Microparticle-associated tissue factor activity correlates with plasma levels of bacterial lipopolysaccharides in meningococcal septic shock

Introduction

The plasma level of bacterial lipopolysaccharides (LPS) is associated with activation of the coagulation system, inhibition of fibrinolysis and the nature of the clinical presentation and outcome in patients with meningococcal disease. Tissue factor (TF)-bearing microparticles (MPs) appear to contribute to the pathogenesis of disseminated intravascular coagulation (DIC). The aim of this study was to investigate the relationship between MP-associated TF activity and the level of bacterial LPS in plasma from patients with meningococcal septic shock and meningitis.

Materials and methods

MPs isolated from citrated plasmas were assessed for TF-dependent activity with both a plasma-based thrombin generation assay (CAT) and whole blood-based thromboelastometry (ROTEM). The LPS level was measured using a chromogenic Limulus amebocyte lysate assay.

Results

MPs obtained from patients with meningococcal septic shock initiated significantly more efficient and TF-dependent thrombin generation in the CAT assay compared to MPs from patients with meningococcal meningitis. Differences in MP-associated TF activity between the septic shock patients and the meningitis patients were also evident when MPs were added to whole blood using ROTEM. The level of plasma LPS in patients with septic shock (range 2–2,100 EU/mL) was correlated with thrombogram parameters in the CAT assay; lagtime (r_s = − 0.84), time to peak (r_s = − 0.83), peak (r_s = 0.85) and ETP (r_s = 0.83).

Conclusions

MPs obtained from patients with meningococcal septic shock displayed more efficient TF-dependent thrombin generation and clot formation compared to MPs from meningitis patients. MP-associated TF activity was closely associated with plasma LPS levels in the septic shock group.     

The effect of chronic exposure to highly aggressive mice on hippocampal gene expression of non-aggressive subordinates

Exposure to a chronic psychosocial stressor changes the behavioral and neuroendocrine response pattern and causes structural changes in the rodent hippocampus. However, the underlying molecular mechanism of these changes induced by chronic stress is largely unknown. Recently, it was shown that exposure to a dominant highly aggressive mouse in the sensory contact model induced long-lasting stress symptoms in subordinate mice genetically selected for long attack latency (LAL mice). The aim of the present study was to study the effect of chronic stress on hippocampal gene expression in these subordinate LAL mice. GeneChips (Affymetrix) were used to compare gene expression profiles of LAL mice exposed to a sensory contact stressor for 25 days and their controls (one array per mouse, n=5 per line). After this stress paradigm, 131 genes were found differentially expressed (P<0.01). Strikingly, all of these genes showed a subtle downregulation in response to a chronic stressor. Interestingly, a significant overrepresentation of genes encoding structural components of ribosomes were found, suggesting diminished protein biosynthesis in the hippocampus of chronically stressed LAL mice. In addition, several genes of the NFkappaB signaling cascade, a pathway crucially involved in neuronal viability and neurite growth, were found to be downregulated. Together, we hypothesize that reduced NFkappaB signaling and diminished protein biosynthesis form part of the molecular mechanisms by which a chronic psychosocial stressor induces structural alterations in hippocampus of LAL mice.     

The Antioxidant System β-D(+) Glucose–Glucose Oxidase–Catalase: Tests for Pyrogenicity and Antigenicity

Purpose: β-D(+) glucose–glucose oxidase (E.C.# 1.1.3.4)–catalase (E.C.# 1.11.1.6) (GO–CAT) is being investigated as a new antioxidant system for use in pharmaceutical solutions. This study reports the results of tests for pyrogenicity and antigenicity of GO–CAT derived from Aspergillus niger when used parenterally in autoclaved preparations. Methods: The Limulus amebocyte lysate (LAL) method was used to test the pyrogenicity of native GO–CAT. Pyrogenicity/antigenicity was evaluated in vivo by injecting autoclaved GO–CAT into New Zealand white rabbits. Antigenicity was also evaluated by Ouchterlony and Western blotting. Results: None of the native GO–CAT concentrations tested (up to 30.83 u/ml) produced a positive gel clot in the LAL test, thereby suggesting its non-pyrogenicity. The rabbits, which received seven injections of autoclaved GO–CAT over a period of eleven weeks, remained healthy during and after the GO–CAT injections. All Ouchterlony and Western blot assays using sera from rabbits injected with autoclaved GO–CAT were negative. Furthermore, autoclaved GO–CAT could not be detected in Ouchterlony assays using a mouse monoclonal antibody (GO40 mAb) to native A. niger glucose oxidase. Control samples containing native GO–CAT produced an antigen–antibody complex reaction in Ouchterlony assays against the GO40 mAb. Antigen-antibody complexes could be detected by non-denaturing PAGE in samples containing native GO–CAT/GO40 and boiled GO–CAT/GO40, but not in samples containing autoclaved GO–CAT/GO40. These results indicate autoclaved GO–CAT is neither pyrogenic nor antigenic. Conclusions: Based on these results, there is potential for the use of β-D(+) glucose–glucose oxidase–catalase as an antioxidant system in pharmaceutical solutions, particularly in terminally autoclaved aqueous formulations for parenteral use.     

Long-term neurobiological consequences of ecstasy: A role for pre-existing trait-like differences in brain monoaminergic functioning?

This study investigated whether trait-like differences in brain monoaminergic functioning relate to differential vulnerability for the long-term neurochemical depletion effects of MDMA. Genetically selected aggressive (SAL) and non-aggressive (LAL) house-mice differing in baseline serotonergic and dopaminergic neurotransmission were administered MDMA. An acute binge-like MDMA injection protocol (three times, using either of the dosages of 0, 5, 10 and 20 mg/kg i.p. with 3 h interval) was employed. Three and 28 days after treatment with MDMA induced a dose-dependent depletion of striatal dopamine and its metabolites that did not differ between SAL and LAL mice. Similarly, the dose-dependent MDMA-induced serotonergic depletion did not differ between lines 3 days after treatment. Interestingly, 28 days after MDMA in LAL mice, 5-HT and 5-HIAA levels were still significantly depleted after treatment with 3 × 10 mg/kg, while in SAL mice 5-HT depletion was only seen after the highest dosage. Surprisingly, LAL mice did not show any long-term 5-HT depletion after treatment with the highest dose. In conclusion, only LAL mice are able to restore initial severe loss of MDMA-evoked 5-HT and 5-HIAA levels. SAL and LAL mice are differentially susceptible for the long-term but not short-term MDMA-induced serotonergic depletion in the striatum. The differentiation between both lines in the long-term striatal serotonergic response to MDMA seems to depend on the capacity of the brain to adapt to the short-term depletion of monoaminergic levels and may somehow be related to individual, trait-like characteristics of brain monoaminergic systems.     

Neurons associated with the flip‐flop activity in the lateral accessory lobe and ventral protocerebrum of the silkworm moth brain

The lateral accessory lobe (LAL) and the ventral protocerebrum (VPC) are a pair of symmetrical neural structures in the insect brain. The LAL‐VPC is regarded as the major target of olfactory responding neurons as well as the control center for olfactory‐evoked sequential zigzag turns. Previous studies of the silkworm moth Bombyx mori showed that these turns are controlled by long‐lasting anti‐phasic activities of the flip‐flopping descending neurons with dendrites in the LAL‐VPC. To elucidate the neural mechanisms underlying the generation of this alternating activity between the LAL‐VPC units of both hemispheres, we first analyzed the detailed neural architecture of the LAL‐VPC and identified five subregions. We then investigated the morphology and physiological responses of the LAL‐VPC neurons by intracellular recording and staining and morphologically identified three types of bilateral neurons and three types of unilateral neurons. Bilateral neurons showed either brief or cyclic long‐lasting responses. At least some neurons of the latter type produced γ‐aminobutyric acid (GABA). Unilateral neurons linking the LAL and VPC, in contrast, showed long‐lasting or quick alternating activity. Timing analysis of the activity onset of each neural type suggests that quick reciprocal neural transmission between unilateral neurons would be responsible for the generation of long‐lasting activity in one LAL‐VPC unit, which lasts for up to a few seconds. Reciprocal inhibition and excitation by the bilateral neurons with long‐lasting activities would mediate the alternating long‐lasting activity between both LAL‐VPC units, which might last for up to 20 seconds. J. Comp. Neurol. 518:366–388, 2010. © 2009 Wiley‐Liss, Inc.     

LAL途径特异性转录因子研究进展

链霉菌可产生许多具有重要应用价值的次级代谢产物,如抗生素、免疫调节剂、抗肿瘤药物等,它们已经广泛应用于临床医药中。次级代谢产物的生物合成受到转录因子的调控,转录因子通过控制结构基因的转录激活或阻遏解除,决定了表达方式和程度。Lux R家族大ATP结合调控因子(large ATP-binding regulators of the Lux R family,LAL)是一类新的次级代谢途径转录因子,由900~1000个氨基酸残基组成,它的N-端具有Walker A和Walker B模体的ATP/GTP结构域,C-端具有保守的Lux R家族螺旋-转角-螺旋模体的DNA结合域。已在40余种次级代谢产物生物合成基因簇中发现LAL,本文对LAL途径特异性转录调控的研究进展进行综述。     

应用LAL动态浊度法测定复方苦参注射液中细菌内毒素含量

目的：建立应用美洲鲎试剂LAL的动态浊度法定量检测复方苦参注射液中细菌内毒素试验方法。方法：采用《中国药典》2015年版(四部)中的细菌内毒素检查法。结果：复方苦参注射液用细菌内毒素定量法检测无干扰因素影响,内毒素回收率在50%～200%范围内。结论：使用LAL进行动态浊度法定量检测复方苦参注射液的细菌内毒素是可行的,可用细菌内毒素方法代替兔热原检查法,同时又避免了限量法内毒素检测时的干扰。     

Successful long‐term outcome of liver transplantation in late‐onset lysosomal acid lipase deficiency

Late‐onset LAL deficiency, previously referred to as cholesteryl ester storage disorder, is a rare lysosomal storage disorder characterized by accumulation of cholesteryl esters. It has a heterogeneous clinical phenotype including abdominal pain, poor growth, hyperlipidemia with vascular complications and hepatosplenomegaly. End‐stage liver disease may occur, but there are few reports of successful LT. There are also concerns that systemic manifestations of the disease might persist post‐LT. We report a case with excellent outcome eight yr following LT. The subject was noted to have asymptomatic hepatosplenomegaly during an intercurrent illness, and LAL deficiency was confirmed with compound heterozygosity in the LIPA. Despite dietary fat restriction, he developed signs of progressive liver disease and subsequently developed hepatopulmonary syndrome. He underwent cadaveric LT at the age of nine and a half yr and recovered with prompt resolution of hepatopulmonary syndrome. Eight yr post‐transplant he has normal growth, normal lipid profile, and liver and renal function tests. Liver histology showed no evidence of disease recurrence at this stage. LT in this subject resulted in an excellent functional correction of late‐onset LAL deficiency.