Important Considerations in Pediatric Heart Failure.

PURPOSE OF REVIEW: The goal of this paper is to provide an overview of contemporary knowledge specific to the causes, management, and outcome of heart failure in children. RECENT FINDINGS: While recently there have been subtle improvements in heart failure outcomes in children, these improvements lag significantly behind that of adults. There is a growing body of literature suggesting that pediatric heart failure is a unique disease process with age- and disease-specific myocardial adaptations. In addition, the heterogenous etiologies of heart failure in children contribute to differential response to therapies and challenge the ability to obtain meaningful results from prospective clinical trials. Consideration of novel clinical trial designs with achievable but clinically relevant endpoints and focused study of the mechanisms underlying pediatric heart failure secondary to cardiomyopathies and structural heart disease are essential if we hope to advance care and identify targeted and effective therapies.

Systematic comparison of 2A peptides for cloning multi-genes in a polycistronic vector.

Cloning of multiple genes in a single vector has greatly facilitated both basic and translational studies that require co-expression of multiple factors or multi-units of complex protein. Many strategies have been adopted, among which 2A "self-cleaving" peptides have garnered increased interest for their polycistronic nature, small size and high "cleavage" efficiency. However, broad application of 2 A peptides is limited by the lack of systematic comparison of different 2As alone or in combination. Here we characterized the effect of varying gene position and 2As on the expression of proteins encoded in bi-, tri-, or quad-cistronic constructs. Using direct cardiac reprogramming as an example, we further determined the effect of varied 2As on the efficiency of fluorescent cell labeling and cell fate conversion. We found that the expression of fluorophores decreased as it was moved towards the end of the construct while reprogramming was most efficient with the fluorophore at the second position. Moreover, quad-cistronic TPE2A constructs resulted in more efficient reprogramming than 3P2A or PTE2A constructs. We expect that the bi-, tri-, and quad-cistronic vectors constructed here and our results on protein expression ratios from different 2A constructs could serve to guide future utilization of 2A peptides in basic research and clinical applications.