Defective triglyceride biosynthesis in CETP-deficient SW872 cells

Abstract

with adipose tissue CETP mRNA levels in both man and hamster ( 6,7 ), showing the importance of this tissue in both lipid storage and the modulation of intravascular lipid metabolism.While most CETP is secreted by cells, a portion is retained ( 8-10 ). Much of the intracellular CETP pool is associated with the endoplasmic reticulum (ER), with increased concentrations near lipid droplets ( 8,11 ). Both cytoplasmic and lipid-droplet pools of CETP have been observed as well ( 11 ). Although the mechanism for CETP release into the cytoplasm is unknown, the escape of other ER luminal proteins to the cytoplasm has been reported (12)(13)(14).In addition to its role in plasma lipoprotein metabolism, multiple studies suggest that CETP has other functions. Cell-associated CETP promotes CE uptake from HDL, stimulates the cell's ability to effl ux cholesterol, and infl uences the storage of CE in cells (15)(16)(17)(18)(19). In addition to altering cellular cholesterol metabolism, both overexpression and under-expression of CETP in SW872 cells, a human cell line commonly used as an adipocyte model, reduce the capacity of these cells to store TG ( 10, 11 ). A role for CETP in adipocyte lipid storage is further supported by observations in transgenic mice. Adipose tissue-specifi c expression of human CETP in mice results in smaller adipocytes containing less TG and cholesterol, and signifi cantly reduces the expression of key lipogenic genes ( 20 ). In hypertriglyceridemic mice, CETP expression normalizes subcutaneous adipose depots and visceral adipocyte size ( 21 ). And in humans, a CETP gene variant that affects the coding sequence of CETP is associated with increased adiposity following long-term overfeeding ( 22 ).In this study, we have examined pathways involved in TG homeostasis in order to understand the metabolic Abstract We previously reported that reducing the expression of cholesteryl ester transfer protein (CETP) disrupts cholesterol homeostasis in SW872 cells and causes an ‫ف‬ 50% reduction in TG. The causes of this reduced TG content, investigated here, could not be attributed to changes in the differentiation status of CETP-defi cient cells, nor was there evidence of endoplasmic reticulum (ER) stress. In short-term studies, the total fl ux of oleate through the TG biosynthetic pathway was not altered in CETP-defi cient cells, although mRNA levels of some pathway enzymes were different. However, the conversion of diglyceride (DG) to TG was impaired. In longer-term studies, newly synthesized TG was not effectively transported to lipid droplets, yet this lipid did not accumulate in the ER, apparently due to elevated lipase activity in this organelle. DG, shown to be a novel CETP substrate, was also ineffi ciently transferred to lipid droplets. This may reduce TG synthesis on droplets by resident diacylglycerol acyltransferase. Overall, these data suggest that the decreased TG content of CETP-defi cient cells arises from the reduced conversion of DG to TG in the ER and/or on the lipid droplet surface, and enhanced ...

Exon 9-deleted CETP inhibits full length-CETP synthesis and promotes cellular triglyceride storage

“…5A). The CETP-specific antibody TP2 reduced transfer by E9-deleted CETP to baseline levels, similar to that previously reported for FL CETP (24). In this experiment, similar levels of FL and E9-deleted CETP mass, based on Western blot analysis, were used for the assay.…”

“…We have previously shown that the extent of 3 H-oleate incorporation under these conditions mirrors the mass of these lipids present in SW872 cells (14,15). In our earlier studies, we demonstrated that reducing both FL and E9-deleted CETP expression caused newly synthesized TG to accumulate in an ER-rich fraction instead of lipid droplets, the natural TG storage site (15,24). A similar study was performed in cells overexpressing E9-deleted CETP to determine whether their increased TG content could arise from increased TG synthesis and/or altered incorporation of this TG into lipid storage droplets.…”

“…Reducing cellular levels of both FL and E9-deleted CETP impairs the movement of TG into lipid droplets (15,24).…”

The lipid substrate preference of CETP controls the biochemical properties of HDL in fat/cholesterol-fed hamsters

“…This remains to be tested experimentally. Alternatively, several studies have shown that human CETP also functions inside cells to control the accumulation of CE and TG in lipid droplets ( 11 , 42 , 43 ). The expression of human CETP may modify this process in the livers of fat-fed hamsters causing cholesterol to accumulate.…”

Low‐density lipoprotein receptor is required for cholesteryl ester transfer protein to regulate triglyceride metabolism in both male and female mice

“…Data presented here could support the first hypothesis if intracellular CETP alters lipid delivery from LDLR‐derived endosomes to TG metabolic and nuclear receptor targets. In vitro work supports a role for intracellular CETP in regulating TG metabolism and LDL uptake in adipocytes and liposarcoma cells (Greene et al, 2015; Izem et al, 2015; Izem & Morton, 2007). The second hypothesis, however, may more easily explain the broad functional impacts of LDLR deletion on CETP function in TG metabolism.…”