Conditional growth defect of Bordetella pertussis and Bordetella bronchiseptica ferric uptake regulator (fur) mutants

Outer-membrane vesicles (OMVs) are promising tools in the development of novel vaccines against the respiratory pathogens Bordetella pertussis and Bordetella bronchiseptica. Unfortunately, vesiculation by bordetellae is too low for cost-effective vaccine production. In other bacteria, iron limitation or inactivation of the fur gene has been shown to increase OMV production, presumably by downregulation of the mla genes, which encode machinery for maintenance of lipid asymmetry in the outer membrane. Here, we followed a similar approach in bordetellae. Whereas a fur mutant was readily obtained in B. bronchiseptica, a B. pertussis fur mutant could only be obtained in iron-deplete conditions, indicating that a fur mutation is conditionally lethal in this bacterium. The fur mutants displayed a growth defect in iron-replete media, presumably because constitutive expression of iron-uptake systems resulted in iron intoxication. Accordingly, expression of the Escherichia coli ferritin FtnA to sequester intracellularly accumulated iron rescued the growth of the mutants in these media. The fur mutations led to the constitutive expression of novel vaccine candidates, such as the TonB-dependent receptors FauA for the siderophore alcaligin and BhuR for heme. However, neither inactivation of fur nor growth under iron limitation improved vesiculation, presumably because the expression of the mla genes appeared unaffected

The association between neuroticism and self-reported common somatic symptoms in a population cohort

OBJECTIVE: To test the hypotheses that (1) neuroticism is associated with self-reported somatic symptoms; (2) this association is especially found with regard to psychosomatic symptoms; and (3) it is not solemnly explained by somatic reflections of psychological distress. METHODS: We studied the cross-sectional association between neuroticism (as measured by EPQ-RSS-N), psychological distress (as measured by GHQ-12 sum score), and the occurrence of 22 common somatic symptoms by linear and logistic regression analyses in a population cohort of 6894 participants. RESULTS: Neuroticism is more strongly associated with the total number of somatic symptoms reported (beta=.32) than GHQ-12 sum score (beta=.15) and well-established risk markers such as gender (beta=.11) and age (beta=.04). Neuroticism was associated with all symptoms in individual logistic regressions controlled for age, gender, and psychological distress. Neuroticism is significantly more strongly related to psychosomatic symptoms (beta=.36) than to infectious/allergic symptoms (beta=.28). CONCLUSION: In a large, population-based cohort, we confirmed that neuroticism is associated with self-reported somatic symptoms. The associations were not attributable to somatic reflections of psychological distress associated with neuroticism and were relatively strong with respect to psychosomatic symptoms. Future studies should include both objective and subjective measures of health to study the mechanisms that connect neuroticism and ill health

Large-scale purification of factor VIII by affinity chromatography: optimization of process parameters

The optimization of a new process for the extraction of human coagulation factor VIII (FVIII) from plasma with the tailor-made affinity matrix dimethylaminopropylcarbamylpentyl-Sepharose CL-4B (C3---C5 matrix) is described. First, plasma is applied to DEAE-Sephadex A-50 anion exchanger in order to separate a number of proteins, including coagulation factors II, IX and X (prothrombin complex), from FVIII. Subsequently, the unbound fraction of the ion exchanger, containing FVIII, is contacted with the C3---C5 affinity matrix. Optimization of the FVIII affinity chromatographic procedure is accomplished in terms of the ligand density of the matrix, adsorption mode (batch-wise versus column-wise adsorption and matrix to plasma ratio), and conditions of pH and conductivity to be applied on washing and desorption. In scale-up experiments, by processing 20 1 of plasma, the recovery (340 U VIII:C/kg plasma) and the specific activity (s.a.) (1.2 U VIII:C/mg protein) are better than those obtained by cryoprecipitation (recovery 300 U VIII:C/kg plasma, s.a. O.3 U VIII:C/mg protein). The newly developed process using the specially designed C3---C5 affinity matrix has potential application in the process-scale purification of FVIII

Urban planning in Lahore:a confrontation with real development

Groningen: FRW-RU

Psychiatric comorbidity and causal disease models

In psychiatry, comorbidity is the rule rather than the exception. Up to 45% of all patients are classified as having more than one psychiatric disorder. These high rates of comorbidity have led to a debate concerning the interpretation of this phenomenon. Some authors emphasize the problematic character of the high rates of comorbidity because they indicate absent zones of rarities. Others consider comorbid conditions to be a validator for a particular reclassification of diseases. In this paper we will show that those at first sight contrasting interpretations of comorbidity are based on similar assumptions about disease models. The underlying ideas are that firstly high rates of comorbidity are the result of the absence of causally defined diseases in psychiatry, and second that causal disease models are preferable to non-causal disease models. We will argue that there are good reasons to seek after causal understanding of psychiatric disorders, but that causal disease models will not rule out high rates of comorbidity-neither in psychiatry, nor in medicine in general. By bringing to the fore these underlying assumptions, we hope to clear the ground for a different understanding of comorbidity, and of models for psychiatric diseases. (C) 2012 Elsevier Inc. All rights reserved.</p

Preconditioning in globally ischemic isolated rat hearts: effect on function and metabolic indices of myocardial damage

We assessed the effects of ischemic preconditioning on heart recovery and metabolic indices of damage following global ischemia and reperfusion, in relationship to post-ischemic lactate release. Three groups of Langendorff rat hearts were studied: (1) A control group of 40 min global ischemia and 45 min reperfusion; (2) preconditioning by 5 min global ischemia and 15 min reperfusion prior to sustained ischemia and reperfusion; (3) Preconditioning by three episodes of brief ischemia-re

One-year follow-up of a randomised controlled trial on added splinting to eccentric exercises in chronic midportion Achilles tendinopathy

OBJECTIVE: The study examined whether the addition of a night splint to eccentric exercises is beneficial for functional outcome in chronic mid-portion Achilles tendinopathy. DESIGN: One-year follow-up of a randomised controlled single blinded clinical trial. SETTING: Sports medicine department in a general hospital. PATIENTS: 58 patients (70 tendons) were included. INTERVENTIONS: All patients completed a 12-week heavy load eccentric training programme. One group received a night splint in addition to eccentric exercises. Main outcome measurements: Outcome scores were: Victorian Institute of Sport Assessment - Achilles (VISA-A) score, subjective patient satisfaction and neovascularisation score measured with Power Doppler Ultrasonography. RESULTS: For both groups the VISA-A score increased significantly (from 50 to 76 (P < 0.01) in the eccentric group and from 49 to 78 (P < 0.01) in the night splint group). No significant differences in VISA-A score were found between the groups from baseline to one year (P = 0.32). Presence of neovessels at baseline did not predict change in VISA-A score after one year in the whole group (P = 0.71). CONCLUSION: Eccentric exercises with or without a night splint improved functional outcome at one-year follow-up. At follow-up there

Synergistic activation of non-rectifying small-conductance chloride channels by forskolin and phorbol esters in cell-attached patches of the human colon carcinoma cell line HT-29cl.19A

Cell-attached patch-clamp studies with the human colon carcinoma HT-29cl.19A cells revealed a small chloride channel with a unitary conductance of 6.5 pS at 70 mV and 4.6 pS at -70 mV clamp potential after cAMP was increased by activation of adenylyl cyclase by forskolin. Usually channels inactivated upon patch excision, but in a few excised patches the channels stayed active and displayed a linear I/V relation in symmetrical (150 mmol/l) chloride solutions with a conductance of 7.5 pS. A 16-fold increase in channel incidence was observed when forskolin and phorbol 12,13-dibutyrate (PDB) were present together. The open probability was voltage-independent and was not different in the presence of forskolin plus PDB or with forskolin alone. The conductance sequence of the channel as deduced from outward currents carried by five different anions including chloride was: Cl->Br->NO3 ->gluconate > I-. The permeability sequence deduced from the reversal potentials was NO3 -≥Br->Cl->I->gluconate. With iodide in the pipette the conductance decreased strongly. Moreover, the inward current was reduced by 61%, indicating a strong inhibition of the chloride efflux by iodide. Similarly, the forskolin-induced increase of the short-circuit current (Isc) in confluent filter-grown monolayers was strongly reduced by iodide in the apical perfusate. Iodide also increased the fractional resistance of the apical membrane and repolarized the membrane potential, indicating an inhibitory action on the forskolin-induced increase of the apical chloride conductance. The PDB-induced Isc was also reduced by iodide, suggesting that the same chloride conductance is involved in the forskolin and in the PDB response. The results suggest that forskolin via cAMP-dependent protein kinase and PDB via protein kinase C regulate the same non-rectifying small-conductance chloride channels in the HT-29cl.19A cells