In vivo porcine left atrial wall stress: Effect of ventricular tachypacing on spatial and temporal stress distribution

Animal models of ventricular tachypacing (VTP) have been successfully used to reproduce the relevant features observed in patients with atrial fibrillation, such as increased atrial pressure and volume, ion-channel alterations and fibrosis. After performing VTP on a healthy Yorkshire pig, we measured an increase in volume of 60%, a two-fold rise in pressure, and a complex pattern of local mechanical, histological and biochemical changes, including a generalized stiffening of the wall. A protocol recently developed was employed to generate computational models of the porcine left atrium mechanics in healthy conditions and after VTP. Comparison of the stress distribution in the healthy vs. VTP case provided a map of how pressure overload affects and modifies left atrium mechanics. Overall, a positive increase in stress was computed after the VTP treatment. Regions of large increase in the stresses post-VTP were the appendage boundaries, the area around the lower pulmonary vein and the area in the front of the atrium towards the appendage. Due to the elevated stress, the back of the atrium mainly modified its mechanical response, while the appendage remodeled both its shape and its mechanical properties. Large changes in the shape of the mitral valve annulus could be observed as a consequence of the remodeling in the front of the atrium. The relation between local mechanical stress and remodeling that emerges from the results is in agreement with our hypothesis that the structural changes in the atrium are a consequence of a stress-mediated mechanism.     

Serum antibody levels to the Pneumocystis jirovecii major surface glycoprotein in the diagnosis of P. jirovecii pneumonia in HIV+ patients

Background

Pneumocystis jirovecii remains an important cause of fatal pneumonia (Pneumocystis pneumonia or PcP) in HIV+ patients and other immunocompromised hosts. Despite many previous attempts, a clinically useful serologic test for P. jirovecii infection has never been developed.

Methods/Principal Findings

We analyzed serum antibody responses to the P. jirovecii major surface glycoprotein recombinant fragment C1 (MsgC1) in 110 HIV+ patients with active PcP (cases) and 63 HIV+ patients with pneumonia due to other causes (controls) by an enzyme-linked immunosorbent assay (ELISA). The cases had significantly higher IgG and IgM antibody levels to MsgC1 than the controls at hospital admission (week 0) and intervals up to at least 1 month thereafter. The sensitivity, specificity and positive predictive value (PPV) of IgG antibody levels increased from 57.2%, 61.7% and 71.5% at week 0 to 63.4%, 100%, and 100%, respectively, at weeks 3–4. The sensitivity, specificity and PPV of IgM antibody levels rose from 59.7%, 61.3%, and 79.3% at week 0 to 74.6%, 73.7%, and 89.8%, respectively, at weeks 3–4. Multivariate analysis revealed that a diagnosis of PcP was the only independent predictor of high IgG and IgM antibody levels to MsgC1. A high LDH level, a nonspecific marker of lung damage, was an independent predictor of low IgG antibody levels to MsgC1.

Conclusions/Significance

The results suggest that the ELISA shows promise as an aid to the diagnosis of PCP in situations where diagnostic procedures cannot be performed. Further studies in other patient populations are needed to better define the usefulness of this serologic test.     

Quantification of angiogenesis in estrogen receptor-positive and negative breast carcinoma

Background

The objective of this study was to evaluate angiogenesis according to CD34 antigen expression in estrogen receptor (ER)-positive and negative breast carcinomas.

Methods

This study comprised 64 cases of infiltrating ductal carcinoma in postmenopausal women divided into two groups: Group A: ER-positive, n = 35; and Group B: ER-negative, n = 29. The anti-CD34 monoclonal antibody was used as a marker for endothelial cells. Microvessel count was carried out in 10 fields per slide using a 40× objective lens (magnification 400×). Statistical analysis of the data was performed using Student's t-test (p < 0.05).

Results

The mean number of vessels stained with the anti-CD34 antibody in the estrogen receptor-positive and negative tumors was 23.51 ± 1.15 and 40.24 ± 0.42, respectively. The number of microvessels was significantly greater in the estrogen receptor-negative tumors (p < 0.001).

Conclusion

ER-negative tumors have significantly greater CD34 antigen expression compared to ER-positive tumors.

     

Regulation of arachidonic acid metabolism by cytochrome P-450 in rabbit kidney.

Renal microsomal cytochrome P-450-dependent arachidonic acid metabolism was correlated with the level of cytochrome P-450 in the rabbit kidney. Cobalt, an inducer of haem oxygenase, reduced cytochrome P-450 in both the cortex and medulla in association with a 2-fold decrease in aryl-hydrocarbon hydroxylase, an index of cytochrome P-450 activity, and a similar decrease in the formation of cytochrome P-450-dependent arachidonic acid metabolites by renal microsomes (microsomal fractions). Formation of the latter was absolutely dependent on NADPH addition and was prevented by SKF-525A, an inhibitor of cytochrome P-450-dependent enzymes. Arachidonate metabolites of cortical microsomes were identified by g.c.-m.s. as 20- and 19-hydroxyeicosatetraenoic acid, 11,12-epoxyeicosatrienoic acid and 11,12-dihydroxyeicosatrienoic acid. The profile of arachidonic acid metabolites was the same for the medullary microsomes. Induction of cytochrome P-450 by 3-methylcholanthrene and beta-naphthoflavone increased cytochrome P-450 content and aryl-hydrocarbon hydroxylase activity by 2-fold in the cortex and medulla, and this correlated with a 2-fold increase in arachidonic acid metabolites via the cytochrome P-450 pathway. These changes can also be demonstrated in cells isolated from the medullary segment of the thick ascending limb of the loop of Henle, which previously have been shown to metabolize arachidonic acid specifically via the cytochrome P-450-dependent pathway. The specific activity for the formation of arachidonic acid metabolites by this pathway is higher in the kidney than in the liver, the highest activity being in the outer medulla, namely 7.9 microgram as against 2.5 micrograms of arachidonic acid transformed/30 min per nmol of cytochrome P-450 for microsomes obtained from outer medulla and liver respectively. These findings are consistent with high levels of cytochrome P-450 isoenzyme(s), specific for arachidonic acid metabolism, primarily localized in the outer medulla.     

Enriched prostaglandin E-9 ketoreductase activity in outer medullary cells of the rabbit kidney

PGE2 metabolism was examined in rabbit renal slices and cell suspensions from the outer medulla, enriched (TALH) and depleted (OMC) for the thick ascending limb of Henle's loop. Metabolism was negligible in intact cells, either OMC or TALH fractions. However, in OMC and TALH homogenates, transformation of PGE2 to PGF2 alpha by NADPH-dependent prostaglandin E-9 ketoreductase (PGE-9KR) was observed at a PGE2 concentration of 4 X 10(-9) M. This activity was not reversible and was enriched ten-fold in the TALH with 41% of PGE2 transformed to PGF2 alpha after 30 min incubation. PGF2 alpha formation from PGE2 could not be detected in homogenates of cortex, medulla or papilla. PGE-9KR activity, particularly in the thick ascending limb, may be a source of PGF2 alpha in urine.     

Effects of lipid environment on the conformational changes of an ABC importer

In order to shuttle substrates across the lipid bilayer, membrane proteins undergo a series of conformation changes that are influenced by protein structure, ligands, and the lipid environment. To test the effect of lipid on conformation change of the ABC transporter MolBC, EPR studies were conducted in lipids and detergents of variable composition. In both a detergent and lipid environment, MolBC underwent the same general conformation changes as detected by site-directed EPR spectroscopy. However, differences in activity and the details of the EPR analysis indicate conformational rigidity that is dependent on the lipid environment. From these observations, we conclude that native-like lipid mixtures provide the transporter with greater activity and conformational flexibility as well as technical advantages such as reconstitution efficiency and protein stability.     

Distribution of cell surface saccharides on pancreatic cells

We describe here a simple, general procedure for the purification of a variety of lectins, and for the preparation of lectin-ferritin conjugates of defined molar composition and binding properties to be used as probes for cell surface saccharides. The technique uses a “universal” affinity column for lectins and their conjugates, which consists of hog sulfated gastric mucin glycopeptides covalently coupled to agarose. The procedure involes: (a) purification of lectins by chromatography of aqueous extracts of seeds or other lectin-containing fluids over the affinity column, followed by desorption of the desired lectin with its hapten suge; (b) iodination of the lectin to serve as a marker during subsequent steps; (c) conjugation of lectin to ferritin with glutaraldehyde; (d) collection of active lectin-ferritin conjugates by affinity chromatography; and (e) separation of monomeric lectin-ferritin conjugates from larger aggregates and unconjugated lectin by gel chromatography. Based on radioactivity and absorbancy at 310 nm for lectin and ferritin, respectively, the conjugates consist of one to two molecules of lectin per ferrritin molecule. Binding studies of native lectins and their ferritin conjugates to dispersed pancreatic acinar cells showed that the conjugation procedure does not significantly alter either the affinity constant of the lectin for its receptor on the cell surface or the number of sites detected.     

Potential Cost-Effectiveness of a New Infant Tuberculosis Vaccine in South Africa - Implications for Clinical Trials: A Decision Analysis

Novel tuberculosis vaccines are in varying stages of pre-clinical and clinical development. This study seeks to estimate the potential cost-effectiveness of a BCG booster vaccine, while accounting for costs of large-scale clinical trials, using the MVA85A vaccine as a case study for estimating potential costs. We conducted a decision analysis from the societal perspective, using a 10-year time frame and a 3% discount rate. We predicted active tuberculosis cases and tuberculosis-related costs for a hypothetical cohort of 960,763 South African newborns (total born in 2009). We compared neonatal vaccination with bacille Calmette-Guérin alone to vaccination with bacille Calmette-Guérin plus a booster vaccine at 4 months. We considered booster efficacy estimates ranging from 40% to 70%, relative to bacille Calmette-Guérin alone. We accounted for the costs of Phase III clinical trials. The booster vaccine was assumed to prevent progression to active tuberculosis after childhood infection, with protection decreasing linearly over 10 years. Trial costs were prorated to South Africa''s global share of bacille Calmette-Guérin vaccination. Vaccination with bacille Calmette-Guérin alone resulted in estimated tuberculosis-related costs of $89.91 million 2012 USD, and 13,610 tuberculosis cases in the birth cohort, over the 10 years. Addition of the booster resulted in estimated cost savings of $7.69–$16.68 million USD, and 2,800–4,160 cases averted, for assumed efficacy values ranging from 40%–70%. A booster tuberculosis vaccine in infancy may result in net societal cost savings as well as fewer active tuberculosis cases, even if efficacy is relatively modest and large scale Phase III studies are required.     

Autoantibodies directed to novel components of the PM/Scl complex, the human exosome

The autoantigenic polymyositis/scleroderma (PM/Scl) complex was recently shown to be the human homologue of the yeast exosome, which is an RNA-processing complex. Our aim was to assess whether, in addition to targeting the known autoantigens PM/Scl-100 and PM/Scl-75, autoantibodies also target recently identified components of the PM/Scl complex. The prevalence of autoantibodies directed to six novel human exosome components (hRrp4p, hRrp40p, hRrp41p, hRrp42p, hRrp46p, hCsl4p) was determined in sera from patients with idiopathic inflammatory myopathy (n = 48), scleroderma (n = 11), or the PM/Scl overlap syndrome (n = 10). The sera were analyzed by enzyme-linked immunosorbent assays and western blotting using the affinity-purified recombinant proteins. Our results show that each human exosome component is recognized by autoantibodies. The hRrp4p and hRrp42p components were most frequently targeted. The presence of autoantibodies directed to the novel components of the human exosome was correlated with the presence of the anti-PM/Scl-100 autoantibody in the sera of patients with idiopathic inflammatory myopathy (IIM), as was previously found for the anti-PM/Scl-75 autoantibody. Other clear associations between autoantibody activities were not found. These results further support the conception that the autoimmune response may initially be directed to PM/Scl-100, whereas intermolecular epitope spreading may have caused the autoantibody response directed to the associated components.