Periodic Reporting for period 2 - GLUCOBAT (In vivo fuel utilization and metabolic regulation by brown adipose tissue)
Période du rapport: 2023-10-01 au 2024-09-30
Brown adipose tissue (BAT) thermogenesis has gained significant clinical interest as a strategy against obesity and diabetes because of its potential to expend excess calories as heat. However, our understanding of BAT metabolism on an organismal level is still rudimentary, thus, to advance BAT-targeted therapies, we must first comprehensively define how BAT utilizes its key fuels, such as glucose in vivo. It is widely assumed that the major role of glucose during thermogenesis is to provide energy for the TCA cycle and subsequent uncoupling by the key thermogenic protein in BAT, uncoupling protein 1 (UCP1). However, recent literature reveal that this assumption is oversimplified since glucose has many fates once it enters cells, feeding into additional metabolic pathways beyond the TCA cycle, which are necessary for optimum thermogenesis.
The overall goal of this project is to investigate if and how glucose supports multiple metabolic pathways, beyond providing energy for the TCA cycle, that are required for thermogenesis, and moreover, how the fate of glucose-derived metabolites potentially varies depending on the thermogenic stimulus and duration. In addition, the role of cold-regulated solute carrier (SLC)-transporters in regulating glucose metabolism and thermogenesis will be investigated.
This project will expand our understanding of how thermogenic adipocytes utilize glucose and will potentially reveal novel therapeutic strategies to enhance thermogenesis.
The overall goal of this project is to investigate if and how glucose supports multiple metabolic pathways, beyond providing energy for the TCA cycle, that are required for thermogenesis, and moreover, how the fate of glucose-derived metabolites potentially varies depending on the thermogenic stimulus and duration. In addition, the role of cold-regulated solute carrier (SLC)-transporters in regulating glucose metabolism and thermogenesis will be investigated.
This project will expand our understanding of how thermogenic adipocytes utilize glucose and will potentially reveal novel therapeutic strategies to enhance thermogenesis.
In this project, I have used state-of-the-art glucose-tracing technology that utilizes mass spectrometry, combined with novel CRISPR/Cas9 genome editing strategies to precisely define how thermogenic adipose tissue utilizes glucose in vivo.
I have performed tracing experiments in brown adipocytes in vitro to investigate the importance of glucose for thermogenesis, to validate findings performed in vivo. Glucose tracing in mice acclimated to different temperatures (TN (30 °C), RT (22 °C) or to SC (6 °C)) and acutely to cold showed and confirmed my hypothesis that glucose carbons are rapidly incorporated into glycolysis and TCA cycle, as well as several auxiliary pathways, including NADPH, nucleotide, and phospholipid synthesis pathways. Furthermore, I have investigated with other carbon tracers if other carbon sources could be utilized by the BAT as energy for the TCA to drive thermogenesis. I have optimized and performed stable isotopic infusion tracing experiments to assess minimum-perturbative glucose tracing in mice acclimated to different temperatures and in UCP1-loss-of-function mice.
In this project I have investigated the role of solute carrier transporters of the SCL family for glucose utilization and thermogenesis in BAT. To investigate the most cold-regulated SLC-transporters in BAT, I assessed the expression of one of the transporters after acclimation to different temperatures and in different tissues. The mRNA expression was induced in BAT and in inguinal white adipose tissue in cold, which indicate a function of this transporter for thermogenesis. I furthermore investigated the loss-of-function by performing siRNA transfections in mature brown adipocytes and observed changed oxygen consumption measured on the Seahorse XF analyser. CRISPR/Cas9 gene editing technology has been used to investigate the loss-of-function in vivo.
I have performed tracing experiments in brown adipocytes in vitro to investigate the importance of glucose for thermogenesis, to validate findings performed in vivo. Glucose tracing in mice acclimated to different temperatures (TN (30 °C), RT (22 °C) or to SC (6 °C)) and acutely to cold showed and confirmed my hypothesis that glucose carbons are rapidly incorporated into glycolysis and TCA cycle, as well as several auxiliary pathways, including NADPH, nucleotide, and phospholipid synthesis pathways. Furthermore, I have investigated with other carbon tracers if other carbon sources could be utilized by the BAT as energy for the TCA to drive thermogenesis. I have optimized and performed stable isotopic infusion tracing experiments to assess minimum-perturbative glucose tracing in mice acclimated to different temperatures and in UCP1-loss-of-function mice.
In this project I have investigated the role of solute carrier transporters of the SCL family for glucose utilization and thermogenesis in BAT. To investigate the most cold-regulated SLC-transporters in BAT, I assessed the expression of one of the transporters after acclimation to different temperatures and in different tissues. The mRNA expression was induced in BAT and in inguinal white adipose tissue in cold, which indicate a function of this transporter for thermogenesis. I furthermore investigated the loss-of-function by performing siRNA transfections in mature brown adipocytes and observed changed oxygen consumption measured on the Seahorse XF analyser. CRISPR/Cas9 gene editing technology has been used to investigate the loss-of-function in vivo.
Since data analysis is still ongoing, the final impact of the project remains to be determined, but by increasing our knowledge on the glucose utilization in BAT, this project could provide bases for the development of targeted therapies.