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The effect of fish oil supplements on atherogenesis is controversial, especially when fish oil does not lower plasma cholesterol. Some studies in swine have shown that a fish oil supplement to a butter-cholesterol diet reduces atherogenesis. The fish oil supplement also frequently reduces average plasma cholesterol levels. The reduction in lesion size has been shown to be greater than can be expected from average plasma cholesterol reductions, if a linear relationship between lesion size and plasma cholesterol was assumed. However, in an experiment in which we equalized time-weighted average plasma cholesterol levels, there was no significant reduction in lesion size in the fish oil supplemented group. This led us to question the validity of the linear relationship between lesion size and plasma cholesterol level. In this study we have combined the results of eight study blocks with a total of 76 swine fed a similar hyperlipidemic, butter-cholesterol diet. Of these, 24 received a fish oil supplement (BT+fish oil) and 52 swine received no fish oil supplement (BT). The average lesion size as measured by nuclear profiles per cross section of a fixed site in the abdominal aorta (ABNpCx) was 7704 +/- 778 (mean +/- SEM) for the BT group and 2360 +/- 1145 for the BT+fish oil group. Total plasma cholesterol levels were measured at the outset and at monthly intervals until sacrifice. For each animal we obtained a time-weighted average based on the trapezoidal rule.(ABSTRACT TRUNCATED AT 250 WORDS)

We have investigated in swine the effect of fish oil additives to a butter-cholesterol hyperlipidemic diet (BT) on atherogenesis and thrombogenesis when average plasma cholesterol levels were kept similar in fish oil-treated and untreated BT groups. The studies included evaluation of lesion sizes and cell numbers, counts of adherent monocytes over lesions, and counts of platelet clumps (microthrombi) over lesions either attached directly to endothelium or to adherent monocytes. Anatomic sites studied for lesion development were the left anterior descending coronary artery (LAD), the distal 1/5 of the abdominal aorta, and a proximal portion of the thoracic aorta. Counts of attached monocytes and platelet clumps were made by scanning electron microscopy only for the LAD and expressed per mm2 of surface. The most striking new result was in regard to the platelet clumps. These were reduced by the fish oil from 996 +/- 295/mm2 in the untreated BT group to 313 +/- 59 and 364 +/- 105 in BT+cod liver oil and BT+menhaden oil groups, respectively. Most of the platelet clumps were adherent to attached monocytes in all groups and the number of attached monocytes were greatly reduced by the fish oil additive. Thus there were close relationships among platelet clumps, monocytes, and lesion endothelium. Numbers of attachments over nonlesion endothelium were much less than those over lesions in all dietary groups. The most surprising result was the lack of retardation of lesion growth by the fish oil additives in spite of the reduction in attached monocytes and platelet clumps. In previous studies where the high plasma cholesterol levels in the BT swine had been modestly reduced (about 25%) there had been a marked retardation of lesion growth. The current result suggests that plasma cholesterol is the major factor controlling lesion growth in this model through under milder conditions and longer observation periods other factors might become apparent.

In this study, young growing swine were made hypercholesterolemic (~300 mg/dl) by feeding milk and eggs for 7 wk. They were then divided into three groups (untreated, clofibrate-treated, and cholestyramine-treated), and the diet was continued for an additional 3-4 wk. A cholesterol balance study was carried out in the terminal week. When the swine were killed, the total carcass content of cholesterol was determined, as well as contents of individual tissues. Both drugs caused a 50% reduction in serum cholesterol levels. The total carcass cholesterol contents were significantly lower in both treatment groups than in the untreated group. The difference was due largely to lower concentrations in the plasma and in bulk tissues. [4-(14)C] Cholesterol was fed 7 days before the animals were killed, and specific activities of cholesterol in individual tissues were determined terminally. These gave a broad spectrum of values in tissues (excluding central nervous system) ranging rather evenly from 33% of plasma specific activity in the aorta to 100% in some tissues. The balance data suggest that cholestyramine reduces the enterohepatic bile acid pool and cholesterol absorption but increases fecal output of bile acids and total body cholesterol synthesis. The balance data, limited to the terminal week, give no indication of the mode of action of clofibrate. Even synthesis was not significantly lower than in the untreated swine.

Three groups of young swine were fed semi-purified diets enriched in fat and cholesterol for 90 days. In one group 40% of calories were from butter; in another 40% were from corn oil; in a third group 20% were from corn oil. Mean serum cholesterol values were respectively 530, 458 and 400 mg/dl. Amounts of atherosclerosis in the abdominal aortas expressed as mean number of nuclear profiles per cross-section were respectively 1611, 734 and 585; expressed as mean lesion area per cross section 46.7, 21.4 and 19.4 mm2 X 10(2). All differences between the butter group and either of the two corn oil groups were statistically significant. We found that intimal response to the butter containing diet was significantly greater than to the corn oil containing diet for equivalent hypercholesterolemic response. We present detailed statistical validation of the disproportionately larger intimal response to butter containing diet. Furthermore, we suggest that estimates of comparative atherogenic risk of food fats based on response of serum cholesterol may mask important atherogenic differences.

Studies of the interaction of cholesterol absorption, excretion, synthesis, and retention were carried out with growing swine through the application of sterol and isotopic balance procedures to a non-steady state situation. The animals were fed either a low-fat commercial mash diet (MA), a commercial mash diet plus 2.4 g/day of crystalline cholesterol (MAC), or a high-fat milk diet containing 2.4 g/day cholesterol (MKC). [(14)C]Cholesterol and beta-[(3)H]sitosterol were used to measure fecal neutral steroid losses. Final plasma cholesterol levels averaged 91, 85, and 195 mg/100 ml for MA, MAC, and MKC animals, respectively. Total carcass dissection followed by cholesterol measurement revealed that accumulation of cholesterol in most body tissues was directly related to increase in body size in the rapidly growing swine. The only notable exceptions were plasma, liver, and bone with marrow of the MKC group, where cholesterol concentrations were significantly elevated. Absorption of cholesterol was significantly greater in animals fed MKC than MAC-fed swine (726 mg/day vs. 142 mg/day), whereas synthesis of cholesterol was significantly greater in MAC animals than in MKC animals (863 mg/day vs. 112 mg/day). Excretion of neutral and acid steroids into the intestine was not significantly different between MAC and MKC groups, but both classes of steroids were more efficiently reabsorbed in MKC animals. The overall differences between swine fed mash-cholesterol and those fed milk-cholesterol diets appear to result from more efficient absorption of both neutral and acid steroids in the milk-cholesterol group only partially compensated for by decreased cholesterol synthesis.

Aquaporin-0 (AQP0) tetramers form square arrays in lens membranes through a yet unknown mechanism, but lens membranes are enriched in sphingomyelin and cholesterol. Here, we determined electron crystallographic structures of AQP0 in sphingomyelin/cholesterol membranes and performed molecular dynamics (MD) simulations to establish that the observed cholesterol positions represent those seen around an isolated AQP0 tetramer and that the AQP0 tetramer largely defines the location and orientation of most of its associated cholesterol molecules. At a high concentration, cholesterol increases the hydrophobic thickness of the annular lipid shell around AQP0 tetramers, which may thus cluster to mitigate the resulting hydrophobic mismatch. Moreover, neighboring AQP0 tetramers sandwich a cholesterol deep in the center of the membrane. MD simulations show that the association of two AQP0 tetramers is necessary to maintain the deep cholesterol in its position and that the deep cholesterol increases the force required to laterally detach two AQP0 tetramers, not only due to protein–protein contacts but also due to increased lipid–protein complementarity. Since each tetramer interacts with four such ‘glue’ cholesterols, avidity effects may stabilize larger arrays. The principles proposed to drive AQP0 array formation could also underlie protein clustering in lipid rafts.