How to diagnose heart failure with preserved ejection fraction: the HFA–PEFF diagnostic algorithm: a consensus recommendation from the Heart Failure Association (HFA) of the European Society of Cardiology (ESC)

Making a firm diagnosis of chronic heart failure with preserved ejection fraction (HFpEF) remains a challenge. We recommend a new stepwise diagnostic process, the ‘HFA–PEFF diagnostic algorithm’. Step 1 (P=Pre‐test assessment) is typically performed in the ambulatory setting and includes assessment for heart failure symptoms and signs, typical clinical demographics (obesity, hypertension, diabetes mellitus, elderly, atrial fibrillation), and diagnostic laboratory tests, electrocardiogram, and echocardiography. In the absence of overt non‐cardiac causes of breathlessness, HFpEF can be suspected if there is a normal left ventricular (LV) ejection fraction, no significant heart valve disease or cardiac ischaemia, and at least one typical risk factor. Elevated natriuretic peptides support, but normal levels do not exclude a diagnosis of HFpEF. The second step (E: Echocardiography and Natriuretic Peptide Score) requires comprehensive echocardiography and is typically performed by a cardiologist. Measures include mitral annular early diastolic velocity (e′), LV filling pressure estimated using E/e′, left atrial volume index, LV mass index, LV relative wall thickness, tricuspid regurgitation velocity, LV global longitudinal systolic strain, and serum natriuretic peptide levels. Major (2 points) and Minor (1 point) criteria were defined from these measures. A score ≥5 points implies definite HFpEF; ≤1 point makes HFpEF unlikely. An intermediate score (2–4 points) implies diagnostic uncertainty, in which case Step 3 (F₁: Functional testing) is recommended with echocardiographic or invasive haemodynamic exercise stress tests. Step 4 (F₂: Final aetiology) is recommended to establish a possible specific cause of HFpEF or alternative explanations. Further research is needed for a better classification of HFpEF.     

Stimulus-Specific Adaptation in Field Potentials and Neuronal Responses to Frequency-Modulated Tones in the Primary Auditory Cortex

In order to structure the sensory environment our brain needs to detect changes in the surrounding that might indicate events of presumed behavioral relevance. A characteristic brain response presumably related to the detection of such novel stimuli is termed mismatch negativity (MMN) observable in human scalp recordings. A candidate mechanism underlying MMN at the neuronal level is stimulus-specific adaptation (SSA) which has several characteristics in common. SSA is the specific decrease in the response to a frequent stimulus, which does not generalize to an interleaved rare stimulus in a sequence of events. SSA was so far mainly described for changes in the response to simple pure tone stimuli differing in tone frequency. In this study we provide data from the awake rat auditory cortex on adaptation in the responses to frequency-modulated tones (FM) with the deviating feature being the direction of FM modulation. Adaptation of cortical neurons to the direction of FM modulation was stronger for slow modulation than for faster modulation. In contrast to pure tone SSA which showed no stimulus preference, FM adaptation in neuronal data differed sometimes between upward and downward FM. This, however, was not the case in the local field potential data recorded simultaneously. Our findings support the role of the auditory cortex as the source for change-related activity induced by FM stimuli by showing that dynamic stimulus features such as FM modulation can evoke SSA in the rat in a way very similar to FM-induced MMN in the human auditory cortex.     

To beat the heat – engineering of the most thermostable pyruvate decarboxylase to date

Pyruvate decarboxylase (PDC) is a key enzyme for the production of ethanol at high temperatures and for cell-free butanol synthesis. Thermostable, organic solvent stable PDC was evolved from bacterial PDCs. The new variant shows >1500-fold-improved half-life at 75 °C and >5000-fold-increased half-life in the presence of 9 vol% butanol at 50 °C.

Pyruvate decarboxylase (PDC) is a key enzyme for the production of ethanol at high temperatures and for cell-free butanol synthesis.

For many years, bioethanol has been used widely as an additive in gasoline or as a fuel by itself.¹ To avoid competition with food, production of ethanol from plant-derived starch or sugar (first generation) has shifted to lignocellulosic biomass (second generation). To date, most of the bioethanol produced industrially still utilises yeasts as the main fermentation organism.^1–3 However, in recent years, several efforts have been made to use thermophilic microorganisms as the new framework.^1,3 The main motivating forces behind this are the volatility of ethanol at higher temperatures, which facilitates easy product removal, reduced risk of contamination and feasibility of consolidated bioprocessing (CBP). In this process, cellulolytic ethanologenic thermophilic microorganisms are applied, thus reducing the energy consumption required for heat exchange between biomass pre-treatment, fermentation and product separation and to avoid cellulase inhibition by accumulating glucose.^3,4There are two major pathways for producing ethanol from pyruvate, the central metabolite in glycolysis: non-oxidative decarboxylation via pyruvate decarboxylase (PDC) and oxidative decarboxylation via pyruvate dehydrogenase (PDH) or pyruvate ferredoxin oxidoreductase (PFOR). Non-oxidative ethanol productions are commonly found in yeasts and other ethanologenic bacteria, while PDH and PFOR routes are described in thermophilic microorganisms.^3,4 PDC mode may offer advantages compared with PDH and PFOR, as it is thermodynamically more favourable and will not lead to any other organic acid by-products.^3–5 In fact, all industrially relevant ethanol producing organisms use PDC. Thermostable PDCs are, therefore, suggested to play an important role in creating homo-ethanologenic, thermophilic microorganisms.^5,6Until recently, there were only limited efforts described in literature for improving the thermostability of PDC. Ancestral sequence reconstruction, supposedly a powerful method of resurrecting ancient thermostable enzymes, did not yield more thermostable PDC.⁷ A computational Rosetta-based design resulted in several variants with improved stability, determined by differential interference contrast (DIC) microscopy; unfortunately, the work was not corroborated by kinetic activity and stability data.⁸ Another effort to redirect the specificity of a thermostable, acetolactate synthase to perform a PDC-like reaction by conventional directed evolution did not yield variants either, with activity similar to that of bacterial PDC.⁹ Thus, creating thermostable PDC variants remains as the main challenge towards second generation ethanol production by thermophilic microorganisms.Besides acting as an important enzyme for ethanol production, PDC has also been applied in the in vitro production of n-butanol (Scheme 1).^10,11 Butanol together with other longer-chain alcohols, such as 1-propanol, isobutanol and isopentanol, is regarded as the next-generation biofuel, due to its closer resemblance to traditional gasoline.¹² Thus, there is an increasing demand to produce longer-chain alcohols sustainably.¹² Industrially, butanol is still produced from petroleum-derived propene utilizing Co-catalyst, H₂, and CO via oxo synthesis and Reppe process.¹³ Albeit its efficiency, production of butanol from petroleum is not sustainable.Open in a separate windowScheme 1Simplified pathway to produce ethanol and 1-butanol from lignocellulosic biomass via pyruvate decarboxylase (PDC). There are two major pathways for producing 1-butanol via pyruvate: CoA-dependent or proline-dependent condensation. ADH is an alcohol dehydrogenase.In a more sustainable way, butanol can be produced via an acetone–butanol–ethanol (ABE) process, utilising Clostridium acetobutylicum.^12,14 Recent work on metabolic engineering has demonstrated the possibility of producing butanol in other microorganisms.^15,16 However, one of the major challenges in butanol fermentation is still the toxicity of butanol to many microorganisms at relatively low concentrations (<2 vol%).^17,18 This effect, however, is less pronounced for the enzymes catalysing butanol production in vitro.^10,19 Many enzymes can already tolerate higher organic solvent concentrations than the corresponding microorganisms.²⁰ Furthermore, engineering enzyme stability towards organic solvents is less challenging than evolving microorganisms to withstand the same concentration of organic solvents.^17,21 Therefore, in vitro butanol production emerges as a very promising alternative to traditional butanol fermentation. Thus, in this work, we focused on the engineering of PDC towards higher thermal stability for prospective ethanol production in thermophilic microorganisms and improved stability in butanol for applications in in vitro, artificial butanol synthesis.Prior to engineering a PDC, we characterised two new yeast PDCs from Candida glabrata (CgPDC) and Zygosaccharomyces rouxii (ZrPDC), as well as other PDCs reported in the literature, to determine the most suitable template. As presented in 22 Characterisation of bacterial PDCs showed similar findings to the previous report, with PDC from Acetobacter pasteurianus (ApPDC) demonstrating the highest kinetic stability (T₅₀^{1 h}) and melting temperature (T_m).²²T₅₀^{1 h} is defined as the temperature at which 50% of the enzymes remained active after 1 h incubation, while T_m is defined as the temperature at which 50% of the enzymes are in an unfolded state.²³ In general, bacterial PDCs showed higher activity at 25 °C and overall stability (T₅₀^{1 h} and T_m) than newly characterised CgPDC and ZrPDC.Summary of kinetic characterisations and thermostability of WT PDCs and variants

Combination of in ovo electroporation and time‐lapse imaging to study migrational events in chicken embryos

Background : During embryonic development cell migration plays a principal role in several processes. In past decades, many studies were performed to investigate migrational events, occurring during embryonic organogenesis, neurogenesis, gliogenesis or myogenesis, just to name a few. Although different common techniques are already used for this purpose, one of their major limitations is the static character. However, cell migration is a sophisticated and highly dynamic process, wherefore new appropriate technologies are required to investigate this event in all its complexity. Results and Conclusions: Here we report a novel approach for dynamic analysis of cell migration within embryonic tissue. We combine the modern transfection method of in ovo electroporation with the use of tissue slice culture and state‐of‐the‐art imaging techniques, such as confocal laser scanning microscopy or spinning disc confocal microscopy, and thus, develop a method to study live the migration of myogenic precursors in chicken embryos. The conditions and parameters used in this study allow long‐term imaging for up to 24 hr. Our protocol can be easily adapted for investigations of a variety of other migrational events and provides a novel conception for dynamic analysis of migration during embryonic development. Developmental Dynamics 243:690–698, 2014. © 2013 Wiley Periodicals, Inc.     

On the symmetry of siblings: automated single-cell tracking to quantify the behavior of hematopoietic stem cells in a biomimetic setup

The interplay between hematopoietic stem and progenitor cells (HSPC) and their local microenvironment is a key mechanism for the organization of hematopoiesis. To quantitatively study this process, a time-resolved analysis of cellular dynamics at the single-cell level is an essential prerequisite. One way to generate sufficient amounts of appropriate data is automatic single-cell tracking using time-lapse video microscopy. We describe and apply newly developed computational algorithms that allow for an automated generation of high-content data of single-cell characteristics at high temporal and spatial resolution, together with the reconstruction and statistical evaluation of complete genealogical histories. This methodology has been applied to the particular example of purified primary human HSPCs in bioengineered culture conditions. The combination of genealogical information and dynamic profiles of cellular properties identified a marked symmetry between sibling HSPCs regarding cell cycle time, but also migration speed and growth kinetics. Furthermore, we demonstrate that this symmetry of HSPC siblings can be altered by exogenous cues of the local biomimetic microenvironment. Using the example of HSPC growth in biomimetic culture systems, we show that our approach provides a valuable tool for the quantitative analysis of dynamic single-cell features under defined in?vitro conditions, allowing for integration of functional and genealogical data. The efficiency and accuracy of our approach pave the way for new and intriguing insights into the organizational principles of developmental patterns and the respective influence of exogenous cues not limited to the study of primary HSPCs.     

Childhood maltreatment is associated with an automatic negative emotion processing bias in the amygdala

Major depression has been repeatedly associated with amygdala hyper‐responsiveness to negative (but not positive) facial expressions at early, automatic stages of emotion processing using subliminally presented stimuli. However, it is not clear whether this “limbic bias” is a correlate of depression or represents a vulnerability marker preceding the onset of the disease. Because childhood maltreatment is a potent risk factor for the development of major depression in later life, we explored whether childhood maltreatment is associated with amygdalar emotion processing bias in maltreated but healthy subjects. Amygdala responsiveness to subliminally presented sad and happy faces was measured by means of fMRI at 3 T in N = 150 healthy subjects carefully screened for psychiatric disorders. Childhood maltreatment was assessed by the 25‐item childhood trauma questionnaire (CTQ). A strong association of CTQ‐scores with amygdala responsiveness to sad, but not happy facial expressions emerged. This result was further qualified by an interaction of emotional valence and CTQ‐scores and was not confounded by trait anxiety, current depression level, age, gender, intelligence, education level, and more recent stressful life‐events. Childhood maltreatment is apparently associated with detectable changes in amygdala function during early stages of emotion processing which resemble findings described in major depression. Limbic hyper‐responsiveness to negative facial cues could be a consequence of the experience of maltreatment during childhood increasing the risk of depression in later life. Limitation: the present association of limbic bias and maltreatment was demonstrated in the absence of psychopathological abnormalities, thereby limiting strong conclusions. Hum Brain Mapp 34:2899–2909, 2013. © 2012 Wiley Periodicals, Inc.     

Hypocapnia during hypoxic exercise and its impact on cerebral oxygenation,ventilation and maximal whole body O2 uptake

With hypoxic exposure ventilation is elevated through the hypoxic ventilatory response. We tested the hypothesis that the resulting hypocapnia reduces maximal exercise capacity by decreasing (i) cerebral blood flow and oxygenation and (ii) the ventilatory drive.