Detection of mRNA alterations is a promising approach for identifying biomarkers as means of differentiating benign from malignant lesions. By choosing the KRAS oncogene as a target gene, two types of molecular beacons (MBs) based on either phosphothioated DNA (PS-DNA-MB) or peptide nucleic acid (TO-PNA-MB, where TO = thiazole orange) were synthesized and compared in vitro and in vivo. Their specificity was examined in wild-type KRAS (HT29) or codon 12 point mutation (Panc-1, SW480) cells. Incubation of both beacons with total RNA extracted from the Panc-1 cell line (fully complementary sequence) showed a fluorescent signal for both beacons. Major differences were observed, however, for single mismatch mRNA transcripts in cell lines HT29 and SW480. PS-DNA-MB weakly discriminated such single mismatches in comparison to TO-PNA-MB, which was profoundly more sensitive. Cell transfection of TO-PNA-MB with the aid of PEI resulted in fluorescence in cells expressing the fully complementary RNA transcript (Panc-1) but undetectable fluorescence in cells expressing the K-ras mRNA that has a single mismatch to the designed TO-PNA-MB (HT29). A weaker fluorescent signal was also detected in SW480 cells; however, these cells express approximately one-fifth of the target mRNA of the designed TO-PNA-MB. In contrast, PS-DNA-MB showed no fluorescence in all cell lines tested post PEI transfection. Based on the fast hybridization kinetics and on the single mismatch discrimination found for TO-PNA-MB we believe that such molecular beacons are promising for in vivo real-time imaging of endogenous mRNA with single nucleotide polymorphism (SNP) resolution.

AIM To investigate the number of publications in cardiovascular disease (CVD) in Latin America and the Caribbean over the last decade. METHODS We performed a bibliometric analysis in PubMed from 2001 to 2010 for Latin America and the Caribbean, the United States, Canada, Europe, China, and India. RESULTS Latin America published 4% of articles compared with 26% from the United States/Canada and 42% from Europe. In CVD, Latin America published 4% of articles vs 23% from the United States/Canada and 40% from Europe. The number of publications in CVD in Latin America increased from 41 in 2001 to 726 in 2010. CONCLUSION Latin America, while publishing more articles than previously, lags behind developed countries. Further advances in research infrastructure are necessary to develop prevention strategies for this region.

PURPOSE: To determine the feasibility of using a virtual auditory test material to evaluate reverberation and noise effects on speech recognition of pediatric cochlear implant (CI) users and to compare performance to that of children with normal hearing. METHOD: Virtual test materials representing non-reverberant and reverberant environments were used to measure speech recognition of 7 children with CIs in quiet and in noise, and of 18 children with normal hearing in the quiet condition. Performance of CI users in noise (SNR-50) was compared to normative data from a previous study (Neuman et al. 2010). For CI users, stimuli were sent directly to the CI speech processor via auxiliary input, while children with normal hearing were tested using insert phones. RESULTS: Speech recognition of children with CIs decreased significantly in the reverberant condition. There were individual differences in susceptibility to reverberation. Children with CIs also required higher signal-to-noise ratios than children with normal hearing in the reverberant condition. CONCLUSIONS: Direct connect testing with reverberant test materials allows assessment of speech recognition under conditions typical of classrooms and could be useful in identifying children with CIs whose performance decreases significantly in the presence of reverberation and noise.

PURPOSE: A real time detection of gastric cancer-associated biomarker molecules in the lumen of the stomach could assist in early detection of this multi-step malignancy. METHODS: Employing α1-antitrypsin precursor (A1AT) as a secreted biomarker model, a platform with immunoassay capabilities, comprising sensing and detecting compartments was developed. It was made of a microarray-type functionalized glass, containing a high density of amine groups. Trypsin, the capturing moiety, was immobilized to the glass surface with the aid of a PEG-based spacer mixture, identified as being crucial for both capturing and detecting properties. The detecting compartment contained near infrared fluorescently labeled nanoparticles conjugated to A1AT-specific antibodies, aimed at generating an optical signal, detectable by a conventional endoscope or a video capsule. RESULTS: The specific recognition reaction between the captured A1AT and the immuno-nanoparticles generated a profound fluorescence with a signal to noise ratio (SNR) of 12-32, in a biomarker-concentration dependent manner. Moreover, the optical recognition signal was intense enough to be detected by a video capsule simulator (with optical detection capabilities of a video capsule) with a SNR of 6-20. CONCLUSIONS: This platform could serve as a real time diagnostic kit for early detection of a secreted biomarker of gastric cancer.

Autophagy and apoptosis constitute important determinants of cell fate and engage in a complex interplay in both physiological and pathological settings. The molecular basis of this crosstalk is poorly understood and relies, in part, on "dual-function" proteins that operate in both processes. Here, we identify the essential autophagy protein Atg12 as a positive mediator of mitochondrial apoptosis and show that Atg12 directly regulates the apoptotic pathway by binding and inactivating prosurvival Bcl-2 family members, including Bcl-2 and Mcl-1. The binding occurs independently of Atg5 or Atg3 and requires a unique BH3-like motif in Atg12, characterized by interaction studies and computational docking. In apoptotic cells, knockdown of Atg12 inhibited Bax activation and cytochrome c release, while ectopic expression of Atg12 antagonized the antiapoptotic activity of Mcl-1. The interaction between Atg12 and Bcl-2 family members may thus constitute an important point of convergence between autophagy and apoptosis in response to specific signals.

This is a short summary of the two plenary sessions held at the International Society for Pharmacoeconomics and Outcomes Research (ISPOR) Latin American Conference in Mexico City (Mexico) in September 2011, with 477 registrants and 235 accepted abstract submissions. The first asked how attainable universal coverage is in the face of rising costs of health technologies; and the second considered the value of health outcomes research to decision-makers. This conference provided a scientific forum where researchers, health technology producers and public and private decision-makers shared their experiences and research in the field of health economic evaluations, health technology assessment and patient-reported outcomes/health-related quality of life studies. It was the third biennial regional meeting in Latin America, the next one being in Buenos Aires (Argentina) in 2013.

A novel approach to the prevention of colorectal cancer (CRC) recurrence by the local, luminal application of the combined therapies: Nutlin-3 (NUT) and the liposomal preparation of doxorubicin, Doxil(*)(Doxil) is presented here. The two drug entities were loaded into calcium alginate beads, engineered to erode upon exposure to a de-crosslinking agent, to allow for the controlled, concomitant release of the two. The synchronized release-driven improved cytotoxicity of NUT and Doxil was tested in vitro in RKO (wild-type p53) and HT-29 (mutant p53) CRC cells, by measuring intracellular expression of p53, p21 and Mdm2, as well as monitoring cell proliferation and viable cell numbers. NUT treatment alone was identified to be cytotoxic exclusively towards RKO cells. However, coadministration of NUT enhanced Doxil's anti-proliferative effects and cell death induction in a synergistic manner in both cell types. It was also identified that combinatorial treatment in a wt p53 context affected the p53 pathway by elevating the expression of p53 and its target p21. The capability of the formulation to erode in the presence of a de-crosslinking agent was demonstrated in vivo in the cecum of the anesthetized rat using indomethacin as a poorly water-soluble PK probe.

Genome-scale screening studies are gradually accumulating a wealth of data on the putative involvement of hundreds of genes in various cellular responses or functions. A fundamental challenge is to chart the molecular pathways that underlie these systems. ANAT is an interactive software tool, implemented as a Cytoscape plug-in, for elucidating functional networks of proteins. It encompasses a number of network inference algorithms and provides access to networks of physical associations in several organisms. In contrast to existing software tools, ANAT can be used to infer subnetworks that connect hundreds of proteins to each other or to a given set of "anchor" proteins, a fundamental step in reconstructing cellular subnetworks. The interactive component of ANAT provides an array of tools for evaluating and exploring the resulting subnetwork models and for iteratively refining them. We demonstrate the utility of ANAT by studying the crosstalk between the autophagic and apoptotic cell death modules in humans, using a network of physical interactions. Relative to published software tools, ANAT is more accurate and provides more features for comprehensive network analysis. The latest version of the software is available at http://www.cs.tau.ac.il/~bnet/ANAT_SI.

Abstract BACKGROUND:Obesity-associated asthma has been proposed to be a distinct entity, differing in immune pathogenesis from atopic asthma. Both obesity-mediated inflammation and increase in adiposity are potential mechanistic factors that are poorly defined among children. We hypothesize that pediatric obesity-associated asthma will be characterized by T helper 1(Th1), rather than the Th2 polarization associated with atopic asthma. Moreover, we speculate that Th1 biomarkers and anthropometric measures will correlate with pulmonary function tests (PFTs) in obese asthmatics. METHODS:We recruited 120 children with 30 in each of the 4 study groups: obese asthmatics, non-obese asthmatics, obese non-asthmatics and non-obese non-asthmatics. All underwent pulmonary function testing. Blood was collected for measurement of serum cytokines. T-cell responses to mitogen, phorbol 12-myristate 13-acetate (PMA), or antigens tetanus toxoid or Dermatophagoides farinae, were obtained by flow cytometric analysis of intracellular cytokine staining for interferon-gamma (IFNγ)(Th1) or IL-4 (Th2) within the CD4 population. RESULTS:Obese asthmatics had significantly higher Th1 responses to PMA (p<0.01) and tetanus toxoid (p<0.05) and lower Th2 responses to PMA (p<0.05) and D. farinae (p<0.01) compared to non-obese asthmatics. Th cell patterns did not differ between obese asthmatics and obese non-asthmatics. Obese asthmatics had lower FEV(1)/FVC (p<0.01) and RV/TLC ratios (p<0.005) compared to the other study groups, which negatively correlated with serum interferon inducible protein-10 (IP-10) and IFNγ levels, respectively. PFTs, however, did not correlate with BMI z-score or waist hip ratio. CONCLUSIONS:We found that pediatric obesity-associated asthma differed from atopic asthma and was characterized by Th1 polarization. The altered immune environment inversely correlated with PFTs in obese asthmatics.