Preparation of immunomagnetic iron-dextran nanopartides and application in rapid isolation of E.coli O157:H7 from foods

AIM: To prepare a kind of magnetic iron-dextran nanopartides that was coated with anti-E.coli O157:H7 IgG, analyze its application conditions, and try to use it to isolate E.coli O157:H7 from foods. METHODS: Magnetic iron-dextran nanopartides were prepared by the reaction of a mixture of ferric and ferrous ions with dextran polymers under alkaline conditions. The particles were coated with antiserum against E.coli O157: H7 by the periodate oxidation-borohydride reduction procedure. The oxidation time, amount of antibody coating the particles, amount of nanoparticles, incubation time and isolation time were varied to determine their effects on recovery of the organisms. Finally, the optimum conditions for isolating E.coli O157:H7 from food samples were established. RESULTS: E.coli O157:H7 can be isolated from samples within 15 min with the sensitivity of 101 CFU/mL or even less. In the presence of 108 CFU/mL of other organisms, the sensitivity is 101-102 CFU/mL. Nonspecific binding of other bacteria to the particles was not observed. Two and a half hours of enrichment is enough for the particles to detect the target from the food samples inoculated with 1 CFU/g. CONCLUSION: Isolation of target bacteria by immuno magnetic nanoparticles is an efficient method with high sensitivity and specificity. The technique is so simple that it can be operated in lab and field even by untrained personnel.     

老年人血脂水平与载脂蛋白E等位基因的关系

为了解老年人血脂水平与载脂蛋白Ｅ基因多态性的关系，并探讨载脂蛋白Ｅ等位基因对老年人血脂水平的内在影响，利用ＰＣＲ－ＲＦＬＰ法测定了载脂蛋白Ｅ基因型，并用酶学方法对其血脂水平进行了测定。结果发现存在五种不同基因型：与携有ε３等位基因的人群相比，携有ε４等位基因的人群倾向具有较高的总胆固醇和低密度脂蛋白胆固醇水平以及较低的甘油三酯水平，而携有ε２等位基因的人群则有较低的低密度脂蛋白胆固醇水平以及较高的     

人周围血单核细胞载脂蛋白E基因表达的竞争性逆转录-聚合酶链反应定量分析

为建立人周围血单核细胞载脂蛋白E基因表达检测方法,研究载脂蛋白E基因与儿童健康的关系,我们抽取26例健康儿童外周静脉血,分离血单核细胞,抽提RNA,采用逆转录-聚合酶链式反应检测载脂蛋白E基因表达,并以正常人cDNA作定量标准物,待测样品与定量标准物共扩增,计算出待测样本的个体的mRNA量.研究发现载脂蛋白E基因能在健康儿童周围血单核细胞表达,健康儿童载脂蛋白E基因表达量为0.37±0.15 mol/mol mRNA.表明采用竞争性逆转录-聚合酶链反应方法来检测人周围血单核细胞载脂蛋白E基因表达的分析方法快速、简便、灵敏、实用、可靠,且能准确定量,值得推广应用.     

Differing roles of protein kinase C-zeta in disruption of tight junction barrier by enteropathogenic and enterohemorrhagic Escherichia coli

BACKGROUND & AIMS: Enteropathogenic Escherichia coli and enterohemorrhagic E. coli harbor highly homologous pathogenicity islands yet show key differences in their mechanisms of action. Both disrupt host intestinal epithelial tight junctions, but the effects of enteropathogenic E. coli are more profound than those of enterohemorrhagic E. coli. The basis for this is not understood. The atypical protein kinase C isoform, protein kinase C-zeta, associates with and regulates the tight junction complex. The aim of this study was to compare the role of protein kinase C-zeta in the disruption of tight junctions after infection with enteropathogenic E. coli and enterohemorrhagic E. coli. METHODS: Model intestinal epithelial monolayers infected by enteropathogenic E. coli or enterohemorrhagic E. coli were used for these studies. RESULTS: Neither bisindolylmaleimide nor G?6976, which block several protein kinase C isoforms but not protein kinase C-zeta, protected against the decrease in transepithelial electrical resistance after enteropathogenic E. coli infection. Rottlerin at concentrations that block novel and atypical isoforms, including protein kinase C-zeta, significantly attenuated the decrease in transepithelial electrical resistance. The specific inhibitory peptide, myristoylated protein kinase C-zeta pseudosubstrate, also significantly decreased the enteropathogenic E. coli -associated decrease in transepithelial electrical resistance and redistribution of tight junction proteins. In contrast to enteropathogenic E. coli, the level of protein kinase C-zeta enzyme activity stimulated by enterohemorrhagic E. coli was transient and minor, and protein kinase C-zeta inhibition had no effect on the decrease in transepithelial electrical resistance or the redistribution of occludin. CONCLUSIONS: The differential regulation of protein kinase C-zeta by enteropathogenic E. coli and enterohemorrhagic E. coli may in part explain the less profound effect of the latter on the barrier function of tight junctions.     

Valvular heart disease in osteogenesis imperfecta.

Aortic and mitral valve abnormalities have been reported which clearly appear to be related to the underlying connective tissue disorder in two patients, a father and daughter, with osteogenesis imperfecta. Although this appears to occur with a much lower prevalence and lesser severity than in the Marfan syndrome, the true prevalence of cardiac connective tissue involvement is not known, and the orthopedic complications of osteogenesis imperfecta may have overshadowed attention to cardiovascular abnormalities. In evaluating patients with osteogenesis imperfecta, careful attention should be paid to cardiovascular findings and if valvular lesions are noted, patients should be instructed regarding the need for antibiotic prophylaxis for dental and surgical procedures. The valvular lesions can progress, and regular follow-up cardiovascular evaluation should be planned. Finally, despite potential problems with tissue friability and healing and a possible tendency for increased bleeding, successful valve replacement can be carried out if necessitated by cardiac disability.     

Augmentation of tumor necrosis factor family-induced apoptosis by E3330 in human hepatocellular carcinoma cell lines via inhibition of NFκB

AIM: To investigate the reduction of cell viability in human hepatocellular carcinoma (HCC) cell lines induced by inhibition of nuclear factor κB (NFκB).METHODS: HLE, SKHep1, and HepG2 were incubated and E3330 was used to compare the stimulation of some chemotherapeutic drugs with that of TNF family, Fas ligand, TNFα and TNF-related apoptosis-inducing ligand (TRAIL) at the point of the reduction of cell viability by inhibiting NFκB.RESULTS: E3330 decreased NFκB levels in HLE cells stimulated by TNF and TRAIL. The cytotoxicity of the combination of TRAIL, TNFα, Fas ligand, and E3330increased synergistically in a dose-dependent manner compared to either E3330 alone in all HCC cell lines by MTT assay. However, the combination of some chemotherapeutic drugs and E3330 did not decrease the cell viability.CONCLUSION: Inhibition of NFκB sensitizes human HCC cell lines to TNF-mediated apoptosis including TRAIL, and TRAIL-based tumor therapy might be a powerful potential therapeutic tool in the treatment of human HCC.     

Evidence for a flow-independent contribution to the phenomenon of thallium redistribution

Although thallium-201 is known to redistribute slowly into regions of ischemic myocardium after restoration of blood flow, it is not clear to what extent normalization of flow is an essential requirement for the redistribution process. In a search for a flow-independent component of thallium redistribution, 12 dogs with stenosis of the circumflex coronary artery underwent atrial pacing for either 20 minutes (group I, 6 dogs) or 2 hours (group II, 6 dogs). Radioactive thallium and radioactive microspheres, 7 to 10 μ, were injected after 10 minutes of atrial pacing in both groups. Pacing resulted in a 40 percent reduction in subendocardial blood flow to the circumflex-perfused myocardium in both groups I and II. This relative reduction in flow was maintained at a stable level over the 2 hour pacing period in group II. Thallium activity in the relatively ischemic zone was significantly greater in dogs with 2 hours of pacing (group II) than in those with 10 minutes of pacing (group I). Redistribution of thallium occurred despite the continued presence of reduced flow in circumflex-perfused endocardial tissue. These data suggest that a significant component of thallium redistribution may be flow-independent.     

EnterohemorrhagicEscherichia coli (EHEC) in Pediatric Hemolytic-Uremic Syndrome: A Prospedtive Study in Germany and Austria*

Summary Studies from Europe indicate that infections with enterohemorrhagic Escherichia coli (EHEC) non-O157:H7 strains are increasing in frequency as a cause of hemolytic-uremic syndrome (HUS). In 1997 a prospective study was performed in Germany and Austria to assess the distribution of EHEC serotypes, to characterize the clinical course and to examine environmental aspects. 95 children with a diagnosis of HUS were evaluated in Germany and Austria. Diarrhea, which was bloody in 67%, was reported in 97% of patients. Oligo-/anuria occurred in 76% of patients, of which 63% required dialysis. Two patients showed neurological sequelae at the 2-month follow-up, both of them were infected with non-O157;H7 serotypes. Case fatality in the acute stage was 3/95, in two of these patients EHEC was isolated. Stool and serum specimens were analyzed for the presence of EHEC and antibodies against O157 lipopolysaccharide (LPS). Serotype O157:H7 was identified in 36/58 (62%) isolates, 22 strains (38%) belonged to non-O157:H7. Combining stool culture with serology, EHEC infection was documented in 88% of patients, including three patients without diarrhea. Non-O157:H7 serotypes occurred in 77% of children up to 36 months of age and were the most prevalent serotype in children up to 12 months of age. Received: February 9, 1999 · Accepted: September 19, 1999     

Effect of D-600 on ischemic and reperfused rabbit myocardium: relation with timing and modality of administration

Summary In this study we have investigated the possibility that D-600, a phenylalkylamine calcium antagonist, protects the isolated rabbit heart against ischemia and reperfusion-induced damage.D-600 was either subcutaneously injected (2 mg/kg, twice daily for 5 to 6 days) in the rabbit before isolation of the heart, or delivered to the isolated hearts in the perfusate (10^–7 M), either at the onset of ischemia and during reperfusion, or only during post-ischemic reperfusion.Ischemia (90 min) was induced by reducing coronary flow from 25 to 1 ml/min, followed by 30 min of reperfusion. Myocardial damage was determined in terms of mechanical function, release of creatine phosphokinase (CPK) and noradrenaline, mitochondrial function, calcium homeostasis, and endogenous stores of ATP and creatine phosphate (CP). Administration of D-600 to the rabbits or to the isolated hearts at the time of ischemia exerted protection. There are four groups of evidence in support of this conclusion: 1) the rise in diastolic pressure during ischemia was diminished with greater recovery of developed pressure during reperfusion; 2) CPK and noradrenaline release during reperfusion were reduced; 3) the oxygen consumption and ATP generating capacities of mitochondria were better maintained; and 4) associated with this preservation of mitochondrial function was the maintenance of near normal calcium homcostasis and of endogenous ATP and CP stores. The two different modalities of administration did not produce substantially different results.When administered to the isolated hearts after the ischemic period, D-600 failed to improve mechanical recovery and release of endogenous substances. However, it reduced mitochondrial calcium overload and improved ATP production. The mechanism of the protective effect of D-600 seems to be multiple: energy-sparing effect, reduction of the toxicity mediated by endogenous catecholamines, and direct inhibition of mitochondrial calcium transport.