Differences in the hydrophobic properties of discrete alpha-helical domains of rat and human apolipoprotein A-IV.

The primary structure of human apolipoprotein A-IV has been analyzed to identify amphipathic alpha-helical regions and compare their hydrophobic properties with analogous segments of rat apolipoprotein A-IV. Significant differences were found in the mean hydrophobicity and/or mean helical hydrophobic moment of several discrete alpha-helical domains of rat and human apolipoprotein A-IV, which may determine the differences in their biophysical and biological behavior.

Binding of human apolipoprotein A-IV to human hepatocellular plasma membranes.

We have investigated the binding of human apolipoprotein A-IV (apo A-IV) to human hepatocellular plasma membranes. Addition of increasing concentrations of radiolabeled apo A-IV to hepatic plasma membranes, in the presence and absence of a 25-fold excess of unlabeled apo A-IV, revealed saturation binding to the membranes with a KD of 154 nM and a binding maximum of 1.6 ng/microgram of membrane protein. The binding was temperature-insensitive, partially calcium-dependent, abolished when apo A-IV was denatured by guanidine hydrochloride or when the membranes were treated with Pronase and decreased when apo A-IV was incorporated into phospholipid/cholesterol proteoliposomes. In displacement studies using purified apolipoproteins and isolated lipoproteins, only unlabeled apo A-IV, apo A-I and high-density lipoproteins effectively competed with radiolabeled apo A-IV for membrane binding sites. We conclude that human apo A-IV exhibits high-affinity binding to isolated human hepatocellular plasma membranes which is saturable, reversible and specific.

Pro-oxidant effect of vitamin E in cigarette smokers consuming a high polyunsaturated fat diet.

Dietary polyunsaturated fats and vitamin E are associated with reduced risk for atherosclerosis, but in smokers, they could promote lipid oxidation. Therefore, we examined the effects of a high polyunsaturated fat diet and vitamin E supplementation on measures of lipid oxidation in cigarette smokers. Ten subjects who smoked >1 pack of cigarettes per day were sequentially fed the following: a baseline diet in which the major fat source was olive oil, a diet in which the major fat source was high-linoleic safflower oil, and finally, the safflower oil diet plus 800 IU vitamin E per day. LDL oxidation lag time and rate and plasma total F(2)-isoprostanes and prostaglandin F(2alpha) (PGF(2alpha)) were determined after 3 weeks on each diet. The safflower oil diet increased total F(2)-isoprostanes from 53.0+/-7.2 to 116.2+/-11.2 nmol/L and PGF(2alpha) from 3.5+/-0.2 to 5.5+/-0.5 nmol/L, without changing LDL oxidation parameters. Addition of vitamin E prolonged mean LDL oxidation lag time but, paradoxically, further increased F(2)-isoprostanes to 188.2+/-10.9 nmol/L and PGF(2alpha) to 7.8+/-0.4 nmol/L. These data suggest that vitamin E may function as a pro-oxidant in cigarette smokers consuming a high polyunsaturated fat diet.

Short-term parenteral nutrition with glucose and Intralipid: effects on serum lipids and lipoproteins.

We prospectively investigated the effect of a 3-wk course of parenteral nutrition with 20% glucose, 4.25% amino acids, and 10% Intralipid on plasma lipids and lipoproteins in a cohort of 12 nutritionally replete patients with inflammatory bowel disease. Mean total serum cholesterol and mean total serum phospholipids increased in parallel throughout the study; density gradient ultracentrifugation demonstrated these increases were due to the appearance of lipoprotein-X. The mean low-density lipoprotein (LDL) mass on the density gradients decreased during the study and high-density lipoprotein (HDL) cholesterol levels decreased by 28% by the end of the third week. Plasma free fatty acids decreased by 42%. These results demonstrate that the glucose and 10% Intralipid regimen caused modest decreases in serum HDL cholesterol and LDL mass and the prompt appearance of lipoprotein-X with attendant hypercholesterolemia and hyperphospholipidemia. Patients undergoing long-term treatment with glucose and Intralipid should be closely monitored for the occurrence of significant dyslipoproteinemia.

In vitro reciprocal exchange of apoproteins and nonpolar lipids between human high density lipoproteins and an artificial triglyceride-phospholipid emulsion (Intralipid).

To determine the nature of lipid and apoprotein exchange between human high density lipoproteins (HDL) and Intralipid particles of Sf greater than 400 (ILIP) we have studied their in vitro interaction during incubation in aqueous buffer and in lipoprotein-deficient serum (LPDS). We found that ILIP acquires apo A-I, apo A-IV and apo E from LPDS, and that this uptake is inhibited by the presence of HDL, which readily donate C-apoproteins to the ILIP surface. In the absence of LPDS exchange of only polar lipids occurred between ILIP and HDL, with HDL gaining phospholipid from, and donating free cholesterol to this fat emulsion. In the presence of LPDS the exchange of nonpolar lipids occurred between the two particles: in the case of HDL, cholesteryl ester content decreased, accompanied by an increase in triglyceride, causing a decrease in the hydrated density of the lipoprotein and an increase in its molecular weight; in the case of ILIP, reciprocal changes in lipid content were seen as a loss of triglyceride and the appearance of cholesteryl esters. When compared to literature data, our findings indicate that Intralipid Sf greater than 400 particles exhibit an in vitro behavior which is remarkably similar to that of nascent chylomicrons with respect to the exchange of A- and C-apoproteins and surface polar lipids with HDL. We postulate that since ILIP and HDL can participate in a LPDS-dependent exchange of non-polar core lipids, that this process may occur when this fat emulsion is administered in vivo.

Effects of a high polyunsaturated fat diet and vitamin E supplementation on high-density lipoprotein oxidation in humans.

Oxidative modification of high-density lipoproteins (HDL) impairs several biologic functions critical to its role in reverse cholesterol transport. We therefore investigated the effect of dietary polyunsaturated fat and vitamin E on the kinetics of HDL oxidation. Ten subjects were fed sequentially: a baseline diet in which the major fat source was olive oil; a high polyunsaturated fat diet in which the major fat source was safflower oil; and the safflower oil diet plus 800 I.U. vitamin E per day. Plasma lipoprotein levels, vitamin E content, fatty acid composition, and oxidation lag time and rate were determined after 3 weeks on each diet. The polyunsaturated fat diet increased the mean HDL(2) lag time from 45.8+/-12.5 to 83.3+/-11.6 min with no change in oxidation rate. Addition of vitamin E further increased the HDL(2) lag time to 115.6+/-4.4 min and decreased the HDL(2) oxidation rate 10-fold. Neither the polyunsaturated diet alone nor the diet with vitamin E supplementation had any effect on HDL(3) oxidation. We conclude that under conditions of controlled dietary fat intake, a high polyunsaturated fat intake does not increase the oxidation susceptibility of HDL subfractions, and that in this setting, vitamin E supplementation reduces the oxidation susceptibility of HDL(2). These data suggest that antioxidants could influence HDL function in vivo.

Lipid abnormalities in cyclosporine-prednisone-treated renal transplant recipients.

Hyperlipidemia and hypertension, two major risk factors for accelerated atherosclerosis, undoubtedly contribute to the excessive cardiovascular morbidity and mortality experienced by renal transplant recipients. The present survey of posttransplant hyperlipidemia in 500 cyclosporine-treated patients documented a 37.6% incidence of hypercholesterolemia, which occurred within 6 months posttransplant in 82% of patients. An etiologic relation to corticosteroid therapy was suggested by the strong correlation between prednisone doses and cholesterol levels, by the reduced cholesterol levels in patients undergoing steroid withdrawal, and by the reduction in hypercholesterolemia to 13% by 3 years posttransplant when steroid doses were less than 10 mg daily. Hypertriglyceridemia, which was present in 14.7% of the patients, was more severe under CsA-prednisone compared with azathioprine-prednisone therapy. Hypertriglyceridemia, which occurred later in the posttransplant course than hypercholesterolemia, strongly correlated with an excessive percent relative weight and elevated serum creatinine but not with steroid or CsA doses. Increasing age, diabetes mellitus, beta-blockers and nephrotic syndrome contribute to posttransplant hyperlipidemia in the CsA-Pred era as they did in the azathioprine era of immunosuppression.

Bitter taster status predicts susceptibility to vection-induced motion sickness and nausea.

BACKGROUND: Sensitivity to bitter taste and susceptibility to nausea are both protective mechanisms that guard against toxin ingestion, and both these traits vary within and between populations. Thus, we postulated that they may have co-evolved, such that they are associated. METHODS: Bitter taster status was determined in 40 subjects (13 men, 27 women) by measuring the differential perceived taste intensity between salt and n-propylthiouracil using a labeled magnitude scale; susceptibility to vection-induced motion sickness and nausea was assessed using an optokinetic drum, a validated multi-symptom scoring scale, and electrogastrography. KEY RESULTS: Taster status distribution was 25% non-tasters (NT), 40% tasters (T), and 35% supertasters (ST). Gender had no impact on this distribution, but females had a higher mean maximum symptom score than males (12.4 ± 1.4 vs 7.3 ± 2.0). Non-tasters displayed a faster and larger increase in mean symptom scores, had a higher percentage of subjects with high maximum symptom scores, and had a higher mean maximum score than T or ST, (14.8 ± 2.6 vs 7.1 ± 1.8, vs 9.8 ± 2.0). Taster status did not affect the gastric myoelectric frequency response to vection. CONCLUSIONS & INFERENCES: Non-tasters are more susceptible to vection-induced motion sickness and nausea than T or ST, suggesting these two traits may have co-evolved in a reciprocal manner: in environments where the NT trait conferred an evolutionary advantage by enabling intake of fruits and vegetables containing bitter, yet beneficial, phytonutrients, increased nausea susceptibility may have arisen to maintain protection against ingested toxins.

Identification of functional domains in the plasma apolipoproteins by analysis of inter-species sequence variability.

Molecular evolution theory posits that sequence motifs essential for protein function are constrained by selective pressure from changing over long stretches of evolutionary time. Thus, analysis of inter-species amino acid sequence variability, by identifying highly conserved intervals, should predict the location of domains critical for protein function. We have analyzed the amino acid sequences of the mammalian apolipoproteins A-I, A-IV, C-I, C-II, C-III, D, and E with a computer algorithm that calculates numerical residue variability scores. The application of a median sieve filter to the data facilitated identification of the exact boundaries of highly conserved domains, which coincided with the location of known structural features and functional domains in this family of proteins. The analysis also identified highly conserved intervals in every apolipoprotein whose function is unknown at present, but which are candidates for regions with specific functional roles.

Apolipoprotein A-IV-2 allele: association of its worldwide distribution with adult persistence of lactase and speculation on its function and origin.

Apolipoprotein A-IV (apo A-IV) is a 46-Kd plasma glycoprotein that may play a major role in intestinal lipid absorption. A genetic polymorphism in the apo A-IV gene, apo A-IV-2, encodes a His-->Gln substitution at codon 360 that alters the biological function of this apolipoprotein. As the worldwide distribution of the apo A-IV-2 allele appeared similar to the frequency of a genetic polymorphism that determines the persistence of lactase into adulthood, we examined the relationship between the apo A-IV-2 and lactase persistence polymorphisms by compiling the prevalence of adult lactase persistence in all populations in which the frequency of the apo A-IV-2 allele has been determined. Across 29 groups, there was an extremely strong correlation (4 = 0.937, P < 0.000001) between apo A-IV-2 allele frequency and the prevalence of adult lactase persistence. Apo A-IV-2 allele frequency was highest in Iceland, an ancient Viking colony, and decreased across Europe in a north-to-south and west-to-east gradient, generally following hypothetical isoclines for the lactase persistence gene. There were no correlations between the population frequencies of the apo E2, E3, or E4 alleles and either the prevalence of lactase persistence or the frequency of the apo A-IV-2 allele. In light of the effects of the apo A-IV-2 polymorphism on lipid metabolism, we speculate that the apo A-IV-2 allele may have originated in ancient Scandinavia, spread by conferring a nutritional advantage in the setting of a lifelong high milkfat intake, and was later carried southwards by the Viking incursions into Europe.

Employment of master's-level psychologists.

Compared results from a survey of employment of Master's-level psychologists in the states of Michigan, Indiana, Ohio, and Kentucky to two previous studies recently published. It was found that results in all three studies are comparable; 87.6% of those mental health centers, state hospitals and other mental health organizations surveyed reported that they employ Master's-level psychologists. Duties of subdoctoral psychologists include testing, therapy, and community consultation. The finding that future Master's-level psychologists may find greater employment potential in towns with a population of less than 50,000 justifies future research in the area of Master's-level employment to consider demographic variables.

Exposure and electronic interaction of tyrosine and tryptophan residues in human apolipoprotein A-IV.

We have investigated the exposure and electronic interaction of tyrosine and tryptophan residues in human apolipoprotein A-IV (apo A-IV). Differential absorption spectroscopy and chemical titration demonstrated that human apo A-IV contains six tyrosine residues, four of which are buried in the hydrophobic interior of the protein and two of which are exposed on the protein surface. Denaturation of the protein by guanidinium chloride caused progressive exposure of the buried tyrosines. The fluorescence emission spectra of apo A-IV were characterized by a blue-shifted tryptophan emission with a low relative quantum yield of 0.37 and a tyrosine emission with a relative quantum yield of 0.62. Fluorescence quenching studies demonstrated a low fractional exposure of tryptophan in the native state. Denaturation of apo A-IV was accompanied by an increase in the relative quantum yield which peaked at the denaturation midpoint. Fluorescence excitation techniques demonstrated energy transfer from tyrosine residues with a transfer efficiency of 0.40 in the native state; the efficiency was conformation dependent and decreased with protein unfolding. Fluorescence studies of tetranitromethane-modified apo A-IV suggested that a significant fraction of energy transfer proceeds from the exposed tyrosine residues. These data demonstrate the existence of intramolecular fluorescence energy transfer and tryptophan quenching in human apolipoprotein A-IV and suggest that the amino terminus of this protein is situated in a hydrophobic domain within energy-transfer range of nonvicinal tyrosine residues.

The self-association of human apolipoprotein A-IV. Evidence for an in vivo circulating dimeric form.

We have investigated the self-association properties of human apolipoprotein A-IV using several complementary physical techniques. Sedimentation equilibrium analysis demonstrated that human apolipoprotein A-IV formed oligomeric species in aqueous solution at physiologic pH. Computer analysis established that the best model of self-association is a monomer-dimer-tetramer scheme, with an unusually large monomer-dimer association constant of 2.9 X 10(5) liters/mol. Fluorescence spectroscopy and electrophoretic analysis demonstrated that the rate of monomer-oligomer interconversion is sufficiently slow that a stable population of dimeric protein exists in solution, even at low total protein concentrations, and that the extent of dimerization is minimally influenced by pH. Moreover, these techniques established that the dissociation of oligomeric forms and the unfolding of the monomeric form are discrete and sequential events. In experiments where apolipoprotein A-IV was incubated with human high density lipoproteins, fractionated by gradient gel electrophoresis, and localized by immunoblotting, dimer formation occurred, but very little binding to lipoproteins was observed. Immunoblots of human serum fractionated on acrylamide gradient gels and isopycnic density gradients demonstrated an apolipoprotein A-IV band of size and density consistent with a circulating dimeric form, unassociated with lipid. We conclude that human apolipoprotein A-IV undergoes high affinity self-association in aqueous solutions, and that such self-association likely occurs in vivo. Self-association may thus be important in determining the biologic behavior of human apolipoprotein A-IV by influencing both the kinetics and distribution of its association with plasma lipoproteins.