Induction of apoptosis by remoxipride metabolites in HL60 and CD34+/CD19- human bone marrow progenitor cells: potential relevance to remoxipride-induced aplastic anemia.

The antipsychotic agent, remoxipride [(S)-(-)-3-bromo-N-[(1-ethyl-2-pyrrolidinyl)methyl]-2,6-dimethoxybenz amide] has been associated with acquired aplastic anemia. We have examined the ability of remoxipride, three pyrrolidine ring metabolites and five aromatic ring metabolites of the parent compound to induce apoptosis in HL60 cells and human bone marrow progenitor (HBMP) cells. Cells were treated for 0-24 h with each compound (0-200 microM). Apoptosis was assessed by fluorescence microscopy in Hoechst 33342- and propidium iodide stained cell samples. Results were confirmed by determination of internucleosomal DNA fragmentation using gel electrophoresis for HL60 cell samples and terminal deoxynucleotidyl transferase assay in HBMP cells. The catechol and hydroquinone metabolites, NCQ436 and NCQ344, induced apoptosis in HL60 and HBMP cells in a time- and concentration dependent manner, while the phenols, NCR181, FLA873, and FLA797, and the derivatives formed by oxidation of the pyrrolidine ring, FLA838, NCM001, and NCL118, had no effect. No necrosis was observed in cells treated with NCQ436 but NCQ344 had a biphasic effect in both cell types, inducing apoptosis at lower concentrations and necrosis at higher concentrations. These data show that the catechol and hydroquinone metabolites of remoxipride have direct toxic effects in HL60 and HBMP cells, leading to apoptosis, while the phenol metabolites were inactive. Similarly, benzene-derived catechol and hydroquinone, but not phenol, induce apoptosis in HBMP cells [Moran et al., Mol. Pharmacol., 50 (1996) 610-615]. We propose that remoxipride and benzene may induce aplastic anemia via production of similar reactive metabolites and that the ability of NCQ436 and NCQ344 to induce apoptosis in HBMP cells may contribute to the mechanism underlying acquired aplastic anemia that has been associated with remoxipride.     

Identification of Molecular Markers of Delayed Graft Function Based on the Regulation of Biological Ageing

IntroductionDelayed graft function is a prevalent clinical problem in renal transplantation for which there is no objective system to predict occurrence in advance. It can result in a significant increase in the necessity for hospitalisation post-transplant and is a significant risk factor for other post-transplant complications.MethodologyThe importance of microRNAs (miRNAs), a specific subclass of small RNA, have been clearly demonstrated to influence many pathways in health and disease. To investigate the influence of miRNAs on renal allograft performance post-transplant, the expression of a panel of miRNAs in pre-transplant renal biopsies was measured using qPCR. Expression was then related to clinical parameters and outcomes in two independent renal transplant cohorts.ResultsHere we demonstrate, in two independent cohorts of pre-implantation human renal allograft biopsies, that a novel pre-transplant renal performance scoring system (GRPSS), can determine the occurrence of DGF with a high sensitivity (>90%) and specificity (>60%) for donor allografts pre-transplant, using just three senescence associated microRNAs combined with donor age and type of organ donation.ConclusionThese results demonstrate a relationship between pre-transplant microRNA expression levels, cellular biological ageing pathways and clinical outcomes for renal transplantation. They provide for a simple, rapid quantitative molecular pre-transplant assay to determine post-transplant allograft function and scope for future intervention. Furthermore, these results demonstrate the involvement of senescence pathways in ischaemic injury during the organ transplantation process and an indication of accelerated bio-ageing as a consequence of both warm and cold ischaemia.     

Class 1 outer membrane protein of Neisseria meningitidis: epitope analysis of the antigenic diversity between strains, implications for subtype definition and molecular epidemiology

The VR1 and VR2 regions of the class 1 protein have been sequenced from a number of meningococcal strains, including non-subtypable strains and strains of apparently identical subtype. The amino acid sequences have been used to construct synthetic peptides for mapping subtype-specific epitopes. The majority of epitopes was found to be located in VR2 at the apex of a predicted surface-exposed loop. A single amino acid change within an epitope, or an amino acid deletion outside an epitope, were both associated with loss of subtype specificity, resulting from a change in the predicted conformation at the apex of the loop structure. Analysis of the sequence information combined with knowledge of defined epitopes also revealed considerable additional information not demonstrated by current subtyping procedures.     

Spatial Distribution of an Uranium-Respiring Betaproteobacterium at the Rifle,CO Field Research Site

The Department of Energy’s Integrated Field-Scale Subsurface Research Challenge Site (IFRC) at Rifle, Colorado was created to address the gaps in knowledge on the mechanisms and rates of U(VI) bioreduction in alluvial sediments. Previous studies at the Rifle IFRC have linked microbial processes to uranium immobilization during acetate amendment. Several key bacteria believed to be involved in radionuclide containment have been described; however, most of the evidence implicating uranium reduction with specific microbiota has been indirect. Here, we report on the cultivation of a microorganism from the Rifle IFRC that reduces uranium and appears to utilize it as a terminal electron acceptor for respiration with acetate as electron donor. Furthermore, this bacterium constitutes a significant proportion of the subsurface sediment community prior to biostimulation based on TRFLP profiling of 16S rRNA genes. 16S rRNA gene sequence analysis indicates that the microorganism is a betaproteobacterium with a high similarity to Burkholderia fungorum. This is, to our knowledge, the first report of a betaproteobacterium capable of uranium respiration. Our results indicate that this microorganism occurs commonly in alluvial sediments located between 3-6 m below ground surface at Rifle and may play a role in the initial reduction of uranium at the site.     

Presynaptic NMDARs in the hippocampus facilitate transmitter release at theta frequency

A rise in [Ca(2+)](i) provides the trigger for neurotransmitter release at neuronal boutons. We have used confocal microscopy and Ca(2+) sensitive dyes to directly measure the action potential-evoked [Ca(2+)](i) in the boutons of Schaffer collaterals. This reveals that the trial-by-trial amplitude of the evoked Ca(2+) transient is bimodally distributed. We demonstrate that "large" Ca(2+) transients occur when presynaptic NMDA receptors are activated following transmitter release. Presynaptic NMDA receptor activation proves critical in producing facilitation of transmission at theta frequencies. Because large Ca(2+) transients "report" transmitter release, their frequency on a trial-by-trial basis can be used to estimate the probability of release, p(r). We use this novel estimator to show that p(r) increases following the induction of long-term potentiation.     

Long-Term Survival and Dialysis Dependency Following Acute Kidney Injury in Intensive Care: Extended Follow-up of a Randomized Controlled Trial

Background

The incidence of acute kidney injury (AKI) is increasing globally and it is much more common than end-stage kidney disease. AKI is associated with high mortality and cost of hospitalisation. Studies of treatments to reduce this high mortality have used differing renal replacement therapy (RRT) modalities and have not shown improvement in the short term. The reported long-term outcomes of AKI are variable and the effect of differing RRT modalities upon them is not clear. We used the prolonged follow-up of a large clinical trial to prospectively examine the long-term outcomes and effect of RRT dosing in patients with AKI.

Methods and Findings

We extended the follow-up of participants in the Randomised Evaluation of Normal vs. Augmented Levels of RRT (RENAL) study from 90 days to 4 years after randomization. Primary and secondary outcomes were mortality and requirement for maintenance dialysis, respectively, assessed in 1,464 (97%) patients at a median of 43.9 months (interquartile range [IQR] 30.0–48.6 months) post randomization. A total of 468/743 (63%) and 444/721 (62%) patients died in the lower and higher intensity groups, respectively (risk ratio [RR] 1.04, 95% CI 0.96–1.12, p = 0.49). Amongst survivors to day 90, 21 of 411 (5.1%) and 23 of 399 (5.8%) in the respective groups were treated with maintenance dialysis (RR 1.12, 95% CI 0.63–2.00, p = 0.69). The prevalence of albuminuria among survivors was 40% and 44%, respectively (p = 0.48). Quality of life was not different between the two treatment groups. The generalizability of these findings to other populations with AKI requires further exploration.

Conclusions

Patients with AKI requiring RRT in intensive care have high long-term mortality but few require maintenance dialysis. Long-term survivors have a heavy burden of proteinuria. Increased intensity of RRT does not reduce mortality or subsequent treatment with dialysis.

Trial registration

www.ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00221013","term_id":"NCT00221013"}}NCT00221013 Please see later in the article for the Editors'' Summary