alkyl-siloxane monolayers supported on silicone rubber: effect of alkyl chain length and ionic functionality.

Self-assembled monolayers of alkylsiloxanes supported on poly(dimethylsiloxane) (PDMS) rubber were used as model systems to study the relation between blood compatibility and surface composition. The inner lumen of PDMS tubes were first treated with an oxygen plasma. The resultant oxidized surfaces were post-derivatized by reaction with alkyltrichlorosilanes to form the monolayer films. The alkyl chain lengths used were slightly longer than in a previous study, and this may alter the phase-state of the monolayer from liquid-like to crystalline. The chemical properties of the monolayer were controlled by varying the chemical composition of the alkyltrichlorosilanes used. Terminal functionalities included -CH3, -CF3, -COOH, -SO3H and -(CH2CH2O)4OH. Surface derivatization was verified with static contact angle measurements and X-ray photoelectron spectroscopy. Blood compatibility was evaluated using a canine ex vivo arterio-venous series shunt model. Surfaces grafted with hydrophobic head groups such as -CH3 and -CF3 were significantly less thrombogenic than the surfaces composed of ionic head groups such as -COOH and -SO3H. Surfaces enriched in -(CH2CH2O)4OH had an intermediate thrombogenicity. Silastic pump grade tubing and polyethylene tubing, used as controls, were found to be the least thrombogenic of all the surfaces tested.     

Novel pH-sensitive chitosan-based hydrogel for encapsulating poorly water-soluble drugs

Carboxymethyl–hexanoyl chitosan (CHC) is an amphiphilic chitosan derivative with excellent swelling ability and water solubility under natural conditions. In this work, the influence of the degree of carboxymethyl and hexanoyl substitution on the pH-sensitive swelling behavior, drug release behavior, and antiadhesion behavior of CHC hydrogels (cross-linked with genipin) were studied. It was found that the pH sensitivity was more pronounced in CHC than in N,O-carboxymethyl chitosan because the hexanoyl group altered the state of water in CHC by inhibiting intermolecular hydrogen bonding. In addition, greater pH sensitivity was observed in samples bearing longer hydrophobic chains (carboxymethyl–palmityl chitosan). Interestingly, when used with ibuprofen (a poorly water-soluble therapeutic agent used here as a model drug), the bursting release of the drug was less prominent in the CHC samples having a high degree of carboxymethyl substitution. The CHC hydrogel also demonstrated good cell compatibility and its antiadhesive ability after grafting was altered by changes in the degree of hexanoyl substitution.     

Synthesis, surface characterization, and platelet reactivity evaluation for the self-assembled monolayer of alkanethiol with sulfonic acid functionality

Owing to the capability of fabricating a well-defined chemical structure on the surface, self-assembled alkanethiols with a variety of terminal functionalities were prepared on the gold substrate for investigating the interactions between the biological environment and synthetic surface. In this study, we report the synthesis of the sulfonic acid terminated long-chain alkanethiol, 10-mercaptodecane-sulfonic acid, for direct preparation of a self-assembled monolayer (SAM) with -SO(3)H functionality. Nuclear magnetic resonance (NMR) and elemental analysis studies indicated that a high purity of sulfonic acid terminated alkanethiol was obtained. Surface characterization results showed that the -SO(3)H terminated SAM is hydrophilic and has a slightly higher hysteresis value, possibly because of the slower chain mobility of the bound sulfonic acid alkanethiol. Electron spectroscopy for chemical analysis (ESCA) analysis demonstrated that the -SO(3)H terminal group is situated in the outermost layer of the monolayer, as previous alkanethiol SAM structure models proposed. The platelet reactivity of the -SO(3)H SAM was higher than that of -OH SAM but less than the -CH(3) terminated one in vitro, whereas similar platelet reactivity was noticed between the -SO(3)H and -COOH SAMs. The higher platelet reactivity found on the -SO(3)H SAM could be caused by the higher surface functional group density inherent in the SAM structure and/or the composition and conformation state of the adsorbed protein layer.     

壳聚糖球形多孔微载体的血液相容性评价

背景：文献报道的微载体大多为实心的、大孔的,虽然较二维微载体的比表面积有明显的增加,但距理想的三维微环境相差甚远。 目的：构建壳聚糖球形多孔微载体,通过溶血实验、凝血实验、血小板计数及聚集实验评价其血液相容性。 方法：利用液氮冷冻干燥技术成功构建浓度为1%,2%,3%的壳聚糖球形多孔微载体。选择健康成年新西兰兔为宿主,采用溶血实验、凝血实验、血小板计数及其聚集实验评价壳聚糖球形多孔微载体的血液相容性。 结果与结论：浓度为1%,2%,3%的壳聚糖球形多孔微载体的溶血率分别为1.56%,2.07%,2.31%,均小于5%,均无致溶血性;3种浓度壳聚糖球形多孔微载体样本材料对兔血时间无明显影响,三者与生理盐水阴性对照组间也无明显差别(P > 0.05);3种浓度壳聚糖球形多孔微载体样本材料对兔血小板计数无明显影响,注入浸提液前后比较和组间比较差异均无显著性意义(P > 0.05)。证实壳聚糖球形多孔微载体无致溶血性、无凝血性和无血小板聚集性,表明壳聚糖球形多孔微载体具有良好的血液相容性。     

6-羧基壳聚糖的制备及血液相容性研究

将壳聚糖溶于0.5mol/L的乙酸,再以NO2选择性氧化壳聚糖分子中氨基葡萄糖单元的6位羟甲基成羧基,制备了6-羧基壳聚糖.红外光谱分析表明该反应选择性较高,产物保持了壳聚糖基本的链结构;扫描电镜和DSC分析发现,6-羧基壳聚糖与壳聚糖的微纤结构不同,其分子构象卷曲,热稳定性较壳聚糖差.抗凝血测试、溶血测试及扫描电镜观察表明：与壳聚糖相比,6-羧基壳聚糖抗凝血性能大大提高,溶血率明显降低,6-羧基壳聚糖具有很好的血液相容性,其血液相容性随壳聚糖氧化度的提高而提高.     

烷基化壳聚糖的制备及止血效果

BACKGROUND: 18-Alkylated-chitosan has been proved to exhibit a good coagulation capacity through hydrophobic modification. OBJECTIVE: To improve the adhesion between the material and the wound through the introduction of alkylated-chitosan molecules. METHODS: The alkyl group was introduced in the chitosan by potentiodynamic method to prepare the alkylated-chitosan with different substitution degrees (7%,16%, 26% and 40%), and then their structure, viscosity, porosity, water absorption, contact angle and in vitro coagulation capacity were detected. Thirty New Zealand rabbits were enrolled, modeled into the left femoral artery hemorrhage, and randomized into six groups, including four experimental groups, positive control and control groups, followed by treated with alkylated-chitosan sponges with different degrees of substitution, absorbable hemostatic gauze and chitosan sponge, respectively. Afterwards, the blood loss and clotting time were observed. RESULTS AND CONCLUISON: With the degree of substitution increased, the viscosity of alkylated-chitosan increased firstly and then decreased, the porosity and water absorption decreased gradually, and the contact angle on a rise. Compared with chitosan, different alkylated-chitosans showed a better henostatic effect in vitro, especially the alkylated-chitosan with 16% substitution degree. The blood loss and clotting time in the experimental groups except that the alkylated-chitosan with 7% substitution degree were significantly lower than those in the control group (P < 0.05); the bold loss in the alkylated-chitosan with 40% substitution degree group was significantly lower than that in the positive control group (P < 0.05). These findings suggest that the alkylated-chitosan with 40% substitution degree exhibits the best hemostatic effect in vivo.     

Synthesis of metal incorporated low molecular weight polyurethanes from novel aromatic diols, their characterization and bactericidal properties

Low molecular weight polyurethanes with sites for metal complexation were synthesized. The -SO(2) and the -COOH groups were incorporated into the polyurethane chain by the reactions of tri-functional monomers, 4,4'-bis(hydroxyphenyl)sulfone and 4,4'-bis(hydroxyphenyl) valeric acid with hexa-methylene di-isocyanate (HMDI) and 2,4 toluene di-isocyanate (TDI), respectively. The reaction was monitored from the disappearance of the -OH group and results show that the -NCO groups of the isocyanate reacted with the -OH group preferentially when compared to the other reactive groups. The pristine tri-functional monomers themselves formed stable complexes with the metals and so were chosen to be incorporated into the polyurethane chain. These polymers were characterized by IR and NMR spectroscopy. The free -SO(2) and the -COOH groups were used for metal complexation using non-toxic metals like zinc and silver. The anti-bacterial studies conducted on the six polymers in film form showed interesting results as elaborated in the paper.     

The influence of hydrogel functional groups on cell behavior

This study provides information on the behavior of macrophages on the surface of the subcutaneously implanted hydrogel strips. Hydrogel containing -OH, -CO-NH-, and (CH3)2N- groups induced a spreading of macrophages on the implants. The materials containing -SO3H groups slightly, and materials containing -COOH groups more intensively, inhibited spreading of the macrophages. The fusion of macrophages into multinucleate cells was inhibited on the surface of materials containing acidic groups (-SO3H, -COOH) and increased on the hydrogels containing 30 mol% of alkaline (CH3)2N- groups in comparison with hydrogels containing -OH or -CO -NH- groups. The differences of the behavior of macrophages on the surface of individual types of hydrogels are probably independent on the adsorption of plasma fibronectin onto the hydrogels. The correlation between the macrophages spreading and fusion and surface charge of the hydrogel implant can be hypothetically explained by electrostatic interaction between macrophages cell membrane and implant.     

Adsorption of bilirubin by amine-containing crosslinked chitosan resins

Crosslinked chitosan resin, which has good blood compatibility, was prepared by crosslinking chitosan solution with glutaraldehyde. Polymeric adsorbents for bilirubin have been synthesized by chemical modification of crosslinked chitosan resins with di-, tri-, tetra-, and pentammines. The adsorption of bilirubin on the adsorbents was investigated in detail. The results indicated that electrostatic interaction and hydrophobic interactions are the main driving forces for the adsorption. Compared with crosslinked chitosan resin, the functionalized chitosan beads have higher adsorption capacity for unconjugated and conjugated bilirubin.     

Surface characterization and platelet compatibility evaluation of surface-sulfonated chitosan membrane

The effect of various sulfonated derivatives of chitosan on platelet activation and blood coagulation was examined. The surface properties of artificial biomaterials have been thought as the key factors which mediate the interactions between the biological environment and biomaterial itself. In this study, the sulfonation was directly performed on the chitosan membrane surface. The chitosan membrane was surface-sulfonated by reactions with sulfur-pyridine trioxide complex (SO3/pyridine) in H2O solution and N,N-sulfur-dimethylformamide trioxide complex (SO3/DMF) in DMF. Blood compatibility was evaluated by an in vitro platelet adhesion assay. The surface reaction of SO3/pyridine in aqueous acid medium yields N,O-sulfated chitosan with cationic NH3+ groups. After neutralization, this surface has been shown to induce a low degree of platelet adhesion and activation. When the surface-sulfonation is performed in an aqueous alkaline medium, although the degree of sulfonation is lower than the samples above, the N-sulfated chitosan significantly reduced the adhesion and activation of platelets. For the acidic SO3/DMF reaction system, N,O-sulfated chitosan was obtained with a high extent of sulfonation and cationic NH3+ groups. On this surface fully spread platelets and some platelet aggregates were found instead. This may be attributed to the ionic interactions between the platelets membrane surface and the cationic groups on the modified chitosan membrane.