Introducing shear stress in the study of bacterial adhesion

During bacterial infections a sequence of interactions occur between the pathogen and its host. Bacterial adhesion to the host cell surface is often the initial and determining step of the pathogenesis. Although experimentally adhesion is mostly studied in static conditions adhesion actually takes place in the presence of flowing liquid. First encounters between bacteria and their host often occur at the mucosal level, mouth, lung, gut, eye, etc. where mucus flows along the surface of epithelial cells. Later in infection, pathogens occasionally access the blood circulation causing life-threatening illnesses such as septicemia, sepsis and meningitis. A defining feature of these infections is the ability of these pathogens to interact with endothelial cells in presence of circulating blood. The presence of flowing liquid, mucus or blood for instance, determines adhesion because it generates a mechanical force on the pathogen. To characterize the effect of flowing liquid one usually refers to the notion of shear stress, which is the tangential force exerted per unit area by a fluid moving near a stationary wall, expressed in dynes/cm(2). Intensities of shear stress vary widely according to the different vessels type, size, organ, location etc. (0-100 dynes/cm(2)). Circulation in capillaries can reach very low shear stress values and even temporarily stop during periods ranging between a few seconds to several minutes (1). On the other end of the spectrum shear stress in arterioles can reach 100 dynes/cm(2)(2). The impact of shear stress on different biological processes has been clearly demonstrated as for instance during the interaction of leukocytes with the endothelium (3). To take into account this mechanical parameter in the process of bacterial adhesion we took advantage of an experimental procedure based on the use of a disposable flow chamber (4). Host cells are grown in the flow chamber and fluorescent bacteria are introduced in the flow controlled by a syringe pump. We initially focused our investigations on the bacterial pathogen Neisseria meningitidis, a Gram-negative bacterium responsible for septicemia and meningitis. The procedure described here allowed us to study the impact of shear stress on the ability of the bacteria to: adhere to cells (1), to proliferate on the cell surface (5)and to detach to colonize new sites (6) (Figure 1). Complementary technical information can be found in reference 7. Shear stress values presented here were chosen based on our previous experience(1) and to represent values found in the literature. The protocol should be applicable to a wide range of pathogens with specific adjustments depending on the objectives of the study.     

Use of a centrifugal bioreactor for cartilaginous tissue formation from isolated chondrocytes

Although a centrifugal bioreactor (CCBR) supports high-density mammalian suspension cell cultures by balancing drag, buoyancy, and centrifugal forces, to date anchorage-dependent cultures have not been tried. Also, steady or intermittent hydrostatic pressures of 8 to 500 kPa, and shears of 0.02 to 1.4 N/m(2) can be simultaneously applied in the CCBR. This article demonstrates the use of a CCBR to stimulate chondrogenesis in a high-density culture. At 3 weeks, histological results show even distribution of glycosaminoglycan (GAG) and collagen, with 1,890 ± 270 cells/mm(2) cell densities that exceed those of 1,470 ± 270 in pellet cultures. Analysis of collagen content reveals similar levels for all treatment groups; 6.8 ± 3.5 and 5.0 ± 0.4 μg collagen/μg DNA for 0.07 and 0.26 MPa CCBR cultures, respectively, in contrast to 6.6 ± 1.9 values for control pellet cultures. GAG levels of 5.6 ± 1.5 and 4.1 ± 0.9 μg GAG /μg DNA are present for cultures stressed at 0.07 and 0.26 MPa, respectively, in comparison to control pellet cultures at the 8.4 ± 0.9 level. Although results to date have not revealed mechanical stress combinations that stimulate chondrogenesis over unstressed controls, system advantages include continuous culture at cell densities above those in the pellet, precise medium control, the ability to independently vary multiple mechanical stresses over a broad range, and the flexibility for integration of scaffold features for future chondrogenesis stimulation studies.     

iCare眼压计测量小鼠眼压值与真实眼压的相关性研究

目的:通过前房插管技术调节小鼠眼压,研究iCare眼压计测量小鼠眼压值与小鼠真实眼压之间的相关性。方法:取20只12周龄的C57BL/6J小鼠,麻醉后行前房穿刺置管,通过改变BSS液瓶高度调节其眼压。使用iCare眼压计测量其眼压值。采用SPSS软件分析iCare眼压计测量值和小鼠真实眼压之间的相关性。结果:iCare眼压计测量值与真实眼压之间相差较大,较真实眼压低约54%-71%,然而两者之间遵循如下线性回归方程:真实眼压(mmHg)=3.3988×(iCare眼压计测量值)-5.6292,rZ=0.9506,P<0.05。结论:iCare眼压计测量值较小鼠真实眼压低,然而在应用线性回归公式的情况下,iCare眼压计的测量修正值可以反映小鼠实际眼压值。     

不同时间服用比索洛尔对非杓型高血压血压的影响

目的:观察不同给药时间分别给予比索洛尔对非杓型原发性高血压患者的降压疗效和血压节律恢复的影响。方法:选取60例非杓型高血压患者,采取随机平行对照试验,观察比索洛尔(n=30)每日早晨(8:00)给药2.5-10mg、比索洛尔(n=30)每日夜间(20:00)给药2.5-10mg治疗8周后的降压疗效。结果:两种给药方法均能降低非杓型高血压患者的全天血压(P<0.05)。两种给药方法在白天的血压控制上无显著性差异(P>0.05),但在夜间血压的控制上夜间服药降压效果具有显著性差异(P<0.05)。夜间服药在血压节律恢复方面优于早晨服药(P<0.05),早晨服药组有10例恢复杓型,夜间服药组有19例恢复杓型。结论:比索洛尔的两种给药方式均能安全有效的降压,但对于非杓型高血压患者夜间服药优于早晨服药,更有利于血压节律的恢复。     

结缔组织生长因子在慢性阻塞性肺疾病血管重建中表达研究

目的:探讨结缔组织生长因子(CTGF)在慢性阻塞性肺疾病(COPD)血管重建中的表达及意义。方法:将30例有吸烟史的男性鳞癌需要手术的患者按其肺功能结果分成二组,对照组:(肺功能正常组);COPD稳定期组:(肺功能异常组),每组15例,标本来自于癌旁的肺组织,肺血管重塑的形态学观察行HE和MASSON三色染色,行免疫组化来观察CTGF蛋白、PCNA蛋白在肺血管平滑肌中的表达。结果:(1)COPD组肺动脉管壁面积/管总面积(WA%)、管壁的胶原厚度、肺动脉平滑肌中CTGF蛋白及PCNA蛋白的表达与对照组相比差异有统计学意义。(2)CTGF与管壁面积/管总面积(WA%)、管壁的胶原厚度及血管平滑肌中PCNA表达呈正相关(,r值分别为0.81、0.68、0.86,P<0.05)。吸烟指数与管壁面积/管总面积及PCNA的表达呈正相关(r=0.73,0.99,P<0.01)。结论:单纯吸烟者即有血管重建,吸烟伴COPD者血管重建更加严重,CTGF在COPD患者肺血管中的表达较对照组高,可能参与了COPD血管重建过程。     

循环纤维细胞及其在肺纤维化中的作用

循环纤维细胞(crculating fibrocytes CF)是一类骨髓来源的间质性细胞,可以同时表达造血细胞、单核细胞和成纤维细胞系的细胞标记。现有越来越多动物实验证据表明,CF的分化、转移功能在慢性炎症反应和胶原过度沉积中起着重要作用。近年来的实验表明,对CF的计数可以作为纤维化类疾病发展过程中的生物标记物,尤其在肺纤维化中更加明显。特发性肺纤维化是一种慢性肺部功能失调疾病,确诊后病人5年生存率低于30%。特发性肺纤维化是一种慢性肺部功能失调疾病,确诊后病人5年生存率低于30%。传统的免疫学疗法疗效很差。最近人们开始把注意力集中到对始祖干细胞的免疫调节机制上来。随着对循环纤维细胞研究的不断深入,对特发性纤维化的发病原因以及治疗研究方面有了较大发展。     

支气管激发试验对咳嗽变异性哮喘诊断和治疗的新思考

目的:通过分析我院经行支气管激发试验明确诊断的咳嗽变异性哮喘(CVA)的临床和肺功能特点,为诊断和治疗CVA提供一些有益的参考。方法:临床资料采用回顾性分析,共收集142例支气管激发试验阳性而经临床诊断明确为咳嗽变异型哮喘的病例,将其按年龄分为5组,分别对不同年龄组在分布例数、症状特点、肺通气功能、激发剂量进行对比分析。结果:在142例CVA患者中,<30岁组病例42例(29.6%),30~40岁组30例(21.1%),40~50岁组28例(19.7%),50~60岁组26例(18.3%),>60岁组16例(11.3%)。在症状方面,除均有慢性咳嗽(>3周)外,随着年龄的增大,各组中出现胸闷、气促等症状的病例比例逐渐增多。在肺通气功能方面,<30岁组的肺通气功能测定明显好于>60岁组。而另外3组之间组与组之间无明显差异,但各组与<30岁组及>60岁组之间均有明显差异。结论:咳嗽变异型哮喘患者以中青年患者居多,老年患者较少,随着年龄的增长,其临床表现及肺通气功能越来越接近典型支气管哮喘,由此推想,若有条件的医院能广泛开展支气管激发试验,对咳嗽变异型哮喘患者进行早期诊断。     

脐血干细胞与创面修复

脐血干细胞是一类具有多向分化潜能的原始祖细胞,具备自我更新和增殖的能力,在特定条件诱导下可以分化为不同细胞,逐渐作为临床组织工程的来源细胞。近年来随着对脐血干细胞的不断研究,发现其在创面修复中具有明显的优势,成为创面临床治疗的一条新途径。本文从脐血干细胞的生物学特性、采集与冻存、体外扩增等方面对创面修复的研究进行综述。     

支原体肺炎合并胸腔积液、肺不张27 例临床分析

目的:探讨肺炎支原体肺炎合并胸腔积液、肺不张的诊断和治疗问题。方法:回顾性分析27例MPP合并胸腔积液、肺不张患儿的临床特征、诊治过程的临床资料,并结合文献进行讨论。结果:在27例MPP患儿中,肺CT表现为胸腔积液17例,肺不张10例;24例治愈,1例胸膜肥厚粘连,2例遗留闭塞性细支气管炎。结论:对MPP合并胸腔积液、肺不张患儿应早诊断,早治疗,除应用大环内酯类药物外,应联合应用头孢菌素,激素及丙种球蛋白,疗效肯定。     

HA纳米载体介导转hGM-CSF基因的HepG2疫苗诱导的 抗瘤效应研究

目的:观察HA纳米颗粒载体介导转染hGM-CSF基因的HepG2细胞疫苗体外抗肿瘤效应,为hGM-CSF基因修饰的HepG2细胞疫苗的临床应用提供依据。方法:HA纳米颗粒载体介导hGM-CSF基因转染HepG2细胞制备转GMCSF基因的HepG2细胞疫苗。密度梯度离心法分离人PBMC,体外诱导人PBMC。WST-1法测定PBMC的增殖活性及对HepG2细胞的杀伤效应,流式细胞术分析CD4+和CD8+的阳性表达率,ELISA法测定INF-γ的分泌。结果:WST-1结果显示,转基因HepG2组疫苗能诱导PBMC增殖,其增殖率优于野生型疫苗(p<0.05);其诱导的PBMC对HepG2的杀伤率高于各野生型疫苗组和各空白对照组(p<0.05)。FCM结果显示,转基因HepG2疫苗组PBMC中CD4+和CD8+阳性表达率均高于各野生型疫苗组和各空白对照组(p<0.05)。ELISA结果显示,转基因组PBMC培养上清中IFN-γ含量为1989.76±254.21pg/ml,高于各野生型疫苗组和各空白对照组(p<0.05)。结论:HA纳米颗粒载体介导转染hGM-CSF基因能增加HepG2细胞疫苗的免疫原性,转hGM-CSF基因HepG2细胞疫苗可有效诱导PBMC增殖、分化,增加INF-γ的分泌,提高其对HepG2细胞的杀伤作用。