Routine differential diagnosis of proteinurias by capillary electrophoresis.

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.

Partially reversible renal tubular damage in patients with obstructive jaundice.

Decreased serum uric acid levels resulting from renal urate wasting have been occasionally encountered in jaundiced patients. However, in these cases, there are no data concerning the underlying renal tubular defects. In the present study, we investigated the renal tubular function in 35 patients with obstructive jaundice of various severity and causes (11 with lithiasis, 17 with carcinoma, and 7 with intrahepatic cholestasis). A detailed study of the renal tubular function was performed. Beyond the conventional methods, (1)H-NMR spectroscopy of urine was used to evaluate noninvasively renal damage by the characteristic perturbation in the excretion pattern of low-molecular weight endogenous metabolites. On admission, patients with obstructive jaundice had significantly lower serum uric acid and phosphate levels and higher bile acid concentrations compared with 40 age- and sex-matched controls. Serum uric acid levels presented a negative correlation with the total and direct bilirubin as well as the fractional excretion of uric acid. Furthermore, a great number of the patients studied developed one or more proximal tubular dysfunction manifestations beyond uricosuria, such as renal glucosuria, phosphaturia, and increased excretion of alpha(1)-microglobulin. (1)H-NMR spectroscopy of the urine showed decreased levels of citrate and hippurate and increased levels of 3-hydroxybutyrate and acetate. In 12 patients partial or complete remission of jaundice was followed by an improvement of the proximal renal tubular damage. In conclusion, obstructive jaundice can cause a partially reversible generalized proximal tubular dysfunction.

Evaluation of five methods for determining low-density lipoprotein cholesterol (LDL-C) in hemodialysis patients(1).

OBJECTIVES: Current recommendations for the management of dyslipidemia are largely based on the concentration of LDL-C. Most clinical laboratories estimate the concentration of LDL-C by the recommended routine method, the equation of Friedewald, in specimens from fasting subjects and with TG concentrations < 4.52 mmol/L. Because of the limitations of the Friedewald calculation, direct methods for an accurate quantification of LDL-C are needed. DESIGN AND METHODS: In the present study we evaluated the accuracy of the following 5 different procedures for LDL-C in 98 patients on hemodialysis: the Friedewald equation, where LDL-C is calculated from HDL-C, measured either by the precipitation procedure with dextran sulfate-Mg(2+) (Method 1), or by a direct HDL-C assay (Method 2), the Direct LDL assay (Method 3), the homogeneous N-geneous LDL assay (Method 4) and the calculated LDL-C values deriving from the ApoB based equation: 0.41TC - 0.32TG + 1.70ApoB - 0.27, (Clin Chem 1997;43:808-815) (Method 5). RESULTS: All five LDL-C methods were found to be in good agreement with ultracentrifugation/dextran sulfate-Mg(2+) precipitation with the coefficients of correlation of the assays to ranging between 0.93-0.95. However, significant differences in the mean values and biases vs. the reference method were observed. The Friedewald equation and the Direct assay were less affected by high LDL-C levels, and they presented higher sensitivity and higher negative predictive value. The N-geneous assay and the ApoB derived calculation were less affected by high triglyceride levels, and they presented higher specificity and higher positive predictive value. At the diagnostic LDL-C level of 3.37 mmol/L, both Friedewald calculations correctly classified 82/92 patients; Direct assay 86/98; N-geneous assay 88/98; and ApoB derived calculation 88/98. At the diagnostic LDL-C level of 2.98 mmol/L, Friedewald calculations (Method 1 and Method 2) correctly classified 82/92 and 81/92 patients, respectively; Direct assay (LDL-3) 87/98; N-geneous assay (LDL-4) 91/98; and ApoB derived calculation (LDL-5) 91/98. CONCLUSIONS: Among hemodialysis patients, who commonly present "average" LDL-C concentrations and high TG levels, the N-geneous assay and the apoB derived calculation seem to yield more acceptable results for the estimation of LDL-C.

Estimation of LDL cholesterol based on the Friedewald formula and on apo B levels.

OBJECTIVES: The plasma apolipoprotein B (apo B) concentrations have been considered to be a more accurate representation of atherogenic particles and it has been proposed that the formula LDL-C (mmol/L) = 0.41TC - 0.32TG + 1.70apo B - 0.27 is reliable for the estimation of LDL-C (Clin Chem 1997; 43: 808-15). We undertook the present study to investigate the reliability of this formula in a large number of hyperlipidemic patients. DESIGN AND METHODS: 1) The Friedewald formula (LDL-F) and the apo B-based formula (LDL-B) were compared with the beta-quantification reference procedure in 130 individuals with a wide range of total cholesterol (TC) and triglyceride (TG) levels, and 2) the LDL-C levels obtained by the Friedewald formula were compared with those calculated by the apo B-based formula in 1010 individuals attending our outpatient lipid clinic. RESULTS: The LDL-F and the LDL-B formulae for LDL-C estimation were found to be in good agreement with the beta-quantification (r = 0.96 and 0.97, respectively). The bias of each method plotted as a function of TG (up to 4.52 mmol/L) was found positive for the LDL-F, whereas the LDL-B was independent of the concentrations of TG. When a large number of individuals were examined, a good correlation between the two equations was found (n = 1010, r = 0.98). The difference between the two methods was not correlated with serum TG levels. However, it was correlated to serum TC, and apo B levels. CONCLUSIONS: The LDL-B formula is a more reliable and accurate method than the LDL-F formula, especially at TG levels >2.26 mmol/L, although it underestimates LDL-C concentrations. Furthermore, this equation can be used in hypertriglyceridemic patients (TG >4.52 mmol/L) in whom the Friedewald equation is inaccurate.

Nuclear distribution of prothymosin alpha and parathymosin: evidence that prothymosin alpha is associated with RNA synthesis processing and parathymosin with early DNA replication.

Prothymosin alpha and parathymosin are two ubiquitous small acidic nuclear proteins that are thought to be involved in cell cycle progression, proliferation, and cell differentiation. In an effort to investigate the molecular function of the two proteins, we studied their spatial distribution by indirect immunofluorescence labeling and confocal scanning laser microscopy in relation to nuclear components involved in transcription, translation, and splicing. Results indicate that both proteins exhibit a punctuated nuclear distribution and are excluded by nucleoli. The distribution of prothymosin alpha in the nucleus is related to that of transcription sites, whereas the distribution of parathymosin correlates with early replication sites. This implies that prothymosin alpha and parathymosin are involved in transcription and replication, respectively. In addition to the punctate distribution, prothymosin alpha also is found concentrated in 1-6 nuclear domains per cell. These domains are found in more than 80% of randomly growing T24 human bladder carcinoma cells. They have a diameter of 0.2-2.5 microm, their size being inversely related to the number of domains per cell. The domains disappear during mitosis and the protein is excluded from the metaphase chromosomes. Double-labeling experiments associate these prothymosin alpha domains with PML and CstF64 containing nuclear bodies, but not with hnRNP-I containing domains or coiled bodies.

The effect of thyrotoxicosis on adrenocortical reserve.

OBJECTIVE: Variations in thyroid function are known to be associated with changes in adrenocortical activity. Previous studies in animals have suggested that long-standing hyperthyroidism may be associated with diminished adrenal functional reserve despite a continuing hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis. In humans, there has been no direct assessment of adrenal secretory reserve in clinical thyrotoxicosis. This study aimed to assess adrenocortical reserve in response to low-dose ACTH, following dexamethasone suppression, in patients with severe thyrotoxicosis. DESIGN AND METHODS: Ten patients (four men and six women, 30-45 years) with severe long-standing thyrotoxicosis due to Graves' disease (n=6) or toxic nodular goitre (n=4) were studied at diagnosis and again when in a stable euthyroid state following drug therapy for 8-12 months. All patients underwent ACTH stimulation tests at 0800h with ACTH(1-24) (Cortrosyn; 0.1microg/kg body weight, i.v.) following overnight suppression of the HPA axis with dexamethasone (1mg per os at 2300h). Serum cortisol was assayed at -15, 0, 15, 30, 60 and 90min after the administration of ACTH. RESULTS: The mean (+/-s.d.) peak and delta cortisol responses to ACTH (634.5+/-164nmol/l and 618+/- 196nmol/l respectively), as well as the net area under the response curve (36769+/-12188nmol/lx min) in the hyperthyroid patients were significantly lower compared with the values when the same patients were euthyroid (911+/-157nmol/l, 905+/-160nmol/l and 57652+/-10128nmol/lxmin respectively; P<0.005). Subnormal peak cortisol responses (<500nmol/l) were observed in two severely toxic patients. The findings were independent of the cause of thyrotoxicosis. CONCLUSION: In patients with severe thyrotoxicosis, cortisol secretion in response to low-dose ACTH stimulation, following dexamethasone suppression, is lower in the hyperthyroid than in the euthyroid state. It appears that thyrotoxicosis is associated with subtle impairment of adrenocortical reserve.

Homogeneous HDL-cholesterol assay versus ultracentrifugation/dextran sulfate-Mg2+ precipitation and dextran sulfate-Mg2+ precipitation in healthy population and in hemodialysis patients.

OBJECTIVES: To evaluate the analytical performance of a new homogeneous HDL-cholesterol assay (Olympus Diagnostica). To investigate possibly discrepant results in chronic hemodialysis patients who commonly exhibit quantitative and qualitative lipoprotein abnormalities, responsible for atherogenic complications in these patients. DESIGN AND METHODS: Serum samples were collected from 50 healthy subjects and 65 chronic hemodialysis patients. HDL-C levels measured by the homogeneous assay were compared with the routine dextran sulfate-Mg2+ precipitation method and the ultracentrifugation/dextran sulfate-Mg2+ precipitation as reference method. RESULTS: The homogeneous assay was linear up to at least 220 mg/dL. The analytical precision was estimated with three different sets of commercial controls and one set of human pooled serum control. The within-day CV ranged between 1.7% and 3.8% and the between-day CV ranged between 1.0% and 2.3%. HDL-C values in both populations correlated highly with the dextran sulfate-Mg2+ precipitation method and the ultracentrifugation/dextran sulfate-Mg2+ precipitation method (r > or = 0.96, bias between -0.9 and 2.3 mg/dL). Lipemia up to triglyceride concentration of 600 mg/dL did not alter the HDL-C value. CONCLUSIONS: The homogeneous assay for HDL-C (Olympus) uses much less sample, is accurate and convenient to handle, and allows full automation. The test should considerably facilitate the screening of individuals at an increased risk of cardiovascular disease, including hemodialysis patients.

Purification of a candidate gonadotrophin surge attenuating factor from human follicular fluid.

Gonadotrophin surge attenuating factor (GnSAF) is a new non-steroidal ovarian substance, different from inhibin, which attenuates the pre-ovulatory luteinizing hormone (LH) surge in superovulated women. Human follicular fluid (FF) was used as a source for the isolation of GnSAF, the activity of which was monitored in an in-vitro pituitary bioassay. Primary rat pituitary cells were incubated with test substances for 48 h and subsequently washed and incubated with 0.1 micromol/l gonadotrophin releasing hormone (GnRH) plus test substances for 4 h. GnSAF activity was expressed as the reduction of GnRH-induced LH secretion in the 4 h incubation. GnSAF was purified from 250 ml of FF which was heat-treated at 80 degrees C for 5 min. Heparin-sepharose chromatography, Con-A sepharose chromatography, reversed-phase high-performance liquid chromatography (HPLC) and preparative native gel electrophoresis were used for GnSAF fractionation. Using these purification steps, we have obtained an apparently homogeneous preparation that stains as a single band on sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis. GnSAF has an apparent molecular weight of 12.5 kDa and was identified by amino acid sequence (mass spectrometry) to be the C-terminal fragment of human serum albumin.

Thyroid autoimmunity is associated with higher urinary iodine concentrations in an iodine-deficient area of Northwestern Greece.

Northwestern Greece was identified in the 1960s for its high prevalence of endemic goiter and iodine deficiency. Although iodized salt has been commercially available since then, a recent epidemiological survey of 3916 schoolchildren found that low-grade goiter is still prevalent in endemic proportions (21%). The aim of this study was to further assess the cause of goiter and the severity of iodine deficiency in children from this endemic area of Greece. Of the 800 children with clinically detectable goiter, 97 children (60 girls and 37 boys, 8-15 years) were recruited for determination of urinary iodine excretion, as well as assessment of thyroid volume and function and detection of antithyroid antibodies. The median urinary iodine concentration was 8.4 microg/dL, indicative of a mild iodine deficiency. Thyroid function was normal in all but 11 children who had subclinical hypothyroidism. Sixteen children (16.5%), including all those with subclinical hypothyroidism, were positive for antithyroid antibodies. Their median urinary iodine concentration (20.6 microg/dL) was higher compared to children who were negative for antibodies (7.4 microg/dL; p<0.001). The mean thyroid volume by ultrasonography (12.2+/-4.1 mL) was above the upper limit of normal for this age group. Thyroid volume was inversely related to the urinary iodine content in the children with negative antithyroid antibodies. Iodine deficiency is still prevalent in northwestern Greece although of mild severity and constitutes the primary cause of goiter among schoolchildren. However, it appears that autoimmune thyroiditis is emerging as a frequent cause of goiter in those children with sufficient iodine intake.

Glucose oxidase-produced H2O2 induces Ca2+-dependent DNA damage in human peripheral blood lymphocytes.

DNA of lymphocytes from human peripheral blood was analyzed by using the single cell gel electrophoresis technique (comet assay). The cells were used either as received from the donors or after treatment with various concentrations of the H2O2-generating enzyme glucose oxidase, in order to achieve a continuous flow of H2O2. The formation of single strand breaks (SSB) was dose-related but the time course of the induction of SSB by relatively low concentrations of glucose oxidase was of a biphasic mode with a fast increase 2 to 5 min after the addition of glucose oxidase followed by a gradual decrease toward the original base level during the next 35 to 60 min. This response of the cells appears to be based on the activation of already existing defense system(s) because it was shown that H2O2 is continuously released during the reaction time and the inhibition of protein synthesis does not affect the observed pattern. Supplementation of the growth medium with various antioxidants resulted in substantial protection only when the agents were taken up by the cells. The presence of the intracellular calcium chelator BAPTA protected the cells from H2O2-induced DNA damage in a dose-dependent manner. Only at the higher rate of H2O2-generation considerable DNA damage was observed in the presence of BAPTA. These results suggest that H2O2, at low concentrations induces DNA damage through intracellular Ca2+ -mediated processes, which lead to DNA strand breaks possibly by endonuclease activation.

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 L-carnitine supplementation on lipid parameters in hemodialysis patients.

It has been reported that cumulative carnitine losses through dialysis membranes may worsen hyperlipidemia during long-term hemodialysis. However, carnitine supplementation has not shown a consistent beneficial response. We undertook the present study to determine if there is any hypolipidemic effect of L-carnitine on Greek dialysis patients in concert with the dialysate buffer composition (acetate or bicarbonate). A total of 28 patients (16 male, 12 female), mean age 43 years (range 21-61), with end-stage renal disease on maintenance hemodialysis for a mean period of 25 months (range 7-84) were studied. The dialysis schedule was 4 h, 3 times/week using cuprophane hollow-fiber dialyzers and acetate (n = 14) or bicarbonate (n = 14) dialysate. In all patients L-carnitine (5 mg/kg body weight) was infused intravenously 3 times/week at the end of each hemodialysis session. Blood samples for carnitine and lipid determinations were obtained before treatment, and 3 and 6 months following treatment. Even though L-carnitine did not modify most of the serum lipid levels, a significant decrease in serum triglycerides was evident in the whole group of patients (from 225 +/- 76 to 201 +/- 75 mg/dl, p = 0.03). Furthermore, L-carnitine could decrease serum triglycerides only in hypertriglyceridemic patients (from 260 +/- 64 to 226 +/- 82 mg/dl, p < 0.05). L-Carnitine resulted in a reduction of serum triglycerides in both patients on bicarbonate and on acetate dialysis, while there were no significant differences in the changes of lipid parameters after L-carnitine between the two groups of hemodialysis patients. We conclude that relatively low doses of L-carnitine supplementation could contribute to the management of some hypertriglyceridemic hemodialysis patients.

Prothymosin alpha modulates the interaction of histone H1 with chromatin.

Prothymosin alpha (ProTalpha) is an abundant acidic nuclear protein that may be involved in cell proliferation. In our search for its cellular partners, we have recently found that ProTalpha binds to linker histone H1. We now provide further evidence for the physiological relevance of this interaction by immunoisolation of a histone H1-ProTalpha complex from NIH 3T3 cell extracts. A detailed analysis of the interaction between the two proteins suggests contacts between the acidic region of ProTalpha and histone H1. In the context of a physiological chromatin reconstitution reaction, the presence of ProTalpha does not affect incorporation of an amount of histone H1 sufficient to increase the nucleosome repeat length by 20 bp, but prevents association of all further H1. Consistent with this finding, a fraction of histone H1 is released when H1-containing chromatin is challenged with ProTalpha. These results imply at least two different interaction modes of H1 with chromatin, which can be distinguished by their sensitivity to ProTalpha. The properties of ProTalpha suggest a role in fine tuning the stoichiometry and/or mode of interaction of H1 with chromatin.

Paraquat-induced renal injury studied by 1H nuclear magnetic resonance spectroscopy of urine.

The herbicide paraquat (1,1'-dimethyl-4,4'-bipyridylium dichloride; PQ), is a poison known to cause delayed mortality due to lung and kidney injuries. High-resolution proton nuclear magnetic resonance (1H NMR) spectroscopy has been extensively applied in evaluating nephrotoxicity by the characteristic perturbations in the excretion pattern of low molecular weight endogenous metabolites. The application of the method allows the rapid localization of the renal injury noninvasively. In this study, we report 1H NMR and conventional clinical chemistry urinalysis in two patients suffering from paraquat intoxication after overdose with suicidal intent. The alterations in the urine NMR spectrum suggest necrosis of the pars recta of the proximal renal tubules. The molecule of paraquat is also clearly detected in the same spectrum. In conclusion, the rapid screening of urine by NMR spectroscopy provides information about both the identity of the poison and the abnormal pattern of endogenous metabolites that characterize the location of the injury in renal tubules and reveals alterations in unusual metabolites that are not commonly measured.

Lipid abnormalities in Greek patients with coronary artery disease.

Lipid abnormalities are major risk factors for premature coronary artery disease (CAD). However, the type and prevalence of dyslipidemia in these patients have not been well characterised, especially in some ethnic groups. The purpose of the present work was to determine the lipid disorders in patients of Northwestern Greece with premature CAD. The study population comprised of 132 men and 38 women who underwent elective diagnostic arteriography in our University Hospital. Subjects with > or = 1 lesion that narrowed the lumen of any of the 15 coronary artery segments by > or = 70% were considered to be CAD cases (n=108), whereas those with narrowing < 70% were excluded (n=54). Asymptomatic subjects (n=104) matched for age and sex were taken as controls. Compared with the controls, patients with premature CAD had increased serum levels of total cholesterol, LDL cholesterol, triglycerides, Apo B, and Lp(a), and decreased serum levels of HDL cholesterol and Apo A1. A lipoprotein or apolipoprotein abnormality was identified in 89 CAD patients (82.4%). The increased serum Apo B level (> 130 mg/dl) was the most common lipid abnormality observed in 72 patients. However, the most prevalent dyslipidemic phenotypes in our patients were type IIA followed by hypoalpha and hyperApoB. Compared to the control population, CAD patients had increased incidence of IIA and hypoalpha phenotypes. On the contrary, a normal lipoprotein phenotype was more common in the control population compared to CAD patients (56.7% vs. 17.6%, P<0.001). We conclude that the majority of Greek patients with premature CAD exhibit lipid and lipoprotein abnormalities, which to a large extent can be defined by determining the traditional lipid parameters (total cholesterol, LDL cholesterol, HDL cholesterol and triglycerides). However, in some cases the value of the quantification of other lipid parameters such as apolipoproteins and Lp(a) should be taken into account.

Increased serum levels of interleukin-1beta in the systemic circulation of patients with essential hypertension: additional risk factor for atherogenesis in hypertensive patients?

The dysfunction of the immune system has been implicated in the cause of essential hypertension (EH). On the other hand, interleukin- 1beta (IL-1beta) has strongly been involved in the pathogenesis of atheromatosis, whereas our preliminary experiments in serum samples from hypertensive patients before any drug therapy have shown the presence of high concentrations of IL-1beta and the absence of interleukin-2 (IL-2). The aim of this study was first to confirm our preliminary findings and second to investigate the possible interrelation(s) among the parameters studied, particularly between the immunologic markers and the blood pressure or the lipid parameters, because so far there are no data regarding the possible participation of IL-1beta in the cascade phenomena presented during the process of EH such as atherogenesis. Serum samples from 28 consecutive unselected patients with EH before any drug administration or after discontinuation of the antihypertensive therapy for at least 4 weeks, 31 normotensive patients with familial hypercholesterolemia (FH, disease control group), and 35 healthy individuals In a control group matched for age and sex were investigated for the presence of IL-1beta (commercial enzyme immunoassay), soluble IL-2 receptors (slL-2Rs, sandwich enzyme-linked immunosorbent assay set up in our laboratory), and some of the acute phase proteins by nephelometry. In addition, total cholesterol, triglycerides, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, apolipoproteins A1 and B, and lipoprotein (a) were determined by standard methods. The data were analyzed by unpaired t test, Mann Whitney-U, chi-squared analysis after Yate's correction, analysis of variance, or Kruskal-Wallis where applicable. Correlation coefficient was calculated by simple regression analysis (r) or nonparametric Spearman correlation coefficient (rs). We found that (1) none of the patients had increased concentrations of sIL-2Rs, and (2) the IL-1beta levels significantly differed in the three groups (p = 0.0001). In more detail, the concentrations of IL-1beta were significantly higher in patients with EH compared with those in patients with FH (p < 0.0005) and the healthy control group (p = 0.0001). By contrast, the IL-1beta concentrations did not differ between patients with FH and the healthy control group. (3) Sixteen (57.1%) patients with EH and only 6 (19.4%) patients with FH (p < 0.01) had increased levels of IL-1beta, and (4) the IL-1beta was not correlated with the acute phase reactants or the lipid parameters in the groups studied. However, the group of patients with EH and increased IL-1beta levels had significantly higher mean concentrations of triglycerides (p < 0.05) and significantly lower mean concentrations of high-density lipoprotein cholesterol (p < 0.05) than those who had IL-1beta levels lower than the cutoff point. (5) The IL-1beta concentrations were positively though slightly correlated with the mean blood pressure only in the group of patients with EH (r = 0.38, p < 0.05). This study demonstrated the presence of high concentrations of IL-1beta and the absence of indicators of cellular immune activation in the systemic circulation of patients with EH, suggesting that this cytokine may be involved in the pathogenesis of EH. In addition, this study showed that the high levels of IL-1beta were associated with lipid indicators of atheromatosis only in the group of patients with EH. More studies are required in an attempt to address whether IL-1beta could have a pathogenetic importance in EH. Taking into account these findings, however, it can be suggested that the presence of high IL-1beta levels may be an additional and perhaps independent risk factor for atheromatosis in patients with EH.

Selective interaction between parathymosin and histone H1.

We have studied the molecular associations of parathymosin, an acidic polypeptide with a wide tissue distribution, by means of three approaches; ligand blotting; native electrophoresis; and immunoprecipitation. We report here that parathymosin binds specifically to the linker histone H1. This binding is enhanced by Zn2+ and is dependent on the concentration of parathymosin. Poly(glutamic acid) is able to compete fully with parathymosin for binding to histone H1, suggesting that this interaction is mediated by the acidic domain of the protein. Moreover, we demonstrate that parathymosin interacts with the globular domain of histone H1 under native conditions. Based on these data, we postulate that parathymosin may belong to a group of nuclear acidic proteins that affect histone H1 function.

The beneficial effect of tamoxifen on serum lipoprotein-A levels: an additional anti-atherogenic property.

It has recently been suggested that increased lipoprotein (a) levels are an independent risk factor for coronary heart disease. It has also been reported that tamoxifen can induce changes in blood lipid values. In this prospective study we investigated the long term effects tamoxifen on the lipid profile, focusing on lipoprotein (a) levels. Thirty-eight postmenopausal women with breast cancer treated with tamoxifen at a dose of 20 mg daily were studied. The mean age was 61 years. Serum lipoprotein (a) levels and lipid parameters (total cholesterol, high and low density lipoprotein cholesterol, triglycerides, apolipoprotein A-I and B) were measured after an overnight fast on the 1, 3, 6 and 9 month of tamoxifen treatment. A progressive fall in median lipoprotein(a) levels from 8 mg/dl to 3 mg/dl (p < 0.001) was observed. In addition, a significant decrease in the total and low density lipoprotein cholesterol and in apolipoprotein B, as well as an increase in apolipoprotein A-l were also noticed. It is concluded that our data on serum lipoprotein(a) levels agree with the beneficial anti-atherogenic effects of tamoxifen administration on lipoproteins.

Human placental ATP diphosphohydrolase is a highly N-glycosylated plasma membrane enzyme.

Human placental ATP diphosphohydrolase (ATP-DPH), has been previously characterized as an azide-sensitive, Ca(2+)- or Mg(2+)-dependent triphospho- and diphosphonucleosidase which migrates as an 82 kDa protein band on SDS-PAGE (Christoforidis, S. et al. (1995) Eur. J. Biochem. 234, 66-74). In this paper we have studied the subcellular localization of placental ATP-DPH by differential centrifugation and flotation experiments. Using specific enzymatic markers it was found that ATP-DPH is localized on plasma membrane. ATP-DPH was found to be a highly N-glycosylated protein which is a common post-translational modification of plasma membrane proteins. Extensive incubation of the native pure enzyme with N-glycosidase F resulted in the elimination of the 82 kDa form and the concurrent formation of a deglycosylated product of 57.5 kDa and four other intermediate products, indicating the presence of at least five N-glycosylation sites within the ATP-DPH molecule. The partially deglycosylated sample retained its activity in solution and in native gel electrophoresis and activity staining.