Distribution of alpha-tocopherol in human plasma lipoproteins.

Lipoproteins were removed from human plasma by ultracentrifugation at a density of 1.225. Three classes of lipoproteins were then separated by 4% Agarose-column chromatography: very low density lipoproteins (VLDL), low-density lipoproteins (LDL), and high-density lipoproteins (HDL). A sensitive high performance liquid chromatography method with a fluorescence detector was used to estimate alpha-tocopherol in plasma and in column eluates. Total plasma tocopherol was not significantly different in males (n = 6) and females (n = 6) and almost all of the vitamin was recovered in the isolated lipoproteins. Although LDL and HDL were the main carriers of alpha-tocopherol in both males and females, more tocopherol was found in LDL than in HDL in males but the opposite was true in females. The distribution of alpha-tocopherol in males was: VLDL, 8%; LDL, 59%; and HDL, 33% whereas that in females was VLDL, 2%; LDL, 42%; HDL, 56%. The distribution of protein in lipoprotein from males was: VLDL, 4%, LDL, 37%; and HDL, 59% and in females: VLDL, 2%; LDL, 25%; and HDL, 73%. The alpha-tocopherol concentration (expressed as microgram alpha-tocopherol/mg protein) in lipoproteins differed little between the sexes. The values in males were: VLDL, 7.0; LDL, 4.3; and HDL, 1.5, in females: VLDL, 3.9; LDL, 4.7; and HDL, 2.1. The data suggest that the different distribution of alpha-tocopherol in plasma lipoproteins in males and females is due to the different levels of proteins in those lipoprotein fractions. Overall, tocopherol and protein levels were highly correlated in HDL, a lower correlation was found in LDL.

Comparison of techniques improving the resolution of standard Coulter cell sizing systems.

Latex spheres with nominal diameters of 1.857, 2.02, 3.5, 5.2 and 9.69 mum were used to compare improvement in Coulter resolution gained (a) by a simple hydrodynamic focusing modification and (b) by Coulter edit and high resolution circuits. Since pulse size profiles of focused 9.69-mum spheres appeared quasimonodisperse [geometric SD (GSD) congruent to 0.016 congruent to (1 + Arithmetic SD/arithmetic mean)], unfocused profiles of these particles approximated the error function resulting from inhomogeneities in the electric and hydrodynamic sensing-zone fields. As expected, this field error function differed systematically with differences in orifice diameters (30, 50, 70 and 100 mum) and length to diameter ratios (0.75 or 1.2), the error being reduced slightly by the edit circuit and considerably by the high resolution circuit. Data on the 1.857- and 2.02-mum spheres (focused GSDs congruent to 1.06) and 3.5- and 5.2-mum spheres (focused GSDs congruent to 1.6) indicated that the field error affected closely similar particles similarly but its influence on the unfocused population means and variances differed in the GSD regions of 1.06 and 1.60.

Vitamin C and vitamin E status in the spontaneously diabetic BB rat before the onset of diabetes.

Ascorbic acid (AA), dehydroascorbic acid (DHAA), and vitamin E were measured in tissues and plasma of 30 control and 30 spontaneously diabetic BioBreeding rats (BBdp) during development and before the onset of diabetes. At weaning, rats were fed an AIN-76 semisynthetic diet for 30, 64, or 113 days, after which plasma and tissues from 10 rats of each group were collected and analysed for AA, DHAA, and vitamin E. AA and DHAA levels were significantly increased in plasma and spleen of the diabetes-prone rats compared with those of the control group at 30 and 64 days, but the difference disappeared by 113 days. No differences were observed in liver, adrenals, thymus, and pancreas at any of the time periods. However, lower levels of vitamin E were observed in adrenal gland, thymus, and pancreas of the diabetes-prone rats. It is concluded that BBdp rats have an altered metabolism of AA, DHAA, and vitamin E, before the onset of diabetes. These changes could be due to genetic and physiological factors operating during development of this rat strain.

Kinetics of tissue RRR-alpha-tocopherol depletion and repletion. Effect of cold exposure.

Vitamin E was estimated in plasma and tissues of rats kept for three months on a low vitamin E diet or a high vitamin E diet. Some of the animals from each group were switched to the opposite diet, and the kinetics of uptake and depletion of vitamin E were followed 3, 8, and 15 days after the diet change. Some rats were also submitted to cold exposure (6 degrees C) for three days. During repletion plasma, red blood cells, liver, spleen, and adrenal gland were the only tissues that responded rapidly to the diet change; after three days, their vitamin E levels corresponded to that of the new diet. Heart, brain, lung, muscle, and thymus were slow in reacting to diet change. Fifteen days after the change in diet, white adipose tissue did not respond. The rate of repletion for all tissues was more rapid than the rate of depletion, but liver was the only tissue that after three days had vitamin E levels corresponding to the low-vitamin diet. Cold exposure for three days did not produce any significant change in the vitamin E content of any tissue, indicating that despite high oxygen consumption by the animal, vitamin E was not consumed or mobilized.

Increased plasma and tissue levels of vitamin E in the spontaneously diabetic BB rat.

Increased plasma and tissue levels of vitamin E were found in spontaneously diabetic BB rats (D) as well as asymptomatic/diabetes-prone BB rats (AD) in comparison to levels in non-diabetic control rats (ND). Treatment of D rats with insulin for 30 days returned plasma and tissue values of vitamin E to control levels. The changes reported here could not be explained solely on the basis of variations in total lipid content of plasma. These data suggest the metabolism of vitamin E is altered in asymptomatic and spontaneously diabetic BB rats and this alteration returns to control values following insulin treatment. Furthermore, it might be speculated that these data indicate a relationship between vitamin E and insulin.

Extracorporeal photopheresis for the treatment of cutaneous T-cell lymphoma--the Düsseldorf and Munich experience.

Extracorporeal photopheresis (ECP) using UVA irradiation of enriched lymphocytes in the presence of 8-methoxypsoralen as a photoactivatable substrate was originally introduced as a therapeutic regimen for cutaneous T-cell lymphoma (CTCL). Whereas ECP has previously been reported to be useful primarily for erythrodermic lymphoma, our purpose was to obtain data on safety and efficacy of ECP in patients suffering from different stages of CTCL. We report on 17 patients, 3 with erythroderma and 14 with plaque or tumor stages. In contrast to other studies our patients were treated predominantly with ECP alone; only a few patients received concomitant therapy. These data have not been published previously, except for preliminary data on four patients. Of the 17 patients, 12 (70%) responded to ECP. In seven patients at least 50% of skin lesions disappeared (defined as partial response) and in five patients at least 25% of skin lesions disappeared (defined as minor response). In two patients the disease remained stable and in three patients the disease progressed under the ECP treatment. No complete remission was observed. Partial responses were achieved not only in patients with early CTCL (stage Ib) but also in those with far progressed tumours (stage IVa). After treatment for 6 months partial responders showed an increase in the number of NK cells in their peripheral blood (P < 0.01). We cannot confirm a relationship between this treatment and CD8 cell counts, as reported by others. Overall, our results indicate that ECP is a safe and effective regimen for the treatment of all stages of CTCL.

Malonaldehyde determination in tissues and biological fluids by ion-pairing high-performance liquid chromatography.

A method for the analysis of malonaldehyde by ion pairing high-performance liquid chromatography is described. The method is direct; no thiobarbiturate chromogen formation is required, and sample preparation is simple. After deproteinization with 50% ethanol and removal of particulate by centrifugation samples were passed through a small silica amino column to remove contaminants. Diluted samples (20 microL) were injected onto an octadecylsilane column (25 cm x 4.6 mm ID, 5 micron) which is eluted with 30 mM sodium phosphate buffer, pH 6.5 containing 30% ethanol and 1 mM tetradecyltrimethylammonium bromide. Detection was accomplished by monitoring absorbance at 267 nm. The lower limit for reliable quantification was 5 pmol per injection. The method has been successfully applied to the quantification of malonaldehyde present in plasma, urine and tissues of rats kept under different dietary conditions as well as after in vivo treatment with CCl4 and iron-dextran. The method was also applied to the quantification of malonaldehyde during liver microsomal lipid peroxidation and was compared to the thiobarbituric acid test.

Effects of dietary linseed oil and marine oil on lipid peroxidation in monkey liver in vivo and in vitro.

Diets rich in linoleic acid (CO) from corn oil, or in linoleic acid and either alpha-linolenic acid (LO) based on linseed oil or n-3 fatty acids (MO) from menhaden oil were fed to male and female Cynomolgus monkeys for 15 wk. In the liver a 40% reduction of alpha-tocopherol occurred in the MO group relative to the CO and LO groups followed by increased formation of lipofuscin in vivo. A four-fold increase of alpha-tocopherol in the MO diet (MO + E) brought the level in the liver to that found with CO and LO. The increased peroxidation in the MO group in the liver phospholipids was associated with the replacement of 60% of the n-6 fatty acids by n-3 fatty acids from menhaden oil. Similar fatty acid profiles were found in groups fed MO and MO + E, respectively. Compared to the CO fed group, feeding alpha-linolenic acid only resulted in a slight incorporation of n-3 fatty acids in the liver membranes mainly due to a direct incorporation of alpha-linolenic acid. However, in monkeys fed menhaden oil more than 30% of the total fatty acids in the liver phospholipids were n-3 fatty acids. The various diets did not influence the activity of liver catalase (EC 1.11.1.6) nor superoxide dismutase (EC 1.15.1.1), but glutathione-peroxidase activity (EC 1.11.1.9) was higher in monkeys fed the MO diet. The catalase activity in females was 20% higher than in males.(ABSTRACT TRUNCATED AT 250 WORDS)

Ascorbic and dehydroascorbic acids status in rats fed diets varying in vitamin E levels.

Vitamin E, ascorbic and dehydroascorbic acids were determined in plasma and selected tissues of rats fed for 2-3 months different diets varying in vitamin E content. The diets contained as low as 5 IU for group LE, a normal amount of 50 IU for group NE and as high as 250 IU of vitamin E for group HE. Small increases in total vitamin C were observed only in liver, kidney, spleen and plasma with increased dietary levels of vitamin E, however, this was not followed by a substantial increment in the ascorbate/dehydroascorbate ratio. These differences were only observed between diets LE and HE; there was no correlation between vitamin E and vitamin C levels in the tissue. These data suggest that the interactions that readily take place in vitro between these two vitamins do not occur in vivo, probably due to the complexity of natural membranes.

Modification of sympathetic nervous system activity in rat tissues by dietary vitamin E.

Noradrenaline content and turnover rate were measured in sympathetically innervated tissues of adult male rats that were fed diets varying in alpha-tocopherol for 10 to 11 weeks. At the end of the feeding period, increasing amounts of vitamin E increased the levels of noradrenaline in liver and white adipose tissue but no differences were observed in heart and interscapular brown adipose tissue. By contrast, increased amounts of dietary vitamin E decreased both the rate constant for the decline of specific activity of noradrenaline with time and the noradrenaline turnover rate in heart and brown adipose tissue. No significant differences were observed in these parameters in liver and white adipose tissue. These results indicate that vitamin E in the diet may affect activity of the sympathetic nervous system.

Mediterranean macrothrombocytopenia.

Platelet count, platelet size, and circulating platelet biomass concentration estimates made with an erythrocyte-calibrated electronic sizing system on EDTA-anticoagulated blood samples gave population medians and 95% ranges for 145 asymptomatic Mediterranean and 200 healthy Northern European subjects. The Mediterraneans had lower platelet counts [161,000 (89,000-290,000)/mul compared with 219,000 (148,000-323,000)/mul] and higher arithmetic mean volumes [17.8 (10.8-29.2) cu mum compared with 12.4 (9.9-15.6) cu mum], while the individual lognormal platelet size distribution profiles were comparable [geomatric standard deviations of 1.78 (1.60-1.98) against 1.70 (1.54-1.88)]; and the platelet biomass concentrations, given by count per microliter times mean volume times 10- minus 7 and expressed as a volumetric percentage of whole blood, were almost identical [0.286% (0.216%-0.379%) against 0.272% (0.201%-0.367%)]. Mediterranean macrothrombocytopenia is, therefore, considered a benign morphologic variant that requires differentiation from thrombocytopenias in which the circulating platelet biomass concentration is decreased.

Splenomegaly, macrothombocytopenia and stomatocytosis in healthy Mediterranean subjects (splenomegaly in Mediterranean macrothrombocytopenia).

Spleen size, stomatocytosis, macrothrombocytopenia, haemoglobin level, white cell count, and abdominal pain episodes were assessed in a coded study of healthy Mediterranean immigrants to Australia. Spleen size was estimated from a length measurement, L, on a standardized plain abdominal radiograph and expressed both as spleen weight and as a spleen length index, L/square root BSA; the platelet count and size parameters were determined electronically and the presence of stomatocytes was evaluated in stained blood films. In relation to 16 Northern European control women 12 of 25 Mediterranean women had radiographic splenomegaly, 10 had macrothrombocytopenia, 9 had stomatocytosis, but none had episodes of abdominal pain. The median spleen weights of the two groups were estimated as 157 and 247 g with ranges from percentile 2.3 to 97.7 of 75 to 328 and 112 to 669 g. Within the Mediterranean group splenomegaly correlated with macrothrombocytopenia (P less than 0.001) but not with stomatocytosis, haemoglobin values or white cell counts. Thus, mild splenomegaly may be expected in Mediterranean macrothrombocytopenia, Mediterranean stomatocytosis appears unrelated, and all of these apparently benign anomalies may be incidental findings in patients from the Italian and Balkan peninsulas.

Megakaryocyte polyploidy as a grouped geometric distribution obeying the log-normal population law.

Discrete nuclear lobe scores and flow- or image-cytometry DNA values of classically identified megakaryocytes behave as a grouped geometric distribution. This model is fully specified by a geometric mean and standard deviation (GM and GSD), the latter typically being ca 1.16 for volumes of diploid blood cell populations. Via log-normal probability paper, the 30 to 50 megakaryocytes in clinical marrow smears readily yield the ploidy model's GM and GSD which are named MPM and MPD for megakaryocyte polyploidy median and dispersion. In euthrombopoietic outbred mammals, MPMs are ca 12N ploidy units, and MPDs approximate a factor of 1.41. Both are unitless criteria. Thus, the thrombon is characterized by three populations exhibiting the high size dispersion which unmasks canonical operation of the log-normal population law: picoliter megakaryothrombocytes with their MPD ca 1.41, femtoliter thrombocytes with a volume GSD ca 1.74, and the end product of locally delivered pieces of subattoliter platelet dust with a volume GSD ca. 2.0.

Ecdysteroids in the pycnogonid Pycnogonum litorale (STROM) (Arthropoda, Pantopoda).

Two ecdysteroids were isolated from juveniles and adults as well as from eggs of the pycnogonid Pycnogonum litorale Ström and characterized by bioassay, thin layer chromatography, gas liquid chromatography, and high performance liquid chromatography. One of the hormones could be identified as 20-hydroxyecdysone whereas the other behaves in HPLC in a way that is similar to but not identical with ecdysone. This unidentified ecdysone-like hormone exceeded 20-hydroxyecdysone in all developmental stages (1.67:1 to 3:1). Determination of moulting hormone titers by a radioimmunoassay showed highest amounts of hormones in juvenile females (150 ng/mg tissue dry wt). The results are compared with those obtained from other arthropods and are discussed in view of the evolution of hormonal systems.

Tissue discrimination between dietary RRR-alpha- and all-rac-alpha-tocopherols in rats.

The discrimination between the natural form of vitamin E, RRR-alpha-tocopherol, and the synthetic form all-rac-alpha-tocopherol was measured in two experiments for plasma and tissues of rats. In Experiment A, the animals were fed RRR-alpha- or all-rac-alpha-tocopheryl acetate at two different levels, 0.035 and 0.200 g/kg diet, for 2.5 mo. In Experiment B, rats were first depleted of vitamin E for 7 wk then fed a diet containing 0.035 g/kg diet of RRR-alpha- or all-rac-alpha-tocopheryl acetate. In the latter experiment, rats were killed after 3, 7, 17, 31 or 45 d to study the time course of the discrimination. At the end of both experiments, levels of alpha-tocopherol were measured in plasma and tissues. The results showed a preferential uptake of the natural RRR-alpha-tocopherol by the majority of the tissues. The preference is small and in some tissues is not immediately manifested. The greatest discrimination in the tissues examined was found in red blood cells; the levels of alpha-tocopherol accumulated with the RRR-alpha-tocopherol diet were four to six times that accumulated with all-rac-alpha-tocopherol diet. For all other tissues, this ratio was between 1.0 and 1.5. These results indicate that tissues have a mechanism for alpha-tocopherol uptake, but this mechanism is not entirely specific for RRR-alpha-tocopherol.

Effects of high ascorbic acid intake on the metabolism of catecholamines in the rat.

The effect of high dietary ascorbic acid intake on the homeostasis of ascorbic acid and on the metabolism of catecholamine was studied in rats. In order to eliminate changes in serum levels due to stress, blood was obtained without anesthesia through implanted cannulas. Rats with a high intake of ascorbic acid (20 g/kg diet, for 7-9 weeks) had significantly higher levels of ascorbic acid in serum (83%), liver (28%), kidney (47%) and superior cervical ganglia (43%). The values were slightly, but not significantly higher in adrenal gland, brain and interscapular brown adipose tissue. However, no significant differences were observed in the levels of noradrenalin, adrenalin and dopamine in any of the tissues analyzed. Dopamine beta-hydroxylase activity was also unchanged. There was thus no evidence of an effect of high dietary intakes of ascorbic acid on the metabolism of catecholamines in the intact rat.

Alpha- and gamma tocopherol concentrations in human serum.

Vitamin E (alpha- and gamma-tocopherol) and total lipid concentrations were determined in the serum of 451 healthy human male and female subjects ranging in age from 19 to 70 years. Mean serum alpha-tocopherol concentration (microgram/ml) was 8.82 +/- 2.84 (SD) and gamma-tocopherol concentration was 1.35 +/- 0.70. The concentration of alpha-tocopherol increased significantly with age, as well as with total lipid; gamma-tocopherol concentration remained unchanged with age. The data also indicated that subjects with higher serum alpha-tocopherol had lower gamma-tocopherol levels and vice versa for subjects with higher gamma levels. A close interrelationship and competition appeared to exist between these two vitamers.

Levels of noradrenaline in plasma during thermogenesis induced by cold-exposure or by noradrenaline infusion in warm- and in cold-acclimated rats.

Oxygen consumption and concentration of noradrenaline (NA) in arterial plasma have been measured in warm-acclimated (WA) and in cold-acclimated (CA) rats during exposure to cold and in barbital-sedated WA or CA rats receiving intravenous infusions of NA at various doses at room temperature. In WA and in CA rats in the cold, NA in plasma increased rapidly from a resting level of 0.1 to 0.2 ng/ml at thermoneutrality to values 5 to 10 times greater at -19 degrees C. The increases were directly related to the degree of cold exposure and were maintained throughout exposure. In barbital-sedated WA or CA rats infused with NA the plasma levels of NA which stimulated calorigenesis were approximately 10 times greater than those resulting from overflow of NA into the blood during intense cold exposure and were in the range found by others to be effective in stimulating O2 consumption in isolated brown adipocytes.

Ascorbic acid in brown adipose tissue: effect of cold acclimation and high intake of the vitamin.

The ascorbic acid content of brown adipose tissue increased 62% when rats underwent cold acclimation; however, the increase was 126% when rats living in the cold were fed a basal diet supplemented with 2% ascorbic acid. These findings suggest a role for ascorbic acid in this tissue during nonshivering thermogenesis.

Mechanisms of absorption, transport and tissue uptake of RRR-alpha-tocopherol and d-gamma-tocopherol in the white rat.

The metabolism of alpha- and gamma-tocopherol was studied in three groups of rats that were fed a modified AIN-76 diet containing normal (NE, 0.2 g alpha-tocopherol/kg), high (HE, 1.0 g alpha-tocopherol/kg) or low (LE, less than 0.02 alpha-tocopherol/kg) vitamin E for 3 mo. After 1, 2 and 3 d of an oral dose of 20 mg of alpha-tocopherol, gamma-tocopherol or both, the levels of the two vitamers were measured in plasma and tissues and in some cases in isolated microsomal and mitochondrial fractions from liver. Twenty-four hours after an oral dose of 20 mg gamma-tocopherol the levels of alpha-tocopherol in plasma and tissues remained constant and higher levels of gamma-tocopherol were found in tissues in which low alpha-tocopherol levels could be found such as in the LE group. In spite of this, it was enabled to remain there, after 2 and 3 d gamma-tocopherol had decreased levels in all tissues. When given in combination with alpha-tocopherol, the levels of gamma-tocopherol were lower than when gamma-tocopherol was given alone. Microsomes and mitochondria from livers of LE group bound five and nine times more alpha-tocopherol than gamma-tocopherol in rats dosed with equal amount of alpha- or gamma-tocopherol, respectively. These data indicate that the mechanisms that regulate the metabolism of vitamin E are highly specific for alpha-tocopherol. Moreover, the relative amount of alpha-tocopherol determined the levels of gamma-tocopherol in tissues. However, the retention of gamma-tocopherol in tissues did not depend on the presence of alpha-tocopherol.(ABSTRACT TRUNCATED AT 250 WORDS)