DataSheet_1_CD73-dependent generation of extracellular adenosine by vascular endothelial cells modulates de novo lipogenesis in adipose tissue.pdf

Next to white and brown adipocytes present in white and brown adipose tissue (WAT, BAT), vascular endothelial cells, tissue-resident macrophages and other immune cells have important roles in maintaining adipose tissue homeostasis but also contribute to the etiology of obesity-associated chronic inflammatory metabolic diseases. In addition to hormonal signals such as insulin and norepinephrine, extracellular adenine nucleotides modulate lipid storage, fatty acid release and thermogenic responses in adipose tissues. The complex regulation of extracellular adenine nucleotides involves a network of ectoenzymes that convert ATP via ADP and AMP to adenosine. However, in WAT and BAT the processing of extracellular adenine nucleotides and its relevance for intercellular communications are still largely unknown. Based on our observations that in adipose tissues the adenosine-generating enzyme CD73 is mainly expressed by vascular endothelial cells, we studied glucose and lipid handling, energy expenditure and adaptive thermogenesis in mice lacking endothelial CD73 housed at different ambient temperatures. Under conditions of thermogenic activation, CD73 expressed by endothelial cells is dispensable for the expression of thermogenic genes as well as energy expenditure. Notably, thermoneutral housing leading to a state of low energy expenditure and lipid accumulation in adipose tissues resulted in enhanced glucose uptake into WAT of endothelial CD73-deficient mice. This effect was associated with elevated expression levels of de novo lipogenesis genes. Mechanistic studies provide evidence that extracellular adenosine is imported into adipocytes and converted to AMP by adenosine kinase. Subsequently, activation of the AMP kinase lowers the expression of de novo lipogenesis genes, most likely via inactivation of the transcription factor carbohydrate response element binding protein (ChREBP). In conclusion, this study demonstrates that endothelial-derived extracellular adenosine generated via the ectoenzyme CD73 is a paracrine factor shaping lipid metabolism in WAT.</p

Image1_Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice.tif

Brown adipose tissue (BAT) has emerged as an appealing therapeutic target for cardio metabolic diseases. BAT is a heat-producing organ and upon activation substantially lowers hyperlipidemia. In response to cold exposure, not only the uptake of lipids into BAT is increased but also the Cyp7b1-mediated synthesis of bile acids (BA) from cholesterol in the liver is triggered. In addition to their role for intestinal lipid digestion, BA act as endocrine signals that can activate thermogenesis in BAT. When exposed to cold temperatures, Cyp7b1−/− mice have compromised BAT function along with reduced fecal bile acid levels. Here, we aim to evaluate the role of Cyp7b1 for BAT-dependent lipid clearance. Using metabolic studies with radioactive tracers, we show that in response to a cold stimulus, BAT-mediated clearance of fatty acids derived from triglyceride-rich lipoproteins (TRL), and their remnants are reduced in Cyp7b1−/− mice. The impaired lipid uptake can be explained by reduced BAT lipoprotein lipase (LPL) levels and compromised organ activity in Cyp7b1−/− mice, which may be linked to impaired insulin signaling. Overall, our findings reveal that alterations of systemic lipoprotein metabolism mediated by cold-activated BAT are dependent, at least in part, on CYP7Β1.</p

Image3_Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice.tif

Brown adipose tissue (BAT) has emerged as an appealing therapeutic target for cardio metabolic diseases. BAT is a heat-producing organ and upon activation substantially lowers hyperlipidemia. In response to cold exposure, not only the uptake of lipids into BAT is increased but also the Cyp7b1-mediated synthesis of bile acids (BA) from cholesterol in the liver is triggered. In addition to their role for intestinal lipid digestion, BA act as endocrine signals that can activate thermogenesis in BAT. When exposed to cold temperatures, Cyp7b1−/− mice have compromised BAT function along with reduced fecal bile acid levels. Here, we aim to evaluate the role of Cyp7b1 for BAT-dependent lipid clearance. Using metabolic studies with radioactive tracers, we show that in response to a cold stimulus, BAT-mediated clearance of fatty acids derived from triglyceride-rich lipoproteins (TRL), and their remnants are reduced in Cyp7b1−/− mice. The impaired lipid uptake can be explained by reduced BAT lipoprotein lipase (LPL) levels and compromised organ activity in Cyp7b1−/− mice, which may be linked to impaired insulin signaling. Overall, our findings reveal that alterations of systemic lipoprotein metabolism mediated by cold-activated BAT are dependent, at least in part, on CYP7Β1.</p

Image2_Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice.tif

Brown adipose tissue (BAT) has emerged as an appealing therapeutic target for cardio metabolic diseases. BAT is a heat-producing organ and upon activation substantially lowers hyperlipidemia. In response to cold exposure, not only the uptake of lipids into BAT is increased but also the Cyp7b1-mediated synthesis of bile acids (BA) from cholesterol in the liver is triggered. In addition to their role for intestinal lipid digestion, BA act as endocrine signals that can activate thermogenesis in BAT. When exposed to cold temperatures, Cyp7b1−/− mice have compromised BAT function along with reduced fecal bile acid levels. Here, we aim to evaluate the role of Cyp7b1 for BAT-dependent lipid clearance. Using metabolic studies with radioactive tracers, we show that in response to a cold stimulus, BAT-mediated clearance of fatty acids derived from triglyceride-rich lipoproteins (TRL), and their remnants are reduced in Cyp7b1−/− mice. The impaired lipid uptake can be explained by reduced BAT lipoprotein lipase (LPL) levels and compromised organ activity in Cyp7b1−/− mice, which may be linked to impaired insulin signaling. Overall, our findings reveal that alterations of systemic lipoprotein metabolism mediated by cold-activated BAT are dependent, at least in part, on CYP7Β1.</p

Image4_Cold-Induced Lipoprotein Clearance in Cyp7b1-Deficient Mice.tif

Brown adipose tissue (BAT) has emerged as an appealing therapeutic target for cardio metabolic diseases. BAT is a heat-producing organ and upon activation substantially lowers hyperlipidemia. In response to cold exposure, not only the uptake of lipids into BAT is increased but also the Cyp7b1-mediated synthesis of bile acids (BA) from cholesterol in the liver is triggered. In addition to their role for intestinal lipid digestion, BA act as endocrine signals that can activate thermogenesis in BAT. When exposed to cold temperatures, Cyp7b1−/− mice have compromised BAT function along with reduced fecal bile acid levels. Here, we aim to evaluate the role of Cyp7b1 for BAT-dependent lipid clearance. Using metabolic studies with radioactive tracers, we show that in response to a cold stimulus, BAT-mediated clearance of fatty acids derived from triglyceride-rich lipoproteins (TRL), and their remnants are reduced in Cyp7b1−/− mice. The impaired lipid uptake can be explained by reduced BAT lipoprotein lipase (LPL) levels and compromised organ activity in Cyp7b1−/− mice, which may be linked to impaired insulin signaling. Overall, our findings reveal that alterations of systemic lipoprotein metabolism mediated by cold-activated BAT are dependent, at least in part, on CYP7Β1.</p

EpCAM expression of human malignant cells.

<p>(A) The mRNA of the malignant cell lines 5061, 5072 (pancreatic cancer), LNCAP, PC3 (prostate cancer), FemX-1, MEWO (melanoma), T47D, MCF7 (breast cancer), HT29 (colon cancer) and OH-1 (small cell lung cancer) were relatively quantified by qPCR, using GAPDH for normalization. 5072, LNCAP, PC3, T47D, MCF7, Caco2 and HT29 showed high expression levels of EpCAM mRNA. (B) EpCAM could be detected by Western blot analysis of HT29 cell lysate with a specific binding to antibody MOC31. Beta-actin was used as loading control. (C) EpCAM could positively be detected by flow cytometry analysis with MOC31 on all cancer cell lines, except of FemX-1 and MEWO. Isotype controls are shown as dotted lines.</p

Blood half life of mAbs T84.1 and IgG1.

<p>The blood half life of [¹²⁵I]-labelled T84.1 and nonspecific [¹²⁵I]-labelled IgG1 antibodies were determined in FemX-1 tumour bearing SCID mice. The blood half life of specific and control antibody were identical (T84.1 = 10.3 h (n = 4), IgG1 = 10.4 h (n = 4)).</p

MRI of HT29 xenograft.

<p>HT29 colon carcinoma (a) grown at the subcutaneous injection site above muscles (b) appear hyper-intense in two-dimensional turbo spin-echo (TSE) sequence (MR images in axial orientation).</p

CEACAM in vivo binding.

<p>[¹²⁵I]-Labelled T84.1 and nonspecific [¹²⁵I]-labelled IgG1 antibody were used for CEACAM <i>in vivo</i> binding in FemX-1 melanoma bearing SCID mice. There is a significant difference between specific and control antibody by FemX-1 melanoma, but not in other organs including blood (Two-way ANOVA, P<0.001, n = 6).</p

EpCAM <i>in vivo</i> binding.

<p>[125I]-Labelled specific anti EpCAM MOC31 and non-specific [125I]-labelled IgG1 antibody were used for EpCAM <i>in vivo</i> binding in HT29 carcinoma bearing SCID mice. There is a significant difference between specific and control antibody by HT29 carcinoma, but except for spleen and blood not in other organs (two-way ANOVA, P<0.001, n = 3). Standard deviations are indicated by bars.</p