内毒素结合肽模拟肽P11的体外活性研究

目的对从噬菌体展示随机肽库筛选获得的内毒素结合肽模拟肽进行体外拈抗内毒素活性鉴定。方法采用FMOC固相合成法化学合成内毒素结合肽模拟肽P11,并进行拮抗内毒素活性和细胞毒性测定。结果亲和ELISA检测显示P11与LPS有较高的亲和力,通过生长曲线和流式细胞学分析细胞周期显示P11对人U937细胞生长无明显影响。流式细胞检测显示P11呈剂量依赖性抑制FITC—LPS与人外周血单核细胞（PBMC）结合。细胞因子生成抑制实验显示10μg/mlP11可显著抑制LPS诱导PBMC和U937细胞TNF—αmRNA转录和蛋白表达。结论体外活性鉴定结果表明化学合成的模拟肽P11可抑制LPS诱导的炎性反应。     

N-terminal brain natriuretic propeptide levels correlate with procalcitonin and C-reactive protein levels in septic patients

The aim of this study was to find the relationship between N-terminal brain natriuretic propeptide (NT-proBNP), procalcitonin (PCT) and C-reactive protein (CRP) plasma concentrations in septic patients. This was a prospective study, performed at Medical University Hospital No. 5 in łódź. Twenty patients with sepsis and severe sepsis were included in the study. N-terminal brain natriuretic propeptide, procalcitonin and C-reactive protein concentrations, and survival were evaluated. In the whole studied group (128 measurements), the mean NT-proBNP, procalcitonin and C-reactive protein concentrations were, respectively: 140.80±84.65 pg/ml, 22.32±97.41 ng/ml, 128.51±79.05 mg/l. The correlations for the NT-proBNP level and procalcitonin and C-reactive protein levels were 0.3273 (p<0.001) and 0.4134 (p<0.001), respectively. NT-proBNP levels correlate with PCT and CRP levels in septic patients. In the survivor subgroup, the mean NT-proBNP plasma concentrations were significantly lower than in the non-survivor subgroup.     

Quantitative lipopolysaccharide analysis using HPLC/MS/MS and its combination with the limulus amebocyte lysate assay

Quantitation of plasma lipopolysaccharides (LPSs) might be used to document Gram-negative bacterial infection. In the present work, LPS-derived 3-hydroxymyristate was extracted from plasma samples with an organic solvent, separated by reversed phase HPLC, and quantitated by MS/MS. This mass assay was combined with the limulus amebocyte lysate (LAL) bioassay to monitor neutralization of LPS activity in biological samples. The described HPLC/MS/MS method is a reliable, practical, accurate, and sensitive tool to quantitate LPS. The combination of the LAL and HPLC/MS/MS analyses provided new evidence for the intrinsic capacity of plasma lipoproteins and phospholipid transfer protein to neutralize the activity of LPS. In a subset of patients with systemic inflammatory response syndrome, with documented infection but with a negative plasma LAL test, significant amounts of LPS were measured by the HPLC/MS/MS method. Patients with the highest plasma LPS concentration were more severely ill. HPLC/MS/MS is a relevant method to quantitate endotoxin in a sample, to assess the efficacy of LPS neutralization, and to evaluate the proinflammatory potential of LPS in vivo.     

Strategies for Correcting Very Long Chain Acyl-CoA Dehydrogenase Deficiency

Very long acyl-CoA dehydrogenase (VLCAD) deficiency is a genetic pediatric disorder presenting with a spectrum of phenotypes that remains for the most part untreatable. Here, we present a novel strategy for the correction of VLCAD deficiency by increasing mutant VLCAD enzymatic activity. Treatment of VLCAD-deficient fibroblasts, which express distinct mutant VLCAD protein and exhibit deficient fatty acid β-oxidation, with S-nitroso-N-acetylcysteine induced site-specific S-nitrosylation of VLCAD mutants at cysteine residue 237. Cysteine 237 S-nitrosylation was associated with an 8–17-fold increase in VLCAD-specific activity and concomitant correction of acylcarnitine profile and β-oxidation capacity, two hallmarks of the disorder. Overall, this study provides biochemical evidence for a potential therapeutic modality to correct β-oxidation deficiencies.     

Scaffold compound L971 exhibits anti‐inflammatory activities through inhibition of JAK/STAT and NFκB signalling pathways

JAK/STAT and NFκB signalling pathways play essential roles in regulating inflammatory responses, which are important pathogenic factors of various serious immune‐related diseases, and function individually or synergistically. To find prodrugs that can treat inflammation, we performed a preliminary high‐throughput screening of 18 840 small molecular compounds and identified scaffold compound L971 which significantly inhibited JAK/STAT and NFκB driven luciferase activities. L971 could inhibit the constitutive and stimuli‐dependent activation of STAT1, STAT3 and IκBα and could significantly down‐regulate the proinflammatory gene expression in mouse peritoneal macrophages stimulated by LPS. Gene expression profiles upon L971 treatment were determined using high‐throughput RNA sequencing, and significant differentially up‐regulated and down‐regulated genes were identified by DESeq analysis. The bioinformatic studies confirmed the anti‐inflammatory effects of L971. Finally, L971 anti‐inflammatory character was further verified in LPS‐induced sepsis shock mouse model in vivo. Taken together, these data indicated that L971 could down‐regulate both JAK/STAT and NFκB signalling activities and has the potential to treat inflammatory diseases such as sepsis shock.     

EB病毒抗体和DNA联合检测可提高儿童传染性单核细胞增多症的诊断灵敏度

目的评估EB病毒抗体VCA-IgM、VCA-IgG、EA-IgG、EBNA-1-IgG及EBV-DNA载量检测在儿童传染性单核细胞增多症(传单)中的诊断意义。方法用ELISA方法检测70例传单患儿和25例健康儿童血清中EBV四种抗体及PCR荧光定量法检测外周血单个核细胞EBV-DNA载量。结果传单患儿组EBV-DNA的阳性率为87.14%(61/70),对照组阳性率为8.00%(2/25),传单组与对照组EBV-DNA的阳性率比较差异有统计学意义(P<0.01)。EBV抗体检测中,传单组的VCA-IgM阳性率最高,达91.43%(64/70),对照组VCA-IgM全部阴性。传单组EB病毒VCA-IgM和EBV-DNA联合检测的阳性率97.1%。结论 EBV抗体和EBV-DNA载量检测对儿童传单的诊断有极高的价值,尤其是VCA-IgM抗体和EBV-DNA联合检测,可提高儿童传单的临床诊断的敏感性。     

乌司他丁联合血必净治疗重症脓毒症的研究

目的：评估乌司他丁联合血必净注射液治疗重症脓毒症的临床疗效。方法：将符合入选标准的脓毒症患者40例,按照随机分组原则分为观察组和对照组,每组20例。所有患者给予常规抗感染治疗（根据培养结果给予敏感抗生素）和必要的对症支持治疗。观察组患者加用血必净注射液50ml静脉滴注,1次／12h,乌司他丁60万u静脉滴注,1次／12h,连用7d。记录治疗前后所有患者的体温、脉搏、呼吸、血白细胞（W33C）、降钙素原（PCT）、c反应蛋白（CRP）、急性生理学与慢性健康状况评分（APACHEII）、7天病死率。结果：疗程结束时观察组脉搏、呼吸、WBC、PCT、CRP、APACHEII评分、7天病死率均较治疗前明显下降,与对照组相比差异有统计学意义（P〈0．05）。结论：血必净注射液联合乌司他丁对重症脓毒症有明确疗效,改善临床症状,降低病死率。     

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.     

一氧化氮合酶与脓毒症的研究进展

脓毒症在外科临床工作中较常见,治疗相当困难;本文主要概述了一氧化氮合酶的基因定位、结构特点以及一氧化氮合酶与脓毒症的关系,进一步说明由一氧化氮合酶介导的一氧化氮生成与脓毒症关系密切,而选择性一氧化氮合酶抑制剂在脓毒症各阶段恰当的应用可能是有效治疗脓毒症、降低病死率的一个重要途径,也将成为今后研究的热点。