Uptake of phospholipid-depleted chylomicrons by the perfused rat liver.

1. Rat lymph chylomicrons were depleted of their surface phospholipids by treatment with pure phospholipase A2 from Crotalus adamanteus venom. 2. About 80% of the phospholipids could be removed from the chylomicrons without any apparent effect on their size, neutral lipid composition or qualitative profile of their tetramethylurea-soluble apoproteins. 3. Phospholipid-depleted chylomicrons were rapidly taken up whole by liver cells when perfused through isolated rat liver preparations. The rate of uptake was dependent on the extent of phospholipid depletion and reached a maximum (4-6.5-fold greater than control chylomicrons) when 80% of the phospholipids had been removed. 4. It is speculated that the hepatic uptake of phospholipid-depleted chylomicrons occurs by a mechanism to that of chylomicron-remnants uptake.     

Action of liproprotein lipase on apoprotein-depleted chylomicrons.

1. Rat lymph chylomicrons were exposed to soluble and to immobilized trypsin. This treatment caused no detectable changes in the chylomicron structure or lipid composition, but did result in virtually total depletion of all their tetramethylurea-soluble apoproteins. 2. The capacity of these apoprotein-depleted chylomicrons to act as substrate for lipoprotein lipase in vitro and in situ (i.e. isolated perfused rat heart) was decreased by about 90 and 75% respectively, compared with intact chylomicrons. 3. On incubation with rat plasma high-density lipoproteins, trypsin-treated chylomicrons readily acquired a full apoprotein complement. This resulted in the complete restoration of their capacity to act as substrate for lipoprotein lipase both in vitro and in situ. 4. It is suggested that with the use of try,sin-treated chylomicrons it is now possible for the first time to investigate the physiological role that individual apoproteins play in the catabolism of triacylglycerol-rich lipoproteins by lipoprotein lipase.     

Hepatic uptake of phospholipid-depleted chylomicrons in vivo. Comparison with the uptake of chylomicron remnants.

1. Rats pretreated with Triton WR-1339 to prevent the formation of remnants were injected with [3H]cholesterol-labelled remnants, intact chylomicrons or chylomicrons depleted of most of their surface phospholipids by treatment with phospholipase A2. Within 5 min about 80% of the injected label of remnants and phospholipid-depleted chylomicrons was incorporated into the livers compared with less than 10% of the injected radioactivity of intact chylomicrons. A similar rapid hepatic uptake of radioactivity occurred when rats not pretreated with Triton were injected with [3H]cholesterol-labelled phospholipid-depleted chylomicrons. This rapid hepatic uptake of phospholipid-depleted chylomicrons occurred apparently without any alteration in the apoprotein composition of the particles. 2. The participation of hepatocytes in the uptake of remnants and phospholipid-depleted chylomicrons was examined. Both types of particles were taken up by the hepatocytes. However, small chylomicrons (Sf less than 400) were taken up more efficiently than were large chylomicrons (Sf greater than 400), but neither was taken up as efficiently as the remnants. 3. The results of this study lend support to the hypothesis that phospholipid-depleted chylomicrons and chylomicron remnants are taken up by the liver by a similar mechanism, which depends on the loss of surface phospholipids.     

Cideb facilitates the lipidation of chylomicrons in the small intestine

Cell death-inducing DFF45-like effector b (Cideb), an endoplasmic reticulum (ER)- and lipid droplet (LD)-associated protein, has been shown to play a critical role in maintaining hepatic lipid homeostasis by promoting the lipidation and maturation of VLDL particles. Here, we observed that Cideb is expressed in the jejunum and ileum sections of the small intestine, and its expression was induced by high-fat diet. Intragastric gavage with lipids resulted in the retention of lipids in the intestine in Cideb-deficient mice. In addition, we observed that mice with Cideb deficiency exhibited reduced intestinal chylomicron-TG secretion and increased lipid accumulation in the enterocytes. The sizes of chylomicrons secreted from the small intestine of Cideb-deficient mice were also smaller than those from wild-type mice. Furthermore, the overexpression of Cideb increased TG secretion and reduced lipid accumulation in the enterocyte-like Caco-2 cells. In addition, we proved that Cideb was localized to the ER and LDs and could interact with ApoB48 in Caco-2 cells. Overall, these data revealed that Cideb plays an important role in controlling intestinal chylomicron lipidation.     

Intracellular events in the assembly of chylomicrons in rabbit enterocytes

The aim of this study was to determine the intracellular events in chylomicron assembly in adult villus enterocytes. We have used novel methods for separation of the intracellular components of the secretory compartment [rough and smooth endoplasmic reticulum (RER and SER, respectively) and Golgi], and their membrane and luminal components, from villus enterocytes isolated from rabbit small intestine. The steady state composition of the components of the secretory compartment and the intracellular pools of newly synthesized apolipoprotein B-48 (apoB-48) and triacylglycerol (TAG) was determined. The observations indicate that the SER is the main site of TAG synthesis and of chylomicron assembly. Newly synthesized apoB-48 and TAG accumulate in the SER membrane and are transferred into the lumen in a microsomal triglyceride transfer protein-dependent step. In enterocytes isolated from chow-fed rabbits, in which fat absorption is relatively slow, transfer of apoB-48 and TAG from the SER membrane into the lumen appears to be rate limiting. In enterocytes from fat-fed rabbits, TAG accumulates in the lumen of the SER, suggesting that movement out of the SER lumen becomes rate limiting, when chylomicron secretion is markedly stimulated. In these cells, the cytosolic TAG also increased to 450 microgram/g enterocytes, compared with 12 microgram/g enterocytes from chow-fed rabbits, indicating that transfer of TAG from the SER membrane into the secretory pathway can become saturated, so that newly synthesized TAG moves into the cytosol.     

Metabolism of chylomicrons by the isolated rat liver

Isolated livers perfused with washed corn oil chylomicrons labeled in vivo with palmitic acid-1-(14)C removed a large proportion of the chylomicrons. Slices from these livers oxidized chylomicron fatty acid esters to both carbon dioxide and acetoacetate. The liver slices also generated free fatty acids from chylomicron lipids and converted chylomicron triglycerides to phospholipids. Similar activities were observed in rat liver slices prepared shortly after the intravenous administration of chylomicrons to intact rats. The observed chylomicron uptake and lipid conversions were similar in livers from both fed and fasted rats. Fasting increased the oxidation of chylomicron fatty acid esters by livers labeled in vivo and by perfusion. In livers removed from intact rats given labeled chylomicrons, the triglyceride-(14)C to phospholipid-(14)C ratio was high, a finding unexpected if the liver had acquired this (14)C by removal of circulating fatty acids formed by extrahepatic lipolysis. These results demonstrate the ability of the liver to remove and utilize chylomicrons directly and suggest that direct removal accounts for a significant portion of the chylomicron fatty acids utilized by the liver of intact rats.     

Degradation of apolipoprotein B-100 in human chylomicrons

The purpose of this study was to investigate the molecular forms of apolipoprotein B (ApoB) in human chylomicrons under well-preserved conditions. To this end, plasma and serum were collected from the same normal subjects after ingestion of a fatty meal. The samples were divided into three or four aliquots before the addition of various preservative mixtures, including antibiotics, antioxidants and proteinase inhibitors. The chylomicrons were isolated immediately, and all steps were carried out at or below 4 degrees C. Changes in the molecular weight of ApoB in chylomicrons were followed by a time study using 3.3% polyacrylamide gel electrophoresis containing SDS. ApoB from chylomicrons analyzed within 5 h of blood collection showed a single band with mobility identical to that of ApoB (ApoB-100) in low-density lipoproteins. When analyzed after 1-2 days, satellite bands smaller than ApoB-100 were observed, and a very faint band with Mr 200,000 appeared, which comigrated with intestinal ApoB (ApoB-48). Upon storage, the molecular weight of ApoB was smaller in chylomicrons subjected to a higher number of reflotations than those in chylomicrons washed less frequently, suggesting that purified chylomicrons degrade faster. A longer storage time at 4 degrees C (i.e., 7 or 14 days) revealed a stepwise degradation of ApoB, yielding Mr 200,000 band as the prominent form. The degradation of ApoB-100 was slower when both proteinase inhibitors, leupeptin and epsilon-amino caproic acid, were employed, and the appearance of Mr 200,000 band was quicker when the chylomicrons were processed at higher temperature (15-25 degrees C) in the absence of a proteinase inhibitor. Immunoblotting shows that the segment removed from ApoB-100 was the carboxyl-terminal portion. These results suggest the possible presence of a proteinase(s), which copurified with chylomicrons, and which converts ApoB-100 from a large to a smaller molecular form. Although the stop codon has been discovered recently in intestinal ApoB mRNA, which explains the mechanism for direct synthesis of ApoB-48, apparently ApoB-100 is also synthesized in the intestine of all eight subjects studied here, and the ApoB-100 degrades to a form which is ApoB-48-like.