bluuebunny/crossref_metadata_2025 huggingface dataset:
Specific sorbent of apolipoprotein B-containing lipoproteins for plasmapheresis. Characterization and experimental use in hypercholesterolemic rabbits.
Authors: Yokoyama S , Hayashi R , Kikkawa T , Tani N , Takada S , Hatanaka K , Yamamoto A
Published: June 2011

Dextran sulfate was covalently bound to cellulose beads (LA01) and used as the specific sorbent of apolipoprotein B-containing lipoproteins. The binding profiles of very low density, low density, and high density lipoproteins to the material were consistent with simple Langmuir adsorption isotherms. Although the affinities of all lipoproteins for the sorbent were roughly similar, the surface area available for high density lipoprotein binding was much smaller than that for very low and low density lipoproteins (29, 927, and 1934 m2/liter of swollen beads, respectively). When human plasma was passed through a sorbent column, very low and low density lipoproteins were selectively adsorbed, while almost all the high density lipoproteins were recovered from the column together with the other major plasma components. An extracorporeal circulation system with this sorbent was used for the treatment of hypercholesterolemic rabbits (diet-induced and homozygous WHHL). With a 25 ml sorbent column, very low and low density lipoproteins were selectively removed from rabbit plasma, resulting in a reduction of plasma cholesterol concentration by 300 mg/dl.
Cardiac and peripheral lymph lipoproteins in dogs fed cholesterol and saturated fat.
Authors: Julien P , Fong B , Angel A
Published: June 2011

In normal dogs fed diets containing 1% cholesterol and 30% saturated fat, cholesterol concentration increased in plasma, cardiac lymph, and peripheral lymph. The increase in cardiac lymph cholesterol always exceeded that of peripheral lymph. At plateau, LDL cholesterol increased in plasma and peripheral lymph but not in cardiac lymph; HDL2 cholesterol increased in all compartments and HDLc appeared in plasma (86.6 +/- 22.2 mg cholesterol/dl) with cholesterol feeding. In cardiac and peripheral lymph, a lipoprotein fraction with ultracentrifugal and pevikon electrophoretic characteristics of plasma HDLc (designated HDL 1.063) was isolated and contained 9.4 +/- 1.3 and 11.9 +/- 6.8 mg cholesterol/dl respectively. The lymph/plasma (L/P) ratios of lipoproteins were generally higher in cardiac lymph compared to peripheral lymph. In cholesterol-fed dogs, the L/P ratios of LDL were reduced compared to control dogs while HDL2 was unaffected, indicating selective interstitial distribution of lipoproteins. On agarose electrophoresis, lymph lipoproteins migrated faster than plasma lipoproteins, suggesting subtle changes in the structure and/or composition, findings consistent with the suggestion that lipoproteins undergo modification in the interstitial space. Thus, each interstitial space, as reflected by lymph, has a characteristic mixture of lipoproteins that can be altered by a high cholesterol diet in a manner that is unique to each particular tissue compartment.
Characterization of antigenic determinants of human apolipoprotein B. Distribution on tryptic fragments of low density lipoprotein.
Authors: Theolis R , Weech P K , Marcel Y L , Milne R W
Published: June 2011

In the hope of obtaining useful probes to study the structure of human apolipoprotein B (apo B), we characterized monoclonal antibodies against low density lipoprotein (LDL). We examined the distribution of their corresponding antigenic determinants on tryptic fragments of LDL separated by monodimensional (SDS) or two-dimensional electrophoresis. Each antibody reacted with several different fragments even when the proteolysis was apparently complete. A peptide of 125,000 daltons was the smallest fragment recognized by all the antibodies. The antibody, 2D8, which cross-reacts with apo B-48 and 3A8 which blocks the LDL pathway both reacted with the same 43,000 dalton fragment. Two other antibodies, 3F5 and 4G3, previously shown to be close together in LDL, also appeared close together in the primary structure of apo B. A determinant present on apo B-26 (1D1) was dissociated from all others examined on fragments of less than 125,000. Similarities in the patterns of reactivities with LDL-tryptic fragments between certain monoclonal antibodies and the lectins Concanavalin A and Limax flavus agglutinin indicated the proximity of the corresponding antigenic determinants to carbohydrate moieties. Competition studies suggested that the two major carbohydrate chains of LDL do not participate in the determinants themselves.
Double pre-beta lipoprotein. Isolation and characterization of the two population of very low density lipoproteins by zonal ultracentrifugation.
Authors: Manzato E , Pagnan A , Ziron L , Gasparotto A , Braggion M
Published: June 2011

Two electrophoretic populations of very low density lipoprotein are frequently observed in both normal and hyperlipidemic human sera. This gives the appearance of two pre-beta bands, called the "double pre-beta lipoprotein phenomenon." The slow and fast bands forming the double pre-beta lipoprotein were isolated by ultracentrifugation in the zonal rotor under rate flotation conditions. On the basis of the effluent position, the slow band showed a lower flotation coefficient than the fast one. The lipid and apoprotein composition of the two very low density lipoprotein populations isolated by zonal ultracentrifugation was in close agreement with those obtained by preparative agarose gel electrophoresis. The slow and fast pre-beta fractions had the same triglyceride fatty acid composition. Both fractions contained only higher molecular weight apo B-100, while the slow fraction was relatively enriched in apo E and apo C-III. Since the slow pre-beta fraction showed the typical properties of the remnant particle, studying this fraction could clarify the possible relationship between remnant lipoproteins and atherogenesis. Zonal rotor ultracentrifugation may be useful to characterize remnant particles in normal and hyperlipidemic subjects.
Mobilization of stored triglycerides from macrophages as free fatty acids.
Authors: von Hodenberg E , Khoo J C , Jensen D , Witztum J L , Steinberg D
Published: June 2011

Because many or most lipid-laden foam cells in atheromas and in xanthomas derive from macrophages, it is important to understand how they accumulate lipids and how they can divest themselves of lipids. The mobilization of stored triglycerides from macrophages was studied in cell cultures. Mouse resident peritoneal macrophages and J774 macrophages increased their triglyceride content six- to tenfold during a 24-hour incubation with free fatty acids complexed to albumin. Subsequent incubation in fresh medium containing free fatty acid-poor albumin was accompanied by a fall in cell triglyceride content (50% in 20 hours) and a corresponding increase in medium-free fatty acid. Release of free fatty acid was linear as a function of time, provided fresh medium was added hourly. When medium was not changed, release rates fell off rapidly, probably due to re-uptake of released free fatty acid. Chloroquine did not affect the rate of free fatty acid release. The results suggest that macrophages-foam cells can reduce their triglyceride stores via the action of a nonlysosomal (presumably cytoplasmic) neutral triglyceride lipase.
Lesion regression and protein synthesis in rabbits after removal of dietary cholesterol.
Authors: Langner R O , Bement C L
Published: June 2011

This study was conducted to determine whether atherosclerotic lesions regressed in rabbits that were placed on a normal, low cholesterol diet. Rabbits (n = 16) were fed a 2% cholesterol diet for 90 days; nine rabbits were killed, and the remaining seven rabbits were fed a low cholesterol diet for 7 months.. Appropriate noncholesterol-fed controls were used with each group. After 90 days of cholesterol feeding, the rabbits' thoracic aortas showed severe lesions, accompanied by significantly elevated rates of collagen and noncollagen protein synthesis and significantly increased amounts of cholesterol and soluble collagen. Cholesterol-fed rabbits that were fed a normal diet for 7 months had normal serum cholesterol levels, but exhibited extensive aortic lesions with elevated cholesterol content. Biochemically, the rate of aortic collagen synthesis remained significantly elevated above control values; however, the collagen content was not different from controls. This lack of collagen accumulation in the presence of a prolonged increase in synthetic rate was surprising. One possible explanation is that in the diseased aorta there is a rapid turnover of collagen, resulting in a redistribution or remodeling of the connective tissue, rather than an increased accumulation.
Serum lipoprotein and lipoprotein lipase in overweight, type II diabetics during and after supplemented fasting.
Authors: Vessby B , Selinus I , Lithell H
Published: June 2011

Poorly controlled, obese, Type II diabetics were studied before, during, and 3 months after a weight reduction program that used supplemented fasting (200 kcal or 0.9 MJ/day). During fasting, the very low density lipoprotein (VLDL) triglycerides (TG) decreased, as did the adipose tissue lipoprotein lipase (AT-LPLA) and skeletal muscle lipoprotein lipase (SM-LPLA) activities. Three months later VLDL TG remained low (-59%), while high density lipoprotein cholesterol was higher (+11%) and blood glucose control improved compared with values on admission. The fractional removal rate (K2) at the i.v. fat tolerance test (IVFTT) and the SM-LPLA were unchanged, while AT-LPLA (expressed per gram of wet weight, but not as whole-body AT-LPLA) increased by 25%. There were no significant correlations between AT-LPLA and the lipoprotein TG concentrations or K2-IVFTT, although there were significant positive correlations between SM-LPLA and K2-IVFtt, both on admission and after body weight stabilization. This may indicate that SM-LPLA is more directly related to the capacity to remove lipoprotein TG, at least in obese diabetic patients. K2-IVFTT was inversely correlated to the VLDL TG and cholesterol concentrations both before and 3 months after fasting. Because both SM-LPLA and K2-IVFTT were unchanged after body weight reduction, the change in VLDL TG may be mainly due to a reduced rate of lipoprotein synthesis.
Prostacyclin production by endothelial cells. Effects of sera from normal and hyperlipidemic subjects.
Authors: Tremoli E , Jaffe E A , Goldman K T , Weksler B B
Published: June 2011

The influence of hyperlipidemic sera on prostacyclin (PGI2) production by cultured endothelial cells was assessed by comparing sera from three types of hyperlipidemias with sera from normal subjects. Sera prepared from normal whole blood (WBS), platelet-rich plasma (PRPS), and platelet-poor plasma (PPPS) were also compared. Bovine aortic endothelial cells (BAEC) incubated with 25% WBS increased PGI2 synthesis significantly within 1 hour, with little further increase by 16 hours; human umbilical vein endothelial cells (HUEC) incubated with 25% WBS for 1 hour showed no elevation in PGI2, whereas PGI2 levels increased substantially after 16 hours. PPPS and PRPS stimulated PGI2 synthesis by BAEC equally at 1 hour. However, there was no rise in PGI2 after PPPS in HUEC; PGI2 rose after 16 hours with PRPS and rose further with WBS after 16 hours. Since WBS best enhanced PGI2 production in human endothelial cells, it was chosen for comparison of the effects of hyperlipidemic and normolipidemic sera. PGI2 synthesis by HUEC significantly increased upon incubation with WBS from Types IIb and IV patients in comparison to WBS from Type IIa hypercholesterolemic patients or normal controls. In contrast, WBS from all these hyperlipidemic subjects stimulated PGI2 synthesis by BAEC similarly to WBS from controls. We conclude that incubation of human endothelial cells with WBS containing high levels of atherogenic lipoproteins does not reduce PGI2 formation by the cells. Moreover, the time course and the contribution of lipid, plasma, or cellular factors to PGI2 formation vary according to the cell type tested. Caution should be exercised in extrapolating results achieved with serum and cells from the same species to other settings.
Differential effects of human interleukin-1 on growth of human fibroblasts and vascular smooth muscle cells.
Authors: Libby P , Wyler D J , Janicka M W , Dinarello C A
Published: June 2011

Monocyte products probably play a role in the initiation of smooth muscle cell proliferation in the arterial wall early in atherogenesis. Several groups have described mitogenic activity for arterial smooth muscle cells that is elaborated by mononuclear phagocytes (macrophage-derived growth factor). However, the biochemical nature of this mitogenic activity is unknown. Interleukin-1 (IL-1) is a well-characterized monocyte product that activates the growth of mitogen-stimulated lymphocytes and promotes the growth of fibroblasts. We tested whether IL-1 also affects the growth of arterial smooth muscle cells and might account for some of the mitogenic activity produced by activated monocytes. Highly purified human IL-1 did stimulate the growth of human fibroblasts of either adult or fetal origin. However, under identical conditions, IL-1 lacked significant mitogenic effects on human, bovine, rabbit, or canine arterial smooth muscle cells. This mediator also failed to stimulate the growth of cultured human or bovine vascular endothelial cells, another cell type that may respond to macrophage-derived growth factor. Interleukin-1 did not render smooth muscle cells competent to divide in the presence of plasma factors such as insulin (10(-6) M), or when growth of muscle cells was limited by incubation in a low (2%) concentration of serum. This monokine also failed to increase the mitogenic effect of purified platelet-derived growth factor on arterial smooth muscle cells incubated in serum-free medium. Thus, cultured human arterial smooth muscle cells differ from fibroblasts and lymphocytes in their response to human IL-1.(ABSTRACT TRUNCATED AT 250 WORDS)
Characterization of a human hepatic receptor for high density lipoproteins.
Authors: Hoeg J M , Demosky S J , Edge S B , Gregg R E , Osborne J C , Brewer H B
Published: June 2011

Characterization of the membrane receptor for the low density lipoproteins (LDL) has led to insights into cellular receptor physiology as well as mammalian lipid transport. Result with LDL have stimulated the search for specific receptors for other plasma lipoproteins. Receptors for high density lipoproteins (HDL) have been identified in human fibroblasts and smooth muscle cells. Specificity for this receptor has been difficult to define since normal HDL contains several apolipoproteins, and particles containing apolipoproteins B and E have been shown to compete for HDL binding. In the present study, we demonstrate that HDL isolated from a patient devoid of apolipoprotein E was bound specifically by human hepatic membranes. This binding reached saturation within 2 hours and was EDTA-resistant. Assuming a single receptor model, we found that 2.9 x 10(15) receptors/mg membrane protein bound with an affinity KD = 3.5 x 10(-7) M at 0 to 4 degrees C and KD = 1.9 x 10(-7) M at 37 degrees C. The binding was effectively competed with intact HDL3, with HDL3 that had undergone selective arginine and lysine residue modification, and with antibodies to apolipoproteins A-I and A-II. However, LDL, asialofetuin, and HDL3 which had undergone tyrosine modification by nitration, and anti-apolipoprotein B did not compete with apo A-I HDL binding. In contrast to LDL binding, the human hepatoma cell line, HEPG2, increased HDL binding with cholesterol loading that was specific for HDL3. Thus, hepatic tissue can modulate its recognition of HDL. Finally, hepatic membranes from a patient lacking normal hepatic LDL receptors bound apo A-I HDL normally. These data indicate that a saturable, specific regulatable receptor for apo E-free HDL is present in human liver.
Postheparin plasma triglyceride lipases. Relationships with very low density lipoprotein triglyceride and high density lipoprotein2 cholesterol.
Authors: Applebaum-Bowden D , Haffner S M , Wahl P W , Hoover J J , Warnick G R , Albers J J , Hazzard W R
Published: June 2011

Hepatic triglyceride (HTGL) and lipoprotein lipase (LPL) probably have major roles in the removal of triglyceride from triglyceride-rich lipoprotein and in the formation of high density lipoprotein (HDL). However, no population-based study of their activity and relationship to lipoprotein lipid levels has been reported. To determine these relationships, we recalled 33 men and 17 women of a randomly selected sample of the Lipid Research Clinics Pacific Northwest Bell Telephone Company Health Survey. The subjects were 53 +/- 7 years old (mean +/- SD) with total triglyceride levels of 120 +/- 57 mg/dl and total cholesterol levels of 224 +/- 35 mg/dl. Postheparin plasma LPL activity (127 +/- 61 nmol/min/ml) was not significantly correlated with either age, sex, or adiposity. In contrast, HTGL activity was significantly higher in men (235 +/- 84 nmol/min/ml) than women (170 +/- 91 nmol/min/ml, p less than 0.02), and was correlated with age in men and with adiposity in women. In both men and women, HTGL activity was related positively with VLDL triglyceride and inversely with HDL2 cholesterol. When the association between HTGL activity and VLDL triglyceride was examined with values from men and women pooled, the relationship was not weakened after adjustment for the linear effect of sex, adiposity, LPL, or HDL2 cholesterol.
Effects of apoprotein E polymorphism on serum lipoprotein concentration.
Authors: Robertson F W , Cumming A M
Published: June 2011

Serum concentrations for the cholesterol and triglyceride content of very low density lipoprotein (VLDL) and the cholesterol content of intermediate density lipoprotein (IDL), low density lipoprotein (LDL), and the high density lipoprotein fractions (HDL2), HDL3) were measured in 337 men and women with the apo E phenotype who were born in the Grampian Region of North East Scotland. The subjects' ages ranged from 45 to 60 years (mean, 53 years). Cigarette smoking and alcohol consumption were recorded and were shown to have minor, but suggestive, effects on serum lipoprotein concentrations. The LDL concentration of E-3/2 subjects was compared with the commonest category, E-3/3, and was found to be about 20% lower in men and about 12% lower in women, in whom the VLDL and IDL concentrations were substantially elevated. In E-4/3 women, the serum LDL concentration was about 10% higher than in E-3/3 women, and the other differences were minor. The effect of allelic substitutions differed between the sexes. The relations between C-peptide ratios and serum lipoprotein concentration differed by gender and phenotype. A comparison with earlier estimates of LDL heritability suggests that in this population, a substantial fraction of the genetic variance of LDL is due to segregation at the apo E locus. The implications of this significant finding are discussed.
Pulsatile flow and atherosclerosis in the human carotid bifurcation. Positive correlation between plaque location and low oscillating shear stress.
Authors: Ku D N , Giddens D P , Zarins C K , Glagov S
Published: June 2011

Fluid velocities were measured by laser Doppler velocimetry under conditions of pulsatile flow in a scale model of the human carotid bifurcation. Flow velocity and wall shear stress at five axial and four circumferential positions were compared with intimal plaque thickness at corresponding locations in carotid bifurcations obtained from cadavers. Velocities and wall shear stresses during diastole were similar to those found previously under steady flow conditions, but these quantities oscillated in both magnitude and direction during the systolic phase. At the inner wall of the internal carotid sinus, in the region of the flow divider, wall shear stress was highest (systole = 41 dynes/cm2, diastole = 10 dynes/cm2, mean = 17 dynes/cm2) and remained unidirectional during systole. Intimal thickening in this location was minimal. At the outer wall of the carotid sinus where intimal plaques were thickest, mean shear stress was low (-0.5 dynes/cm2) but the instantaneous shear stress oscillated between -7 and +4 dynes/cm2. Along the side walls of the sinus, intimal plaque thickness was greater than in the region of the flow divider and circumferential oscillations of shear stress were prominent. With all 20 axial and circumferential measurement locations considered, strong correlations were found between intimal thickness and the reciprocal of maximum shear stress (r = 0.90, p less than 0.0005) or the reciprocal of mean shear stress (r = 0.82, p less than 0.001). An index which takes into account oscillations of wall shear also correlated strongly with intimal thickness (r = 0.82, p less than 0.001). When only the inner wall and outer wall positions were taken into account, correlations of lesion thickness with the inverse of maximum wall shear and mean wall shear were 0.94 (p less than 0.001) and 0.95 (p less than 0.001), respectively, and with the oscillatory shear index, 0.93 (p less than 0.001). These studies confirm earlier findings under steady flow conditions that plaques tend to form in areas of low, rather than high, shear stress, but indicate in addition that marked oscillations in the direction of wall shear may enhance atherogenesis.
Regulation of high density lipoprotein binding activity of aortic endothelial cells by treatment with acetylated low density lipoprotein.
Authors: Brinton E A , Kenagy R D , Oram J F , Bierman E L
Published: June 2011

High density lipoprotein (HDL) binding to human fibroblasts and arterial smooth muscle cells is up-regulated when sterol is delivered to cells in the form of nonlipoprotein cholesterol or low density lipoprotein (LDL). Results from the present study show that the HDL binding activity of aortic endothelial cells is up-regulated when cholesterol in the form of acetylated LDL (AcLDL) is delivered to cells via the "scavenger" lipoprotein receptor pathway. AcLDL treatment led to a dose-dependent, but saturable, increase in HDL binding to cultured bovine aortic endothelial cells that was reversed when cells were treated with lipoprotein-deficient serum. The AcLDL-mediated enhancement in HDL binding activity was inhibited by cycloheximide, suggesting the involvement of protein synthesis. This enhancement was associated with an increased cell cholesterol content, a suppressed rate of cholesterol synthesis, and an increased rate of cholesterol ester formation. Kinetic analysis of HDL binding showed that AcLDL treatment caused an increase in the apparent number of high-affinity binding sites (Kd approximately 3 micrograms/ml HDL protein). Competition and direct binding studies revealed that the inducible binding sites exhibited relative specificity for HDL over LDL and AcLDL. Thus, aortic endothelial cells appear to possess specific receptors for HDL that may function to facilitate HDL-mediated removal from cells of excess cholesterol internalized by the scavenger receptor pathway.
Effect of acetylsalicylic acid on pulmonary arteriosclerosis induced by a one-year Dirofilaria immitis infection.
Authors: Rawlings C A , Keith J C , Schaub R G
Published: June 2011

The ability of aspirin to block arteriosclerosis that developed in response to chronic, low-level injury to pulmonary arteries was evaluated in 21 dogs during their 1-year infection with Dirofilaria immitis. Three groups, with seven dogs in each group, were studied before and after sustained injury produced by the transplantation of 28 adult Dirofilaria immitis into each dog. Group A received no treatment and served as controls; Group B received no treatment for 6 months and then received 7 mg/kg of aspirin daily) for 6 months; Group C received 7 mg/kg of aspirin daily for the entire year. The pulmonary arterial response was evaluated by hemodynamic and arteriographic studies at 6 and 12 months and by scanning electron microscopy at the end of the 12-month study. All groups developed a similar, mild pulmonary hypertension. The arteriographic changes of dilation and flow obstruction were worse in Groups A and B than in Group C at 6 months, and at 12 months both Groups B and C were less obstructed than Group A. Scanning electron microscopy revealed large, complex myointimal proliferations in Group A, whereas the two aspirin-treated groups had smaller, less complex lesions that covered a much smaller surface area. We concluded that: 1) aspirin markedly reduced the microscopic and macroscopic arteriosclerosis in Groups B and C; 2) aspirin in Group B not only arrested further development but also permitted resolution of arteriosclerosis while the arteries were still being injured.
Aortic atherosclerosis in pigs with heterozygous von Willebrand disease. Comparison with homozygous von Willebrand and normal pigs.
Authors: Badimon L , Steele P , Badimon J J , Bowie E J , Fuster V
Published: June 2011

We have reported that pigs with severe homozygous von Willebrand disease (vWd) are resistant to spontaneous and high fat, high cholesterol, diet-induced atherosclerosis. In this study we report the quantitation of aortic atherosclerotic plaques in three groups of pigs fed with a high fat, high cholesterol (2%) diet from age 3 to 9 months. Nine normal pigs (normal factor VIII antigen, VIII R:AG, and ristocetin co-factor, VIII:RWF) had a mean of 21% atherosclerotic involvement of the distal aortic surface and a 4.5% mean involvement of the entire aorta. Five homozygous vWd pigs (undetected VIII R:AG and VIII:RWF) had a mean of 4.2% atherosclerotic involvement of the distal aortic surface and 1.2% involvement of the entire aorta (p less than 0.01, rank sum test). Five heterozygous vWd pigs (approximately 35% VIII R:AG and VIII:RWF) had a mean of 25% atherosclerotic involvement of the distal aortic surface and 6% involvement of the entire aorta; the results were not significantly different from those in the normal pigs. We concluded that resistance to atherosclerosis is not found in animals with moderate reduction of VIII R:AG and VIII:RWF. This may have implications for humans, since in human vWd both factors are almost always present.
Apo E-containing lipoproteins in low or high density lipoprotein deficiency.
Authors: Gibson J C , Rubinstein A , Brown W V , Ginsberg H N , Greten H , Norum R , Kayden H
Published: June 2011

Apolipoprotein (apo) E-containing subfractions of very low density lipoprotein (VLDL), intermediate density lipoprotein (IDL), and high density lipoprotein (HDL) have been described in normolipidemic and hyperlipidemic subjects. These lipoproteins exist, however, in the presence of large amounts of apo A-I- and apo B-containing lipoproteins so that it has been difficult to assess the independence of these apo E-containing subclasses from the major lipoprotein classes. The present study has approached this question by taking advantage of three hypolipidemic states in which one or more of the major apolipoproteins is deficient or absent. After separating lipoproteins from whole plasma by molecular sieve chromatography followed by radioimmunoassay of column fractions, we found that two subjects with abetalipoproteinemia had no apo E-containing lipoproteins the size of VLDL or IDL and all the plasma apo E was in a fraction of large HDL. Two subjects with Tangier disease and two with familial apo A-I/C-III deficiency had extremely low levels of HDL cholesterol and of apo A-I-containing lipoproteins. In spite of the absence of classical HDL, a major fraction of apo E-containing lipoproteins was reproducibly observed at the elution volume characteristic of large HDL and was identical to that found in normal subjects. These data thus suggest the existence of apo E-containing lipoproteins that are the size of HDL and are not dependent upon the presence of either apo B or apo A-I. While studies in normal subjects indicate that apo E is associated with other apolipoproteins in HDL, further investigations will be needed to determine the full composition of these apo E-containing lipoproteins in the lipoprotein-deficient patients described in this report.
Topographic study of sudanophilic lesions in cholesterol-fed minipigs by image analysis.
Authors: Cornhill J F , Barrett W A , Herderick E E , Mahley R W , Fry D L
Published: June 2011

A new morphometric technique using image analysis has been developed to express the topographic distribution of atherosclerotic lesions in unambiguous statistical terms. Computer-stored images of opened Sudan IV-stained aortas and iliac and coronary arteries from hypercholesterolemic minipigs (n = 39) were used in this study. The image processing methods included transformation of the data to standard templates, automated image segmentation, and creation of probability-of-occurrence maps. These maps have shown that sudanophilic lesions are localized with a characteristic topography along the aortas and iliac and coronary arteries. Areas of high probability are associated with the entrance regions of vessels and the lateral leading edges of the major flow dividers. Regions immediately distal to large branches were found to be areas of low probability. Despite the association of areas of sudanophilia with entrance regions and branch points, a major portion of sudanophilic lesions was not associated with any orifice region (e.g., ductus scar, dorsolateral surface of abdominal aorta, and ventral surface of terminal aorta). The present study provides the necessary information for the development of a rational sampling strategy for the experimental study of the distribution of localizing factors (e.g., hemodynamic, biochemical, cellular, mass transport, histological) and their relationships to putative atherogenic mechanisms.
Platelet interactions with Dacron vascular grafts. A model of acute thrombosis in baboons.
Authors: Hanson S R , Kotze H F , Savage B , Harker L A
Published: June 2011

We have developed a model of acute Dacron graft thrombosis in baboons in order to assess platelet alterations secondary to arterial thrombus formation. In this model, thrombus formation was initiated by Dacron vascular grafts inserted as extension segments into chronic arteriovenous Silastic shunts. Following platelet labeling with 111In-oxine, platelet deposition was measured for 1 hour following blood contact under arterial flow conditions using a scintillation camera. Graft platelet activity rapidly increased 40- to 50-fold, plateauing by 1 hour. All grafts produced equivalent reductions in circulating platelet count and blood 111In-platelet radioactivity, demonstrating that the labeled cells were functionally equivalent to the total platelet population. After graft placement, the remaining platelets survived normally. Acute platelet deposition was equivalent on grafts placed 1, 24, 48, and 72 hours following injection of the labeled cells, indicating that a variable delay between platelet labeling and graft imaging was without detectable consequence. Platelet destruction by the graft produced a tenfold increase in plasma levels of platelet factor 4 (PF4) and beta-thromboglobulin (beta TG) but did not modify either the alpha-granule (PF4, beta TG) or dense granule (ADP, ATP) contents of circulating platelets.
Selective removal of low density lipoprotein by plasmapheresis in familial hypercholesterolemia.
Authors: Yokoyama S , Hayashi R , Satani M , Yamamoto A
Published: June 2011

Plasma lipoproteins were selectively removed from familial hypercholesterolemic patients by using two types of plasmapheresis: double-membrane filtration and selective adsorption of very low density lipoproteins (VLDL) and low density lipoproteins (LDL). In both techniques, plasma was separated from blood cells by using hollow-fiber filters, and 100% of the VLDL and LDL was recovered in the filtrate. In double-membrane filtration, the second hollow-fiber filter trapped 84% of LDL + VLDL, 48% of high density lipoprotein (HDL), 24% of albumin, and 46% of the remaining plasma protein. By treating 3 liters of plasma from a patient weighing 60 kg, 60% of the LDL and 30% to 40% of the HDL were removed as a result of an exponential decay of each component with the respective trapping coefficients. When dextran sulfate-cellulose was used as a LDL sorbent, there was only loss of LDL and VLDL, and no loss of any other major plasma component or of HDL. The sorbent column (400 ml) was saturated with 7.5 g of LDL cholesterol by treatment with 3.5 liters of plasma; the maximum reduction of LDL cholesterol was thus about 300 mg/dl for the patient weighing 60 kg. No serious side effects were observed during the long-term trials (19 to 27 months for four patients on double-membrane filtration and 10 months for the two patients on dextran sulfate-cellulose treatment.
Subendothelial proteins and platelet adhesion. von Willebrand factor and fibronectin, not thrombospondin, are involved in platelet adhesion to extracellular matrix of human vascular endothelial cells.
Authors: Houdijk W P , de Groot P G , Nievelstein P F , Sakariassen K S , Sixma J J
Published: June 2011

Endothelial cell matrix contained von Willebrand factor (VWF), fibronectin, and thrombospondin. The role of these proteins in the adhesion of platelets was investigated by preincubation of the matrix with specific antibodies and subsequent perfusion with human blood. When perfusions were performed with platelets in a human albumin solution (HAS) platelet adhesion was similar to that with normal plasma, indicating that proteins in the matrix can fully support adhesion. Preincubation of the matrix with a monoclonal antibody to VWF and perfusion with HAS showed a nearly complete inhibition of platelet adhesion at 1300 s-1, indicating a role for matrix-bound VWF at high shear rates and no requirement for VWF in plasma. Preincubation of the matrix with antihuman fibronectin F(ab')2 showed a slight inhibition of adhesion. The same result was obtained with perfusions with fibronectin-free plasma, and an untreated matrix. Preincubation with antifibronectin F(ab')2 and perfusion with fibronectin-free plasma showed a significant inhibition of platelet adhesion at all shear rates. These results indicate that fibronectin is required for adhesion at all shear rates. Preincubation of the matrix with different antibodies against human platelet thrombospondin showed no inhibition of platelet adhesion at all wall shear rates. Thrombospondin in the matrix is evidently not required for platelet adhesion.
Effects of platelet-derived growth factor on 3-hydroxy-3-methylglutaryl coenzyme A reductase activity in cultured human fibroblasts.
Authors: Fairbanks K P , Witte L D , Goodman D S
Published: June 2011

Studies were conducted to delineate in detail the effects of platelet-derived growth factor (PDGF) upon 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity in cultured human fibroblasts. PDGF has a variety of coordinated effects on low density lipoprotein (LDL) and cholesterol metabolism and on DNA synthesis in these cells. We have previously shown that there is a critical time period, 10 to 20 hours after PDGF addition to quiescent cells, when mevalonate (MVA) is required for DNA synthesis to occur. The present studies demonstrate that PDGF produces a biphasic stimulation of reductase activity in cultured fibroblasts: a peak at 4 to 6 hours, followed by a decline, and then a second smaller peak at 24 hours, concurrent with DNA synthesis. The stimulation of both peaks was PDGF concentration-dependent, although quantitative differences were observed. Proportionally similar, LDL concentration-dependent reductions in both peaks of reductase activity were also seen. PDGF stimulated cholesterol synthesis from acetate in intact cells reaching peak values after 24 to 28 hours. The two peaks of reductase activity stimulated by PDGF neither coincide with the critical time period when mevalonate is needed for later DNA synthesis to occur, nor reflect the pattern of cholesterol synthesis within the cells.
Chlorpromazine inhibits arterial ACAT and reduces arterial cholesterol and cholesteryl ester accumulation in cholesterol-fed rabbits.
Authors: Bell F P , Schaub R G
Published: June 2011

Rabbits were fed an atherogenic diet concurrently with chlorpromazine for 2 weeks (10 mg/kg/day in the diet) or 12 weeks (20 mg/kg/day given orally in a single capsule). After 2 weeks, arterial acyl CoA: cholesterol acyltransferase (ACAT) activity tended to be reduced by chlorpromazine treatment with no affect on net arterial cholesterol. After 12 weeks of treatment, arterial ACAT activity was significantly reduced and was paralleled by a reduction in net arterial cholesterol, a reduction in the esterified cholesterol/unesterified cholesterol ratio, and a reduction in lipid staining intensity as determined histologically with oil red O staining of aortic cross sections. Paradoxically, there was no histological evidence for a reduction in the size of atheromatous lesions with chlorpromazine treatment as determined by morphometric analysis of tissue cross sections. The results support the hypothesis that the increased esterification of cholesterol that characteristically accompanies the atherogenic process serves as a biochemical trapping mechanism for cholesterol entering the vessel wall and suggest that regulation of the enzyme in vivo can reduce the net accumulation of arterial cholesterol.
Presence of multiple subpopulations of lipoproteins of intermediate density in normal subjects.
Authors: Musliner T A , Giotas C , Krauss R M
Published: June 2011

Analysis of human plasma by gradient gel electrophoresis (GGE) revealed multiple distinct bands in the size range between very low density (VLDL) and low density lipoproteins (LDL). GGE of intermediate density lipoproteins (IDL) from normal subjects consistently showed two major bands with particle diameters in the range of 275 to 300 A. The larger, usually predominant subspecies was designated IDL-1, and the smaller, IDL-2. GGE also demonstrated two sizes of subpopulations in the VLDL density range in all subjects. A discontinuous nonequilibrium density gradient ultracentrifugation technique was devised to isolate a series of fractions containing progressively smaller lipoproteins. Successive fractions showed progressive enrichment in cholesteryl-esters, depletion of triglyceride, slower migration on agarose, and depletion of apo E and the C apolipoproteins relative to apo B. The IDL-1 and IDL-2 subspecies appeared to represent two distributions of lipoproteins overlapping in size (particle diameter 280-300 A and 268-284 A, respectively) and buoyant density (d = 1.008-1.022 g/ml and d = 1.013-1.028 g/ml, respectively). Plasma from two patients with dysbetalipoproteinemia showed predominant increases in fractions normally containing small VLDL and IDL-1, while patients with severe hypertriglyceridemia had increases primarily in levels of larger VLDL. Heterogeneity of lipoprotein precursors could account for the discrete subspecies of LDL observed in human plasma.
Expression of factor V on human umbilical vein endothelial cells is modulated by cell injury.
Authors: Annamalai A E , Stewart G J , Hansel B , Memoli M , Chiu H C , Manuel D W , Doshi K , Colman R W
Published: June 2011

Since human endothelial cells synthesize Factor V but do not secrete it into the medium, we studied the effects of cell injury on the availability of Factor V at the surface of these cells. Human umbilical vein endothelial cells (HUVEC), grown to confluency and incubated with human 125I Factor Va, specifically bound 5000 to 7000 molecules per cell. In the absence of added Va, no antigen was detected on adherent HUVEC with either labeled anti-V(Va) monoclonal or polyclonal IgG. However, exogenous Va, not V, prebound to these cells allows binding of labeled 125I anti-V(Va). Immunodectectibility of bovine Factor V contributed by fetal calf serum in the concentration used in cultures is less than 0.1% of that detected in human plasma. HUVEC, suspended by scraping from dishes, specifically bound 4000 molecules/cell of 125/I monoclonal IgG against V(Va). Although undisturbed cells excluded trypan blue, dye uptake by many of the suspended HUVEC indicated cell injury. Quantitation of injury by 51Cr release after scraping followed by multiple passages through an 18 g needle showed that 51Cr release increased with number of manipulations up to 60% and was observed almost immediately after manipulation. We suggest that little Factor V(Va) is present on the surface of intact adherent HUVEC. However, mechanical injury to HUVEC released or exposed endogenous Factor V(Va), resulting in expression of V that might mediate Factor Xa binding as well as activation of protein C by thrombin. Thus, injured, but not intact, HUVEC could participate in both promoting and limiting blood coagulation.
Endothelial cell monolayer integrity. I. Characterization of dense peripheral band of microfilaments.
Authors: Wong M K , Gotlieb A I
Published: June 2011

Although the endothelial cell (EC) cytoskeleton has been studied both in vitro and in vivo, little is known about its role in endothelial integrity. We have previously suggested that specific EC microfilament (MF) structure, which we have termed the "dense peripheral band (DPB)," may play a major role in this process. We have extended our studies to characterize this structure in pig aortic ECs in vitro. During the growth of EC cultures, the DPB appears only when the cultures have attained confluency. Using double fluorescent labeling, we found that alpha-actinin, myosin, and tropomyosin colocalized with the F-actin making up the DPB. Occasional microtubules were present in this region, although there was no preferred association between microtubules and the DPB. Colocalization studies revealed vinculin plaques at the cell-cell interface. Thin MFs extended from the DPB into the cytoplasmic side of these plaques. The DPB was completely disrupted by low dose cytochalasin B within 30 minutes, whereas many central MF bundles were still present at 24 hours. The results of this study suggest that the DPB is a distinct structure in the confluent EC monolayer and is closely associated with the ability of ECs to form and maintain the EC monolayer. The disruption of the DPB as an important initial event in the pathogenesis of vascular diseases such as atherosclerosis is discussed.
Immunochemical characterization of apolipoprotein A-I from normal human plasma. In vitro modification of apo A-I antigens.
Authors: Milthorp P , Weech P K , Milne R W , Marcel Y L
Published: June 2011

Two series of monoclonal antibodies (MAB) directed against apolipoprotein A-I (apo A-I) reacted differentially with freshly prepared sera or plasma. MAB from Series 1 were obtained after immunization and screening with purified apo A-I, while those from Series 2 were obtained by immunization with high density lipoprotein (HDL) and screening with both HDL and apo A-I. Series 2 MAB 5F6, 3G10, and 4F7 reacted significantly better with fresh material than material stored at 4 degrees C for longer than 1 month. Conversely, Series 1 MAB 3D4, and 6B8, and Series 2 MAB 2F1 reacted more strongly with the stored preparations. Series 2 MAB 4H1 reacted equally with stored or fresh material. The inability of 3D4 and 6B8 to react with fresh sera or plasma was not due to an inhibitor in the fresh material. Preparation of HDL from serum or plasma and delipidation of this material had no effect on the above phenomena, which appears related directly to apo A-I and not to the interaction of apo A-I with lipids. The variation of immunoreactivity with storage at 4 degrees C was also shown to be unrelated to proteolysis of apo A-I. Alkaline treatment of freshly prepared sera, HDL, or apo HDL with sodium hydroxide simulated the effect of storage, allowing this material to react strongly with MAB 3D4, 6B8. As expected, there was a decrease in the reactivity of MAB 5F6 with alkali-treated fresh material compared to untreated fresh material. However, further investigation showed that alkaline treatment does not completely mimic the effect of storage. MAB 2F1, which reacts poorly with fresh material, reacted very poorly with alkali-treated fresh material, and MAB 3G10 and 4F7, which react well with fresh material, reacted even better with alkali-treated fresh material. While isoelectric focusing of apo HDL prepared from stored serum showed a redistribution of apo A-I toward more acidic isoforms compared to fresh serum, alkaline treatment of apo HDL resulted in partial cleavage of apo A-I and in the generation of isoforms more acidic than known apo A-I isomorphs. Therefore, alkaline treatment of serum is only a partial model for antigenic modification of apo A-I upon storage. These results demonstrate that in vitro conditions (storage or alkaline treatment) selectively modify certain antigenic sites of apo A-I, but not others. This phenomenon, which is probably related to deamidation, may affect apo A-I immunoassays with either monoclonal or polyclonal antibodies.
Blood flow through vasa vasorum of coronary arteries in atherosclerotic monkeys.
Authors: Heistad D D , Armstrong M L
Published: June 2011

Morphologic studies and postmortem arteriograms provide qualitative evidence that atherosclerosis induces neovascularization in coronary arteries. In this study we obtained the first measurements of blood flow through vasa vasorum in atherosclerotic coronary arteries. We measured blood flow with microspheres to intima media and to adventitia in normal and atherosclerotic cynomolgus monkeys. Blood flow to adventitia was not altered by atherosclerosis. The blood flow to intima media, however, was increased more than fivefold by atherosclerosis: flow was 3 +/- 1 in normal animals and 16 +/- 5 in atherosclerotic monkeys (p less than 0.05). Blood flow was threefold greater in atherosclerotic than in normal coronary arteries during adenosine-induced vasodilatation, which suggests that proliferation of new vessels, not dilatation of existing vessels, accounts for the increase in flow through vasa in intima media. Histologic studies indicated that vasa vasorum to intima media in atherosclerotic coronary arteries originate from adventitial vasa. We conclude that: 1) there is a large increase in blood flow through vasa vasorum to intima media, but not to adventitia, in atherosclerotic coronary arteries; and 2) increased flow is produced primarily by proliferation of new vessels that originate from adventitial vasa.
Increased lipid transfer activities in hyperlipidemic rabbit plasma.
Authors: Son Y S , Zilversmit D B
Published: June 2011

Three models of hypercholesterolemia in rabbits were positively associated with plasma cholesteryl ester and triglyceride transfer activities. In cholesterol-fed rabbits, the transfer activities increased two- to three-fold when plasma cholesterol concentration reached about 1500 mg/dl. In hypercholesterolemia induced by feeding a cholesterol-free semisynthetic casein-sucrose diet, plasma cholesterol increased up to three-fold and the increase in cholesteryl ester and triglyceride transfer activities per unit of plasma cholesterol increment was similar to that in cholesterol-fed rabbits. Watanabe heritable hyperlipidemic rabbits, in which hypercholesterolemia is induced by a genetic defect of the LDL receptor, had significantly higher transfer activities than normolipidemic rabbits. However, fasting, which also induced hypercholesterolemia, lowered transfer activities, whereas refeeding returned them to prefasting levels. In rats, hypercholesterolemia induced by a high cholesterol diet did not increase cholesteryl ester and triglyceride transfer activities. Although hypercholesterolemia in rabbits may not be the primary cause of increased lipid transfer activities, some mechanism related to hypercholesterolemia appears to be associated with increased cholesteryl ester and triglyceride transfer activities.