| Titel | Zeit | Ort | Dozent |
|---|---|---|---|
| Theory & discussion | 18.01.2021 09:00 - 12:00 (Mo) | Zeigerer, Anja | |
| Hepatocyte isolation | 18.01.2021 13:00 - 14:00 (Mo) | Zeigerer, Anja | |
| Metabolic measurements (body weight, fed blood glucose) in HFD/LFD mice Tissue culture plate preparation | 18.01.2021 14:00 - 17:00 (Mo) | Zeigerer, Anja | |
| Overview of the day Hepatocyte plating | 19.01.2021 09:00 - 12:00 (Di) | Zeigerer, Anja | |
| Imaging of steatotic liver tissue | 19.01.2021 13:00 - 14:00 (Di) | Zeigerer, Anja | |
| Hepatocyte culturing and fatty acid treatments | 19.01.2021 14:00 - 17:00 (Di) | Zeigerer, Anja | |
| Start starvation of mice | 20.01.2021 08:00 - 09:00 (Mi) | Zeigerer, Anja | |
| Overview of the day & FFA measurements in mouse serum | 20.01.2021 09:30 - 13:00 (Mi) | Krones-Herzig, Anja | |
| Glucose tolerance test in HFD and LFD mice | 20.01.2021 14:00 - 16:00 (Mi) | Zeigerer, Anja | |
| Overview of the day Seahorse assays in fatty acid treated primary hepatocytes | 21.01.2021 09:00 - 11:00 (Do) | Zeigerer, Anja | |
| Primary hepatocyte starvation for endocytosis assay | 21.01.2021 11:00 - 12:00 (Do) | Zeigerer, Anja | |
| EGF/LDL endocytosis assays, fixation & mounting Imaging of pre-stained mitochondria | 21.01.2021 13:00 - 16:00 (Do) | Zeigerer, Anja | |
| Microscopy and image analysis of endocytosis assay | 22.01.2021 09:00 - 12:00 (Fr) | Zeigerer, Anja | |
| Summary & exam | 22.01.2021 13:00 - 16:00 (Fr) | Zeigerer, Anja |
Prerequisites:
Basic knowledge in metabolism and liver structure
Knowledge and experience in pipette handling, scientific calculations and sterile work
Knowledge in cell culture is a plus
Level (Basic/Advanced): Intermediate
Description of Contents:
This course will focus on how diet induced changes affect metabolic homeostasis and can lead to obesity development and glucose intolerance. We will use in vivo approaches in high fat diet treated mice to investigate the consequences for glucose tolerance and serum lipid parameters. In addition, we will investigate, how excess fatty acids (FAs) lead to liver steatosis in vivo as well as in vitro. For the last experiment, primary mouse hepatocytes will be isolated, plated and treated with FAs in culture for 3 days to observe lipid droplet formation in vitro. The influence of hepatic steatosis on intracellular transport will be followed by high-‐resolution confocal microscopy and quantitative imaging. The plasticity of the metabolically active hepatocytes is visualized by measurements of the oxygen consumption rate for mitochondria function and immunofluorescence staining for mitochondria morphology. Altogether, this course will teach how high fat diet treatments affect hepatocellular energy metabolism and endocytosis as well as systemic glucose and lipid homeostasis.
Learning Objectives:
Metabolic control and responses to lipid overload in a mouse model of diet induced obesity leading to glucose intolerance.
Liver structure/function relationship and its importance for physiology
Isolation, plating and culturing of primary mouse hepatocytes for cell biological and metabolic assays
Immunohistochemistry, fluorescent uptake assays and seahorse analysis for mitochondria analysis in primary hepatocytes
Confocal microscopy and quantitative imaging
Elisa and plate reader assays for serum free fatty acids
Exam:
Documentation of the course in form of a scientific protocol
Final test about the content of the course