[Show abstract][Hide abstract] ABSTRACT: Severe infections in intensive care patients show high morbidity and mortality rates. Linezolid is an antimicrobial drug frequently used in critically ill patients. Recent data indicates that there might be high variability of linezolid serum concentrations in intensive care patients receiving standard doses. This study was aimed to evaluate whether standard dosing of linezolid leads to therapeutic serum concentrations in critically ill patients.
Critical care (London, England) 07/2014; 18(4):R148. · 4.72 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In major depressive disorder, changes in cytokine levels have been reported to play a role in pathogenesis. Therefore, we sought to investigate a broad range of cytokines in depression. We compared serum levels of interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, IL-13, granulocyte macrophage colony-stimulating factor (GM-CSF), interferon (INF-γ) and tumor necrosis factor (TNF)-α in 64 subjects with current depression and 206 non-depressed subjects. Depressed patients had higher levels of IL-2, IL-5, IL-12, IL-13, GM-CSF, INF-γ and TNF-α, compared to non-depressed subjects. Splitting groups into non-obese (BMI < 30) and obese (BMI ≥ 30), the non-obese depressed patients (n = 40) showed elevated IL-5, IL-12, IL-13, GM-CSF, INF-γ and TNF-α levels compared to non-obese and non-depressed subjects (n = 85). The obese and depressed patients (n = 24) showed elevated levels of IL-5, IL-12 and INF-γ compared to obese but not depressed subjects (n = 121). Levels of several cytokines were found to be associated with physical activity, employment status and presence of daily naps. The results support over-expression of pro-inflammatory cytokines in depression and extend the range of cytokines potentially associated with depression to include GM-CSF, IL-5 and IL-13. Changes in these cytokines may contribute to co-morbidity between depression and allergic and asthmatic diseases. The results also suggest inflammatory processes associated with obesity, and support an interaction between cytokine serum concentrations and behavioral aspects of both obesity and depression.
Journal of Psychiatric Research 05/2014; · 4.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Numerous high-throughput sequencing studies focus on detecting conventionally spliced mRNAs in RNA-seq data. However, non-standard RNAs arising through gene fusion, circularization, or trans-splicing are often neglected. We introduce a novel, unbiased algorithm to detect splice junctions from single-end cDNA sequences. In contrast to other methods, our approach accommodates multi-junction structures. Our method compares favorably with competing tools on conventionally spliced mRNAs and, with a gain of up to 40\% of recall, systematically outperforms them on reads with multiple splits, trans-splicing and circular products. The algorithm is integrated into our mapping tool segemehl (www.bioinf.uni-leipzig.de/Software/segemehl/).
[Show abstract][Hide abstract] ABSTRACT: Plant sterols are supplemented in foods to reduce cardiovascular risk. Randomized controlled trials show 2 g of plant sterols a day reduce serum cholesterol by about 10%. This reduction in serum cholesterol levels is achieved at the expense of increased serum plant sterol levels. Findings in patients with phytosterolemia, in experimental studies and in clinical trials have lead to speculations that plant sterols might be atherogenic. In view of emerging safety issues the role of plant sterols in cardiovascular prevention has become controversial. This review reflects the ongoing controversial scientific debate and points out recent developments in guidelines of national and international societies.
Biochemical and Biophysical Research Communications 01/2014; · 2.41 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Endolysosomal organelles play a key role in trafficking, breakdown and receptor-mediated recycling of different macromolecules such as low-density lipoprotein (LDL)-cholesterol, epithelial growth factor (EGF) or transferrin. Here we examine the role of two-pore channel (TPC) 2, an endolysosomal cation channel, in these processes. Embryonic mouse fibroblasts and hepatocytes lacking TPC2 display a profound impairment of LDL-cholesterol and EGF/EGF-receptor trafficking. Mechanistically, both defects can be attributed to a dysfunction of the endolysosomal degradation pathway most likely on the level of late endosome to lysosome fusion. Importantly, endolysosomal acidification or lysosomal enzyme function are normal in TPC2-deficient cells. TPC2-deficient mice are highly susceptible to hepatic cholesterol overload and liver damage consistent with non-alcoholic fatty liver hepatitis. These findings indicate reduced metabolic reserve of hepatic cholesterol handling. Our results suggest that TPC2 plays a crucial role in trafficking in the endolysosomal degradation pathway and, thus, is potentially involved in the homoeostatic control of many macromolecules and cell metabolites.
[Show abstract][Hide abstract] ABSTRACT: Abstract Background: Liquid biobanking is an important tool for laboratory diagnostics in routine settings and clinical studies. However, the current knowledge about adequate storage conditions for different classes of biomarkers is incomplete and, in part, contradictory. Here, we performed a comprehensive study on the effects of different storage conditions on the stability of various biomarkers in human serum and plasma. Methods: Serum and citrated plasma were aliquoted and stored at 4°C, -20°C, -80°C, and <-130°C for 0, 7, 30, and 90 days, respectively (5-10 pools/condition). Additionally, frozen aliquots were temporarily exposed to higher temperatures during storage to simulate removing individual samples. Stability was tested for 32 biomarkers from 10 different parameter classes (electrolytes, enzymes, metabolites, inert proteins, complement factors, ketone bodies, hormones, cytokines, coagulation factors, and sterols). Results: Biobanking at -80°C and <-130°C for up to 90 days did not lead to substantial changes (defined as >3 interassay coefficients of variation and p<0.01) of any biomarker concentration. In contrast, storage at 4°C and -20°C induced substantial changes in single biomarker concentrations in most classes. Such substantial changes were increases (<20%) in electrolytes, metabolites, and proteins, and decreases (<96%) in enzymes, ketone bodies, cytokines, and coagulation factors. Biomarker stability was minimally affected by occasional short-term thermal exposure. Conclusions: Based on these results, we provide recommendations for storage conditions of up to 90 days for several biomarkers. Generally, storage at ≤-80°C for at least 90 days including occasional short-term thermal exposure is an excellent storage condition for most biomarkers.
Clinical Chemistry and Laboratory Medicine 12/2013; · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Abstract Background: Linezolid serum concentrations have been shown to be highly variable in critically ill patients with often sub-therapeutic drug levels regarding minimal inhibitory concentrations for relevant pathogens. Consequently, therapeutic drug monitoring of linezolid must be considered, requiring a reliable and convenient analytical method. We therefore developed and validated an LC-MS/MS method applying isotope dilution internal standardization and on-line solid phase extraction for serum linezolid quantification. Methods: Sample preparation was based on protein precipitation and on-line solid phase extraction with two-dimensional liquid chromatography and column switching. Three-fold deuterated linezolid was used as the internal standard. The method was validated involving two separate LC-MS/MS systems covering the concentration range of 0.13-32 mg/L. The run time was 4 min. Results: Validation revealed good analytical performance, with inaccuracy <6% and imprecision of <7.3% (CV) for six quality control samples (0.38-16.0 mg/L). The method was found to be robust during the validation process and during a pharmacokinetic study so far involving 600 samples. Comparative measurements on two LC-MS/MS systems revealed close agreement. Conclusions: This LC-MS/MS assay described herein is a convenient, robust and reliable method for linezolid quantification in serum which can be routinely applied using different LC-MS/MS systems. The method can be used for clinical studies and subsequent TDM of linezolid.
Clinical Chemistry and Laboratory Medicine 10/2013; · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Since 2007, genome-wide association studies (GWAS) have led to the identification of numerous loci of atherosclerotic cardiovascular disease. The majority of these loci harbor genes previously not known to be involved in atherogenesis. In this review, we summarize the recent progress in understanding the pathophysiology of genetic variants in atherosclerosis.
Fifty-eight loci with P < 10 have been identified in GWAS for coronary heart disease and myocardial infarction. Of these, 23 loci (40%) overlap with GWAS loci of classical risk factors such as lipids, blood pressure, and diabetes mellitus, suggesting a potential causal relation. The vast majority of the remaining 35 loci (60%) are at genomic regions where the mechanism in atherogenesis is unclear. Loci most frequently found in independent GWAS were at Chr9p21.3 (ANRIL/CDKN2B-AS1), Chr6p24.1 (PHACTR1), and Chr1p13.3 (CELSR2, PSRC1, MYBPHL, SORT1). Recent work suggests that Chr9p21.3 exerts its effects through epigenetic regulation of target genes, whereas mechanisms at Chr6p24.1 remain obscure, and Chr1p13.3 affects plasma LDL cholesterol.
Novel GWAS loci indicate that our understanding of atherosclerosis is limited and implicate a role of hitherto unknown mechanisms, such as epigenetic gene regulation in atherogenesis.
Current opinion in lipidology 10/2013; 24(5):410-8. · 6.13 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Rationale: QTL mapping of an intercross between C57.Apoe(-/-) and FVB.Apoe(-/-) mice revealed an atherosclerosis locus controlling aortic root lesion area on proximal chromosome 10, Ath11. In previous work subcongenic analysis showed Ath11 to be complex with proximal (10a) and distal (10b) regions. Objective: To identify the causative genetic variation underlying the atherosclerosis modifier locus Ath11 10b. Methods and Results: We now report subcongenic J, which narrows the 10b region to five genes, Myb, Hbs1L, Aldh8a1, Sgk1, and Raet1e. Sequence analysis of these genes revealed no amino acid coding differences between the parental strains. However, comparing aortic expression of these genes between F1.Apoe(-/-) Chr10SubJ((B/F)) and F1.Apoe(-/-) Chr10SubJ((F/F)) uncovered a consistent difference only for Raet1e with decreased, virtually background, expression associated with increased atherosclerosis in the latter. The key role of Raet1e was confirmed by showing transgene-induced aortic over-expression of Raet1e in F1.Apoe(-/-) Chr10SubJ((F/F)) mice decreased atherosclerosis. Promoter reporter constructs comparing C57 and FVB sequences identified an FVB mutation in the core of the major aortic transcription start site abrogating activity. Conclusions: This non-biased approach has revealed Raet1e, an MHC class 1-like molecule expressed in lesional aortic endothelial cells and macrophage-rich regions, as a novel atherosclerosis gene, and represents one of the few successes of the QTL strategy in complex diseases.
Circulation Research 08/2013; · 11.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To investigate the role of secretory phospholipase A2-(sPLA2)-IIA in cardiovascular disease.
Higher circulating levels of sPLA2-IIA mass or sPLA2 enzyme activity have been associated with increased risk of cardiovascular events. However, it is not clear if this association is causal. A recent phase-III clinical trial of an sPLA2 inhibitor (varespladib) was stopped prematurely for lack of efficacy.
We conducted a Mendelian randomization meta-analysis of 19 general population studies (8021 incident, 7513 prevalent major vascular events (MVE) in 74,683 individuals) and ten acute coronary syndrome (ACS) cohorts (2520 recurrent MVE in 18,355 individuals) using rs11573156, a variant in PLA2G2A encoding the sPLA2-IIA isoenzyme, as an instrumental variable.
PLA2G2A rs11573156 C-allele associated with lower circulating sPLA2-IIA mass (38-44%) and sPLA2 enzyme activity (3-23%) per C allele. The odds ratio (OR) for MVE per rs11573156 C-allele was 1.02(95%CI:0.98,1.06) in general populations and 0.96(95%CI:0.90,1.03) in ACS cohorts. In the general population studies, the OR derived from the genetic instrumental variable analysis for MVE for a 1 log unit lower sPLA2-IIA mass was 1.04(95%CI:0.96,1.13), and differed from the non-genetic observational estimate (OR0.69;95%CI:0.61,0.79). In the ACS cohorts, both the genetic instrumental variable and observational ORs showed a null association with MVE. Instrumental variable analysis failed to show associations between sPLA2 enzyme activity and MVE.
Reducing sPLA2-IIA mass is unlikely to be a useful therapeutic goal for preventing cardiovascular events.
Journal of the American College of Cardiology 07/2013; · 14.09 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The chromosome 9p21 (Chr9p21) locus of coronary artery disease has been identified in the first surge of genome-wide association and is the strongest genetic factor of atherosclerosis known today. Chr9p21 encodes the long non-coding RNA (ncRNA) antisense non-coding RNA in the INK4 locus (ANRIL). ANRIL expression is associated with the Chr9p21 genotype and correlated with atherosclerosis severity. Here, we report on the molecular mechanisms through which ANRIL regulates target-genes in trans, leading to increased cell proliferation, increased cell adhesion and decreased apoptosis, which are all essential mechanisms of atherogenesis. Importantly, trans-regulation was dependent on Alu motifs, which marked the promoters of ANRIL target genes and were mirrored in ANRIL RNA transcripts. ANRIL bound Polycomb group proteins that were highly enriched in the proximity of Alu motifs across the genome and were recruited to promoters of target genes upon ANRIL over-expression. The functional relevance of Alu motifs in ANRIL was confirmed by deletion and mutagenesis, reversing trans-regulation and atherogenic cell functions. ANRIL-regulated networks were confirmed in 2280 individuals with and without coronary artery disease and functionally validated in primary cells from patients carrying the Chr9p21 risk allele. Our study provides a molecular mechanism for pro-atherogenic effects of ANRIL at Chr9p21 and suggests a novel role for Alu elements in epigenetic gene regulation by long ncRNAs.
[Show abstract][Hide abstract] ABSTRACT: Dyslipidemia is a frequent component of the metabolic disorder of diabetic patients contributing to organ damage. Herein, in low-density lipoprotein receptor-deficient hyperlipidemic and streptozotozin-induced diabetic mice, hyperglycemia and hyperlipidemia acted reciprocally, accentuating renal injury and altering renal function. In hyperglycemic-hyperlipidemic kidneys, the accumulation of Tip47-positive lipid droplets in glomeruli, tubular epithelia, and macrophages was accompanied by the concomitant presence of the oxidative stress markers xanthine oxidoreductase and nitrotyrosine, findings that could also be evidenced in renal biopsy samples of diabetic patients. As liver X receptors (LXRα,β) regulate genes linked to lipid and carbohydrate homeostasis and inhibit inflammatory gene expression in macrophages, the effects of systemic and macrophage-specific LXR activation were analyzed on renal damage in hyperlipidemic-hyperglycemic mice. LXR stimulation by GW3965 up-regulated genes involved in cholesterol efflux and down-regulated proinflammatory/profibrotic cytokines, inhibiting the pathomorphology of diabetic nephropathy, renal lipid accumulation, and improving renal function. Xanthine oxidoreductase and nitrotyrosine levels were reduced. In macrophages, GW3965 or LXRα overexpression significantly suppressed glycated or acetylated low-density lipoprotein-induced cytokines and reactive oxygen species. Specifically, in mice, transgenic expression of LXRα in macrophages significantly ameliorated hyperlipidemic-hyperglycemic nephropathy. The results demonstrate the presence of lipid droplet-induced oxidative mechanisms and the pathophysiologic role of macrophages in diabetic kidneys and indicate the potent regulatory role of LXRs in preventing renal damage in diabetes.
American Journal Of Pathology 01/2013; · 4.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A current challenge in the era of genome-wide studies is to determine the responsible genes and mechanisms underlying newly identified loci. Screening of the plasma proteome by high-throughput mass spectrometry (MALDI-TOF MS) is considered as a promising approach for identification of metabolic and disease processes. Therefore, plasma proteome screening might be particularly useful for identifying responsible genes when combined with analysis of variation in the genome. Here, we describe a proteomic quantitative trait locus (pQTL) study of plasma proteome screens in an F2 intercross of 455 mice mapped with 177 genetic markers across the genome. 69 of 176 peptides revealed significant LOD-scores (≥5.35) demonstrating strong genetic regulation of distinct components of the plasma proteome. Analyses were confirmed by mechanistic studies and MALDI-TOF/TOF, LC-MS/MS analyses of the two strongest pQTLs: A pQTL for m/z 3494 (LOD 24.9, D11Mit151) was identified as the N-terminal 35 amino acids of hemoglobin subunit A (Hba) and caused by genetic variation in Hba. Another pQTL for m/z 8713 (LOD 36.4; D1Mit111) was caused by variation in apolipoprotein A2 (Apoa2) and co-segregated with HDL-cholesterol. Taken together, we show that genome-wide plasma proteome profiling in combination with genome-wide genetic screening aids in the identification of causal genetic variants affecting abundance of plasma proteins.
[Show abstract][Hide abstract] ABSTRACT: Common marmosets are suitable non-human primate models for many human diseases. Standard values for blood parameters are required to evaluate physiological and pathological situations. Two studies were conducted: study I to determine standard values and study II to examine these under changed housing conditions. In study I, all parameters for clinical chemistry were similar in range for both genders with these specifics: male marmosets had significantly higher total and LDL cholesterol levels than females, whereas the mean corpuscular volume and the mean corpuscular haemoglobin were significantly lower than in females. In study II, glucose, lymphocytes and salivary cortisol were significantly lower, and faecal cortisol was increased during the change of housing conditions. In conclusion, standard values for haematology and clinical chemistry for the common marmoset were determined. Further on, parameters that are influenced by relocation stress and its importance for experimental results are described.
Journal of Medical Primatology 07/2012; 41(4):241-50. · 1.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Many human diseases are modulated by intrauterine environment, which is called prenatal programming. This study investigated effects of prenatal glucocorticoids on the lipid metabolism of three filial generations of common marmosets.
Pregnant primates were treated with dexamethasone during pregnancy. Body weight and blood lipid parameters of adult female offspring (F1: n = 5, F2: n = 6, F3: n = 3) were compared with age-related female controls (n = 12).
F1, F2, and F3 offspring showed significantly lower percentage of plasma n3 fatty acids than controls. F2 and F3 presented higher cholesterol levels, with significantly more LDL cholesterol, significantly less HDL triglycerides and an enhanced cholesterol/HDL cholesterol ratio. Body weight was not significantly affected.
Prenatal dexamethasone led to higher amounts of cardiovascular risk factors and less protective parameters in female F1-F3 offspring. The intergenerational consequences suggest prenatal programming through epigenetic effects.
Journal of Medical Primatology 07/2012; 41(4):231-40. · 1.11 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In the framework of a case report on a patient suffering from major depression and inflammatory bowel disease we address the pharmacotherapeutical options in case of subtherapeutic mirtazapine levels.
We applied therapeutic drug monitoring (TDM) and cytochrome P450 2D6 genotyping, and switched to an orodispersible tablet.
Thus, mirtazapine plasma levels could be raised and clinical improvement of the depressive symptoms was achieved.
[Show abstract][Hide abstract] ABSTRACT: Muscle wasting occurs in both chronic heart failure (CHF) and normal aging and contributes to exercise intolerance and increased morbidity/mortality. However, the molecular mechanisms of muscle atrophy in CHF and their interaction with aging are still largely unknown. We therefore measured the activation of the ubiquitin-proteasome system and the lysosomal pathway of intracellular proteolysis in muscle biopsies of CHF patients and healthy controls in two age strata and assessed the age-dependent effects of a 4-week endurance training program on the catabolic-anabolic balance.
Sixty CHF patients (30 patients aged ≤55 years, mean age 46±5 years; 30 patients aged ≥65 years, mean age 72±5 years) and 60 healthy controls (30 subjects aged ≤55 years, mean age 50±5 years; 30 subjects aged ≥65 years, mean age 72±4 years) were randomized to 4 weeks of supervised endurance training or to a control group. Before and after the intervention, vastus lateralis muscle biopsies were obtained. The expressions of cathepsin-L and the muscle-specific E3 ligases MuRF-1 and MAFbx were measured by real-time polymerase chain reaction and confirmed by Western blot. At baseline, MuRF-1 expression was significantly higher in CHF patients versus healthy controls (mRNA: 624±59 versus 401±25 relative units; P=0.007). After 4 weeks of exercise training, MuRF-1 mRNA expression was reduced by -32.8% (P=0.02) in CHF patients aged ≤55 years and by -37.0% (P<0.05) in CHF patients aged ≥65 years.
MuRF-1, a component of the ubiquitin-proteasome system involved in muscle proteolysis, is increased in the skeletal muscle of patients with heart failure. Exercise training results in reduced MuRF-1 levels, suggesting that it blocks ubiquitin-proteasome system activation and does so in both younger and older CHF patients.
URL: http://www.clinicaltrials.gov. Unique identifier: NCT00176319.
[Show abstract][Hide abstract] ABSTRACT: Proliferation signal inhibitors/mTOR-inhibitors have been shown to reduce de novo development of hypercholesterolemic atherosclerosis in animal models. However, their effect on pre-existing atherosclerosis has not yet been studied.
Feeding LDL-R-KO mice a high cholesterol diet for 12 weeks resulted in formation of moderate fibroatheroma (induction phase). Sixty mice received either everolimus (1 or 5 mg/kg) or no everolimus for further 12 weeks (treatment phase). Everolimus significantly enhanced hypercholesterolemia (plasma cholesterol +45%, p<0.001). Atherosclerosis progressed obstructively in treated and non-treated mice. Everolimus (5 mg/kg) tended to reduced progression in aortic root lesions (0.28±0.02 vs. 0.33±0.03 mm(2), p=ns) and brachiocephalic lesions (0.044±0.006 vs. 0.066±0.012 mm(2), p=ns) but without significance. Everolimus (5mg/kg) resulted in an arrest of CD68 positive plaque area (p=0.03) and nearly halved CD68 fraction (p=0.05) in aortic root lesions but not in brachiocephalic lesions. Taken together, despite a trend to reduced progression and inflammatory cell content there was less conclusive net effect of everolimus treatment than expected.
A higher potential of everolimus in the treatment of atherosclerosis might be obscured by its concomitant hypercholesterolemia. Considering stronger effects in previous studies we suggest that everolimus might exert more potent anti-atherogenic properties in earlier stages of atherogenesis than in advanced atherosclerosis.
[Show abstract][Hide abstract] ABSTRACT: Post-traumatic dysfunction of the immune system is a major source of morbidity and mortality in patients with multiple trauma. The underlying intracellular mechanisms are still incompletely understood. Previous mRNA expression studies in monocytes suggested an involvement of the MAP kinases p38 and JNK and of the transcription factor c-Jun. Therefore, it was the aim of this study to elucidate whether alterations in the protein expression p38 MAPK, JNK, and c-Jun could be linked to PRBC substitution and survival.
Thirty-seven patients with blunt multiple injuries and an ISS > 16 points were enrolled in our study. Blood was drawn on admission and 6, 12, 24, 48, and 72 h after the traumatic event. Monocytes were isolated immediately after sample collection and nuclear protein was extracted and phosphoprotein concentrations were measured. The resulting data were statistically analyzed.
An increased activation of MAP kinases and c-Jun could be shown in patients who died from their injuries. Additionally, patients who received PRBC substitution ≥10 units exhibited increased expression of activated MAP kinases and c-Jun.
We present a serial, sequential investigation in human monocytes of major trauma patients evaluating the activation of p38 MAPK, JNK and c-Jun in the post-traumatic period. We show that death after trauma and massive PRBC substitution are associated with activation of this pathway. The p38 MAPK, JNK, and c-Jun have well established proinflammatory properties. Therefore, it appears likely that this pathway is involved in the systemic hyperinflammatory states seen after massive PRBC transfusion and multiple trauma.
Journal of Surgical Research 03/2012; 178(1):385-9. · 2.02 Impact Factor