Excision of Sleeping Beauty transposons: parameters and applications to gene therapy

A major problem in gene therapy is the determination of the rates at which gene transfer has occurred. Our work has focused on applications of the Sleeping Beauty (SB) transposon system as a non-viral vector for gene therapy. Excision of a transposon from a donor molecule and its integration into a cellular chromosome are catalyzed by SB transposase. In this study, we used a plasmid-based excision assay to study the excision step of transposition. We used the excision assay to evaluate the importance of various sequences that border the sites of excision inside and outside the transposon in order to determine the most active sequences for transposition from a donor plasmid. These findings together with our previous results in transposase binding to the terminal repeats suggest that the sequences in the transposon-junction of SB are involved in steps subsequent to DNA binding but before excision, and that they may have a role in transposase-transposon interaction. We found that SB transposons leave characteristically different footprints at excision sites in different cell types, suggesting that alternative repair machineries operate in concert with transposition. Most importantly, we found that the rates of excision correlate with the rates of transposition. We used this finding to assess transposition in livers of mice that were injected with the SB transposon and transposase. The excision assay appears to be a relatively quick and easy method to optimize protocols for delivery of genes in SB transposons to mammalian chromosomes in living animals. Copyright (C) 2004 John Wiley Sons, Ltd

Physicochemical, Microbiological, and Toxicological Characterization of Pâté Prepared from the Meat and Liver of Bullfrog (Aquarana catesbeiana) Carcasses

The development of balanced, healthy, ready-to-consume, and easy-to-prepare products has led to the development of new food technologies. Despite their high commercial value, bullfrog (Aquarana catesbeiana) carcasses result in low yields, with the thighs being the most marketed in comparison to other carcass portions. In this sense, liver pâté is a traditional food consumed worldwide, mainly in European countries, and may be prepared by incorporating bullfrog meat by-products and certain viscera. In this context, the aim of the present study was to develop a pâté product based on a mixture comprising 50% grounded bullfrog torso meat and 50% liver paste, with each treatment incorporating 10% liver paste increments, totaling five final mixtures. The nutritional compositions and physicochemical, microbiological, and toxicological characteristics of each mixture were assessed. The dry matter percentage of the prepared product was determined to be 27.00%, while mineral content was 1.45%, lipid content was 4.00%, and total protein content was 20.00%. Finally, microbiological counts were in agreement with current food safety regulations. The developed pâté serves as a standard, recycling underused industrial materials, adding value to the production chain at low operational costs, creating a more accessible market, and promoting the popularization of this type of meat

Minimally Invasive Approach for Diagnosing TMJ Osteoarthritis

This study’s objectives were to test correlations among groups of biomarkers that are associated with condylar morphology and to apply artificial intelligence to test shape analysis features in a neural network (NN) to stage condylar morphology in temporomandibular joint osteoarthritis (TMJOA). Seventeen TMJOA patients (39.9 ± 11.7 y) experiencing signs and symptoms of the disease for less than 10 y and 17 age- and sex-matched control subjects (39.4 ± 15.2 y) completed a questionnaire, had a temporomandibular joint clinical exam, had blood and saliva samples drawn, and had high-resolution cone beam computed tomography scans taken. Serum and salivary levels of 17 inflammatory biomarkers were quantified using protein microarrays. A NN was trained with 259 other condyles to detect and classify the stage of TMJOA and then compared to repeated clinical experts’ classifications. Levels of the salivary biomarkers MMP-3, VE-cadherin, 6Ckine, and PAI-1 were correlated to each other in TMJOA patients and were significantly correlated with condylar morphological variability on the posterior surface of the condyle. In serum, VE-cadherin and VEGF were correlated with one another and with significant morphological variability on the anterior surface of the condyle, while MMP-3 and CXCL16 presented statistically significant associations with variability on the anterior surface, lateral pole, and superior-posterior surface of the condyle. The range of mouth opening variables were the clinical markers with the most significant associations with morphological variability at the medial and lateral condylar poles. The repeated clinician consensus classification had 97.8% agreement on degree of degeneration within 1 group difference. Predictive analytics of the NN’s staging of TMJOA compared to the repeated clinicians’ consensus revealed 73.5% and 91.2% accuracy. This study demonstrated significant correlations among variations in protein expression levels, clinical symptoms, and condylar surface morphology. The results suggest that 3-dimensional variability in TMJOA condylar morphology can be comprehensively phenotyped by the NN

Recuperação de áreas degradadas do Estado do Rio de Janeiro.

bitstream/CNPAB-2010/27273/1/doc076.pd

Asystolic Cardiac Arrest from Near Drowning Managed with Therapeutic Hypothermia

[West J Emerg Med. 2014;15(4):369–371.

Asystolic Cardiac Arrest from Near Drowning Managed with Therapeutic Hypothermia

[West J Emerg Med. 2014;15(4):369–371.