On the search for the right definition of heart failure with preserved ejection fraction

The definition of heart failure with preserved ejection fraction (HFpEF) has evolved from a clinically based “diagnosis of exclusion” to definitions focused on objective evidence of diastolic dysfunction and/or elevated left ventricular filling pressures. Despite advances in our understanding of HFpEF pathophysiology and the development of more sophisticated imaging modalities, the diagnosis of HFpEF remains challenging, especially in the chronic setting, given that symptoms are provoked by exertion and diagnostic evaluation is largely conducted at rest. Invasive hemodynamic study, and in particular — invasive exercise testing, is considered the reference method for HFpEF diagnosis. However, its use is limited as opposed to the high number of patients with suspected HFpEF. Thus, diagnostic criteria for HFpEF should be principally based on non-invasive measurements. As no single non-invasive variable can adequately corroborate or refute the diagnosis, different combinations of clinical, echocardiographic, and/or biochemical parameters have been introduced. Recent years have brought an abundance of HFpEF definitions. Here, we present and compare four of them: 1) the 2016 European Society of Cardiology criteria for HFpEF; 2) the 2016 echocardiographic algorithm for diagnosing diastolic dysfunction; 3) the 2018 evidence-based H2FPEF score; and 4) the most recent, 2019 Heart Failure Association HFA-PEFF algorithm. These definitions vary in their approach to diagnosis, as well as sensitivity and specificity. Further studies to validate and compare the diagnostic accuracy of HFpEF definitions are warranted. Nevertheless, it seems that the best HFpEF definition would originate from a randomized clinical trial showing a favorable effect of an intervention on prognosis in HFpEF

Dynamic NMR Structures of [Rp]- and [Sp]-Phosphorothioated DNA-RNA Hybrids: Is Flexibility Required for RNase H Recognition?

Chemically modified DNA oligonucleotides have been crucial to the development of antisense therapeutics. High-resolution structural studies of pharmaceutically relevant derivatives have been limited to only a few molecules. We have used NMR to elucidate the structure in solution of two DNA-RNA hybrids with the sequence d(CCTATAATCC)·r(GGAUUAUAGG). The two hybrids contain an unmodified RNA target strand, whereas the DNA strand contains one of two different stereoregular sugar-phosphate backbone linkages at each nucleotide: 1), [Rp]-phosphorothioate or 2), [Sp]-phosphorothioate. Homonuclear two-dimensional spectroscopy afforded nearly complete nonlabile proton assignments. Distance bounds, calculated from the nuclear Overhauser effect (NOE) crosspeak intensities via a complete relaxation matrix approach with the program MARDIGRAS, were used to restrain the structure of the two hybrids during simulations of molecular dynamics. Analysis of restrained molecular dynamics trajectories suggests that both hybrids are flexible, requiring the use of molecular dynamics with time-averaged restraints (MDtar) to generate ensembles of structures capable of satisfying the NMR data. In particular, the deoxyribose sugars of the DNA strand show strong evidence of repuckering. Furthermore, deoxyribose sugar repuckering is accompanied by increased flexibility of overall helical geometry. These observations, together with the analysis of the crystal structure of a hybrid duplex in complex with ribonuclease H (RNase H), suggested that this flexibility may be required for recognition by RNase H

Unusual Thermal Stability of RNA/[R(P)-PS]-DNA/RNA Triplexes Containing a Homopurine DNA Strand

Homopurine deoxyribonucleoside phosphorothioates, as short as hexanucleotides and possessing all internucleotide linkages of R(P) configuration, form a triple helix with two RNA or 2′-OMe-RNA strands, with Watson-Crick and Hoogsteen complementarity. Melting temperature and fluorescence quenching experiments strongly suggest that the Hoogsteen RNA strand is parallel to the homopurine [R(P)-PS]-oligomer. Remarkably, these triplexes are thermally more stable than complexes formed by unmodified homopurine DNA molecules of the same sequence. The triplexes formed by phosphorothioate DNA dodecamers containing 4–6 dG residues are thermally stable at pH 7.4, although their stability increases significantly at pH 5.3. FTIR measurements suggest participation of the C(2)-carbonyl group of the pyrimidines in the stabilization of the triplex structure. Formation of triple-helix complexes with exogenously delivered PS-oligos may become useful for the reduction of RNA accessibility in vivo and, hence, selective suppression/inhibition of the translation process

IDH1R132H in Neural Stem Cells: Differentiation Impaired by Increased Apoptosis.

BACKGROUND:The high frequency of mutations in the isocitrate dehydrogenase 1 (IDH1) gene in diffuse gliomas indicates its importance in the process of gliomagenesis. These mutations result in loss of the normal function and acquisition of the neomorphic activity converting α-ketoglutarate to 2-hydroxyglutarate. This potential oncometabolite may induce the epigenetic changes, resulting in the deregulated expression of numerous genes, including those related to the differentiation process or cell survivability. METHODS:Neural stem cells were derived from human induced pluripotent stem cells following embryoid body formation. Neural stem cells transduced with mutant IDH1R132H, empty vector, non-transduced and overexpressing IDH1WT controls were differentiated into astrocytes and neurons in culture. The neuronal and astrocytic differentiation was determined by morphology and expression of lineage specific markers (MAP2, Synapsin I and GFAP) as determined by real-time PCR and immunocytochemical staining. Apoptosis was evaluated by real-time observation of Caspase-3 activation and measurement of PARP cleavage by Western Blot. RESULTS:Compared with control groups, cells expressing IDH1R132H retained an undifferentiated state and lacked morphological changes following stimulated differentiation. The significant inhibitory effect of IDH1R132H on neuronal and astrocytic differentiation was confirmed by immunocytochemical staining for markers of neural stem cells. Additionally, real-time PCR indicated suppressed expression of lineage markers. High percentage of apoptotic cells was detected within IDH1R132H-positive neural stem cells population and their derivatives, if compared to normal neural stem cells and their derivatives. The analysis of PARP and Caspase-3 activity confirmed apoptosis sensitivity in mutant protein-expressing neural cells. CONCLUSIONS:Our study demonstrates that expression of IDH1R132H increases apoptosis susceptibility of neural stem cells and their derivatives. Robust apoptosis causes differentiation deficiency of IDH1R132H-expressing cells