SK channels regulate excitatory synaptic transmission and plasticity in the lateral amygdala

At glutamatergic synapses, calcium influx through NMDA receptors (NMDARs) is required for long-term potentiation (LTP); this is a proposed cellular mechanism underlying memory and learning. Here we show that in lateral amygdala pyramidal neurons, SK channels are also activated by calcium influx through synaptically activated NMDARs, resulting in depression of the synaptic potential. Thus, blockade of SK channels by apamin potentiates fast glutamatergic synaptic potentials. This potentiation is blocked by the NMDAR antagonist AP5 (D(-)-2-amino-5-phosphono-valeric acid) or by buffering cytosolic calcium with BAPTA. Blockade of SK channels greatly enhances LTP of cortical inputs to lateral amygdala pyramidal neurons. These results show that NMDARs and SK channels are colocalized at glutamatergic synapses in the lateral amygdala. Calcium influx through NMDARs activates SK channels and shunts the resultant excitatory postsynaptic potential. These results demonstrate a new role for SK channels as postsynaptic regulators of synaptic efficacy.     

Reduction of mitral regurgitation using the Coapsys device: a novel ex vivo method using excised recipients' hearts

The purpose of this study was to evaluate the ex vivo effects of the Coapsys device upon functional mitral regurgitation (MR) in human hearts. We used seven excised hearts from patients who underwent cardiac transplantation. All patients had functional MR of grade 2 or greater associated with dilated (n = 3) or ischemic (n = 4) cardiomyopathy. After the aortic valve was removed, the left ventricle was pressurized from the aorta with saline at a constant pressure. The degree of MR was then subjectively graded from the opened left atrium (from 0 to 4). The last three studies included volumetric measurements of MR. By tightening the device, the mean MR grade was reduced from 3.3 +/- 0.8 to 1.1 +/- 0.4 (p = 0.0002). In the quantitative analysis, mean regurgitation volume was reduced from 1,108 +/- 1,134 ml/min to 236 +/- 89 ml/min (p = not significant). The mitral annular septal-lateral dimension decreased from 2.0 +/- 0.3 cm to 1.6 +/- 0.5 (p = 0.043). The Coapsys device reduced functional MR in the ex vivo study using excised dilated hearts.     

Acute metabolic responses to a 24-h ultra-marathon race in male amateur runners

The study was conducted to evaluate the metabolic responses to a 24 h ultra-endurance race in male runners. Paired venous and capillary blood samples from 14 athletes (mean age 43.0 ± 10.8 years, body weight 64.3 ± 7.2 kg, VO_2max 57.8 ± 6.1 ml kg⁻¹ min⁻¹), taken 3 h before the run, after completing the marathon distance (42.195 km), after 12 h, and at the finish of the race, were analyzed for blood morphology, acid–base balance and electrolytes, lipid profile, interleukin-6 (IL-6), high-sensitivity C-reactive protein (hsCRP), and serum enzyme activities. Mean distance covered during the race was 168.5 ± 23.1 km (range 125.2–218.5 km). Prolonged ultra-endurance exercise triggered immune and inflammatory responses, as evidenced by a twofold increase in total leukocyte count with neutrophils and monocytes as main contributors, nearly 30-fold increase in serum IL-6 and over 20-fold rise in hsCRP. A progressive exponential increase in mean creatine kinase activity up to the level 70-fold higher than the respective pre-race value, a several fold rise in serum activities of aspartate aminotransferase and alanine aminotransferase, and a fairly stable serum γ-glutamyl transferase level, were indicative of muscle, but not of liver damage. With duration of exercise, there was a progressive development of hyperventilation-induced hypocapnic alkalosis, and a marked alteration in substrate utilization towards fat oxidation to maintain blood glucose homeostasis. The results of this study may imply that progressive decline in partial CO₂ pressure (hypocapnia) that develops during prolonged exercise may contribute to increased interleukin-6 production.     

A diversity-covering approach to immunization with Plasmodium falciparum apical membrane antigen 1 induces broader allelic recognition and growth inhibition responses in rabbits

Plasmodium falciparum apical membrane antigen 1 (PfAMA1), a candidate malaria vaccine, is polymorphic. This polymorphism is believed to be generated predominantly under immune selection pressure and, as a result, may compromise attempts at vaccination. Alignment of 355 PfAMA1 sequences shows that around 10% of the 622 amino acid residues can vary between alleles and that linkages between polymorphic residues occur. Using this analysis, we have designed three diversity-covering (DiCo) PfAMA1 sequences that take account of these linkages and, when taken together, on average incorporate 97% of amino acid variability observed. For each of the three DiCo sequences, a synthetic gene was constructed and used to transform the methylotrophic yeast Pichia pastoris, allowing recombinant expression. All three DiCo proteins were reactive with the reduction-sensitive monoclonal antibody 4G2, suggesting the DiCo sequences had conformations similar to those of naturally occurring PfAMA1. Rabbits were immunized with FVO strain PfAMA1 or with the DiCo proteins either individually or as a mixture. Antibody titers and the ability to inhibit parasite growth in vitro were determined. Animals immunized with the DiCo mix performed similarly to animals immunized with FVO AMA1 when measured against FCR3 strain parasites but outperformed animals immunized with FVO AMA1 when assessed against other strains. The levels of growth inhibition (approximately 70%) induced by the mix of three DiCo proteins were comparable for FVO, 3D7, and HB3, suggesting that a considerable degree of diversity in AMA1 is adequately covered. This suggests that vaccines based upon the DiCo mix approach provide a broader functional immunity than immunization with a single allele.     

A method for quantifying time dependent changes in MR signal intensity of articular cartilage as a function of tissue deformation in intact joints

A method is proposed to determine accurately the signal intensity changes of the articular cartilage from sectional MR images and its related cartilage deformation under compression in an intact joint. Image processing methods are developed to delineate and register the cartilage boundaries in consecutive MR images in order to track corresponding tissue sectors during the loading experiment. Regions of interest can then be defined and traced during the compression, making a spatial and temporal analysis of signal intensity changes possible. In addition, the cartilage deformation is calculated in the respective tissue sectors and is related to the MR signal changes. Using a fat-suppressed FLASH 3D sequence, the preliminary results showed location-dependent slight changes of the signal intensity varying from individual to individual. The quantitative analysis of the signal intensity changes as a function of cartilage deformation with magnetic resonance imaging (MRI) aims to characterize microstructural properties of the articular cartilage that may lead to a better understanding of degenerative joint disease.     

Serum levels of interferon-gamma, tumour necrosis factor-alpha, soluble interleukin-6R and soluble cell activation markers for monitoring response to treatment of leprosy reactions

Identifying pathogen and host-related laboratory parameters are essential for the early diagnosis of leprosy reactions. The present study aimed to clarify the validity of measuring the profiles of serum cytokines [interleukin (IL)-4, IL-6, IL-10, interferon (IFN)-gamma and tumour necrosis factor (TNF)-alpha], the soluble IL-6 receptor (sIL-6R), soluble T cell (sCD27) and macrophage (neopterin) activation markers and Mycobacterium leprae-specific anti-PGL-I IgM antibodies in relation to the leprosy spectrum and reactions. Serum samples from 131 Indonesian leprosy patients (82 non-reactional leprosy patients and 49 reactional) and 112 healthy controls (HC) from the same endemic region were investigated. Forty-four (89.8%) of the reactional patients had erythema nodosum leprosum (ENL) while only five (10.2%) had reversal reaction (RR). Follow-up serum samples after corticosteroid treatment were also obtained from 17 of the patients with ENL and one with RR. A wide variability in cytokine levels was observed in the patient groups. However, IFN-gamma and sIL-6R were elevated significantly in ENL compared to non-ENL patients. Levels of IFN-gamma, TNF-alpha and sIL-6R declined significantly upon corticosteroid treatment of ENL. Thus, although the present study suggests limited applicability of serial measurement of IFN-gamma, TNF-alpha and sIL-6R in monitoring treatment efficacy of ENL, reactions it recommends a search for a wider panel of more disease-specific markers in future studies.     

Immediate moxifloxacin hypersensitivity: Is there more than currently meets the eye?

Immediate drug hypersensitivity reactions (IDHR) to moxifloxacin constitute a pathomechanistic conundrum and a diagnostic challenge. Our objective was to study whether simultaneous phenotyping and quantification of histamine release might add to our knowledge about the basophil activation properties of moxifloxacin and constitute a reliable diagnostic aid. Fifteen patients with an IDHR to moxifloxacin and nine moxifloxacin challenged controls were selected. All had a basophil activation test (BAT) with moxifloxacin. Flow cytometric analysis of basophil responses implied labeling for CD63, CD203c, and intracellular histamine. Unlike tolerant challenged controls, basophilic upregulation of CD203c in response to moxifloxacin was observed in seven of 15 patients. Only two of these seven patients demonstrated appearance of CD63 and release of histamine. In the remainder eight patients, no basophil responses were demonstrable. In conclusion, immediate hypersensitivity to moxifloxacin might involve mechanisms difficult to capture by traditional CD63‐/CD203c‐based BAT. Deciphering the complexity of quinolone IDHR seems mandatory.     

The novel αB‐crystallin (CRYAB) mutation p.D109G causes restrictive cardiomyopathy

Restrictive cardiomyopathy (RCM) is a rare heart disease characterized by diastolic dysfunction and atrial enlargement. The genetic etiology of RCM is not completely known. We identified by a next‐generation sequencing panel the novel CRYAB missense mutation c.326A>G, p.D109G in a small family with RCM in combination with skeletal myopathy with an early onset of the disease. CRYAB encodes αB‐crystallin, a member of the small heat shock protein family, which is highly expressed in cardiac and skeletal muscle. In addition to in silico prediction analysis, our structural analysis of explanted myocardial tissue of a mutation carrier as well as in vitro cell transfection experiments revealed abnormal protein aggregation of mutant αB‐crystallin and desmin, supporting the deleterious effect of this novel mutation. In conclusion, CRYAB appears to be a novel RCM gene, which might have relevance for the molecular diagnosis and the genetic counseling of further affected families in the future.     

Identification of disease-associated DNA methylation in intestinal tissues from patients with inflammatory bowel disease

Lin Z, Hegarty JP, Cappel JA, Yu W, Chen X, Faber P, Wang Y, Kelly AA, Poritz LS, Peterson BZ, Schreiber S, Fan J‐B, Koltun WA. Identification of disease‐associated DNA methylation in intestinal tissues from patients with inflammatory bowel disease. Overwhelming evidence supports the theory that inflammatory bowel disease (IBD) is caused by a complex interplay between genetic predispositions of multiple genes, combined with an abnormal interaction with environmental factors. It is becoming apparent that epigenetic factors can have a significant contribution in the pathogenesis of disease. Changes in the methylation state of IBD‐associated genes could significantly alter levels of gene expression, potentially contributing to disease onset and progression. We have explored the role of DNA methylation in IBD pathogenesis. DNA methylation profiles (1505 CpG sites of 807 genes) of matched diseased (n = 26) and non‐diseased (n = 26) intestinal tissues from 26 patients with IBD [Crohn's disease (CD) n = 9, ulcerative colitis (UC) n = 17] were profiled using the GoldenGate? methylation assay. After an initial identification of a panel of 50 differentially methylated CpG sites from a training set (14 non‐diseased and 14 diseased tissues) and subsequent validation with a testing set (12 non‐diseased and 12 diseased tissues), we identified seven CpG sites that are differentially methylated in intestinal tissues of IBD patients. We have also identified changes in DNA methylation associated with the two major IBD subtypes, CD and UC. This study reports IBD‐associated changes in DNA methylation in intestinal tissue, which may be disease subtype‐specific.