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ObjectiveTo i) compare incretin responses to oral glucose and mixed meal of diabetic patients with the normoglycaemic population and ii) to investigate whether incretin responses are associated with hypertriglyceridaemia and alanine aminotransferase (ALT) as liver fat marker.DesignA population-based study.MethodsA total of 163 persons with normal glucose metabolism (NGM), 20 with intermediate hyperglycaemia and 20 with type 2 diabetes aged 40–65 years participated. Participants received a mixed meal and oral glucose load on separate occasions. Glucagon-like peptide 1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and glucagon profiles were analysed as total area under the curve (tAUC) and incremental area under the curve.ResultsIn diabetic patients compared with persons with NGM, we found increased GLP-1 secretion (tAUC per hour) following oral glucose (23.2 pmol/l (95% CI 17.7–28.7) vs 18.0 (95% CI 16.9–19.1), PPConclusionThis study confirms that in type 2 diabetes, GLP-1 secretion is generally preserved and that GIP secretion is exaggerated. The mechanism underlying the divergent associations of GLP-1 and GIP metabolism with fat metabolism and liver fat accumulation warrants further study.

Background: Inflammation and oxidative stress are associated with atherosclerosis. Myeloperoxidase (MPO) is linked to both inflammation and oxidative stress by its location in leukocytes and its role in catalyzing the formation of oxidizing agents. Recent evidence suggests that MPO activity precipitates atherogenesis. Measurement of MPO in plasma may therefore contribute to cardiovascular disease (CVD) risk stratification. Content: Cross-sectional studies, case-control studies, and prospective-cohort studies investigating the relation between MPO and CVD have been evaluated. Differences in study populations, sample materials, sample handling, and assays were ascertained. Potential causal mechanisms linking MPO to accelerated atherosclerosis are discussed here. A majority of studies indicate that measurement of MPO in plasma was associated with improved CVD risk stratification above and beyond risk stratification results obtained with markers used in routine clinical practice. However, comparison of these epidemiological studies with regard to MPO and outcome is hampered because the reported MPO concentration depends on the assay method, sampling material, and preanalytical and analytical procedures. The link between MPO and CVD can, at least partly, be explained by MPO-dependent oxidation of LDL and HDL, subsequently leading to cholesterol accumulation in the arterial wall. Furthermore, MPO may reduce the bioavailability of nitric oxide, resulting in endothelial dysfunction. Finally, MPO destabilizes atherosclerotic plaques. Summary: Increasing evidence suggests that MPO is causally linked to atherosclerosis and its measurement may improve CVD risk estimation. Before MPO can be used in routine clinical practice, however, standardization of sampling and laboratory procedures is needed.

Background. Asymmetrical dimethylarginine (ADMA) is capable of inhibiting nitric oxide synthase enzymes, whereas symmetrical dimethylarginine (SDMA) competes with arginine transport. The potential role of inflammation in the metabolism of ADMA has been elucidated in an in vitro model using tumour necrosis factor-, resulting in a decreased activity of the ADMAdegrading enzyme dimethylarginine dimethylaminohydrolase (DDAH). The kidney probably plays a crucial role in the metabolism of ADMA by both urinary excretion and degradation by DDAH. We aimed to further elucidate the role of the kidney in a rat model under basal conditions and during endotoxaemia. Methods. Twenty-five male Wistar rats weighing 275–300 g were used for this study. The combination of arteriovenous concentration differences and kidney blood flow allowed calculation of net organ fluxes. Blood flow was measured using radiolabelled microspheres according to the reference sample method. Concentrations of ADMA, SDMA and arginine were measured by high-performance liquid chromatography. Results. The kidney showed net uptake of both ADMA and SDMA and fractional extraction rates were 35% and 31%, respectively. Endotoxaemia resulted in a lower systemic ADMA concentration (P ¼ 0.01), which was not explained by an increased net renal uptake. Systemic SDMA concentrations increased during endotoxaemia (P ¼ 0.007), which was accompanied by increased creatinine concentrations. Conclusions. The rat kidney plays a crucial role in the regulation of concentrations of dimethylarginines, as both ADMA and SDMA were eliminated from the systemic circulation in substantial amounts. Furthermore, evidence for the role of endotoxaemia in the metabolism of dimethylarginines was obtained as plasma levels of ADMA were significantly lower in endotoxaemic rats.

BACKGROUND: The accuracy by which a number of newly described semen variables can predict either total fertilization failure (TFF) or pregnancy outcome in IVF, has not previously been investigated. The study aim was, therefore, to determine prospectively the predictive value of these variables. METHODS: The semen variables investigated were the post-wash total progressively motile sperm cell count (TPMC(post-wash)), the acrosome index (AI), 'cytoplasmic residues' and normal sperm morphology, evaluated according to the strict criteria ('strict criteria'), as well as the fast and slow total radical trapping antioxidant potential ('fast TRAP' and 'slow TRAP' respectively). RESULTS: The study group (n = 87) showed a mean (+/- SD) number of 10.2 +/- SD retrieved oocytes, 12.6% TFF, a mean fertilization rate of 59.7% and a pregnancy rate of 19.5% (17/87). TFF was significantly predicted by TPMC(post-wash), 'strict criteria', AI and 'cytoplasmic residues' (all P < 0.05). The outcome after embryo transfer was significantly predicted by AI and 'fast TRAP'. Semen samples with an AI

Oxidation of LDL in the subendothelial space of the arterial wall initiates a series of events leading to enhanced uptake of LDL by scavenger receptors on macrophages and subsequent foam cell formation. Therefore, LDL oxidation is supposed to play a role in the development of atherosclerosis (1)(2). The most uniformly accepted procedure for measuring the resistance of LDL to in vitro oxidation is determination of the lag time (LT) for conjugated diene (CD) formation, initiated by catalytic amounts of transition metal ions (3). Another method is based on the oxidation of parinaric acid (PA), a fluorescent and oxidation-sensitive polyunsaturated fatty acid (FA) probe, incorporated into LDL (4). A third method monitors fluorescence development (FD) during LDL oxidation (5), caused by reaction of protein amino groups with aldehydes generated during decomposition of peroxidized FAs (6). The three methods described above measure different stages of the oxidation process. Oxidation of the PA probe represents one of the earliest stages of oxidation. The formation of CDs represents an intermediate stage of the oxidation process, and finally FD represents a late stage of the oxidation process. The methods for measurement of LDL oxidizability were evaluated with respect to reproducibility and potential for automation. Their relationship with LDL constituents, as determinants of LDL oxidation, was also studied. Subjects were 98 normolipidemic type 2 diabetic patients with good glycemic control (mean hemoglobin A1c, 6.2% ± 1.0%). LDL was isolated by ultracentrifugation between densities 1.019 and 1.063 kg/L, as described elsewhere (7). PA was purchased from Molecular Probes. Other reagents used were obtained from Merck or Sigma. Total cholesterol, free cholesterol, triglycerides, and phospholipids were analyzed using enzymatic test kits (Boehringer Mannheim). The cholesterol ester content of LDL was calculated as total minus free cholesterol. LDL lipid constituents were standardized for LDL protein …

We have recently described a technique for measuring LDL size by high performance gel-filtration chromatography (HPGC) with UV detection (Scheffer et al., Clin Chem 1997;43:1904–12). A drawback of this method is the necessity of LDL isolation before chromatography. We now describe a modification of this method based on selective detection of lipoproteins by postcolumn labeling with parinaric acid, a fluorescent lipid probe. Measuring the size of isolated LDL by HPGC in 56 subjects, we obtained diameters of 25.72 ± 0.60 nm with UV detection and of 25.74 ± 0.58 nm with fluorescence detection. The modified method is suitable for LDL size measurement in whole plasma or serum. LDL sizes measured in whole plasma correlated strongly with the respective values in isolated LDL (r = 0.976) but were on average 0.18 nm larger (P < 0.001). CVs for within- and between-series imprecision were

We describe a new technique for measuring LDL size by high-performance gel-filtration chromatography (HPGC). LDL was subjected to chromatography, and the column effluent was monitored at 280 nm. The retention time of the LDL peak was used to calculate the LDL diameter. We compared the HPGC method with gradient gel electrophoresis (GGE) on 2–10% nondenaturing polyacrylamide gels. In a group of 60 non-insulin-dependent diabetes mellitus patients, LDL size as measured by HPGC and GGE was highly correlated (r = 0.88, P

The recent article by Ensign et al. (1), reporting on a disappointingly low agreement among 4 methods to assess LDL particle size and phenotypic classification, casts doubt on the utility of these techniques in clinical practice. A similar poor correlation ( r = 0.39) between LDL size measurements by nuclear magnetic resonance (NMR) and gradient gel electrophoresis (GGE) has been reported by Witte et al. in a study with 324 individuals (2). Ensign et al.(1) advocate a standardization program to reduce the lack of concordance between methods. In addition to standardization problems, however, there is another reason for the observed discrepancy that is not addressed in the Ensign paper and accompanying editorial; the assumption that LDL size is adequately described by a single variable, diameter. We would like to point out that this assumption may not be valid for LDL particle size. The situation is akin to …

Background Endogenous arginine synthesis is important for maintaining normal plasma levels of arginine. Recently it was shown that low plasma levels of arginine are the drive for renal arginine synthesis. In chronic renal insufficiency low plasma levels of arginine are common. To evaluate renal arginine synthesis in acute renal injury, as seen after periods of ischaemia during major vascular surgery, ischaemia–reperfusion of one kidney was performed in a rat model. Methods In this model of unilateral renal ischaemia–reperfusion male Wistar rats were used. Arginase infusion (ASE) was used to lower arginine plasma levels to 50 per cent of normal and control rats received saline infusion (SAL). After an ischaemic period of 90 min, the kidney was perfused for 150 min (IR kidney). The contralateral kidney was left in situ and served as a control (CL kidney). Blood flow measurement was performed at the end of the experiment using radiolabelled microspheres. Blood samples were taken for amino acid analysis (high-pressure liquid chromatography). Uptake or release of arginine (flux) was calculated from flow and the arteriovenous concentration difference. Results Infusion of arginase efficiently decreased arginine plasma levels (ASE: mean(s.d.) 45·3(4·8) μmol/dm3 versus SAL: 107·5(6·0); P < 0·0001). In SAL rats, in both IR and CL kidneys a net uptake of arginine was observed (arginine flux CL kidney: + 12·5(4·9), IR kidney: + 23·6(5·8) μmol/dm3; P not significant). Lowering arginine plasma levels by arginase resulted in production of arginine in the (non-ischaemic) CL kidney. In contrast, in the IR kidney a net uptake of arginine was seen (arginine flux CL kidney: − 5·3(2·2) μmol/dm3, IR kidney: + 10·3(3·7) μmol/dm3; P < 0·01). Conclusion Synthesis of arginine by the kidney was impaired after a period of acute ischaemia–reperfusion. Ischaemia–reperfusion injury of the kidney is a condition often seen after major vascular reconstruction and impaired arginine synthesis might limit substrate for the l-arginine nitric oxide pathway.

Background The fundamental role of the l-arginine–nitric oxide pathway has not been established for maintaining renal blood flow. Plasma levels of arginine appear to be rate limiting for nitric oxide synthesis in various cell types. Low plasma levels of arginine are seen after major surgery, trauma and sepsis. The present study was designed to establish the effects of low arginine plasma levels on renal blood flow during the initial phase of renal ischaemia–reperfusion. Methods In this unilateral renal ischaemia–reperfusion model, male Wistar rats were used. Arginase infusion (ASE) was used to lower arginine plasma levels to 50 per cent of normal; control rats received saline infusion (SAL). After an ischaemic period of 90 min, the kidney was perfused for 150 min (IR kidney). The contralateral kidney was left in situ and served as a control (CL kidney). Blood flow measurement was performed at the end of the experiment using radiolabelled microspheres. Blood samples were taken for amino acid analysis (high-pressure liquid chromatography). Results Infusion of arginase efficiently decreased plasma levels of arginine (ASE mean(s.d.) 45·3(4·8) μmol/dm3 versus SAL 107·5(6·0) μmol/dm3; P < 0·0001). In both SAL and ASE rats, a significantly lower blood flow was found in the IR kidney compared with the (non-ischaemic) CL kidney. Lowering arginine plasma levels by arginase did not influence renal blood flow compared with SAL, in both IR and CL kidneys. Conclusion In this acute, unilateral renal ischaemia–reperfusion model, low arginine plasma levels did not further aggravate renal blood flow.