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Biomedical subjects

L C Smith

Publications and source records attributed to L C Smith.

At least 181 records · Page 10Linked to original sources
PubMed1980

Effects of HDL subclasses on 3-hydroxy-3-methylglutaryl coenzyme A reductase in human fibroblasts.

Monolayer cultures of normal human fibroblasts were used to study the effects of the main subclasses of high-density lipoproteins, HDL2 and HDL3, on 3-hydroxy-3-methylglutaryl CoA reductase (EC 1.1.1.34) activity. In this system, HDL3 (d = 1.125-1.210 g/cm3) specifically induced HMG-CoA reductase activity. Evaluation of culture dynamics revealed that enzyme induction was restricted to the stationary phase of growth. When the cells were incubated with HDL2 (d = 1.063-1.125 g/cm3), suppression of reductase activity was observed. Mixtures of HDL2 and HDL3 suppressed reductase activity when HDL2 was greater than 35% of the total HDL. The suppressive effects of HDL2 were abolished by treatment with cyclohexanedione and restored by regeneration of the arginyl residues, suggesting an apoprotein-mediated suppressive mechanism. These observations show that the cellular effects of HDL depend upon the stage of cell growth and the ratio of HDL subclasses in HDL as usually isolated.

Cell Cycle↗
PubMed1980

Transport of lipid across capillary endothelium.

Fatty acids, monoacylglycerol, cholesterol, and phospholipids are taken up from blood by many different tissues. These substances, which are strongly amphipathic at physiological pH, are poorly soluble in water and neutral lipids (tyriacylglycerol nd cholesteryl ester). They are transported in blood as components of lipoprotein particles or, in the case of fatty acids, s monomers bound to albumin. Fatty acids derived from the diet are carried as triacylglycerol in chylomicrons and very low density lipoprpoteins (VLDL). Phospholipids and cholesterol absorbed from intestines are also transported in chylomicrons and VLDL. We propose that transport of fatty acids and monoacylglycerol from chylomicrons Tand VLDL) across capillary endothelium in extrahepatic tissues requires 1) conversion of chylomicron triacylglycerol to amphipathic lipids (fatty acids and monoacylglycerol) by lipoprotein lipase at the capillary surface, 2) location and lateral movement of ipolytic products in a continuous interface composed of the chylomicron surface film and the external leaflet of plasma and intracellular membranes of endothelial and parenchymal cells, and 3) removal of lipolytic products from the interface in endoplasmic reticulum where they are reesterified to traicylglycerol and accumulate between leaflets of endoplasmic reticulum. We suggest that cholesterol and phospholipids from chylomicrons, and fatty acids from plasma albumin, also cross capillary endothelium by lateral movement in cell membranes.

Capillary Permeability↗
PubMed1979

Differences in neural organization between individuals with inverted and noninverted handwriting postures.

Levy's hypothesis that movements of the distal musculature are controlled by ipsilateral motor projections in subjects with inverted writing posture was tested in a reaction-time experiment with lateralized auditory, tactual, and visual stimulation. Subjects were required to depress a response key with the left or right index finger when they detected a stimulus in either the left or right sensory field. Writers with noninverted posture responded quickest to stimuli on the same side as the responding hand in all modalities tested, whereas inverted writers showed this pattern only in auditory and tactual modalities. In the visual modality, they responded quickest to stimuli on the side opposite the responding hand. Because Levy's hypothesis predicts the latter effect in all modalities for inverted writers, it is challenged by our results, which suggest that inverted writers may be characterized by anomalous visual or visuomotor organization.

Behavior↗
PubMed1979

Suppression of 3-hydroxy-3-methylglutaryl-CoA reductase by low density lipoproteins produced in vitro by lipoprotein lipase action on nonsuppressive very low density lipoproteins.

Very low density lipoproteins (VLDL), Sf60 to 400, from normolipemic individuals do not suppress 3-hydroxy-3-methylglutaryl-CoA reductase activity in cultured normal human fibroblasts at concentrations 20-fold higher than those of low density lipoproteins (LDL) that give total suppression. To determine if these VLDL contain all of the structural elements necessary for receptor-mediated suppression, they were converted in vitro with bovine milk lipoprotein lipase to low density lipoproteins. These LDL-like lipoproteins were as effective in suppression as LDL isolated directly from plasma, with half-maximal and complete suppression at 1 and 4 microgram of cholesterol ml-1. Neither native LDL nor LDL produced in vitro suppressed receptor-negative fibroblasts. We conclude that action of lipoprotein lipase on VLDL leads to a rearrangement of lipoprotein components that permits interaction of LDL produced in vitro with the LDL-specific cell surface receptor of fibroblasts and subsequent suppression of 3-hydroxy-3-methylglutaryl-CoA reductase.

Cells, Cultured↗
PubMed1979

Reconstituted low density lipoprotein: a vehicle for the delivery of hydrophobic fluorescent probes to cells.

Previous studies have shown that the cholesteryl ester core of plasma low density lipoprotein (LDL) can be extracted with heptane and replaced with a variety of hydrophobic molecules. In the present report we use this reconstitution technique to incorporate two fluorescent probes, 3-pyrenemethyl-23, 24-dinor-5-cholen-22-oate-3 beta-yl oleate (PMCA oleate) and dioleyl fluorescein, into heptane-extracted LDL. Both fluorescent lipoprotein preparations were shown to be useful probes for visualizing the receptor-mediated endocytosis of LDL in cultured human fibroblasts. When normal fibroblasts were incubated at 37 degrees C with either of the fluorescent LDL preparations, fluorescent granules accumulated in the perinuclear region of the cell. In contrast, fibroblasts from patients with the homozygous form of familial hypercholesterolemia (FH) that lack functional LDL receptors did not accumulate visible fluorescent granules when incubated with the fluorescent reconstituted LDL. A fluorescence-activated cell sorter was used to quantify the fluorescence intensity of individual cells that had been incubated with LDL reconstituted with dioleyl fluorescein. With this technique a population of normal fibroblasts could be distinguished from a population of FH fibroblasts. The current studies demonstrate the feasibility of using fluorescent reconstituted LDL in conjunction with the cell sorter to isolate mutant cells lacking functional LDL receptors.

Cells, Cultured↗
PubMed1979

A review of the unique features of HDL apoproteins.

The human plasma high density lipoproteins (HDL) are a heterogeneous ensemble of five proteins associated with both neutral and polar lipids. The sequence of all five proteins are known. ApoA-I and apoA-II are the major protein components; apoC-I, apoC-II and apoC-III are the minor protein components. All these apoproteins spontaneously recombine with phospholipids to give stable lipid-protein complexes and freely exchange between the two major HDL subclasses, HDL2 and HDL3. In addition, ApoC-I, apoC-II, and apoC-III exchange between HDL and very low density lipoproteins. Furthermore, certain HDL apoproteins are activators for plasma enzymes that are important in lipid metabolism. ApoA-I and apoC-I activate lecithin/cholesterol acyltransferase; apoC-II is an activator of lipoprotein lipase. The regions of apoC-I and apoC-II that are involved in the activation of these enzymes have been localized with synthetic peptides. Studies of synthetic and native fragments of apoA-II, apoC-I, apoC-II, and apoC-III as well as model lipid-binding peptides have identified specific regions with structural features common to lipid-binding proteins. These special properties, which include helical potential, sequences with a critical amphipathic length, and high hydrophobicity of the nonpolar side of the amphipathic helix, are the determinants of HDL structure and metabolism.

Amino Acid Sequence↗
PubMed1978

Kinetics of rac-1-oleyl-2-[4-(3-pyrenyl)butanoyl]glycerol transfer between high density lipoproteins.

The mechanism of transfer of diglyceride between high density lipoproteins (HDL) was investigated with a pyrene-containing analogue whose fluorescent properties depend on the microscopic concentration in the lipoprotein. Transfer rates were first order, rapid (3.5s-1), and invariant over a 100-fold range of HDL concentration and over a 10-fold range of rac-1-oleyl-2-[4-(3-pyrenyl)butanoyl]glycerol concentrations. Similar behavior of the probe was observed with HDL3, although the rate was 40% slower. These results support a mechanism in which rate-limiting dissociation of the diglyceride analogue from one HDL particle into the aqueous phase precedes rapid diffusion and subsequent uptake by another such particle.

Diglycerides↗
PubMed1978

Control of 3-hydroxy-3-methylglutaryl-CoA reductase activity in cultured human fibroblasts by very low density lipoproteins of subjects with hypertriglyceridemia.

Very low density lipoproteins (VLDL) and low density lipoproteins (LDL) from human normolipemic plasma, and the VLDL, the intermediate density lipoprotein (IDL), and LDL from patients with Type III hyperlipoproteinemic plasma were tested for their abilities to suppress the activity of 3-hydroxy-3-methylglutaryl-Coenzyme A (HMG-CoA) reductase in cultured human fibroblasts from normal subjects and a Type III patient. Regulation of cholesterol synthesis in the fibroblasts of a patient with Type III hyperlipoproteinemia appears to be normal. VLDL from normal subjects, isolated by angle head ultracentrifugation (d < 1.006) or by gel filtration on BioGel A-5m, were about 5 times less effective than LDL in suppressing HMG-CoA reductase activity, based on protein content, in agreement with previous reports with normal fibroblasts. Zonal centrifugation of normal VLDL isolated by both methods showed that the VLDL contained IDL. Normal VLDL from the angle head rotor, refractionated by the zonal method, had little, if any, ability to suppress the HMG-CoA reductase activity in either normal or Type III fibroblasts. VLDL, IDL, and LDL fractionated by zonal ultracentrifugation from Type III plasma gave half-maximum inhibition at 0.2-0.5 mug of protein/ml, indistinguishable from the suppression caused by normal LDL. Type III VLDL did not suppress HMG-CoA reductase in mutant LDL receptor-negative fibroblasts. Zonally isolated VLDL obtained from one Type IV and one Type V patient gave half-maximal suppression at 5 and 0.5 mug of protein/ml, respectively. Molecular diameters and apoprotein compositions of the zonally isolated normal and Type III VLDL were similar; the major difference in composition was that Type III VLDL contained more cholesteryl esters and less triglyceride than did normal VLDL. The compositions and diameters of the Type IV and Type V VLDL were similar to normal VLDL. These findings show that the basic defect in Type III hyperlipoproteinemia is qualitatively different from the cellular defect found in familial hypercholesterolemia, since the regulation of HMG-CoA reductase activity is normal in Type III fibroblasts. The metabolic defect in hypertriglyceridemia is related to the triglyceriderich lipoproteins which, free of other lipoproteins, have an enhanced ability to interact with cultured fibroblasts to regulate HMG-CoA reductase activity. These studies suggest that, in hypertriglyceridemia, there is a mechanism for direct cellular catabolism of VLDL which is not functional for normal VLDL.

Adult↗
PubMed1978

Quantification of apoC-II and apoC-III of human very low density lipoproteins by analytical isoelectric focusing.

ApoC-II and apoC-III of human very low density lipoproteins (VLDL) have been quantified by analytical isoelectric focusing (IEF) between pH 4 and 6 in polyacrylamide gels containing 8 M urea. The isoelectric point of apoC-III0 is pH 4.93; apoC-II, pH 4.78; apoC-III1, pH 4.72, and apoC-III2, pH 4.54. ApoC-I is not found in the pH range between pH 4 and 6. Two minor peptides, apoC-IV and apoC-V, with isoelectric points of pH 4.61 and 4.44, respectively, are apoproteins not previously identified. The sensitivity (5--40 microgram) and reproducibility (+/- 8%) of this method allow quantitative analysis of apoC-II and apoC-III distribution in VLDL.

Apolipoproteins↗
PubMed1978

Lecithin:cholesterol acyltransferase activation and lipid binding by synthetic fragments of apolipoprotein C-I.

Peptide fragments of apolipoprotein C-I (apoLP-C-I) have been synthesized by solid phase methodology. After purification, each peptide showed the correct amino acid analysis and was a single band by polyacrylamide gel electrophoresis. In density gradient ultracentrifugation with vesicles of dimyristoyl phosphatidylcholine, peptide fragments 32-57, 24-57, and 17-57 formed stable complexes while 39-57 did not. With mixed vesicles of dimyristoyl phosphatidylcholine-cholesterol 20 micrometer of the fragments 32-57, 24-57 and 17-57 stimulated lecithin:cholesterol acyltransferase (LCAT) activity 50, 60, and 100%, respectively, of the value found for apolipoprotein C-I, while fragment 39-57 was inactive. The results indicate that residues 17-57 contain the structural requirements for LCAT activation by apoLP-C-I, and that residues 32-57 represent one of the major phospholipid-binding regions of apoLP-C-I.

Amino Acid Sequence↗