Associations between blood glucose and carotid intima-media thickness disappear after adjustment for shared risk factors: the KORA F4 study.

The association between blood glucose and carotid intima-media thickness (CIMT) is considered to be established knowledge. We aimed to assess whether associations between different measures of glycaemia and CIMT are actually independent of anthropometric variables and metabolic risk factors. Moreover, we checked published studies for the adjustment for shared risk factors of blood glucose and CIMT. Fasting glucose, 2-hour glucose, HbA1c, and CIMT were measured in 31-81-years-old participants of the population-based Cooperative Health Research in the Region of Augsburg (KORA) F4 study in Southern Germany (n = 2,663). CIMT was assessed according to the Rotterdam protocol. Linear and logistic regression models with adjustment for age, sex, anthropometric measures, hypertension, and dyslipidaemia were fitted to assess the association between continuous measures of glycaemia, and categories of glucose regulation, respectively, with CIMT. We found a 0.10 mm increase (95%-confidence interval: 0.08-0.12) in CIMT in subjects with compared to subjects without diabetes in crude analysis. This increase was not significant in age-sex adjusted models (p = 0.17). Likewise, neither impaired fasting glucose (p = 0.22) nor impaired glucose tolerance (p = 0.93) were associated with CIMT after adjustment for age, sex, and waist circumference. In multivariable adjusted models, age, sex, hypertension, waist circumference, HDL and LDL cholesterol, but neither fasting glucose nor 2-hour glucose nor HbA1c were associated with elevated CIMT. Literature findings are inconclusive regarding an independent association of glucose levels and CIMT. CIMT is highly dependent on traditional cardiovascular risk factors, but no relationships between blood glucose and CIMT were found after adjustment for age, sex, and anthropometric variables

Harmonization of growth hormone measurements with different immunoassays by data adjustment

Background: The aim of our study was to evaluate the between-assay variability of commercially available immunoassays for the measurement of human growth hormone (hGH). In addition, we asked whether the comparability of the diagnosis of childhood onset growth hormone deficiency could be improved by adjusting hGH results by statistical methods, such as linear regression, conversion factors, and quantile transformation. Methods: In archived sera from 312 children and adolescents (age: 17 days-17 years) hGH values between 0.01 and 16.5 ng/mL were determined by using the following immunoassays: AutoDELFIA (PerkinElmer), BC-IRMA (Beckman-Coulter), ELISA (Mediagnost), IMMULITE 2000 (Siemens), iSYS (IDS), Liaison (DiaSorin), UniCel DxI 800 Access (BeckmanCoulter) and "In house"-RIA (Tubingen). Results: The assays differed in median hGH concentrations by as much as 5.44 ng/mL (Immulite), and as little as 2.67 ng/mL (BC-IRMA). The mean difference between assays ranged from 0.35 to 2.71 ng/mL, whereas several samples displayed differences up to 11.4 ng/mL. The best correlation (r=0.992) was found between AutoDELFIA and Liasion, the lowest (r=0.864) was between an in-house RIA and iSYS. The between-assay CV (mean +/- SD) of values within the cut-off range was 24.3%+/- 7.4%, resulting in an assay-dependent diagnosis of growth hormone deficiency (GHD) in more than 27% of patients. Yet, adjustment of this data by linear regression or a conversion factor reduced the CV below 14%, and the ratio of assay-dependent diagnoses below 8%. Using quantile transformation, the CV and ratio were reduced to 11.4% and < 1%, respectively. Conclusions: hGH measurements using different assays vary significantly. Linear regression, conversion factors, or particularly quantile transformation are useful tools to improve comparability in the diagnostic procedure for the confirmation of GHD in childhood and adolescence

Metabolomics of dietary fatty acid restriction in patients with phenylketonuria

Patients with phenylketonuria (PKU) have to follow a lifelong phenylalanine restricted diet. This type of diet markedly reduces the intake of saturated and unsaturated fatty acids especially long chain polyunsaturated fatty acids (LC-PUFA). Long-chain saturated fatty acids are substrates of mitochondrial fatty acid oxidation for acetyl-CoA production. LC-PUFA are discussed to affect inflammatory and haemostaseological processes in health and disease. The influence of the long term PKU diet on fatty acid metabolism with a special focus on platelet eicosanoid metabolism has been investigated in the study presented here. 12 children with PKU under good metabolic control and 8 healthy controls were included. Activated fatty acids (acylcarnitines C6-C18) in dried blood and the cholesterol metabolism in serum were analyzed by liquid chromatographic tandem mass spectrometry (LC-MS/MS). Fatty acid composition of plasma glycerophospholipids was determined by gas chromatography. LC-PUFA metabolites were analyzed in supernatants by LC-MS/MS before and after platelet activation and aggregation using a standardized protocol. Patients with PKU had significantly lower free carnitine and lower activated fatty acids in dried blood compared to controls. Phytosterols as marker of cholesterol (re-) absorption were not influenced by the dietary fatty acid restriction. Fatty acid composition in glycerophospholipids was comparable to that of healthy controls. However, patients with PKU showed significantly increased concentrations of y-linolenic acid (C18:3n-6) a precursor of arachidonic acid. In the PKU patients significantly higher platelet counts were observed. After activation with collagen platelet aggregation and thromboxane B(2) and thromboxane B(3) release did not differ from that of healthy controls. Long-term dietary fatty acid restriction influenced the intermediates of mitochondrial beta-oxidation. No functional influence on unsaturated fatty acid metabolism and platelet aggregation in patients with PKU was detected

Potential of Dried Blood Self-Sampling for Cyclosporine C2 Monitoring in Transplant Outpatients

Background. Close therapeutic drug monitoring of Cyclosporine (CsA) in transplant outpatients is a favourable procedure to maintain the long-term blood drug levels within their respective narrow therapeutic ranges. Compared to basal levels (C0), CsA peak levels (C2) are more predictive for transplant rejection. However, the application of C2 levels is hampered by the precise time of blood sampling and the need of qualified personnel. Therefore, we evaluated a new C2 self-obtained blood sampling in transplant outpatients using dried capillary and venous blood samples and compared the CsA levels, stability, and clinical practicability of the different procedures. Methods. 55 solid organ transplant recipients were instructed to use single-handed sampling of each 50 μL capillary blood and dried blood spots by finger prick using standard finger prick devices. We used standardized EDTA-coated capillary blood collection systems and standardized filter paper WS 903. CsA was determined by LC-MS/MS. The patients and technicians also answered a questionnaire on the procedure and sample quality. Results. The C0 and C2 levels from capillary blood collection systems (C0 [ng/mL]: 114.5 ± 44.5; C2: 578.2 ± 222.2) and capillary dried blood (C0 [ng/mL]: 175.4 ± 137.7; C2: 743.1 ± 368.1) significantly (P < .01) correlated with the drug levels of the venous blood samples (C0 [ng/mL]: 97.8 ± 37.4; C2: 511.2 ± 201.5). The correlation at C0 was ρcap.-ven. = 0.749, and ρdried blood-ven = 0.432; at C2: ρcap.-ven. = 0.861 and ρdried blood-ven = 0.711. The patients preferred the dried blood sampling because of the more simple and less painful procedure. Additionally, the sample quality of self-obtained dried blood spots for LC-MS/MS analytics was superior to the respective capillary blood samples. Conclusions. C2 self-obtained dried blood sampling can easily be performed by transplant outpatients and is therefore suitable and cost-effective for close therapeutic drug monitoring

Management of high-risk patients with hypertension and left ventricular hypertrophy in Germany: differences between cardiac specialists in the inpatient and outpatient setting

BACKGROUND: Among patients with hypertension, those with established left ventricular hypertrophy (LVH) represent a high risk cohort with poor prognosis. We aimed to investigate differences in characteristics and health care management of such patients treated as inpatients or outpatients by cardiac specialists. METHODS: Prospective cross-sectional study in patients with hypertension and LVH who were referred to either inpatient care (rehabilitation hospitals) or to outpatient care (cardiology practices). RESULTS: A total of 6358 inpatients (59.6% males; mean age 66.6 years) and 2246 outpatients (59.5% males; mean age 63.2 years) were followed up for a mean of 23 vs. 52 days, respectively. Inpatients compared to outpatients had a significantly higher prevalence of coronary heart disease, history of stroke, renal failure or diabetes. Mean blood pressure of inpatients compared to outpatients was significantly lower both at entry (150/84 vs. 161/93 mmHg) and at end of follow-up (129/75 vs. 139/83 mmHg). After adjustment for baseline blood pressure and a propensity score, differences between out- and inpatients at end of follow-up were 8.0/5.1 mmHg in favour of inpatients. Blood pressure goals as specified by guidelines were not met by 32% of inpatients and 55% of outpatients. CONCLUSION: Inpatients had a higher rate of comorbidities and more advanced atherosclerotic disease than outpatients. Control of hypertension of inpatients was already better on admission than in outpatients, and treatment intensity in this group was also higher during the observation period. While blood pressure lowering was substantial in both groups, there were still a high proportion of patients who did not achieve treatment goals at discharge

Comparison of Whole Blood RNA Preservation Tubes and Novel Generation RNA Extraction Kits for Analysis of mRNA and MiRNA Profiles

Background: Whole blood expression profiling is frequently performed using PAXgene (Qiagen) or Tempus (Life Technologies) tubes. Here, we compare 6 novel generation RNA isolation protocols with respect to RNA quantity, quality and recovery of mRNA and miRNA. Methods: 3 PAXgene and 3 Tempus Tubes were collected from participants of the LIFE study with (n=12) and without (n=35) acute myocardial infarction (AMI). RNA was extracted with 4 manual protocols from Qiagen (PAXgene Blood miRNA Kit), Life Technologies (MagMAX for Stabilized Blood Tubes RNA Isolation Kit), and Norgen Biotek (Norgen Preserved Blood RNA Purification Kit I and Kit II), and 2 (semi-) automated protocols on the QIAsymphony (Qiagen) and MagMAX Express-96 Magnetic Particle Processor (Life Technologies). RNA quantity and quality was determined. For biological validation, RNA from 12 representative probands, extracted with all 6 kits (n=72), was reverse transcribed and mRNAs (matrix metalloproteinase 9, arginase 1) and miRNAs (miR133a, miR1), shown to be altered by AMI, were analyzed. Results: RNA yields were highest using the Norgen Kit I with Tempus Tubes and lowest using the Norgen Kit II with PAXgene. The disease status was the second major determinant of RNA yields (LIFE-AMI 11.2 vs. LIFE 6.7 mu g, p < 0.001) followed by the choice of blood collection tube. (Semi-) automation reduced overall RNA extraction time but did not generally reduce hands-on-time. RNA yields and quality were comparable between manual and automated extraction protocols. mRNA expression was not affected by collection tubes and RNA extraction kits but by RT/qPCR reagents with exception of the Norgen Kit II, which led to mRNA depletion. For miRNAs, expression differences related to collection tubes (miR30b), RNA isolation (Norgen Kit II), and RT/qRT reagents (miR133a) were observed. Conclusion: We demonstrate that novel generation RNA isolation kits significantly differed with respect to RNA recovery and affected miRNA but not mRNA expression profiles

P-576: Differential effects of selective cyclooxygenase-2 inhibitors on endothelial function in salt-induced hypertension

In view of the ongoing controversy of potential differences in cardiovascular safety of selective COX-2 inhibitors (coxibs), we compared the effects of two different coxibs and a traditional NSAID on endothelial dysfunction, a well established surrogate of cardiovascular disease, in salt-induced hypertension. Salt-sensitive (DS) and salt-resistant (DR) Dahl rats were treated with a high-sodium diet (4% NaCl) for 56 days. From days 35 to 56, diclofenac (6 mg/kg/d; DS-diclofenac), rofecoxib (2 mg/kg/d; DS-rofecoxib), celecoxib (25 mg/kg/d; DS-celecoxib) or placebo (DS-placebo) were added to the chow. Vascular reactivity of isolated aortic rings was assessed by isometric tension recording. Blood pressure increased with high sodium diet in the DS-groups which was more pronounced after diclofenac and rofecoxib treatment (p<0.005 vs DS-placebo), but slightly blunted by celecoxib (p<0.001 vs DS-placebo). Sodium diet markedly reduced NO-mediated endothelium-dependent relaxations to acetylcholine (ACh, 10−10−10−5 mol/L) in untreated hypertensive rats (p<0.0001 vs DR-placebo). Relaxation to ACh improved after celecoxib (p<0.005 vs DS-placebo and DS-rofecoxib), but remained unchanged after rofecoxib and diclofenac treatment. Vasoconstriction after NOS inhibition with Nω-Nitro-L-Arginine Methyl Ester (10-4 mol/L) was blunted in DS rats (p<0.05 vs DR-placebo), normalized by celecoxib, but not affected by rofecoxib or diclofenac. Protein expression of eNOS was decreased in DS rats with a trend for increased eNOS levels in the DS-celecoxib group (97.8±25.6 vs 54.8±2.8 %, p=0.088 vs DS-placebo). Indicators of oxidative stress, 8-isoprostane levels, were elevated in untreated DS rats on 4% NaCl (6.55±0.58 vs 3.65±1.05 ng/ml, p<0.05) and normalized by celecoxib only (4.29±0.58 ng/ml), while SOD protein expression was decreased in DS rats and not affected by any treatment. Plasma levels of prostaglandines did not change during high sodium diet or any treatment. These data show that celecoxib, but not rofecoxib or diclofenac, improves endothelial dysfunction and reduces oxidative stress, thus pointing to differential effects of coxibs in salt-sensitive hypertension. Am J Hypertens (2004) 17, 243A-243A; doi: 10.1016/j.amjhyper.2004.03.65

Serum amino acid profiles and their alterations in colorectal cancer

Mass spectrometry-based serum metabolic profiling is a promising tool to analyse complex cancer associated metabolic alterations, which may broaden our pathophysiological understanding of the disease and may function as a source of new cancer-associated biomarkers. Highly standardized serum samples of patients suffering from colon cancer (n=59) and controls (n=58) were collected at the University Hospital Leipzig. We based our investigations on amino acid screening profiles using electrospray tandem-mass spectrometry. Metabolic profiles were evaluated using the Analyst 1.4.2 software. General, comparative and equivalence statistics were performed by R 2.12.2. 11 out of 26 serum amino acid concentrations were significantly different between colorectal cancer patients and healthy controls. We found a model including CEA, glycine, and tyrosine as best discriminating and superior to CEA alone with an AUROC of 0.878 (95% CI 0.815-0.941). Our serum metabolic profiling in colon cancer revealed multiple significant disease-associated alterations in the amino acid profile with promising diagnostic power. Further large-scale studies are necessary to elucidate the potential of our model also to discriminate between cancer and potential differential diagnoses. In conclusion, serum glycine and tyrosine in combination with CEA are superior to CEA for the discrimination between colorectal cancer patients and control

Rationale and Design of the Leipzig (LIFE) Heart Study: Phenotyping and Cardiovascular Characteristics of Patients with Coronary Artery Disease

We established the Leipzig (LIFE) Heart Study, a biobank and database of patients with different stages of coronary artery disease (CAD) for studies of clinical, metabolic, cellular and genetic factors of cardiovascular diseases.The Leipzig (LIFE) Heart Study (NCT00497887) is an ongoing observational angiographic study including subjects with different entities of CAD. Cohort 1, patients undergoing first-time diagnostic coronary angiography due to suspected stable CAD with previously untreated coronary arteries. Cohort 2, patients with acute myocardial infarction (MI) requiring percutaneous revascularization. Cohort 3, patients with known left main coronary artery disease (LMCAD).We present preliminary results of demographics and phenotyping based on a 4-years analysis of a total of 3,165 subjects. Cohort 1 (n=2,274) shows the typical distribution of elective coronary angiography cohorts with 43% cases with obstructive CAD and 37% normal angiograms. Cohorts 2 and 3 consist of 590 and 301 subjects, respectively, adding patients with severe forms of CAD. The suitability of the database and biobank to perform association studies was confirmed by replication of the CAD susceptibility locus on chromosome 9p21 (OR per allele: 1.55 (any CAD), 1.54 (MI), 1.74 (LMCAD), p<10(-6), respectively). A novel finding was that patients with LMCAD had a stronger association with 9p21 than patients with obstructive CAD without LMCAD (OR 1.22, p=0.042). In contrast, 9p21 did not associate with myocardial infarction in excess of stable CAD.The Leipzig (LIFE) Heart Study provides a basis to identify molecular targets related to atherogenesis and associated metabolic disorders. The study may contribute to an improvement of individual prediction, prevention, and treatment of CAD