Measurement of Subcellular CA2+ Redistribution in Cardiac Muscle In Situ: Time Resolved Rapid Freezing and Electron Probe Microanalysis

To directly assess the physiological roles of sarcoplasmic reticulum (SR) and miitochondria (MT), we have utilized energy dispersive electron probe microanalysis (EPMA) on ultrathin freeze-dried cryosections from isolated papillary muscles, rapidly frozen at precise time points of the contractile cycle. Using this approach, we can detect redistribution of subcellular Ca2+ during the cardiac contractile cycle. Changes in Ca2+ of less than 1.0 mmol/kg dry wt can be detected. By determining the variability of the Ca2+ measurements in preliminary experiments, we have also demonstrated that it is possible to optimize experimental design, i.e., to predict the number of animals per treatment group and the number of X-ray spectra per animal that are required in order to detect a specified Ca2+ difference. Quantitative EPMA of rapidly frozen contracting papillary muscle has also allowed us to correlate the Ca2+ content of SR and MT with the contractile state of the muscle. Our results show a decrease of 40% in the amount of Ca2+ stored in the junctional SR during a cardiac muscle twitch, thus providing direct evidence for a role of the SR as a primary site of Ca2+ release. In addition, we have demonstrated dissociation between MT Ca2+ uptake and activation of regulatory enzymes, such as pyruvate dehydrogenase, indicating that MT Ca2+ uptake is not required for activation of MT metabolism

Classifying Severity of Cystic Fibrosis Lung Disease Using Longitudinal Pulmonary Function Data

Rationale: The study of genetic modifiers in cystic fibrosis (CF) lung disease requires rigorous phenotyping. One type of genetic association study design compares polymorphisms in patients at extremes of phenotype, requiring accurate classification of pulmonary disease at varying ages

Correlation of high levels of antibodies to multiple pre-erythrocytic Plasmodium falciparum antigens and protection from infection

High levels of antibodies to multiple antigens may be more strongly associated with protection from infection than antibodies to a single antigen. Antibody-associated protection against Plasmodium falciparum infection was assessed in a cohort of 68 adults living in an area of holoendemic malaria in Kenya. Antibodies to the pre-erythrocytic antigens circumsporozoite protein (CSP), liver-stage antigen-1 (LSA-1), thrombospondin-related adhesive protein (TRAP), and blood-stage antigens apical membrane antigen-1 (AMA-1), erythrocyte binding antigen-175 (EBA-175), and merozoite surface protein 1 (MSP-1) were tested. Peptides were used for CSP (NANP repeat) and LSA-1 (central repeat), and recombinant antigens were used for TRAP (aa D(48)-K(394)), AMA-1 (ectodomain, non-glycosylated), EBA-175 (non-glycosylated), and MSP-1 (MSP-1(19)). Weekly microscopy testing for P. falciparum infection was performed over a 12-week period after drug-mediated clearance of P. falciparum parasitemia. Individuals with high levels of IgG antibodies (\u3e 2 arbitrary units) to CSP, LSA-1, and TRAP had a 57% decrease in the risk of infection (95% confidence interval = 20-77%, P = 0.016). This decreased risk remained significant after adjustment for age, prior parasitemia, bed net use, sickle cell trait, and village of residence. In contrast, protection against infection did not correlate with high levels of IgG antibodies to blood-stage antigens or IgM antibodies to pre-erythrocytic or blood-stage antigens. High levels of IgG antibodies to CSP, LSA-1, and TRAP may be useful immune correlates of protection against P. falciparum infection in malaria-endemic populations

Reduction in Asthma Morbidity in Children as a Result of Home Remediation Aimed at Moisture Sources

OBJECTIVE: Home dampness and the presence of mold and allergens have been associated with asthma morbidity. We examined changes in asthma morbidity in children as a result of home remediation aimed at moisture sources. DESIGN: In this prospective, randomized controlled trial, symptomatic, asthmatic children (n = 62), 2–17 years of age, living in a home with indoor mold, received an asthma intervention including an action plan, education, and individualized problem solving. The remediation group also received household repairs, including reduction of water infiltration, removal of water-damaged building materials, and heating/ventilation/air-conditioning alterations. The control group received only home cleaning information. We measured children’s total and allergen-specific serum immuno-globulin E, peripheral blood eosinophil counts, and urinary cotinine. Environmental dust samples were analyzed for dust mite, cockroach, rodent urinary protein, endotoxin, and fungi. The follow-up period was 1 year. RESULTS: Children in both groups showed improvement in asthma symptomatic days during the preremediation portion of the study. The remediation group had a significant decrease in symptom days (p = 0.003, as randomized; p = 0.004, intent to treat) after remodeling, whereas these parameters in the control group did not significantly change. In the postremediation period, the remediation group had a lower rate of exacerbations compared with control asthmatics (as treated: 1 of 29 vs. 11 of 33, respectively, p = 0. 003; intent to treat: 28.1% and 10.0%, respectively, p = 0.11). CONCLUSION: Construction remediation aimed at the root cause of moisture sources and combined with a medical/behavioral intervention significantly reduces symptom days and health care use for asthmatic children who live in homes with a documented mold problem

Bullying of extremely low birth weight children: Associated risk factors during adolescence

Preterm children have many risk factors which may increase their susceptibility to being bullied. AIMS: To examine the prevalence of bullying among extremely low birth weight (ELBW, <1kg) and normal birth weight (NBW) adolescents and the associated sociodemographic, physical, and psychosocial risk factors and correlates among the ELBW children

Topical Application of Activity-based Probes for Visualization of Brain Tumor Tissue

Several investigators have shown the utility of systemically delivered optical imaging probes to image tumors in small animal models of cancer. Here we demonstrate an innovative method for imaging tumors and tumor margins during surgery. Specifically, we show that optical imaging probes topically applied to tumors and surrounding normal tissue rapidly differentiate between tissues. In contrast to systemic delivery of optical imaging probes which label tumors uniformly over time, topical probe application results in rapid and robust probe activation that is detectable as early as 5 minutes following application. Importantly, labeling is primarily associated with peri-tumor spaces. This methodology provides a means for rapid visualization of tumor and potentially infiltrating tumor cells and has potential applications for directed surgical excision of tumor tissues. Furthermore, this technology could find use in surgical resections for any tumors having differential regulation of cysteine cathepsin activity