Objective-We investigated the role of adipocyte differentiation-related protein (ADRP) in triglyceride turnover and in the secretion of very low-density lipoprotein (VLDL) from McA-RH7777 cells and primary rat hepatocytes. Methods and Results-An increase in the expression of ADRP increased triglyceride accumulation in cytosolic lipid droplets and prevented the incorporation of fatty acids into secretable triglycerides, thereby reducing the secretion of triglycerides as well as of apolipoprotein B-100 (apoB-100) and apoB-48 VLDL. The ability of ADRP to block the secretion of apoB-100 VLDL1 decreased with increasing quantities of fatty acids in the medium, indicating a saturable process and emphasizing the importance of sequestering of fatty acids for the effect of ADRP on VLDL secretion. Knockdown (small interfering RNA) of ADRP decreased the pool of cytosolic lipid droplets but increased only the secretion of apoB-48 VLDL1. Additionally, there was an increased flow of fatty acids into ␤-oxidation. Conclusions-ADRP is essential for the accumulation of triglycerides in cytosolic lipid droplets. An increase in ADRP prevents the formation of VLDL by diverting fatty acids from the VLDL assembly pathway into cytosolic triglycerides, whereas a decrease of the protein increases the sorting of fatty acids to ␤-oxidation and promotes the secretion of apoB-48 VLDL1. Key Words: adipose differentiation-related protein Ⅲ cytosolic lipid droplets Ⅲ apolipoproteins B Ⅲ ␤-oxidation Ⅲ small interfering RNA C ytosolic lipid droplets are ubiquitous organelles involved in the storage and turnover of neutral lipids such as triglycerides. Several proteins have been identified on these droplets, the most well known being the PAT domain proteins, 1-3 including the perilipins, adipocyte differentiationrelated protein (ADRP or adipophilin) and Tip 47. ADRP, which is ubiquitously expressed, 4 has a central role in the formation of lipid droplets. 5 These droplets are assembled at the microsomal membrane by an insulin-dependent process 6 that requires phospholipase D1, extracellular signal regulated kinase 2, and the motor protein dynein. 6,7 The assembly process involves the formation of small primordial droplets, 7 which grow in size by a fusion process that is dependent on intact microtubules 8 and dynein. 6 The assembly of very-low density lipoproteins (VLDLs) 9 -12 starts with the cotranslational lipidation of apolipoprotein B-100 (apoB-100), forming a pre-VLDL particle. VLDL2 (Svedberg flotation [sf] units 20 to 60) is formed from pre-VLDL by additional lipidation, 13 whereas VLDL1 (sf 60 to 80) is formed from VLDL2 by a mechanism that is dependent on an ADP ribosylation factor 1-controlled sorting/transport process 14 and involves the addition of a bulk load of lipids to the particle. 12,13 The triglycerides used in this assembly process are largely derived from triglycerides in cytosolic lipid droplets. 15,16 In this article, we demonstrate that an increase in ADRP promotes the storage of triglycerides in cytosolic lipid dropl...

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Receptor-dependent productive uptake of GLP1-conjugated antisense oligonucleotides occurs selectively in pancreatic β-cells.

In this study, we tested the hypothesis that two separate pathways, the two-step process and an apolipoprotein B (apoB) size-dependent lipidation process, give rise to different lipoproteins. Expression of apoB-100 and C-terminally truncated forms of apoB-100 in McA-RH7777 cells demonstrated that VLDL particles can be assembled by apoB size-dependent linear lipidation, resulting in particles whose density is inversely related to the size of apoB. This lipidation results in a LDL-VLDL 2 particle containing apoB-100. VLDL 1 is assembled by the two-step process by apoB-48 and larger forms of apoB but not to any significant amount by apoB-41. The major amount of intracellular apoB-80 and apoB-100 banded with a mean density of 1.10 g/ml. Its formation was dependent on the sequence between apoB-72 and apoB-90. This dense particle, which is retained in the cell, possibly by chaperones or association with the microsomal membrane, is a precursor of secreted VLDL 1. The intracellular LDL-VLDL 2 particles formed during size-dependent lipidation appear to be the precursors of intracellular VLDL 1. We propose that the dense apoB-100 intracellular particle is converted to LDL-VLDL 2 by size-dependent lipidation. LDL-VLDL 2 is secreted or converted to VLDL 1 by the uptake of the major amount of triglycerides. -Stillemark-Billton, P., C. Beck, J. Immunoelectron microscopy (1) and kinetic studies (2, 3) indicate that VLDLs are assembled in two major steps (4, 5). The first step occurs during the translation of apolipoprotein B (apoB) and gives rise to a premature particle (2, 6) we refer to as a primordial lipoprotein. Major amounts of lipid are added in the second step, resulting in bona fide VLDL (2, 7, 8); a second precursor of VLDL, an apoB-free "lipid droplet" in the smooth endoplasmic reticulum (1, 9) whose assembly requires microsomal triglyceride transfer protein (MTP), may also be involved (10). VLDLs are secreted in two forms: large, triglyceride-rich VLDL 1 and smaller, triglyceride-poor VLDL 2. Overproduction of VLDL 1 is linked to conditions such as insulin resistance and type II diabetes (11).The lengths of C-terminally truncated forms of apoB-100 are inversely related to the amount of lipid in the lipoproteins they assemble; assembly with apoB-100 results in VLDL (12). This size-dependent lipidation of apoB does not fit the two-step model; in particular, it cannot explain why apoB-48 has the ability to assemble VLDL but apoB-40 lacks this ability (8).In this study, we tested the hypothesis that two separate pathways, the two-step process and an apoB size-dependent lipidation process, give rise to different lipoproteins. Our results indicate that although apoB can assemble VLDL 1 once it reaches the size of apoB-48, the size-dependent process gives rise to LDL-VLDL 2 particles first when apoB-100 is reached. In contrast to apoB-48 (2), the major intracellular form of apoB-100 is much denser than expected from the size/density relation, and it is retained in the secretory pathway. The sequence between apoB-7...