Neonatal screening for hyperlipoproteinemia. Methods for direct estimation of cord serum VLDL + LDL.

The early detection of hyperlipoproteinemia in newborn infants has so far been based upon estimation of cord blood total lipids (cholesterol and triglyceride) and lipoprotein-lipids (VLDL-, LDL- and HDL-cholesterol). To be able to make a direct estimation of cord serum beta-lipoproteins (VLDL + LDL) two quite different methods were modified, one immunological and the other turbidimetric. Good correlations were found to VLDL- + LDL-cholesterol isolated in the ultracentrifuge (r = 0.848 and 0.831, respectively). If neonatal screening for hyperlipoproteinemia is considered, we recommend the very easy and inexpensive turbidimetric method. Furthermore, using cord serum, two conventional precipitation methods with heparin-CaCl2 and heparin-MnCl2 were compared by ultracentrifugation and high correlations were found (r = 0.923 and 0.899, respectively). A clamping study showed that following early clamping of the cord, the concentration of cord serum lipids and lipoproteins did not change markedly within the first five minutes. Storing experiments showed that serum should be separated within the first 12 h to avoid unpredictable changes in the concentration of cord serum lipids and lipoproteins.     

Effect of the composition of very low and low density lipoproteins on the rate of cholesterylester transfer from high density lipoproteins in man, studied in vitro

The process of cholesterylester (CE) transfer is supposed to be a regulatory factor in the distribution of CE between lipoproteins. In addition to the activity of CE transfer protein, this process may be affected by acceptor lipoprotein characteristics. In this study the effect of the composition of different very low density lipoproteins (VLDL) and low density lipoproteins (LDL) on the ability to accept CE from HDL in vitro was investigated. [3H]-CE high density lipoprotein (HDL) (100 nmol CE) from one batch was incubated with VLDL (75 nmol CE), isolated from fifteen subjects for 4 h and separately with LDL (250 nmol CE), isolated from thirteen subjects for 16 h, both in the presence of lipoprotein-free plasma providing a source of cholesterylester transfer protein. The CE transfer rate of VLDL (range 1.34-2.84% [3H]-CE transferred h-1) was correlated to the triacylglycerol (TG):CE molar ratio (r: 0.63, P less than 0.05), to the phospholipid (PL):CE molar ratio (r: 0.75, P less than 0.01), to the protein (Pr):CE ratio (expressed in g nmol-1) (r: 0.72, P less than 0.01) and to the free cholesterol (FC):CE molar ratio (r: 0.69, P less than 0.01), but not to the FC:PL molar ratio (r: -0.08, NS). The CE transfer rate to LDL (range 1.18-3.59 nmol CE h-1) was correlated to the Pr:CE ratio (r: 0.72, P less than 0.01) and inversely to the FRC:PL molar ratio (r: -0.88, P less than 0.001), but not to the TG:CE molar ratio (r: 0.40, NS), nor to the FC:CE molar ratio (r: -0.37, NS).(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparison of phosphotungstate and dextran sulfate-Mg2+ precipitation procedures for determination of high density lipoprotein cholesterol

Because of the controversy over the best method for assaying high density lipoprotein (HDL) cholesterol in the clinical laboratory, a commonly used phosphotungstate method for precipitating low density and very low density lipoproteins (LDL and VLDL) was compared with a recently recommended dextran sulfate precipitation method. The accuracy and precision of HDL cholesterol determinations were similar for both methods. Either of these procedures would appear to be equally satisfactory for the assay of HDL cholesterol in the clinical laboratory.     

Effects of plant stanol and sterol esters on serum phytosterols in a family with familial hypercholesterolemia including a homozygous subject

We studied the concentrations and ratios to cholesterol of noncholesterol sterols reflecting absorption (eg, campesterol) or synthesis (eg, lathosterol) of cholesterol off and on plant sterol and stanol ester spreads in serum and in different lipoproteins of a family with familial hypercholesterolemia, including heterozygous parents receiving no treatment and their homozygous offspring undergoing long-term treatment with statins and apheresis. Serum cholesterol levels were similar in the homozygous and heterozygous individuals, but the concentrations of sterols reflecting cholesterol absorption were as much as 10 times greater in the homozygous child than in the heterozygous parents, whereas the respective markers of cholesterol synthesis only tended to be higher. About 70% of squalene in the homozygous individual (60% in the heterozygous family members) and 85% to 90% of noncholesterol sterols (60%-80% in the heterozygous subjects) were transported by low-density lipoprotein. The ratios of absorption sterols to cholesterol were higher in high-density lipoprotein (HDL) than in very low-density lipoprotein (VLDL), whereas those of synthesis markers and plant stanols were highest in VLDL. The ratios of absorption sterols in serum were mostly lower than those in HDL but higher than in VLDL, whereas the ratios of synthesis sterols in serum were lower than they were in VLDL. Both spreads reduced serum total cholesterol by about 14% in the heterozygous family members and 9% in the homozygous individual. The sterol ester spread increased serum plant sterol concentrations (eg, campesterol in the homozygous family member increased from 5 to 9 mg/dL) and the ratios to cholesterol, but the stanol ester spread decreased them. Plant sterol esters seemed to similarly decrease serum cholesterol in this family with familial hypercholesterolemia, but the clinical role of increased plant sterol concentrations, almost doubled in the LDL of homozygous individuals, is not known.     

Preparation and application of Procion Yellow Starch for amylase assay

A new starch derivative for the determination of amylase activity has been synthesized by coupling Procion Yellow dye with starch. The product of this reaction is intensely yellow and is easily suspended in water, in neutral buffer solution, and also in acidic solution. Amylase from pancreatin, saliva, urine and serum readily hydrolyzes this chromogenic substrate. A method for determining amylase activity based on the use of the Procion Yellow Starch substrate is described. The procedure requires 0.1 ml of sample and an incubation time of 30 min. The soluble chromogen, which is liberated by enzymatic hydrolysis, is measured at 420 nm and the amylase activity-absorbance relationship at this wavelength is linear. The normal range for this method is 50–150 Somogyi units/dl.     

Human postheparin plasma hepatic lipase activity against triacylglycerol and phospholipid substrates

Recent evidence has suggested that the major physiological substrate of the heparin-releasable (postheparin plasma) hepatic lipase is HDL₂-phospholipid. However, all the current assay methods for this enzyme are based on the use of triacylglycerol substrate. Even though both lipolytic activities of hepatic lipase are likely to be due to a single enzyme it is possible that the use of unphysiological lipid as a substrate may give misleading results. Therefore we did parallel assays of the activity of postheparin plasma hepatic lipase using triacylglycerol, monoacylglycerol and phospholipid substrates. The correlation coefficients between the three lipolytic activities were 0.95–0.98, indicating that identical results are obtained using any of these three lipids in the assay of postheparin plasma hepatic lipase.     

Association of visceral and subcutaneous fat with glucose intolerance, insulin resistance, adipocytokines and inflammatory markers in Asian Indians (CURES-113)

Objectives:

The aim of the study was to assess the association between visceral and subcutaneous fat with glucose intolerance, adipocytokines, inflammatory markers and carotid IMT in Asian Indians.

Design and methods:

Subjects with NGT (n = 85), IGT (n = 49) and T2DM (n = 93) were randomly selected from CURES. Total abdominal, visceral and subcutaneous fat were measured using Helical CT scan. Adiponectin, hs-CRP, TNF-alpha, oxidized LDL, visfatin and leptin and IMT and insulin resistance were assessed.

Results:

Total abdominal fat (p = 0.041) and the visceral fat (p = 0.039) but not subcutaneous fat progressively increased from NGT, IGT and T2DM subjects. With increasing quartiles of visceral fat, there was a significant increase in insulin resistance (p = 0.040); significant decrease in adiponectin (p = 0.043) and increase in TNF-alpha (p = 0.028), hs-CRP (p = 0.043), OX-LDL (p = 0.034) and visfatin (p = 0.040), and carotid IMT (p = 0.047) was observed.

Conclusion:

Visceral fat levels increased with increasing glucose intolerance and are associated with decreased levels of adiponectin and increased levels of hs-CRP, TNF-alpha, oxidized LDL, visfatin, HOMA-IR and IMT.