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Biomedical subjects

L C Smith

Publications and source records attributed to L C Smith.

At least 163 records · Page 9Linked to original sources
PubMed1983

Interaction of synthetic N-5-dimethylaminonaphthalene-1-sulfonyl-apolipoprotein C-II peptides with lipoprotein lipase.

Synthetic fragments of apo-C-II, specifically labeled on their NH2-terminals with the 5-dimethylaminonaphthalene-1-sulfonyl (dansyl or DNS) fluorophore, have been prepared by solid phase peptide synthesis. When a complex is formed between bovine milk lipoprotein lipase and N-dansyl-apo-C-II peptides, resonance energy transfer occurs from the tryptophan residues of the enzyme to the dansyl-labeled peptides upon excitation at 280 nm. In the absence of lipid, the association constant increases 10-fold when the length of the DNS peptide is increased from apo-C-II-DNS(64-78) (0.04 X 10(6) M-1) to apo-C-II-DNS(60-78) (0.3 X 10(6) M-1). In the presence of lipid, the association constants are dependent on peptide chain length, and increase from 0.4 X 10(6) M-1 for apo-C-II-DNS(64-78) to 2.2 X 10(7) M-1 for apo-C-II-DNS(43-78). The interactions are specific for lipoprotein lipase, are disrupted by guanidinium chloride, are not affected by 1.0 M NaCl, and are competitive with the corresponding nondansylated peptide. Apolipoproteins C-III and A-I, at 5 to 1 molar ratios, had no effect on the interaction. These findings demonstrate the importance of the COOH-terminal region in the lipoprotein lipase-apo-C-II interaction and show that activation of the enzyme involves a specific protein-protein interaction.

Amino Acids↗
PubMed1983

Three patterns of extra-testicular venous drainage in the rabbit. Divergence from a traditional concept.

Seventy-eight male New Zealand white rabbits were autopsied and found to have variable left extra-testicular venous anatomy. Our observations reveal that in the rabbit the left testis is drained in one of three ways, identified as either A (18%), B (30%) or C (52%)--type drainage. The right testicular vein in all cases drained directly into the inferior vena cava immediately superior to the right iliolumbar vein. In type A drainage, the left testicular vein drained directly into the inferior vena cava at the level of the left iliolumbar vein. In type B drainage, the left testicular vein emptied into the left iliolumbar vein, which in turn drained into the inferior vena cava. In type C drainage both the left testicular and iliolumbar veins anastomosed to form a "lumbotesticular" trunk which emptied directly into the left renal vein. These three patterns of left venous vascular anatomy in the rabbit can be explained on the basis of their embryologic development. Our observations suggest that it is the caudal segment of the left pelvic subcardinal vein and its anastomosis with the caudal cardinal complex which persist as the left testicular vein and that the more cranial segment of this vein, heretofore presumed to remain patent, atrophies to the level of the developing left renal vein.

Animals↗
PubMed1983

Transfer of polycyclic aromatic hydrocarbons between model membranes: relation to carcinogenicity.

Polynuclear aromatic hydrocarbons (PAH), some of which are potent carcinogens, are common environmental pollutants. The transport processes for these hydrophobic compounds into cells and between intracellular membranes are diverse and are not well understood. A common mechanism of transport is by spontaneous desorption and transfer through the aqueous phase. From the partitioning parameters, we have inferred that the rate limiting step involves solvation of the transfer species in the interfacial water at the phospholipid surface. Transfer of 10 PAH (pyrene, 3,4-benzophenanthrene, triphenylene, chrysene, 1,2-benzanthracene, 1,1'-binaphthyl, 9-phenylanthracene, 2,2'-binaphthyl, m-tetraphenyl and 1,3,5-triphenylbenzene) out of phosphatidylcholine vesicles has been examined. Our results show that the molecular volume of the PAH is a rate-determining factor. Moreover, high performance liquid chromatography (HPLC) data confirms the hypothesis that the rate of transfer is correlated with the size of the molecule and with the partitioning of the molecule between a polar and hydrocarbon phase. The kinetics and characteristics of the spontaneous transfer of carcinogens are likely to have a major impact on the competitive processes of PAH metabolism within cells.

Biological Transport↗
PubMed1983

A simplified approach to resonance energy transfer in membranes, lipoproteins and spatially restricted systems.

A general model is developed to simulate dipole-dipole resonance energy transfer in spatially restricted systems. At low concentrations of acceptor molecule, the overall quantum yield of a donor population can be defined quantitatively in terms of transfer to multiple defined acceptor regions. Energy transfer at higher acceptor concentrations can be approximated by assuming an exponential dependence of relative quantum yield on the acceptor concentrations. Through geometrical manipulations, this algorithm has been applied using an electronic calculator to systems in which donor-acceptor interaction is limited by unique steric restriction on donor and acceptor distribution within lipid aggregates. The systems that have been analyzed include monomolecular films, bilayer membranes, small discoidal lipid-protein complexes and plasma lipoproteins. The observed energy transfer from N-(2-naphthyl)-23,24-dinor-5-cholen-22-amide-3 beta-ol to N-dansyldimyristoylphosphatidylethanolamine in a dimyristoylphosphatidylcholine bilayer agrees with that predicted by this model.

Dimyristoylphosphatidylcholine↗
PubMed1983

Inverse relationship between blood levels of high density lipoprotein subfraction 2 and magnitude of postprandial lipemia.

Triglyceridemic response to a standard oral fat meal was determined in 28 healthy subjects and related to the levels of several lipids, lipoproteins, and apolipoproteins in the post-absorptive plasma. A fatty test meal was administered orally, and postprandial plasma triglyceride levels were determined. In the fasting blood samples, concentrations of apolipoproteins (apo) A-I, A-II, and B were determined by radioimmunoassay, and those of high density lipoprotein (HDL) subfractions HDL2 and HDL3, by zonal ultracentrifugation. The magnitude of triglyceridemic response showed a negative correlation with the plasma levels of HDL2 (r = -0.860, P less than 0.001), HDL-associated cholesterol (r = -0.605, P less than 0.001), and apoA-I (r = -0.459, P less than 0.02). No correlation was found between the triglyceridemic response and the levels of total cholesterol, HDL3, and apoA-II. Triglyceridemic response was correlated positively with fasting triglyceride concentrations (r = 0.450, P less than 0.02) and apoB levels (r = 0.396, P less than 0.03). In two subjects followed for 3 yr, when HDL2 levels rose or fell, the triglyceridemic response decreased or increased, respectively (r = -0.944; r = -0.863). Our data indicate that normolipidemic individuals with high HDL2 levels in the plasma are able to clear alimentary fat at a faster rate than normolipidemic subjects with low HDL2 levels. The pronounced difference in severity and duration of postprandial lipemia among subjects with varying HDL2 levels may help to explain the negative correlation between the risk of atherosclerosis and HDL cholesterol levels.

Adult↗
PubMed1982

Incorporation of defined cholesteryl esters into lipoproteins using cholesteryl ester-rich microemulsions.

Cholesteryl ester transfer to low density lipoproteins (LDL) was achieved either by incubation of a cholesteryl ester-rich lipid microemulsion with whole plasma or by incubation of the cholesteryl ester-rich microemulsion with lipoprotein-free plasma and LDL. Sequential ultracentrifugal isolation gave LDL that contained between 5 and 37% of the cholesteryl ester derived from the artificial cholesteryl ester donor. The labeling was essentially complete within 2 h of incubation. Resonance energy transfer of LDL, labeled with fluorescent cholesteryl esters and with the surface probe 5-(N-hexadecanoylamino) fluorescein, indicated that the added cholesteryl ester was located in the inner core of the lipoprotein. In addition, biological screening of the labeled LDL in cultured cells in vitro and in the rat in vivo showed that this labeling procedure had no detectable deleterious effect on LDL. This method of labeling lipoproteins allows a rapid and specific introduction of cholesteryl esters into lipoproteins.

Animals↗
PubMed1982

5-(dimethylamino)naphthalene-1-sulfonic acid, a fluorescent probe of the medium chain fatty acid binding site of serum albumin.

Reversible binding of 5-(dimethylamino)-naphthalene-1-sulfonic acid (DNS) to human and bovine serum albumin has been monitored by changes in fluorescence intensity, wavelength maxima, and polarization. DNS has only one major binding site (ka = 5 x 10(6)) and one minor site (ka = 3 x 10(5)) on these proteins. The probe is competitively displaced from its high-affinity site by medium chain fatty acids and by N-acetyl-L-tryptophan. The binding site for DNS is highly hydrophobic and is considerably less polar than the hydrocarbon region of lipid bilayers. Resonance energy transfer indicates that the binding site is located 20.7 +/- 2 A from the single tryptophan residue of human serum albumin.

Binding Sites↗
PubMed1982

Cholesteryl ester-rich microemulsions: stable protein-free analogs of low density lipoproteins.

A method has been devised for the preparation of stable lipid microemulsions containing cholesterol, cholesteryl ester, phosphatidylcholine, and trioleoylglycerol in the relative molar ratios found in low density lipoproteins. Gel permeation chromatography showed these microemulsions to be essentially homogeneous with respect to chemical composition. Omission of triolein or substitution of a diunsaturated phosphatidylcholine for either a disaturated or monosaturated-monounsaturated phosphatidylcholine destroyed the observed homogeneity of the microemulsions. The particle diameter of the negatively-strained relative elution volumes of the cholesteryl ester-rich microemulsion, VLDL2, VLDL3, and LDL indicated a mean diameter of about 35 nm. The cholesteryl ester-rich microemulsion can be used as a cholesteryl ester donor for plasma protein-mediated transfer of cholesteryl ester to plasma lipoproteins and for studying apoprotein-lipid interactions.

Apolipoproteins↗
PubMed1981

Low density lipoproteins reconstituted with steroids containing the nitrobenzoxadiazole fluorophore.

A new cholesterol analog, N-(7-nitrobenz-2-oxa-1,3-diazole)-23,24-dinor-5-cholen-22-amine-3 beta-ol, with fluorescent properties similar to those of fluorescein, has been synthesized. The fluorescence lifetimes, quantum yields, and wavelength maxima of N-(7-nitrobenz-2-oxa-1,3-diazole)-23,24-dinor-5-cholen-22-amine-3 beta-ol and its linoleate ester were solvent-dependent. The cholesterol analog was a satisfactory substrate for lecithin:cholesterol acyltransferase. The fluorescent sterol, added in ethanol, gave half-maximal suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in cultured human fibroblasts at 2.5 microM and was twice as effective as cholesterol. The fluorescent steryl ester, incorporated into low density lipoprotein, was used to demonstrate high affinity cellular uptake and degradation of the reconstituted lipoproteins, intracellular accumulation of the free sterol and simultaneous suppression of 3-hydroxy-3-methylglutaryl coenzyme a reductase in fibroblasts. Half-maximal suppression was achieved at 10 micrograms mL-1 of low density lipoproteins reconstituted with the fluorescent steryl ester, compared to the same degree of suppression produced by 2 micrograms mL-1 of native low density lipoproteins. The interaction of low density lipoproteins reconstituted with N-(7-nitrobenz-2-oxa-1,3-diazole)-23,24-dinor-5-cholen-22-amine-3 beta-ol linoleate with cells was readily visualized by fluorescence microscopy and quantified by fluorimetry. These analogs will facilitate the studies of lipoprotein-cell interactions and phospholipid vesicle-cell interactions, the selection of cell mutants defective in lipoprotein metabolism, and the assessment of the immediate environment of the steroids in cellular membranes.

4-Chloro-7-nitrobenzofurazan↗
PubMed1980

Stimulation and suppression of 3-hydroxy-3-methylglutaryl coenzyme A reductase in normal human fibroblasts by high density lipoprotein subclasses.

Plasma concentrations of high density lipoproteins, (HDL3), a subclass of HDL, are relatively constant, while those of HDL2 are variable. We report that HDL2 suppress, while HDL3 stimulate 3-hydroxy-3-methylglutaryl-CoA reductase (EC 1.1.1.34) activity in normal human fibroblasts. HDL3, which contained no detectable HDL2 or low density lipoproteins, stimulated 3-hydroxy-3-methylglutaryl-CoA reductase activity 2-fold from 60 to 120 pmol/mg per min. The induction, which exhibited saturation kinetics with maximum stimulation at 150 microgram HDL phospholipid/ml medium, occurred only in the late-log and stationary phases of cell growth and was abolished by 0.1 mM actinomycin D or cycloheximide.l Apoliproprotein HDL3 did not stimulate enzyme activity, whereas the total lipid extract of HDL3 was about 1.7 times more potent than were the native HDL3 in stimulating enzyme activity. HDL2 consistently suppressed 3-hydroxy-3-methylglutaryl-CoA reductase in normal fibroblasts by 20-50% at 80 microgram HDL2 protein/ml. Mixtures of HDL2 and HDL3 suppressed when HDL2 were greater than 35% of the total HDL. The suppressive effects of HDL2 were abolished by treatment with 0.1 M cyclohexanedione and restored by regeneration of arginyl residues, suggesting an apolipoprotein-mediated suppressive mechanism. The total lipid extract of HDL2 stimulated 3-hydroxy-3-methylglutaryl-CoA reductase 2-fold at 3 microgram lipid phosphorus/ml. Moreover, HDL2 and HDL3 both stimulated 3-hydroxy-3-methylglutaryl-CoA reductase activity in receptor-negative fibroblasts. HDL2 have both the ability to suppress 3-hydroxy-3-methylglutaryl-CoA reductase activity in cells which possess low density lipoproteins receptors and to activate the enzyme in receptor-negative cells. These results show that variations in culture conditions and differences in the proportions of HDL subclasses must be considered in the interpretation of studies investigating cellular responses to HDL.

Apolipoproteins↗