Konstantin Seferiadis

Evaluation of Methods for the Measurement of Low-Density Lipoprotein Cholesterol

A high concentration of low-density lipoprotein cholesterol (LDL-C) in plasma is one of the strongest risk factors for atherosclerotic cardiovascular disease and mortality. The most common approach to determining LDL-C in the clinical laboratory is the Friedewald calculation. There is an increased interest to improve the accuracy of LDL-C estimated by this equation. The expert panel convened by National Cholesterol Education Program has recommended the development of accurate direct methods to measure LDL-C. Several homogeneous and fully automated methods have been introduced in recent years that show improved precision and accuracy over earlier methods, especially the Friedewald calculation. Each of the atherogenic particles in plasma—very-low, intermediate-, and low-density lipoprotein—as well as lipoprotein (a), contain one molecule of apolipoprotein B (apoB) and thus, plasma total concentration of apoB reflects the number of atherogenic particles. Several studies suggested that the measurement of apoB could improve the prediction of risk of coronary artery disease. Thus, in addition to the newly developed direct assays, alternative calculation procedures have been proposed that also take into consideration total serum apoB concentration for the estimation of LDL-C and the presence of small, dense LDL particles. The new generation of homogenous methods for the measurement of LDL-C and the use of serum apoB concentration for the estimation of LDL-C can contribute to the accurate LDL-C determination.

Isolation and partial sequence of goat spleen prothymosin α

Goat prothymosin α, a highly acidic polypeptide of pl 3.5, 109 amino acid residues, has been isolated from lymphoid and non-lymphoid tissues of young female goats. Unlike rat, murine and porcine prothymosins α, goat prothymosin α appears at a higher concentration in the spleen compared with the thymus. The sequence of segments of the polypeptide involving known mutations has been determined, by automatic sequencing of its tryptic peptide fragments. The acidic amino acid-rich segment in the middle of the molecule, including residues 49–83, has not been sequenced. Goat prothymosin α closely resembles bovine prothymosin α, with only one substitution, proline for alanine at position 85. It also resembles human prothymosin α, with only three substitutions. It differs more significantly from rat and murine prothymosins α, by two deletions and three substitutions. The results show the highly conserved nature of the molecule, with substitutions at given positions only.

Enhancement of Human T Lymphocyte Function by Prothymosin α: Incremed Production of Interleukin-2 and Expression of Interleukin-2 Receptors in Normal Human Peripheral Blood t Lymphocytes

The in vitro incubation of phytohemagglutinin (PHA)- or alloantigen-stimulated peripheral blood T cells with prothymosin alpha (ProT alpha) resulted in a marked and reproducible increase in the production of interleukin-2 (IL-2). Incubation of T cells with ProT alpha, in the absence of PHA or alloantigen, failed to induce any production of IL-2. ProT alpha by itself did not exert any IL-2 activity. Finally, ProT alpha was shown to increase the expression of IL-2 receptors on phytohemagglutinin- or alloantigen-activated T cells. These data provide the basis for understanding the in vitro immunoenhancing effects of ProT alpha in cellular immune systems.

Isolation and partial characterization of prothymosin α from porcine tissues

Prothymosin α, an immunoactive polypeptide of 12 kDa, has been isolated from porcine thymus, spleen, lung and kidney. It lacks aromatic and sulfur‐containing amino acids and has a high content of glutamic and aspartic acids. Tryptic digestion of porcine thymus prothymosin α yielded peptides which on separation, amino acid analysis and alignment with the known sequence of prothymosin α from rat and man showed that the amino terminal portion of the molecule is conserved and the few differences present are confined to the car☐y terminal.

Routine Differential Diagnosis of Proteinurias by Capillary Electrophoresis

Abstract Capillary electrophoresis is a relatively new analytical technique that begins to have an impact on both routine and research in clinical laboratories. Recently, a fully automated system has become commercially available (Paragon CZE 2000, Beckman, USA) for the analysis of human serum proteins. Urine protein analysis, on the other hand, is currently accomplished by electrophoresis of concentrated urine specimens. The method is used to distinguish the glomerular from the tubular proteinuria and for the identification of Bence-Jones proteins. The procedure is labor-intensive and technically demanding. We developed a technique for the serum capillary electrophoresis instrument that can also be applied routinely to the differential diagnosis of proteinurias. Overriding the programmed dilution step of the instrument, we were able to distinguish different types of proteinurias without concentration of specimens with a total protein content of 150–200 mg/l as determined by sulfosalicylic acid. The different electrophoretic patterns obtained by the capillary electrophoresis system for various specimens correlated well with established techniques (Hydragel Proteinurie Kit, Sebia, France). The method is applicable for routine analysis of urinary proteins. It is reliable, less expensive and faster than the conventional methods (electrophoretic or immunonephelometric) used today for the differentiation of proteinurias, and it can be used as a quick screening test.

Changes in gonadotrophin response to gonadotrophin releasing hormone in normal women following bilateral ovariectomy

Pituitary responsiveness to GnRH varies throughout the normal menstrual cycle. We have investigated whether there are differences in the ovarian mechanisms which regulate gonadotrophin secretion between the follicular and the luteal phase of the cycle.

Development and application of competitive ELISA assays for rat LH and FSH.

Rat LH (rLH) and FSH (rFSH) were measured by sensitive and specific competition ELISAs. The rat LH ELISA used rLH-I-9 coated plates, an antiserum against rLH and an antibody against rabbit IgG labeled with peroxidase. Using rLH-RP-3 as a standard, rat LH was determined by binding of the anti-LH antibody to rLH-I-9 coated plates. The sensitivity of the assay was 0.8 ng/mL. Similarly, the rat FSH-ELISA used rFSH-I-8 coated plates, an antiserum against rFSH and an antibody against rabbit IgG labeled with peroxidase. Using rFSH-RP-3 as a standard, the FSH-ELISA was also determined by binding of the anti-FSH antibody to rFSH-I-8 coated plates. The sensitivity of this assay was 1.25 ng/mL. Both rat LH and FSH ELISA assays are highly specific and provide accurate determination of gonadotrophins in buffers, sera, cell culture media, and anterior pituitary extracts. These assays were used for monitoring the gonadotrophin surge-attenuating factor (GnSAF) and inhibin activities present in human follicular fluid (hFF). The 2 new ELISA procedures have practical advantages (safety, convenience, economy) over the RIA methods, and they perform as well as the RIA techniques at the same range of concentrations.

Effect of follicle stimulating hormone or human chorionic gonadotrophin treatment on the production of gonadotrophin surge attenuating factor (GnSAF) during the luteal phase of the human menstrual cycle

OBJECTIVE Although there is much in‐vivo evidence for the existence of a gonadotrophin surge attenuating factor (GnSAF), its source and identity remain unknown. We have studied the control of GnSAF production by FSH and hCG during the luteal phase of the cycle.

Appearance of thymosin α1 in supernatants of monocytes incubated with prothymosin α

Abstract Prothymosin α, a polypeptide of 109 to 111 amino acid residues, contains the entire thymosin α 1 sequence (residues 1–28) at its amino terminal. Human peripheral blood monocytes incubated with prothymosin α release thymosin α 1 in the culture supernatants. In addition total RNA is found to increase. The production of thymosin α 1 involves de novo protein synthesis as shown by the kinetics of this release and its inhibition by actinomycin D and cycloheximide. Thymosin α 1 release, possibly in association with HLA-DR, stimulates the proliferation of the T Cell population.

Statins and homocysteine

To the Editor: Hyperhomocysteinemia confers an independent risk factor for atherosclerosis besides the well-established risk factors, such as hypercholesterolaemia, hypertension, diabetes mellitus, smoking habit and family history [1 /3]. However, the effect of statins (especially the most potent drugs of this class) on homocysteine (Hcy) plasma levels is not yet well established. Five recently published studies examined the effect of statins (simvastatin and atorvastatin) on Hcy plasma levels [4 /8] (Table 1). Dieter Luftjohann et al. reported a significant decrease in Hcy plasma levels after high doses of simvastatin (80 mg daily) for 24 weeks, suggesting a possible contribution to the reduction in cardiovascular events seen with high doses of simvastatin. On the contrary, the other studies (Table 1) failed to detect any changes in Hcy after statin therapy. Since the number of patients participating in the above studies was limited, we examined the effect of atorvastatin (40 mg daily) on Hcy plasma levels in 61 patients with hyperlipidaemia attending our lipid clinic. As shown in Table 2, the administration of atorvastatin (40 mg daily) for 10 weeks did not affect the Hcy plasma levels (10.49/3.6 mmol/l before vs. 10.19/3.4 mmol/l after atorvastatin administration). We suggest that the antiatherogenic properties of statins, other than their cholesterol lowering effect, are not through any change in Hcy plasma levels. References