No abstract

Lipids circulate in the blood in association with plasma lipoproteins and enter the tissues either after hydrolysis or as non-hydrolyzable lipid esters. We studied cardiac lipids, lipoprotein lipid uptake, and gene expression in heart-specific lipoprotein lipase (LpL) knock-out (hLpL0), CD36 knock-out (Cd36 ؊/؊ ), and double knock-out (hLpL0/Cd36 ؊/؊ -DKO) mice. Loss of either LpL or CD36 led to a significant reduction in heart total fatty acyl-CoA (control, 99.5 ؎ 3.8; hLpL0, 36.2 ؎ 3.5; Cd36 ؊/؊ , 57.7 ؎ 5.5 nmol/g, p < 0.05) and an additive effect was observed in the DKO (20.2 ؎ 1.4 nmol/g, p < 0.05). Myocardial VLDL-triglyceride (TG) uptake was reduced in the hLpL0 (31 ؎ 6%) and Cd36 ؊/؊ (47 ؎ 4%) mice with an additive reduction in the DKO (64 ؎ 5%) compared with control. However, LpL but not CD36 deficiency decreased VLDL-cholesteryl ester uptake. Endogenously labeled mouse chylomicrons were produced by tamoxifen treatment of ␤-actin-MerCreMer/ LpL flox/flox mice. Induced loss of LpL increased TG levels >10-fold and reduced HDL by >50%. After injection of these labeled chylomicrons in the different mice, chylomicron TG uptake was reduced by ϳ70% and retinyl ester by ϳ50% in hLpL0 hearts. Loss of CD36 did not alter either chylomicron TG or retinyl ester uptake. LpL loss did not affect uptake of remnant lipoproteins from ApoE knock-out mice. Our data are consistent with two pathways for fatty acid uptake; a CD36 process for VLDLderived fatty acid and a non-CD36 process for chylomicron-derived fatty acid uptake. In addition, our data show that lipolysis is involved in uptake of core lipids from TG-rich lipoproteins.Under normal physiological conditions, myocardial energy demands are predominantly met by fatty acid (FA) 3 oxidation (Ͼ70%) with the remaining energy provided by glucose, lactate, and ketones. The majority of the lipid entering cardiac cells is diverted toward FA utilization (1) with some being stored or used for structural requirements. FAs are delivered to the heart from two sources: (a) FAs esterified as triglyceride (TG) contained in circulating lipoproteins and liberated by lipoprotein lipase (LpL)-mediated lipolysis, and (b) non-esterified FA, referred to as free FAs (FFAs), bound to serum albumin. The mechanisms responsible for the uptake of FAs by the heart, or any organ, are incompletely understood. Specifically, the importance of receptor-mediated uptake versus diffusion of FAs across membranes is under debate. Studies in cardiomyocytes (2-4) and other cells (5, 6) have suggested that uptake of FAs by cultured cells can occur via two pathways (7,8). A low capacity but high affinity uptake pathway thought to represent receptormediated uptake is operative at FFA/albumin ratios normally found in the plasma (5). At higher FFA concentrations, uptake occurs via a lower affinity non-saturable process (9, 10); this has been studied in synthetic membranes and thought to represent non-receptor uptake also referred to as "flip-flop" (11-13). Several proteins including fatty acid translocase (FAT/ ...

Background: Lipoprotein lipase (LpL) is rate-limiting for plasma triglyceride lipolysis, but its importance in adipose development is uncertain. Results: Adipocyte LpL knock-out affected brown but not white fat composition. White fat was reduced when muscle LpL expression was increased. Conclusion: LpL distribution and adipose metabolism affect adipogenesis. Significance: All fat depots are not equally dependent on triglyceride uptake.

Elevated TGs, but not FFAs, postburn are associated with worsened organ function and clinical outcomes.

Severe burn injury causes hepatic dysfunction that results in major metabolic derangements including insulin resistance and hyperglycemia and is associated with hepatic endoplasmic reticulum (ER) stress. We have recently shown that insulin reduces ER stress and improves liver function and morphology; however, it is not clear whether these changes are directly insulin mediated or are due to glucose alterations. Metformin is an antidiabetic agent that decreases hyperglycemia by different pathways than insulin; therefore, we asked whether metformin affects postburn ER stress and hepatic metabolism. The aim of the present study is to determine the effects of metformin on postburn hepatic ER stress and metabolic markers. Male rats were randomized to sham, burn injury and burn injury plus metformin and were sacrificed at various time points. Outcomes measured were hepatic damage, function, metabolism and ER stress. Burn-induced decrease in albumin mRNA and increase in alanine transaminase (p < 0.01 versus sham) were not normalized by metformin treatment. In addition, ER stress markers were similarly increased in burn injury with or without metformin compared with sham (p < 0.05). We also found that gluconeogenesis and fatty acid metabolism gene expressions were upregulated with or without metformin compared with sham (p < 0.05). Our results indicate that, whereas thermal injury results in hepatic ER stress, metformin does not ameliorate postburn stress responses by correcting hepatic ER stress.

Objective-Although epidemiologic data suggest that hypertriglyceridemia and elevated plasma levels of fatty acids are toxic to arteries, in vitro correlates have been inconsistent. To investigate whether increased endothelial cell expression of lipoprotein lipase (LpL), the primary enzyme creating free fatty acids from circulating triglycerides (TG), affects vascular function, we created transgenic mice that express human LpL (hLpL) driven by the promoter and enhancer of the Tie2 receptor. Methods and Results-Mice expressing this transgene, denoted EC-hLpL and L for low and H for high expression, had decreased plasma TG levels compared with wild-type mice (WT): 106Ϯ31 in WT, 37Ϯ17 (line H), and 63Ϯ31 mg/dL (line L) because of a reduction in VLDL TG; plasma cholesterol and HDL levels were unaltered. Crossing a high expressing EC-hLpL transgene onto the LpL knockout background allowed for survival of the pups; TG in these mice was approximately equal to that of heterozygous LpL knockout mice. Surprisingly, under control conditions the EC-hLpL transgene did not alter arterial function or endothelial cell gene expression; however, after tumor necrosis factor (TNF)-␣ treatment, arterial vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and endogenous TNF-␣ mRNA levels were increased and arteries had impaired endothelium-dependent vasodilatation. This was associated with reduced eNOS dimers. Conclusions-Therefore, we hypothesize that excess vascular wall LpL augments vascular dysfunction in the setting of inflammation. LpL is the rate-limiting enzyme for hydrolysis of plasma TG. LpL is synthesized by myocytes and adipocytes but is thought to function primarily on the luminal surface of endothelial cells. Therefore, along the arterial wall much of the exposure of endothelial cells to FAs is modulated by LpL actions.LpL has been suggested to have a dual effect on atherosclerosis development. 6 We and others have demonstrated that LpL contributes to lipid accumulation by stimulating the cellular binding and uptake of atherogenic lipoproteins in different vascular cell types including smooth muscle cells and macrophages. 7,8 Moreover, loss of macrophage LpL reduces atherosclerosis. 9 LpL acts as an activator of macrophage function, inducing TNF alpha (TNF-␣), 10 augmenting monocyte adhesion 11,12 and increasing phagocytosis in low glucose conditions. 13 Although these data suggest that LpL has atherogenic and inflammatory effects, there are contradictory data. LpL is reported to play an antiinflammatory role through generation of PPAR ligands for endothelial cells 14 and macrophages. 15 In human endothelial cells, LpL alone attenuated the inflammatory cascade produced by TNF-␣. 16 Thus, the function and role of LpL in inflammatory responses are not clear, but may vary depending on the in vitro conditions studied.To study the actions of LpL in arteries, we created mice with human LpL (hLpL) expression specifically in endothelial cells using the Tie2 promoter-enhancer 17 to drive an hLpL minigene. These transgenic mic...

The first 24 h following burn injury is known as the ebb phase and is characterized by a depressed metabolic rate. While the postburn ebb phase has been well described, the molecular mechanisms underlying this response are poorly understood. The endoplasmic reticulum (ER) regulates metabolic rate by maintaining glucose homeostasis through the hepatic ER stress response. We have shown that burn injury leads to ER stress in the liver during the first 24 h following thermal injury. However, whether ER stress is linked to the metabolic responses during the ebb phase of burn injury is poorly understood. Here, we show in an animal model that burn induces activation of activating transcription factor 6 (ATF6) and inositol requiring enzyme-1 (IRE-1) and this leads to increased expression of spliced X-box binding protein-1 (XBP-1s) messenger ribonucleic acid (mRNA) during the ebb phase. This is associated with increased expression of XBP-1 target genes and downregulation of the key gluconeogenic enzyme glucose-6-phosphatase (G6Pase). We conclude that upregulation of the ER stress response after burn injury is linked to attenuated gluconeogenesis and sustained glucose tolerance in the postburn ebb phase.

The trauma of a severe burn injury induces a hypermetabolic response that increases morbidity and mortality. Previously, our group showed that insulin resistance post-burn injury is associated with endoplasmic reticulum (ER) stress. Evidence suggests that c-jun N-terminal kinase (JNK) -2 may be involved in ER stress-induced apoptosis. Here, we hypothesized that JNK2 contributes to the apoptotic response after burn injury downstream of ER stress. To test this, we compared JNK2 knockout mice (−/−) to wildtype mice after inducing a 30% total body surface area thermal injury. Animals were sacrificed after 1, 3 and 5 days. Inflammatory cytokines in the blood were measured by multiplex analysis. Hepatic ER stress and insulin signaling were assessed by Western Blotting and insulin resistance was measured by a peritoneal glucose tolerance test. Apoptosis in the liver was quantified by TUNEL staining. Liver function was quantified by AST and ALT activity assays. ER stress increased after burn in both JNK2−/− and wildtype mice, indicating that JNK2 activation is downstream of ER stress. Knockout of JNK2 did not affect serum inflammatory cytokines; however, the increase in IL-6 mRNA expression was prevented in the knockouts. Serum insulin did not significantly increase in the JNK2−/− group. On the other hand, insulin signaling (PI3K/Akt pathway) and glucose tolerance tests did not improve in JNK2−/−. As expected, apoptosis in the liver increased after burn injury in wildtype mice but not in JNK2−/−. AST/ALT activity revealed that liver function recovered more quickly in JNK2−/−. This study indicates that JNK2 is a central mediator of hepatic apoptosis after a severe burn.