Molecular mechanism of poly(ADP-ribosyl)ation by PARP1 and identification of lysine residues as ADP-ribose acceptor sites

Poly(ADP-ribose) polymerase 1 (PARP1) synthesizes poly(ADP-ribose) (PAR) using nicotinamide adenine dinucleotide (NAD) as a substrate. Despite intensive research on the cellular functions of PARP1, the molecular mechanism of PAR formation has not been comprehensively understood. In this study, we elucidate the molecular mechanisms of poly(ADP-ribosyl)ation and identify PAR acceptor sites. Generation of different chimera proteins revealed that the amino-terminal domains of PARP1, PARP2 and PARP3 cooperate tightly with their corresponding catalytic domains. The DNA-dependent interaction between the amino-terminal DNA-binding domain and the catalytic domain of PARP1 increased Vmax and decreased the Km for NAD. Furthermore, we show that glutamic acid residues in the auto-modification domain of PARP1 are not required for PAR formation. Instead, we identify individual lysine residues as acceptor sites for ADP-ribosylation. Together, our findings provide novel mechanistic insights into PAR synthesis with significant relevance for the different biological functions of PARP family member

Are Albumin Levels a Good Predictor of Mortality in Elderly Patients with Neck of Femur Fractures?

Background Neck of femur (NOF) fractures are associated with significant morbidity and mortality in elderly people with multiple co-morbidities; making management of this patient subgroup challenging. Predictors of an increase in morbidity and mortality would therefore provide a useful framework for the assessment and management of this demographic. Within the current literature, hypoalbuminaemia (<35g/dl) has been highlighted as being a good biochemical predictor of short-term mortality (<12 months). Our aims were to assess whether there was an association between low albumin levels and short-term mortality and whether the severity adversely affects outcomes. Materials and Methods Patients admitted to our large district hospital between January 2011 and December 2012 who had sustained a NOF fracture, were over 65 years old and had a pre-operative albumin level were included. The study concluded in July 2014. Demographic and pre-operative function and albumin data was collated retrospectively. Correlation with mortality was made. Results 471 patients had usable data. Mean pre-operative albumin level was 29.5g/dl (SD 6.22g/dl) in patients who died and 32.8g/dl (SD 6.43g/dl) in patients who survived during the study period. Pre-operative albumin level was significantly associated with survival (hazard ratio0.957: 95% CI (0.937, 0.978); p<0.001) A reduction of 1g/dl in pre-operative albumin is associated with an increased hazard of death of 4.3%. Conclusions Early identification of patients with hypoalbuminaemia on admission with a venous blood sample and timely input from orthogeriatrians could optimise these patients pre- and post-operatively. This may enable rates of morbidity and mortality to fall. Hypoalbuminaemia may be a reasonable predictor of shorter-term mortality in this patient subgroup. However, this may reflect existing co-morbidities rather than an isolated cause. This study supports a correlation between hypoalbuminaemia and poorer outcome for patients with NOF fractures

Differential basolateral–apical distribution of scavenger receptor, class B, type I in cultured cells and the liver

The high-density lipoprotein (HDL) receptor, scavenger receptor class B, type I (SR-BI), mediates selective cholesteryl ester uptake into the liver, which finally results in cholesterol secretion into the bile. Despite several reports, the distribution of hepatic SR-BI between the sinusoidal and canalicular membranes is still under debate. We present immunohistological data using specific markers showing that the bulk of SR-BI is present in sinusoidal membranes and, to a lesser extent, in canalicular membranes in murine and human liver sections. In addition, SR-BI was detected in preparations of rat liver canalicular membranes. We also compared the in vivo findings to HepG2 cells, a widely used in vitro hepatocyte model. Interestingly, SR-BI was enriched in bile canalicular-like (BC-like) structures in polarized HepG2 cells, which were cultivated either conventionally to form a monolayer or in Matrigel to form three-dimensional structures. Fluorescently labeled HDL was transported into close proximity of BC-like structures, whereas HDL labeled with the fluorescent cholesterol analog BODIPY-cholesterol was clearly detected within these structures. Importantly, similarly to human and mouse liver, SR-BI was localized in basolateral membranes in three-dimensional liver microtissues from primary human liver cells. Our results demonstrate that SR-BI is highly enriched in sinusoidal membranes and is also found in canalicular membranes. There was no significant basolateral–apical redistribution of hepatic SR-BI in fasting and refeeding experiments in mice. Furthermore, in vitro studies in polarized HepG2 cells showed explicit differences as SR-BI was highly enriched in BC-like structures. These structures are, however, functional and accumulated HDL-derived cholesterol. Thus, biological relevant model systems should be employed when investigating SR-BI distribution in vitro. Electronic supplementary material The online version of this article (doi:10.1007/s00418-014-1251-9) contains supplementary material, which is available to authorized users

A systematic quality rating of available mobile health apps for borderline personality disorder

Background Mobile health apps (MHAs) may offer a mean to overcome treatment barriers in Borderline Personality Disorder (BPD) mental health care. However, MHAs for BPD on the market lack transparency and quality assessment. Methods European app stores were systematically searched, and two independent trained reviewers extracted relevant MHAs. Employed methods and privacy and security details documentation of included MHAs were extracted. MHAs were then assessed and rated using the German version of the standardized Mobile Application Rating Scale (MARS-G). Mean values and standard deviations of all subscales (engagement, functionality, aesthetics, information, and therapeutic gain) and correlations with user ratings were calculated. Results Of 2,977 identified MHAs, 16 were included, showing average quality across the four main subscales (M=3.25, SD=0.68). Shortcomings were observed with regard to engagement (M=2.87, SD = 0.99), potential therapeutic gain (M=2.67, SD=0.83), existing evidence base (25.0% of included MHAs were tested empirically), and documented privacy and security details. No significant correlations were found between user ratings and the overall total score of the MARS-G or MARS-G main subscales. Conclusions Available MHAs for BPD vary in quality and evidence on their efficacy, effectiveness, and possible adverse events is scarce. More substantial efforts to ensure the quality of MHAs available for patients and a focus on transparency, particularly regarding privacy and security documentation, are necessary

PARP1 ADP-ribosylates lysine residues of the core histone tails

The chromatin-associated enzyme PARP1 has previously been suggested to ADP-ribosylate histones, but the specific ADP-ribose acceptor sites have remained enigmatic. Here, we show that PARP1 covalently ADP-ribosylates the amino-terminal histone tails of all core histones. Using biochemical tools and novel electron transfer dissociation mass spectrometric protocols, we identify for the first time K13 of H2A, K30 of H2B, K27 and K37 of H3, as well as K16 of H4 as ADP-ribose acceptor sites. Multiple explicit water molecular dynamics simulations of the H4 tail peptide into the catalytic cleft of PARP1 indicate that two stable intermolecular salt bridges hold the peptide in an orientation that allows K16 ADP-ribosylation. Consistent with a functional cross-talk between ADP-ribosylation and other histone tail modifications, acetylation of H4K16 inhibits ADP-ribosylation by PARP1. Taken together, our computational and experimental results provide strong evidence that PARP1 modifies important regulatory lysines of the core histone tail

Predicting power ramps from joint distributions of future wind speeds

Power ramps are sudden changes in turbine power and must be accurately predicted to minimize costly imbalances in the electrical grid. Doing so requires reliable wind speed forecasts, which can be obtained from ensembles of physical numerical weather prediction (NWP) models through statistical postprocessing. Since the probability of a ramp event depends jointly on the wind speed distributions forecasted at multiple future times, these postprocessing methods must not only correct each individual forecast but also estimate the temporal dependencies among them. Typically though, crucial dependencies are adopted directly from the raw ensemble, and the postprocessed forecast is limited to the tens of members computationally feasible for an NWP model. We extend statistical postprocessing to include temporal dependencies using novel multivariate Gaussian regression models that forecast 24-dimensional distributions of next-day hourly wind speeds at three offshore wind farms. The continuous joint distribution forecast is postprocessed from an NWP ensemble using flexible generalized additive models for the components of its mean vector μ and for parameters defining the forecast error covariance matrix Σ. Modeling these parameters on predictors which characterize the empirical joint distribution of the NWP ensemble allows forecasts for each hour and their temporal dependencies to be adjusted in one step. Wind speed ensembles of any size can be simulated from the postprocessed joint distribution and transformed into power for computing high-resolution ramp predictions that outperform state-of-the-art reference methods.</p

Differential effects of chlorinated and oxidized phospholipids in vascular tissue:implications for neointima formation

The presence of inflammatory cells and MPO (myeloperoxidase) in the arterial wall after vascular injury could increase neointima formation by modification of phospholipids. The present study investigates how these phospholipids, in particular oxidized and chlorinated species, are altered within injured vessels and how they affect VSMC (vascular smooth muscle cell) remodelling processes. Vascular injury was induced in C57BL/6 mice and high fat-fed ApoE-/- (apolipoprotein E) mice by wire denudation and ligation of the left carotid artery (LCA). Neointimal and medial composition was assessed using immunohistochemistry and ESI-MS. Primary rabbit aortic SMCs (smooth muscle cells) were utilized to examine the effects of modified lipids on VSMC proliferation, viability and migration at a cellular level. Neointimal area, measured as intima-to-media ratio, was significantly larger in wire-injured ApoE-/- mice (3.62±0.49 compared with 0.83±0.25 in C57BL/6 mice, n=3) and there was increased oxidized low-density lipoprotein (oxLDL) infiltration and elevated plasma MPO levels. Relative increases in lysophosphatidylcholines and unsaturated phosphatidylcholines (PCs) were also observed in wire-injured ApoE-/- carotid arteries. Chlorinated lipids had no effect on VSMC proliferation, viability or migration whereas chronic incubation with oxidized phospholipids stimulated proliferation in the presence of fetal calf serum [154.8±14.2% of viable cells at 1 μM PGPC (1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphocholine) compared with control, n=6]. In conclusion, ApoE-/- mice with an inflammatory phenotype develop more neointima in wire-injured arteries and accumulation of oxidized lipids in the vessel wall may propagate this effect

Physical activity promotion in physiotherapy practice: a systematic scoping review of a decade of literature

Background: The health benefits of physical activity (PA) have been extensively documented. Globally PA levels are low with only a small proportion of the population reaching recommended levels. Insufficient PA is seen as a major public health problem with high cost to society. Physiotherapists work with people to manage long-term conditions and are well-placed to deliver individual interventions to increase PA. Despite this little is known about the evidence that exists in this field. Methods: This scoping review comprises a comprehensive search of key databases using pre-determined search terms. This is supplemented with a parallel search that incorporated novel social media strands. In-line with current guidance, a robust screening process took place using agreed inclusion and exclusion criteria. Results: Thirty one studies met the inclusion criteria. The number of studies published annually increased over the decade. Ireland and USA yielded the largest number of publications with only one study from the UK. The target populations included physiotherapists and service users from a range of clinical populations. The studies were mainly quantitative and observational in design with a predominance of studies that scoped attitudes, perceptions, barriers and current practice. Conclusions: This reconnaissance has shown the state of the evidence to be sparse and disparate. However, the sharp rise in published work in recent years is encouraging. The predominance of scoping studies and the clear social, economic and political drivers for change in this area highlights a need for more pragmatic, interventional studies that can inform clinical practice

Utility of spherical human liver microtissues for prediction of clinical drug-induced liver injury.

Drug-induced liver injury (DILI) continues to be a major source of clinical attrition, precautionary warnings, and post-market withdrawal of drugs. Accordingly, there is a need for more predictive tools to assess hepatotoxicity risk in drug discovery. Three-dimensional (3D) spheroid hepatic cultures have emerged as promising tools to assess mechanisms of hepatotoxicity, as they demonstrate enhanced liver phenotype, metabolic activity, and stability in culture not attainable with conventional two-dimensional hepatic models. Increased sensitivity of these models to drug-induced cytotoxicity has been demonstrated with relatively small panels of hepatotoxicants. However, a comprehensive evaluation of these models is lacking. Here, the predictive value of 3D human liver microtissues (hLiMT) to identify known hepatotoxicants using a panel of 110 drugs with and without clinical DILI has been assessed in comparison to plated two-dimensional primary human hepatocytes (PHH). Compounds were treated long-term (14 days) in hLiMT and acutely (2 days) in PHH to assess drug-induced cytotoxicity over an 8-point concentration range to generate IC50 values. Regardless of comparing IC50 values or exposure-corrected margin of safety values, hLiMT demonstrated increased sensitivity in identifying known hepatotoxicants than PHH, while specificity was consistent across both assays. In addition, hLiMT out performed PHH in correctly classifying hepatotoxicants from different pharmacological classes of molecules. The hLiMT demonstrated sufficient capability to warrant exploratory liver injury biomarker investigation (miR-122, HMGB1, α-GST) in the cell-culture media. Taken together, this study represents the most comprehensive evaluation of 3D spheroid hepatic cultures up to now and supports their utility for hepatotoxicity risk assessment in drug discovery

Molecular mechanism of poly(ADP-ribosyl)ation by PARP1 and identification of lysine residues as ADP-ribose acceptor sites

Poly(ADP-ribose) polymerase 1 (PARP1) synthesizes poly(ADP-ribose) (PAR) using nicotinamide adenine dinucleotide (NAD) as a substrate. Despite intensive research on the cellular functions of PARP1, the molecular mechanism of PAR formation has not been comprehensively understood. In this study, we elucidate the molecular mechanisms of poly(ADP-ribosyl)ation and identify PAR acceptor sites. Generation of different chimera proteins revealed that the amino-terminal domains of PARP1, PARP2 and PARP3 cooperate tightly with their corresponding catalytic domains. The DNA-dependent interaction between the amino-terminal DNA-binding domain and the catalytic domain of PARP1 increased Vmax and decreased the Km for NAD. Furthermore, we show that glutamic acid residues in the auto-modification domain of PARP1 are not required for PAR formation. Instead, we identify individual lysine residues as acceptor sites for ADP-ribosylation. Together, our findings provide novel mechanistic insights into PAR synthesis with significant relevance for the different biological functions of PARP family members