Year: 2009
Type: dissertation
Abstract: In mammals, adipose tissue is the major lipid storage organ accumulating excess neutral lipids within lipid droplets to provide energy during food deprivation or physical activity.In times of energy demand, triglycerides (TG) are hydrolysed in three sequential steps and catalysed by adipose triglyceride lipase (ATGL), hormonesensitive lipase (HSL) and monoglyceride lipase.TG hydrolysis results in the generation of diglycerides, monoglyceride and glycerol with a constant liberation of fatty acids.Beside adipose tissue, the liver is able to store significant amounts of TG to protect the body from potential toxicity of circulating fatty acids.Additional functions of the liver are synthesis of lipids and TG secretion in form of very lowdensity lipoprotein (VLDL) into the circulation to supply extra hepatic tissues with energy.Fatty acids derived from adipose tissue and liver are the major energy sources of the working muscle.Fatty acids either enter the intramuscular TG storage pool or are directly used for beta-oxidation to provide energy under resting conditions, during exercise and in post-exercise recovery periods.ATGL was identified as an enzyme specifically hydrolysing TG and performing the first and rate-limiting step in lipolysis.The phenotypical appearance of ATGLdeficient mice illustrated that the absence of the enzyme results in TG accumulation in multiple tissues, including liver and skeletal muscle.Mutations in human ATGL cause defective lipase activity leading to Neutral Lipid Storage Disease, a disorder characterised by systemic TG accumulation and the development of myopathy, muscle weakness and hepatic steatosis.Overall, the observations of impaired TG metabolism in the liver and skeletal muscle in mice and humans prompted us to investigate the role of ATGL in these tissues.Although the expression of ATGL is very low in the liver compared to adipose tissue, ATGL deficiency resulted in significantly reduced TG hydrolase activity in the liver.These data indicated that ATGL is involved in hepatic TG metabolism.Additionally, we observed that liver ATGL protein expression is substantially upregulated by fasting, implicating that the enzyme is regulated by the nutritional state.To get a deeper insight into the role of ATGL in hepatocytes, primary hepatocytes were isolated from the liver of wild-type and ATGL-deficient mice.ATGL-deficient hepatocytes exhibited an increased incorporation of fatty acids into TG stores in vitro, suggesting that the lack of ATGL impairs the mobilisation of TG.Consequently, the release of fatty acids and TG into the supernatant was significantly reduced in hepatocytes lacking ATGL.Accordingly, overexpression of the enzyme reduced TG content and inhibited the incorporation of radiolabeled fatty acids into TG stores.Together, these observations suggest that the lack of ATGL or overexpression of the enzyme substantially affects TG storage in primary murine hepatocytes.To investigate the role of ATGL in the liver in vivo, we generated transgenic mice specifically overexpressing ATGL in the liver under the control of the human alpha 1 antitrypsin promoter (liv-ATGL mice).The ATGL transgene was specifically overexpressed in the liver and was enzymatically active.The elevated lipase activity resulted in a 7-fold increase of TG hydrolase activity that caused significantly reduced hepatic TG levels in liv-ATGL mice.Nevertheless, overexpression of ATGL did not affect metabolic blood parameters of fed and fasted animals compared to control mice.Plasma fatty acids and TG levels do not differ from wild-type mice, indicating that ATGL overexpression does not directly affect the concentrations of circulating lipids.However, metabolic in vivo experiments revealed that liv-ATGL mice exhibit decreased VLDL-TG synthesis under moderately fasted conditions.These results suggest that the decreased TG content in the liver of these mice affects VLDL synthesis and secretion under fasting conditions.To investigate whether overexpression of ATGL increases fatty acid oxidation in liver, we determined the mRNA levels of several genes involved in this process.RT-PCR analysis of marker genes involved in beta-oxidation, ketone body synthesis and fatty acid transport revealed no differences in mRNA levels compared to controls.Overall, our studies demonstrate that ATGL overexpression reduces liver TG storage in vivo and does not affect circulating lipoprotein and fatty acid levels.To study the effect of physical exercise on energy metabolism in ATGL-and HSLdeficient mice, we performed exercise experiments using an Exer-6M treadmill (speed of 14 m/min and 0% slope).Metabolic blood energy parameters and liver glycogen levels of resting and exercised mice were determined and compared.ATGL-deficient mice exhibited reduced glycogen stores under resting conditions which were completely depleted after the training.Their plasma fatty acids and blood glucose levels were significantly reduced after exercise compared to wild-type and HSL-deficient mice.These data indicated that the lack of ATGL resulted in an increased use of carbohydrates caused by the inability to hydrolyse/use fatty acids for energy conversion.In contrast, HSL-deficient mice exhibited normal levels of plasma parameters and carbohydrate stores in comparison to wild-type mice.This indicates that other lipases, like ATGL, are able to compensate the deficiency of HSL under the moderate working settings.Furthermore, staining of ATGL-deficient muscle using Sudan III revealed a tremendous accumulation of neutral lipids specifically in type I fibres indicating that ATGL is mainly expressed in this fibre type.Since ATGL-deficient mice accumulate tremendous TG levels in cardiac muscle resulting in cardiac abnormalities and premature death, we wondered whether cardiac insufficiency affects exercise performance.For that purpose, we generated mice overexpressing ATGL in cardiac muscle (wild-type/CM) and crossed them on the ATGL-deficient background (ATGL-deficient/CM).Exercised ATGL-deficient/CM mice revealed significantly reduced plasma fatty acids and glucose levels compared to wild-type/CM controls.ATGL-deficient/CM also exhibited reduced liver glycogen and blood glucose levels.These data indicated that changes in energy metabolism during exercise are not caused by heart abnormalities and predominantly reflect defective mobilisation of TG stores which leads to an undersupply of the working muscle with energy in the form of fatty acids. .1.1.2.Adipose Triglyceride Lipase .......
Author: Schoiswohl, Gabriele Maria
Language: English
Subfield: Biochemistry
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7052292552