Physical and catalytic properties of a peroxidase derived from cytochrome c

Except for its redox properties, cytochrome c is an inert protein. However, dissociation of the bond between methionine-80 and the heme iron converts the cytochrome into a peroxidase. Dissociation is accomplished by subjecting the cytochrome to various conditions, including proteolysis and hydrogen peroxide (H₂O₂)-mediated oxidation. In affected cells of various neurological diseases, including Parkinson's disease, cytochrome c is released from the mitochondrial membrane and enters the cytosol. In the cytosol cytochrome c is exposed to cellular proteases and to H₂O₂ produced by dysfunctional mitochondria and activated microglial cells. These could promote the formation of the peroxidase form of cytochrome c. In this study we investigated the catalytic and cytolytic properties of the peroxidase form of cytochrome c. These properties are qualitatively similar to those of other heme-containing peroxidases. Dopamine as well as sulfhydryl group-containing metabolites, including reduced glutathione and coenzyme A, are readily oxidized in the presence of H₂O₂. This peroxidase also has cytolytic properties similar to myeloperoxidase, lactoperoxidase, and horseradish peroxidase. Cytolysis is inhibited by various reducing agents, including dopamine. Our data show that the peroxidase form of cytochrome c has catalytic and cytolytic properties that could account for at least some of the damage that leads to neuronal death in the parkinsonian brain.     

Adult ADHD and suicide

While suicidal behaviour has been implicated in a plethora of psychiatric disorders including depression, psychoses and substance abuse, its association with adult ADHD is largely under-researched. Given that emotional instability and the high prevalence of comorbid conditions such as mood disorders and alcohol/drug dependence are typical for ADHD, the question of suicide risk must not be neglected in this patient group. A review of the current literature focusing on this issue provides strong evidence that ADHD patients are at a significant risk for experiencing suicidal ideations and committing suicide. For daily clinical practice, it is therefore essential to incorporate this aspect into the diagnostic and therapeutic process and to take preventive measures.     

Biomarker Profiling by Nuclear Magnetic Resonance Spectroscopy for the Prediction of All-Cause Mortality: An Observational Study of 17,345 Persons

Background

Early identification of ambulatory persons at high short-term risk of death could benefit targeted prevention. To identify biomarkers for all-cause mortality and enhance risk prediction, we conducted high-throughput profiling of blood specimens in two large population-based cohorts.

Methods and Findings

106 candidate biomarkers were quantified by nuclear magnetic resonance spectroscopy of non-fasting plasma samples from a random subset of the Estonian Biobank (n = 9,842; age range 18–103 y; 508 deaths during a median of 5.4 y of follow-up). Biomarkers for all-cause mortality were examined using stepwise proportional hazards models. Significant biomarkers were validated and incremental predictive utility assessed in a population-based cohort from Finland (n = 7,503; 176 deaths during 5 y of follow-up). Four circulating biomarkers predicted the risk of all-cause mortality among participants from the Estonian Biobank after adjusting for conventional risk factors: alpha-1-acid glycoprotein (hazard ratio [HR] 1.67 per 1–standard deviation increment, 95% CI 1.53–1.82, p = 5×10⁻³¹), albumin (HR 0.70, 95% CI 0.65–0.76, p = 2×10⁻¹⁸), very-low-density lipoprotein particle size (HR 0.69, 95% CI 0.62–0.77, p = 3×10⁻¹²), and citrate (HR 1.33, 95% CI 1.21–1.45, p = 5×10⁻¹⁰). All four biomarkers were predictive of cardiovascular mortality, as well as death from cancer and other nonvascular diseases. One in five participants in the Estonian Biobank cohort with a biomarker summary score within the highest percentile died during the first year of follow-up, indicating prominent systemic reflections of frailty. The biomarker associations all replicated in the Finnish validation cohort. Including the four biomarkers in a risk prediction score improved risk assessment for 5-y mortality (increase in C-statistics 0.031, p = 0.01; continuous reclassification improvement 26.3%, p = 0.001).

Conclusions

Biomarker associations with cardiovascular, nonvascular, and cancer mortality suggest novel systemic connectivities across seemingly disparate morbidities. The biomarker profiling improved prediction of the short-term risk of death from all causes above established risk factors. Further investigations are needed to clarify the biological mechanisms and the utility of these biomarkers for guiding screening and prevention. Please see later in the article for the Editors'' Summary     

Impact of Hormone-Associated Resistance to Activated Protein C on the Thrombotic Potential of Oral Contraceptives: A Prospective Observational Study

Introduction

The increased thrombotic risk of oral contraceptives (OC) has been attributed to various alterations of the hemostatic system, including acquired resistance to activated protein C (APC). To evaluate to what extent OC-associated APC resistance induces a prothrombotic state we monitored plasma levels of thrombin and molecular markers specific for thrombin formation in women starting OC use. Elevated plasma levels of thrombin have been reported to characterize situations of high thrombotic risk such as trauma-induced hypercoagulability, but have not yet been studied during OC use.

Patients and Methods

Blood samples were collected prospectively from healthy women (n = 21) before and during three menstruation cycles after start of OC. APC resistance was evaluated using a thrombin generation-based assay. Plasma levels of thrombin and APC were directly measured using highly sensitive oligonucleotide-based enzyme capture assay (OECA) technology. Thrombin generation markers and other hemostasis parameters were measured additionally.

Results

All women developed APC resistance as indicated by an increased APC sensitivity ratio compared with baseline after start of OC (p = 0.0003). Simultaneously, plasma levels of thrombin, prothrombin fragment 1+2, and of thrombin-antithrombin complexes did not change, ruling out increased thrombin formation. APC plasma levels were also not influenced by OC use, giving further evidence that increased thrombin formation did not occur.

Conclusions

In the majority of OC users no enhanced thrombin formation occurs despite the development of APC resistance. It cannot be ruled out, however, that thrombin formation might occur to a greater extent in the presence of additional risk factors. If this were the case, endogenous thrombin levels might be a potential biomarker candidate to identify women at high thrombotic risk during OC treatment. Large-scale studies are required to assess the value of plasma levels of thrombin as predictors of OC-associated thrombotic risk.     

Serum tau fragments as predictors of death or poor neurological outcome after out-of-hospital cardiac arrest

Background: Anoxic brain injury is the primary cause of death after resuscitation from out-of-hospital cardiac arrest (OHCA) and prognostication is challenging. The aim of this study was to evaluate the potential of two fragments of tau as serum biomarkers for neurological outcome.

Methods: Single-center sub-study of 171 patients included in the Target Temperature Management (TTM) Trial randomly assigned to TTM at 33?°C or TTM at 36?°C for 24?h after OHCA. Fragments (tau-A and tau-C) of the neuronal protein tau were measured in serum 24, 48 and 72?h after OHCA. The primary endpoint was neurological outcome.

Results: Median (quartile 1 – quartile 3) tau-A (ng/ml) values were 58 (43–71) versus 51 (43–67), 72 (57–84) versus 71 (59–82) and 76 (61–92) versus 75 (64–89) for good versus unfavourable outcome at 24, 48 and 72?h, respectively (p_group = 0.95). Median tau C (ng/ml) values were 38 (29–50) versus 36 (29–49), 49 (38–58) versus 48 (33–59) and 48 (39–59) versus 48 (36–62) (p_group = 0.95). Tau-A and tau-C did not predict neurological outcome (area under the receiver-operating curve at 48?h; tau-A: 0.51 and tau-C: 0.51).

Conclusions: Serum levels of tau fragments were unable to predict neurological outcome after OHCA.     

Mitochondrial haplogroups, control region polymorphisms and malignant melanoma: a study in middle European Caucasians

Background

Because mitochondria play an essential role in energy metabolism, generation of reactive oxygen species (ROS), and apoptosis, sequence variation in the mitochondrial genome has been postulated to be a contributing factor to the etiology of multifactorial age-related diseases, including cancer. The aim of the present study was to compare the frequencies of mitochondrial DNA (mtDNA) haplogroups as well as control region (CR) polymorphisms of patients with malignant melanoma (n = 351) versus those of healthy controls (n = 1598) in Middle Europe.

Methodology and Principal Findings

Using primer extension analysis and DNA sequencing, we identified all nine major European mitochondrial haplogroups and known CR polymorphisms. The frequencies of the major mitochondrial haplogroups did not differ significantly between patients and control subjects, whereas the frequencies of the one another linked CR polymorphisms A16183C, T16189C, C16192T, C16270T and T195C were significantly higher in patients with melanoma compared to the controls. Regarding clinical characteristics of the patient cohort, none of the nine major European haplogroups was associated with either Breslow thickness or distant metastasis. The CR polymorphisms A302CC-insertion and T310C-insertion were significantly associated with mean Breslow thickness, whereas the CR polymorphism T16519C was associated with metastasis.

Conclusions and Significance

Our results suggest that mtDNA variations could be involved in melanoma etiology and pathogenesis, although the functional consequence of CR polymorphisms remains to be elucidated.     

The proprotein convertase PCSK9 induces the degradation of low density lipoprotein receptor (LDLR) and its closest family members VLDLR and ApoER2

The proprotein convertase PCSK9 gene is the third locus implicated in familial hypercholesterolemia, emphasizing its role in cardiovascular diseases. Loss of function mutations and gene disruption of PCSK9 resulted in a higher clearance of plasma low density lipoprotein cholesterol, likely due to a reduced degradation of the liver low density lipoprotein receptor (LDLR). In this study, we show that two of the closest family members to LDLR are also PCSK9 targets. These include the very low density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2) implicated in neuronal development and lipid metabolism. Our results show that wild type PCSK9 and more so its natural gain of function mutant D374Y can efficiently degrade the LDLR, VLDLR, and ApoER2 either following cellular co-expression or re-internalization of secreted human PCSK9. Such PCSK9-induced degradation does not require its catalytic activity. Membrane-bound PCSK9 chimeras enhanced the intracellular targeting of PCSK9 to late endosomes/lysosomes and resulted in a much more efficient degradation of the three receptors. We also demonstrate that the activity of PCSK9 and its binding affinity on VLDLR and ApoER2 does not depend on the presence of LDLR. Finally, in situ hybridization show close localization of PCSK9 mRNA expression to that of VLDLR in mouse postnatal day 1 cerebellum. Thus, this study demonstrates a more general effect of PCSK9 on the degradation of the LDLR family that emphasizes its major role in cholesterol and lipid homeostasis as well as brain development.     

Bundle sheath leakiness and light limitation during C4 leaf and canopy CO2 uptake

Perennial species with the C(4) pathway hold promise for biomass-based energy sources. We have explored the extent that CO(2) uptake of such species may be limited by light in a temperate climate. One energetic cost of the C(4) pathway is the leakiness () of bundle sheath tissues, whereby a variable proportion of the CO(2), concentrated in bundle sheath cells, retrodiffuses back to the mesophyll. In this study, we scale from leaf to canopy level of a Miscanthus crop (Miscanthus x giganteus hybrid) under field conditions and model the likely limitations to CO(2) fixation. At the leaf level, measurements of photosynthesis coupled to online carbon isotope discrimination showed that leaves within a 3.3-m canopy (leaf area index = 8.3) show a progressive increase in both carbon isotope discrimination and as light decreases. A similar increase was observed at the ecosystem scale when we used eddy covariance net ecosystem CO(2) fluxes, together with isotopic profiles, to partition photosynthetic and respiratory isotopic flux densities (isofluxes) and derive canopy carbon isotope discrimination as an integrated proxy for at the canopy level. Modeled values of canopy CO(2) fixation using leaf-level measurements of suggest that around 32% of potential photosynthetic carbon gain is lost due to light limitation, whereas using determined independently from isofluxes at the canopy level the reduction in canopy CO(2) uptake is estimated at 14%. Based on these results, we identify as an important limitation to CO(2) uptake of crops with the C(4) pathway.