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Biomedical subjects

H Reinauer

Publications and source records attributed to H Reinauer.

At least 55 records · Page 3Linked to original sources
PubMed1995

Evaluation of a radioimmunoassay for determination of calcitriol in human sera employing a 125I-labelled tracer.

The performance characteristics of a radioimmunoassay employing a 125I-labelled tracer for determination of calcitriol in human sera are reported. The assay is based on an immunoextraction step employing a monoclonal antibody against calcitriol followed by a radioimmunoassay (using 125I-labelled calcitriol and sheep antiserum against calcitriol). Bound/free separation is performed with an anti-sheep IgG antibody bound to cellulose. Within-run imprecision (n = 20) was 12.8% (mean = 9.4 ng/l) and 11.1% (mean = 48.8 ng/l), between-day imprecision (n = 11) was 27.1% (mean = 9.6 ng/l) and 17.2% (mean = 47.2 ng/l). Linearity of dilution as investigated by mixing pooled sera containing 62.0 ng/l and 4.7 ng/l calcitriol, respectively. The relationship between measured and expected concentrations was characterized by a linear correlation coefficient of r = +0.990. The values obtained with the 125I-based radioimmunoassay were compared with those obtained with a radioreceptor-assay using a 3H-labelled tracer; the regression line was y = 1.091 x -4.545 (n = 84; r = +0.935), where y = calcitriol [125I] [ng/l] and x = calcitriol [3H] [ng/l]. Mixing 9 volumes of sera from patients with renal insufficiency (n = 7) with 1 volume of 'calibrator F' (assigned value: 227 ng/l) yielded recovery rates of 90 +/- 8% (mean +/- SD). The detection limit was 3.0 ng/l. The cross-reactivity of cholecalciferol metabolites was found to be < 0.00003 for 25-hydroxycholecalciferol, 24R,25-dihydroxycholecalciferol and 25S,26-dihydroxycholecalciferol. A preliminary reference interval (5th to 95th percentile) was established in 40 apparently healthy persons (17 males and 23 females; age range: 20-61 [mean: 32] years) (19-74 ng/l). The method presented shows high practicability and may therefore be considered as a useful alternative to cumbersome assays using 3H-labelled tracers.

Adult↗
PubMed1994

Prostacyclin improves glucose utilization in patients with sepsis.

PURPOSE: In patients with sepsis, impaired glucose metabolism and altered microcirculatory blood flow are common findings. Prostacyclin (PGI2) improves tissue oxygenation, indicated by enhanced oxygen delivery (DO2) and oxygen uptake (VO2). The purpose of this study was to explore whether these effects are associated with improved glucose utilization. METHODS: In 7 patients with sepsis, glucose metabolism was analyzed using dideuterated and 13C-labeled glucose and isotope dilution mass spectrometry. All patients received total parenteral nutrition with glucose covering 60% to 70% of the predicted energy expenditure and needed continuous intravenous insulin (40 microU.kg-1.min-1) to keep blood glucose concentrations below 10 mmol.l-1.VO2 and carbon dioxide production (VCO2) were continuously measured directly from the respiratory gases via indirect calorimetry. After equilibration of the infused labeled glucose with body glucose pool, baseline variables of glucose metabolism were assessed: glucose oxidation rate was determined from the enrichment of 13CO2 in the expired gas during primed constant infusion of [U-13C]glucose, glucose turnover rate (Ra) from the plasma enrichment of simultaneously infused [6,6-2H2]glucose. Endogenous glucose production rate was calculated as the difference between Ra and glucose infusion rate. Then, we examined the effect of PGI2 infusion (5 to 12.5 ng.kg-1.min-1) on glucose metabolism and gaseous exchange. RESULTS: The PGI2-induced increase in DO2 (from 15.8 to 17.7 ml.kg-1.min-1; P < .05) resulted in an increase in directly measured VO2 from 5.0 to 5.3 ml.kg-1.min-1 (P < .01) whereas VCO2 remained unchanged. Although glucose turnover and production rates remained constant, glucose oxidation rate increased significantly from 1.21 to 1.38 mg.kg-1.min-1 (P < .02). CONCLUSIONS: Improving tissue perfusion and oxygenation with PGI2 may also modify the impaired glucose metabolism by increasing glucose oxidation rate in patients with sepsis, suggesting enhanced adenosine triphosphate production.

Adenosine Triphosphate↗
PubMed1994

Helicobacter pylori in patients with systemic sclerosis: detection with the 13C-urea breath test and eradication.

In patients with systemic sclerosis peristaltic abnormalities may delay gastric emptying, giving rise to bacterial overgrowth, including possibly Helicobacter pylori (HP). Infection with Helicobacter is an important risk factor for esophageal and gastric diseases, including esophagitis, gastritis and gastric cancer. The purpose of this prospective study was to assess gastric HP infection in patients with systemic sclerosis. In 12 patients with systemic sclerosis the newly introduced breath test with 13C-labelled urea was used for indirect detection of gastric urease activity due to HP infection. Five out of 12 patients gave Helicobacter-positive results (42%); 7 patients were negative for Helicobacter colonization (58%). Thus, the risk for gastric diseases caused by HP infection is enhanced in patients with systemic sclerosis compared with white healthy, asymptomatic persons examined in other studies. Helicobacter-positive patients were treated with 2 x 20 mg omeprazole and 4 x 500 mg amoxicillin over 14 days. Afterwards the 13C-urea breath test was repeated and showed negative results for Helicobacter in all systemic sclerosis patients treated. Dual therapy with omeprazole and amoxicillin therapy effectively eradicated HP. The 13C-urea breath test did not cause any side-effects and is therefore considered to be a non-invasive, non-toxic and safe method for the diagnosis and therapeutic control of Helicobacter-status.

Adult↗
PubMed1993

Regulation of cardiac insulin receptor function by guanosine nucleotides.

The present study examined the effect of GTP-gamma-S on the function of insulin receptors partially purified from adult rat cardiomyocytes by WGA chromatography. GTP-gamma-S increased receptor autophosphorylation about two times and fully mimicked the stimulatory action of insulin on poly(Glu:Tyr) phosphorylation with no additional effect of the hormone. The effect of GTP-gamma-S was specific, dose-dependent, and due to an increase in the Vmax of the kinase. In the presence of ATP or AMP-PNP, insulin significantly enhanced the binding of [35S]GTP-gamma-S to the partially purified insulin receptor. The findings suggest coupling of the insulin receptor to a G-protein which may be involved in the regulation of tyrosine kinase activity.

Animals↗
PubMed1993

Differential insertion of insulin receptor complexes into Triton X-114 bilayer membranes. Evidence for a differential accessibility of the membrane-exposed receptor domain.

In the present study, the Triton X-114 phase-separation system has been used to characterize molecular properties of the membrane-exposed domain of an integral-membrane hormone receptor. This approach provides novel details of the structure/function relationship of insulin receptors. Upon raising the temperature of a micellar Triton X-114 solution above the cloud-point, a detergent enriched phase pellets and coprecipitates 95% of the purified insulin-free (alpha beta)2 receptors. In contrast, 83% of the hormone bound (alpha beta)2 receptor complexes prefer the detergent-depleted phase, exhibiting prominent properties of non-membraneous proteins. Kinetic studies show that, following insulin binding, the amphiphilicity of the receptor complexes is immediately altered. Only monodisperse (alpha beta)2 complexes were detected when receptor/insulin complexes of the detergent-depleted phase were analyzed by detergent-free sucrose density centrifugation in the presence of 10 nM insulin. These results can be explained in the light of the lipid-bilayer-like organization of the precipitating Triton X-114; hormone-induced intramolecular alterations of (alpha beta)2 receptors appear to fundamentally restrict access to the membrane-exposed receptor domain. Basically, different molecular properties are found for alpha beta receptors. Only 67% of the insulin-free receptors coprecipitate with the Triton-X-114-enriched phase; following insulin binding the coprecipitation is only decreased to 42%. In contrast to (alpha beta)2 receptors, formation of noncovalently aggregated receptor complexes, which are detected by sucrose density centrifugation, could account for the exclusion of alpha beta receptor species from Triton X-114 membranes.

Chemical Precipitation↗
PubMed1993

Development of criteria for the evaluation of reference method values.

We present a uniform concept for the requirements of reference methods (RMs) used for target-setting in the German external quality assessment (EQA) scheme. It is proposed that the total allowable error (TAE) of the RMs shall be 0.2 times the acceptance limits in the EQA-scheme. From the TAE are derived limits for maximum RM-bias, RM-imprecision, and the number of measurements.

Chemistry, Clinical↗
PubMed1993

Candidate reference methods for determining target values for cholesterol, creatinine, uric acid, and glucose in external quality assessment and internal accuracy control. II. Method transfer.

We describe the testing of transferability of candidate Reference Methods developed by INSTAND for cholesterol, creatinine, uric acid, and glucose. The methods are based on isotope dilution-gas chromatography--mass spectrometry. The study consisted of two parts: setup of the methods and self-evaluation for readiness in the collaborating laboratory, followed by independent measurements in parallel with INSTAND. Criteria used for judging the transferability and general reliability of the candidate Reference Methods were: the accuracy and precision of the collaborating laboratory and the agreement between the two laboratories. The accuracy was judged from the results on the Standard Reference Material 909 from the National Institute of Standards and Technology. For all analytes except glucose the bias from the certified value was < 0.7%. The mean intralaboratory imprecision ranged from 0.66% to 1.24%. The agreement between the results was tested by an advanced linear-regression analysis and Student's t-test. In general, the results demonstrate that the candidate Reference Methods developed by INSTAND can be successfully transferred without loss of their inherent precision and accuracy.

Blood Glucose↗
PubMed1993

Candidate reference methods for determining target values for cholesterol, creatinine, uric acid, and glucose in external quality assessment and internal accuracy control. I. Method setup.

In Germany, the target values for External Quality Assessment (EQA) and internal accuracy control are determined by Reference Methods for several analytes, including cholesterol, creatinine, uric acid, and glucose. We present candidate Reference Methods for these compounds, based on isotope dilution-gas chromatography--mass spectrometry methods that have been developed at INSTAND, one of the two official Germany EQA reference institutions. Each Reference Method target value is calculated from six independent measurements performed on three different days. The mean method CVs ranged from 0.66% for glucose to 0.96% for creatinine. The inaccuracy (bias) of the methods is < 0.7%, as compared with the Standard Reference Material 909 of the National Institute of Standards and Technology. The maximum total error of a Reference Method value, including the 95% confidence interval and systematic errors, is < 2.3%. The presented candidate Reference Methods have been successfully used to set target values in the German EQA scheme and the internal accuracy control of routine laboratories.

Blood Glucose↗
PubMed1992

Contraction-induced translocation of the glucose transporter Glut4 in isolated ventricular cardiomyocytes.

Field stimulation of isolated adult ventricular cardiomyocytes was used to study the effect of contractile activity on 3-O-methylglucose transport and the subcellular distribution of Glut4. Cells contracting at a frequency of 1 Hz for 30 min exhibited unaltered basal and insulin-stimulated rates of glucose transport when compared to resting cells. However, at 5 Hz 3-O-methylglucose transport increased to 224% of control after 5 min. Under these conditions insulin was unable to produce a significant additional stimulation of glucose transport. Immunoblotting with an anti-Glut4 polyclonal antibody showed that both insulin and contraction (5 Hz) increased the amount of Glut4 in a plasma membrane fraction by about 8-fold with a parallel decrease in an intracellular membrane fraction by 60-65%. These data suggest the existence of an identical insulin- and contraction-recruitable Glut4 transporter pool in cardiomyocytes.

3-O-Methylglucose↗
PubMed1992

Evidence indicating that the multicatalytic proteinase of rabbit reticulocytes is not incorporated as a core enzyme into a 26 S proteinase complex.

We have reinvestigated the recent proposal that the multicatalytic proteinase, together with other components of reticulocyte lysate, may become incorporated into a very large, "26 S" proteinase complex via an ATP-dependent process. Different from these published results, we consistently isolate the multicatalytic proteinase as a 650,000 Da "20 S" multisubunit proteinase. Analysis on nondenaturing polyacrylamide gels of reticulocyte fractions containing the putative complexed form of the multicatalytic proteinase reveal that activity against succinyl-Leu-Leu-Val-Tyr-7-amino-4-methylcoumarin is associated with two groups of protein of different molecular mass. One migrates like multicatalytic proteinase purified to homogeneity, displays, on sodium dodecyl sulfate gels, a set of protein species in the range of 23,000-32,000 Da, characteristic of the multicatalytic proteinase, and is recognized by a monospecific antibody to the enzyme. In contrast, the activity associated with the higher molecular mass (26 S) proteinase complex lacks the typical multicatalytic proteinase subunits and is devoid of antigenic material, when tested with the antibody. These results confirm and extend our recent findings in mouse liver by showing that the multicatalytic proteinase is not a constituent of a 26 S proteinase complex.

Animals↗
PubMed1992

[Diabetic cardiopathy. Pathophysiologic concepts and therapeutic approaches].

Epidemiological data reviewed suggest that diabetes itself increases the cardiac risk of diabetics (types I and II), independently from the development of coronary heart disease and in addition to other risk factors (hypertension, hypercholesterolemia, hypertriglyceridemia, smoking and others), presumably by a specific myocardial disease called "diabetic cardiopathy", or according to the recommendations of the WHO, "diabetic heart muscle disease." Disturbances of the left and right ventricular function as well as the autonomic function of the heart can be understood as signs of this specific cardiopathy. The pathophysiological mechanisms underlying this disease are not yet fully known; however, recent evidence is presented that diabetes leads to a facet of metabolic dysfunctions regarding glucose and energy metabolism, calcium homeostasis and the expression of specific proteins that diminish the ability of the heart to respond to increased workload and increase the vulnerability of the heart in diabetes. Since preliminary experimental data indicate that inhibitors of the angiotensin-converting enzyme can protect the heart in diabetes, it is intriguing to suggest that increased release of angiotensin II plays a significant role in the change from reduced adaptability to irreversible damage of the heart in diabetes.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1991

Diabetes-induced decrease in the mRNA coding for sarcoplasmic reticulum Ca(2+)-ATPase in adult rat cardiomyocytes.

The present study examined the level of the mRNA coding for the sarcoplasmic reticulum (SR) Ca(2+)-ATPase in isolated ventricular myocytes from streptozotocin-diabetic rats and genetically obese (fa/fa) rats using Northern blotting techniques. In both animal models one single transcript with a mean size of 4.12 kb could be detected. In insulin-deficient rats the Ca(2+)-ATPase transcript level decreased by 51% when compared to normal rats. In obese animals a modest decrease in the Ca(2+)-ATPase mRNA content to 77% of lean controls has been detected. Decreased mRNA expression of the SR Ca(2+)-ATPase may partly explain the delayed diastolic relaxation observed in the diabetic heart.

Animals↗
PubMed1991

Inverse regulation of glucose transporter Glut4 and G-protein Gs mRNA expression in cardiac myocytes from insulin resistant rats.

The present study examined the mRNA levels of glucose transporter Glut4 and G-protein Gs alpha-subunit in isolated ventricular myocytes from lean and genetically obese (fa/fa) Zucker rats and streptozotocin-diabetic rats. In obese animals the amount of transcripts coding for Glut4 increased to 122 +/- 6% of lean controls, whereas the mRNA coding for Gs alpha-subunit decreased by 42 +/- 12%. An unaltered level of Gs mRNA was observed in insulin deficient rats. When cardiomyocytes from normal rats were treated with insulin, the Glut4 transcript level increased by 48 +/- 5%, whereas the Gs mRNA level decreased by 55 +/- 8%. The findings suggest that insulin may act as a potential regulator of Glut4 and Gs mRNA expression in the cardiac cell.

Animals↗
PubMed1991

Determination of aldosterone in human serum by isotope dilution gas chromatography/mass spectrometry using a new heptafluorobutyryl derivative.

The formation of a new derivative of aldosterone with heptafluorobutyric anhydride for gas chromatography/mass spectrometry (GC/MS) is presented. The highest and also the most prominent ion of this derivative is observed at m/z 734. The new derivative is, under the described conditions, reproducibly formed, is stable and has good gas chromatographic properties. These characteristics make the new derivative extremely suitable for selective, sensitive, precise and accurate analysis of aldosterone in serum by GC/MS in association with isotope dilution. This is proven by the agreement between the measurement results obtained by two laboratories for samples of three batches of lyophilized control serum. The sample pretreatment procedures used in each laboratory are described.

Aldosterone↗
PubMed1991

Alpha-adrenoceptor-mediated increase in cytosolic free calcium in isolated cardiac myocytes.

The effect of alpha-adrenoceptor stimulation on the concentration of cytosolic free calcium (Cai2+) was determined by measuring indo-l fluorescence in isolated ventricular cardiomyocytes from normal and streptozotocin-diabetic rat; 1.3 x 10(5) alpha 1-adrenoceptors per normal myocyte and an unaltered number of these receptors in cells from diabetic rats were detected using the alpha 1-selective ligand WB-4101. Under basal conditions, Cai2+ was found to be 154 +/- 4 nM (n = 34) reaching a value of 192 +/- 10 nM (n = 15) after stimulation of myocytes with a maximal dose of methoxamine for 5 min. Under the same conditions the leakage of dye produced a significantly smaller increase of basal values of 169 +/- 5 nM (n = 17). Indo-l loaded cells did not respond to beta-stimulation unless in the presence of KCl (50 mM), demonstrating the specificity of methoxamine action. Treatment of cells with nifedipine or chelation of extracellular calcium by EGTA did not modify the alpha-adrenergic response. Experiments with cardiomyocytes from streptozotocin-diabetic rats showed an unaltered modulation of Cai2+ by both alpha- and beta-receptor stimulation. It is concluded that signalling by alpha 1-adrenoceptors in ventricular cardiomyocytes results in mobilization of intracellular calcium stores.

Animals↗
PubMed1991

Induction of insulin resistance in primary cultured adult cardiac myocytes.

Primary cultured cardiac myocytes from adult rats were used to elucidate the role of insulin and catecholamines in the development of insulin resistance in this tissue. Cardiomyocytes exhibited a stable response toward insulin up to at least 48 h in serum-free culture, as determined by measuring the effect of the hormone on initial rates of 2-deoxyglucose uptake. Culturing of cells in the absence of insulin for 6 and 19 h, respectively, resulted in a loss of insulin sensitivity and a reduced (33%) maximal responsiveness after 19 h of insulin deficiency. This was paralleled by a decrease in [14C]phenylalanine incorporation and an unaltered level of insulin binding. Insulin action was completely lost in cells cultured in the presence of cycloheximide for 19 h. When added to the culture medium for 4 h, both isoproterenol and (Bu)2cAMP decreased insulin binding by about 50%. Under these conditions maximal insulin responsiveness was not affected by isoproterenol but was reduced by 46% by (Bu)2cAMP. Nifedipine antagonized the inhibitory action of (Bu)2cAMP, but was ineffective when the culture period was extended to 19 h. Cardiomyocytes cultured in the presence of palmitate exhibited a largely reduced (67%) insulin responsiveness, which was only partly restored by inhibition of fatty acid oxidation. From these data we conclude that: 1) insulin deficiency induces insulin resistance due to decreased protein synthesis; 2) sustained, prolonged elevation of cAMP modulates insulin action by both Ca(++)-dependent and Ca(++)-independent mechanisms; and 3) free fatty acids antagonize insulin action by both metabolic and nonmetabolic pathways.

Animals↗
PubMed1991

Tissue-specific changes of multicatalytic proteinase activity in the fasted rat.

During a three-day fast, followed by four days of refeeding, the content of the multicatalytic proteinase as well as hydrolyzing activity towards Suc-Leu-Leu-Val-Tyr-7-amino-4-methylocoumarin (SLLVT-MCA) was measured in various rat tissues. When compared with normal rats, the MCP content, as determined by immunochemical techniques, was unchanged over the entire experimental period in the three tissues examined: gastrocnemius muscle, thymus and testis. By contrast, a differential response was observed in the three tissues with respect to specific and total SLLVT-MCA splitting activity: for thymus and testis, these values were again unchanged, whereas in gastrocnemius muscle, both specific and total enzyme activity fell by almost 70% on day three of fasting but returned to control values on day four of refeeding. This change in activity was not due to the accumulation or degradation of a specific proteinase inhibitor. Data demonstrate that, in association with the insulin-deficient state of starvation, the activity of the multicatalytic proteinase shows an adaptive behaviour which becomes manifest in some but not in other tissues.

Animals↗