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There are two distinct types of adipose tissue which have different functions within the body, white (WAT) and brown (BAT). Browning of WAT occurs with increases in the WAT sympathetic nervous system (SNS) drive. In this regard we previously reported that melatonin (MEL) stimulation of MEL receptor 1A (MEL1a) within the SNS outflow to the WAT might be implicated in a naturally-occurring reversal of obesity (by ~30% of total body fat). Therefore, in this study we tested the hypothesis that MEL causes browning of WAT through the stimulation of SNS drive to WAT. This was done by comparing specific browning and lipolytic markers in WAT following 10 weeks of MEL treatment, short day housing (SD), and long day housing with saline injections (LD+VEH). Browning effects of a 5 day treatment of a β3-adrenergeric (β3 AR), CL 316, 243, were also measured. We found that CL 316, 243, MEL treatment, and SD housing had increased expressions of browning markers within WAT and lipolytic activity in MEL treated animals was increased in specific WAT.

The normal development of adipose tissue lipolysis as measured by glycerol release was studied in epididymal fat pads of Sprague-Dawley rats between 4 and 16 weeks of age and correlated with changes in fat cell size.For each age group studied, 4 weeks, 8 weeks, and 16 weeks of age, basal (no hormone present) and hormone stimulated lipolytic activity were observed for two concentrations of epinephrine were used, maximal (10,00 nM) and minimal (10 nM). Basal levels of glycerol were not linear. There was an increase between 4 and 8 weeks of age followed by a decrease between 8 and 16 weeks of age. The maximal dosage of hormone evoked a large increase in 9lYcerol production between 4 and 8 weeks, which was followed by a decrease between 8 and 16 weeks of age. The minimal dosage of epinephrine, although not significant, showed a decrease in glycerol production from 4 to 16 weeks of age. Fat cell size continued to increase between 4 and 16 weeks. Both fat cell diameter and volumes underwent a linear increase with age. However, the change was not reflected in epinephrine stimulated glycerol release. Therefore, glycerol release is inversely correlated with fat cell size during early development.The results indicate that age influences hormone stimulated lipolysis and is not dependent on cell size. Although the mechanism for the decreased lipolytic response of the isolated adipocytes was not discovered, it is believed that it may be due in part to a reduced number of receptors and to a reduced sensitivity of the cellular enzymatic system underlying lipolysis. / School of Physical Education

Macrophage foam cells are the major culprits in atherosclerotic lesions, having a prominent role in both lesion initiation and progression. With atherosclerosis being the main factor underlying cardiovascular complications, there is a long-standing interest on finding ways to reverse lipid buildup in plaques. Studies have shown that promoting reverse cholesterol transport (RCT) from macrophage foam cells is anti-atherogenic because it alleviates the cholesterol burden of the plaques. The goal of this thesis was to gain insight into the mechanisms that govern cholesterol efflux from macrophage foam cells. The first part of this study looked at the ability of different oxysterols to promote cholesterol efflux in unloaded as compared to lipid-loaded macrophages, and our major finding here is that epoxycholesterol decreases efflux in lipid-loaded macrophages. It appears that epoxycholesterol does so by impairing the release cholesterol from its cellular storage site, the lipid droplet (LD), where it accumulates in the form of cholesteryl esters (CE). These results highlighted the importance of cholesterol release from LDs for efflux; indeed, this process is increasingly being recognized as the rate-limiting step for RCT in vivo. Subsequent experiments aimed at elucidating the mechanisms that govern LD CE hydrolysis in macrophage foam cells lead to the discovery of a novel pathway involved in cholesterol efflux. Macrophage CE hydrolysis is classically defined as being entirely dependent on neutral CE hydrolases. In the second part of this study, we demonstrate that in addition to the canonical CE hydrolases, which mediate neutral lipid hydrolysis, lysosomal acid lipase (LAL) also participates in the hydrolysis of cytoplasmic CE. Autophagy is specifically triggered in macrophages by atherogenic lipoproteins and delivers LD CE to LAL in lysosomes, thus generating free cholesterol for efflux. This autophagy-mediated cholesterol efflux is a process that is primarily dependant on the ABCA1 transporter and, importantly, is important for whole-body RCT. Overall, the studies presented in this thesis support that macrophage LD CE hydrolysis is rate-limiting for cholesterol efflux and shed light on the mechanisms of cholesterol mobilization for efflux in macrophage foam cells.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2005. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2004. / Härtill 4 uppsatser.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2006. / Härtill 4 uppsatser.

Macrophage foam cells are the major culprits in atherosclerotic lesions, having a prominent role in both lesion initiation and progression. With atherosclerosis being the main factor underlying cardiovascular complications, there is a long-standing interest on finding ways to reverse lipid buildup in plaques. Studies have shown that promoting reverse cholesterol transport (RCT) from macrophage foam cells is anti-atherogenic because it alleviates the cholesterol burden of the plaques. The goal of this thesis was to gain insight into the mechanisms that govern cholesterol efflux from macrophage foam cells. The first part of this study looked at the ability of different oxysterols to promote cholesterol efflux in unloaded as compared to lipid-loaded macrophages, and our major finding here is that epoxycholesterol decreases efflux in lipid-loaded macrophages. It appears that epoxycholesterol does so by impairing the release cholesterol from its cellular storage site, the lipid droplet (LD), where it accumulates in the form of cholesteryl esters (CE). These results highlighted the importance of cholesterol release from LDs for efflux; indeed, this process is increasingly being recognized as the rate-limiting step for RCT in vivo. Subsequent experiments aimed at elucidating the mechanisms that govern LD CE hydrolysis in macrophage foam cells lead to the discovery of a novel pathway involved in cholesterol efflux. Macrophage CE hydrolysis is classically defined as being entirely dependent on neutral CE hydrolases. In the second part of this study, we demonstrate that in addition to the canonical CE hydrolases, which mediate neutral lipid hydrolysis, lysosomal acid lipase (LAL) also participates in the hydrolysis of cytoplasmic CE. Autophagy is specifically triggered in macrophages by atherogenic lipoproteins and delivers LD CE to LAL in lysosomes, thus generating free cholesterol for efflux. This autophagy-mediated cholesterol efflux is a process that is primarily dependant on the ABCA1 transporter and, importantly, is important for whole-body RCT. Overall, the studies presented in this thesis support that macrophage LD CE hydrolysis is rate-limiting for cholesterol efflux and shed light on the mechanisms of cholesterol mobilization for efflux in macrophage foam cells.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22304号 / 医博第4545号 / 新制||医||1040(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 川口 義弥, 教授 坂井 義治, 教授 羽賀 博典 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Obesity and obesity related diseases represent a leading cause of mortality in the United States and worldwide. Our research is oriented towards the role of lipid metabolism in longevity. Adipose triglyceride lipase, or ATGL, is a rate limiting enzyme in the lipolytic pathway. The nematode, C. elegans has many conserved biologic pathways to mammals, and the lipolytic pathway is one of them. The homologues include the insulin receptor (DAF-2), FoxO1 (DAF-16), and ATGL (ATGL-1). In this study, we use C. elegans as a model to study the role of lipolysis in longevity. It has been previously shown in our lab that overexpression of ATGL-1::GFP increases lifespan. To confirm that the increase in longevity was due to the overexpression of ATGL-1, we have used RNA interference to downregulate expression of ATGL-1::GFP. We have corroborated that ATGL-1::GFP worms have longer lifespans, than wildtype N2 worms. We have also found that RNAi control diet does not affect lifespan of ATGL-1::GFP strains. However, ATGL-1::GFP strains on an RNAi GFP diet demonstrate reduced levels of ATGL-1::GFP and have shorter lifespans compared to their control counterparts. Our findings confirm that overexpression of ATGL-1 increases lifespan of C. elegans probably due to its role in reducing fat content.