[Current issues in measurement and reporting of urinary albumin excretion]. / Données actuelles sur le dosage de l'excrétion urinaire de l'albumine.

Urinary excretion of albumin indicates kidney damage and is recognized as a risk factor for progression of kidney disease and cardiovascular disease. The role of urinary albumin measurements has focused attention on the clinical need for accurate and clearly reported results. The National Kidney Disease Education Program and the IFCC convened a conference to assess the current state of preanalytical, analytical, and postanalytical issues affecting urine albumin measurements and to identify areas needing improvement. The chemistry of albumin in urine is incompletely understood. Current guidelines recommend the use of the albumin/creatinine ratio (ACR) as a surrogate for the erro-prone collection of timed urine samples. Although ACR results are affected by patient preparation and time of day of sample collection, neither is standardized. Considerable intermethod differences has been reported for both albumin and creatinine measurement, but trueness is unknown because there are no reference measurement procedures for albumin and no referance materials for either analyte in urine. The recommanded reference intervals for the ACR do not take into account the large intergroup differences in creatinine excretion (e.g., related to differences in age, sex, and ethicity) nor the continuous increase in risk related to albumin excretion. Clinical needs have been identified for standardization of (a) urine collection methodes, (b) urine albumin and creatinine measurements based on a complete reference system, (c) reporting of test results, and (d) reference intervals for the ACR.

Overexpression of acyl-coA binding protein and its effects on the flux of free fatty acids in McA-RH 7777 cells.

Overexpression of acyl-CoA binding protein (ACBP) was induced in a rat hepatoma cell line (McA-RH 7777) by stable integration of rat ACBP cDNA. The transfected cells (ACBP-27) had 3.5-fold higher concentrations of ACBP than control cells (14 vs. 4 ng/microg DNA). Both ACBP-27 and control cells were cultured in the presence of various concentrations of radiolabeled palmitic acid; and the effects of ACBP on lipogenesis and beta-oxidation were studied. Incubation of the cells with 100 microM palmitic acid resulted in 42% greater incorporation of the fatty acid in ACBP-27 cells as compared to that in the control cells. This increased incorporation of the fatty acid was observed predominantly in the triglyceride fraction. Higher concentrations of palmitic acid (200 to 400 microM) were associated with a significant decrease in the production of 14CO2 in the ACBP-27 cell line than in the control cells, while lower concentrations had no effect. Our data suggest a role for ACBP in the partitioning of fatty acids between esterification reactions leading to the formation of neutral lipids and beta-oxidation. ACBP may play a regulatory role by influencing this important branch point in intermediary lipid metabolism.

A study to examine the accuracy of potassium measurements in clinical laboratories across Canada.

OBJECTIVE: To assess the accuracy of potassium measurements in clinical laboratories across Canada. DESIGN AND METHOD: The flame atomic emission spectrophotometry reference method for the determination of potassium was established at the Canadian Reference Laboratory by using National Institute of Science and Technology standard reference materials. The method was subsequently used to assign target values for potassium to Canadian Reference Laboratory's External Quality Assessment human-serum-based testing material. A total of 503 laboratories participated and 9,279 individual External Quality Assessment test results were included in the study. Bias was determined by using difference plots. RESULTS: Clinically significant bias (>1.6%) was observed in 45.9% of the laboratories. Bias ranged from 0.34 mmol/L to -0.54 mmol/L. At low concentrations (<3.5 mmol/L) a positive bias was most frequently observed (14.7% of analytical systems). At high potassium concentrations (>5.1 mmol/L) a negative bias was most frequently observed (31.4% of analytical systems). CONCLUSION: Inaccuracy in potassium results can contribute to test redundancy and mismanagement of patients, while prohibiting the merger of laboratory data from disparate testing sites for the purpose of trending and consolidation within a "universal health record." Inaccurate test results and the lack of standardization among laboratories adversely impact our ability to establish common reference intervals and critical limits. This inability has an adverse effect on medical decisions and patient care.

Laboratory standardization of a large international clinical trial: the DAIS experience. DAIS Project Group. Diabetes Atherosclerosis Intervention Study.

OBJECTIVE: To implement a quality control program for the standardization and harmonization of lipid and lipoprotein analyses as performed at two core laboratories (St. Paul's Hospital, UBC [Vancouver], and NPHI [Helsinki]) for the Diabetes Atherosclerosis Intervention Study (DAIS). DESIGN AND METHODS: A DAISSOFT computer program was designed to minimize the occurrence of data and sample management errors during the course of the study. Fresh human serum was used for the provision of an accuracy based external quality control program that monitored the analytical performance of lipid testing at these two laboratories. A separate program was designed for monitoring hemoglobin A1c (HbA1c). At the outset of the study, allowable total error goals were established for each analyte. Ongoing performance was monitored using bimonthly blinded challenges of fresh human serum. The two EQA programs routinely monitored the analysis of total cholesterol, calculated LDL-cholesterol, HDL-cholesterol, net triglycerides, apoprotein A-1, apoprotein B, and HbA1c. RESULTS: The EQA precision and accuracy data for the measurement of total cholesterol at the two core laboratories over the last 5 years indicated both laboratories operated with good precision, approximately 1% CV over the time period. The accuracy at both laboratories was similar initially. Part way through the study, the accuracy of the cholesterol method at NHPI tended to drift upward with an operating positive bias (+3%) relative to the Abell Kendall reference method. Triglyceride measurements were the most problematic for the study. By EQA cycle 8, the accuracy of the method at UBC had stabilized and was meeting the accuracy goals of the study. NPHI's method was negatively biased relative to the accuracy base of the DAIS study. In spite of recalibrating their method, NPHI found it difficult to maintain consistent accuracy for the measurement of triglycerides during the study. Both laboratories operated their HDL methods with excellent precision. Accuracy at NHPI was well maintained over the course of the study whereas the accuracy of HDL measurements at UBC was more problematic. There was an inconsistent variation in the accuracy of apoprotein A-1 measurements at both laboratories. In most cases, the bias would be corrected by the time of the next EQA challenge. In the case of apo B, one laboratory was standardized to the CDC while the other laboratory was standardized to IFCC/WHO. The discrepancy between these two accuracy bases was >20%. Recalibration to a common accuracy base rectified the problem. Only minor problems were encountered with the precision and accuracy of the DIAMAT assay for hemoglobin A-1c. The two DAIS core laboratories consistently operated within the 9% total error goals of the study for HbA1c. CONCLUSIONS: Through the use of this program, the two DAIS core laboratories were able to maintain their lipid analyses within the limits of allowable total error that had been established for the study.

Point: status of lipid and lipoprotein standardization.

Cholesterol and triglyceride standardization procedures have been used extensively and continuously since the 1950s. Definitive and Reference Methods, as well as primary and secondary standards, have been developed and maintained as the basis for evaluating the accuracy of results by various methods in many laboratories. But, although standardization efforts for apolipoprotein A-I and B measurements have been reported in detail in the scientific literature, much less has been reported in the area of total and lipoprotein cholesterol and triglyceride standardization efforts. Standardized cholesterol and triglyceride concentrations, determined in multiple large epidemiological and clinical studies, have been instrumental to the National Cholesterol Education Program panels that have assessed the lipoprotein values associated with risk of coronary disease, and have determined the cutpoints that are now used extensively by physicians to guide diagnosis and treatment of individual patients.

The effects of 3-hydroxy-3-methylglutaryl-CoA reductase inhibition on tissue levels of carnitine and carnitine acyltransferase activity in the rabbit.

Recently, a new class of lipid lowering agents [3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitors] was introduced into clinical practice. The use of these agents could lead to a secondary deficiency in carnitine, which may manifest clinically as a myalgia/myositis-a side effect that is occasionally seen with this class of drugs. In the present study, we examined the effect of an HMG-CoA reductase inhibitor (lovastatin) on serum and tissue levels of carnitine and carnitine acyltransferase activities in the rabbit. Rabbits (n = 6) were fed chow containing lovastatin (30 mg/d) for 16 wk. Blood was collected and tissues (liver, heart, and skeletal muscle) harvested at sacrifice. Free and total carnitine were measured in serum and tissues by a radioenzymatic method. Carnitine acetyltransferase and carnitine palmitoyltransferase (CPT) activities were determined and expressed relative to DNA. Serum free (24.0 +/- 2.6 vs. 29.4 +/- 3.1 microM) and total (35.1 +/- 4.7 vs. 52.8 +/- 8.8 microM) carnitine levels increased significantly with 16 wk of treatment. This increase in total carnitine was mainly due to an increase in the levels of serum acylcarnitine (12.7 +/- 3.1 vs 26.5 +/- 5.7 microM). Tissue levels of total carnitine were significantly decreased by the treatment. Carnitine acetyltransferase was unaffected by the treatment, whereas there was a significant increase in the activity of CPT in the liver and heart.

Effects of high-fat diet and fasting on levels of acyl-coenzyme A binding protein in liver, kidney, and heart of rat.

Acyl-coenzyme A (CoA) binding protein (ACBP) is a 10-kd protein that binds acyl-CoA moieties and stimulates medium-chain fatty acid synthesis by goat mammary gland fatty acid synthetase. Its exact role in intermediary lipid metabolism has not been fully elucidated. It is hypothesized that ACBP is directly involved in the metabolism of lipid. In the present study, purified rat liver ACBP was used to generate a polyclonal antisera for radioimmunoassay of ACBP in tissue specimens isolated from fasted rats and rats fed normal rat chow and a high-fat diet. In addition, purified ACBP was used to examine its effect on the activity of mitochondrial outer membrane (OM) carnitine palmitoyltransferase (CPT0). Fasting for 24 hours significantly decreased tissue levels of ACBP in the liver (69.0 +/- 7.2 v 46.7 +/- 5.0 pg/ng DNA), whereas feeding of a high-fat diet for 48 hours caused ACBP levels to increase (69.0 +/- 7.2 v 103.9 +/- 18.0). Hepatic levels of this protein continued to increase and remained elevated with prolonged exposure to the high-fat diet (28 days). A similar pattern of change was observed in the kidney, but the magnitude of change was less. Heart ACBP did not respond acutely to the high-fat diet, but did increase after prolonged exposure (28 days). Fasting had no effect on ACBP levels in kidney and heart. Addition of ACBP to an in vitro assay system significantly increased the activity of CPT0 (from 5.2 +/- 0.8 to 72.1 +/- 5.3 nmol palmitoylcarnitine formed.min-1.mg-1 protein) when measured under inhibiting concentrations of palmitoyl-CoA (40 mumol/L).(ABSTRACT TRUNCATED AT 250 WORDS)

Electron transfer flavoprotein: ubiquinone oxidoreductase (ETF:QO) deficiency in an adult.

A 19-year-old woman with mild myopathic symptoms from age 6 and fasting intolerance presented with a Reye-like syndrome and a myopathy. Investigations disclosed a lipid storage myopathy, type II glutaric acidemia, and carnitine deficiency in skeletal muscle. Riboflavin and carnitine treatment corrected the metabolic abnormalities and she improved clinically. She later died from pulmonary complications secondary to aspiration. Subsequent studies established electron transfer flavoprotein: ubiquinone oxidoreductase (ETF:QO) deficiency (fibroblast ETF:QO activity was 2.9 mU/mg, normal range is 14.1 +/- 3.8 mU/mg) as the cause of her illness. This is the first documented case of ETF:QO diagnosed in an adult.

The role of apoproteins in disorders of lipoprotein metabolism.

Apolipoproteins are essential components of plasma lipoproteins. They facilitate the absorption and secretion of fat from the intestine, serve as activators of enzymes of lipoprotein metabolism and act as ligands for lipoprotein receptors on cell surfaces. Changes in the apoprotein quantity or composition affect plasma lipid concentrations. Specific examples of apolipoprotein alterations are known for apo A-I, apo B, and for C and E apolipoproteins. With the availability of both apolipoprotein protein and gene analytical techniques both quantitative and qualitative assays of apolipoproteins are becoming more important in the diagnosis of lipoprotein disorders. Assays of apo A-I, apo B and, less frequently, apo C-II and E apolipoproteins are useful diagnostic tools, if performed properly. Major problems with the standardization and quality control of these assays remain to be solved.

Digoxin-like immunoreactivity, displacement of ouabain and inhibition of Na+/K+ ATPase by four steroids known to be increased in essential hypertension.

An endogenous digoxin-like immunoreactive substance(s) (DLIS, "endoxin") may be of significance in the etiology of essential hypertension (EH). Progesterone, dehydroepiandrosterone sulphate (DHEA-S), 11-deoxycortisol and 18-hydroxy-11-deoxycorticosterone (18-OH-DOC), four steroids known to be increased in essential hypertension, were found to have digoxin-like immunoreactivity at levels 1,000 times higher than physiological concentrations. Of these steroids, progesterone and 18-OH-DOC were the most efficient in displacing 3H-ouabain from canine kidney Na+/K+ ATPase whereas progesterone and 11-deoxycortisol were the most potent inhibitors of this enzyme's activity. Although 18-OH-DOC and DHEA-S cross-reacted with digoxin-specific antibodies, their ability to inhibit Na+/K+ ATPase activity was minimal. Although it is concluded that these steroids may contribute to DLIS as isolated from hypertensive patients, it is unlikely that they would be of physiological significance in the etiology of EH unless they were to accumulate and act synergistically within vascular wall smooth muscle tissues.

A study into the nature and organ source of digoxin-like immunoreactive substance(s) in the perinatal period.

Digoxin-like immunoreactive substance(s) (DLIS) was isolated from sera and autopsy-derived tissue obtained from premature and full-term neonates. The highest tissue level of DLIS was in the small bowel followed by the adrenal, gallbladder and liver. Of the fluids examined, meconium had the highest level of DLIS. Preparative high performance liquid chromatography fractionation of cord blood generated at least six different fractions which not only contained DLIS material but also inhibited canine kidney Na+/K+-ATPase activity. Recovery/inhibition studies indicated that 72% of the canine kidney Na+/K+-ATPase inhibition within one fraction could be accounted for on the basis of progesterone content of the fraction.

L-carnitine treatment in the hyperlipidemic rabbit.

A study was designed to examine the hypolipidemic effect of L-carnitine treatment (4 weeks, 170 mg/kg/d) in rabbits fed a high fat diet (5% corn oil/0.5% cholesterol, w/w). Eight weeks of exposure to the high fat diet significantly increased plasma total cholesterol and triglycerides. VLDL associated triglycerides, cholesterol, apo-B, and total protein were also significantly increased with the diet. There was no change in HDL-cholesterol levels. Plasma concentration of carnitine (free, acyl, and total) all increased significantly with the high fat diet. The content of free, short-chain, and total carnitine were decreased in the liver whereas the content of long-chain acylcarnitines was increased. The diet generated a significant steatosis within the livers of these animals. Four weeks of treatment of L-carnitine reduced the extent of the liver steatosis and significantly decreased plasma total cholesterol, triglycerides, VLDL associated triglycerides, cholesterol, and total protein. HDL-cholesterol levels were unaffected by the treatment. All plasma fractions of carnitine (free, acetyl, acyl, and total) were significantly increased above those levels seen after 8 weeks of the high fat diet alone. The content of liver carnitine and its esters was normalized following treatment. The high fat diet decreased liver HMG-CoA reductase activity and increased the activities of 7-alpha-hydroxylase and acylcholesterol acyltransferase (ACAT). L-Carnitine treatment blunted the magnitude of the diet induced increase in 7 alpha-hydroxylase activity, yet overall the activity still remained elevated relative to controls. ACAT activity increased (1.5 times) with the high fat diet and increased further (4.5 times) following carnitine treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Minimizing analytical interferences from digoxin-like immunoreactive substances (DLIS) in cases of digoxin toxicity.

Recently, the value of therapeutic drug monitoring for digoxin has been called into question by the finding of endogenous digoxin-like immunoreactive substances (DLIS) in the serum of individuals, especially premature and full-term neonates, not being treated with digoxin. In some cases, values have been as high as 10 micrograms/L. Levels as high as 20 micrograms/L and 80 micrograms/g can be found in bile and meconium. Because of the magnitude of this interference, it is essential that methods be developed for measuring digoxin in the presence of DLIS. This is particularly important when such analyses are required in forensic science cases of suspected digoxin toxicity. This report outlines the high performance liquid chromatographic (HPLC) and radioimmunoassay (RIA) methods that we used in assessing the relative contribution made by digoxin, its metabolites, and DLIS to serum and tissue digoxin concentrations obtained by RIA in a forensic pediatric case of suspected digoxin toxicity.