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Membrane dynamics of lipid droplets

Membrane dynamics of lipid droplets in liver regeneration This research line is embedded in the Research programs Tissue Repair of the Faculty of Veterinary Medicine and in Growth & Differentiation of Utrecht University Hepatic stellate cells (HSCs) are non-parenchymal cells that are located perisinusoidally in the space of Disse and comprise about 5-10% of the total liver cell population. At the onset of liver injury or repair, hepatic stellate cells become activated during which process the quiescent HSC undergoes a gradual transformation from a non-dividing lipid storing phenotype into a proliferative myofibroblastic phenotype. HSC activation has been recognized as initial steps in the process of liver cirrhosis and repair. It is therefore important to understand the molecular mechanisms that underlie the activation process of HSCs. Quiescent HSCs have a lipid storing phenotype as indicated by the presence of large lipid droplets. Interestingly, during the process of activation the HSCs rapidly loose their lipid droplets.

0 Days 14 Days
  Bodipy (LDs)
Figure: Freshly isolated HSCs were cultured and fixed after 2 h (left panels) or 14 days (right panels). Morphology and neutral lipid content was analyzed by differential interference contrast microscopy (DIC, top panels) and fluorescence microscopy after Bodipy staining of LDs (bottom panels). Testerink et al (2012) PLoS One.

Lipid droplets exist of a core of neutral lipids surrounded by a phospholipid monolayer. In most cells, the neutral lipids stored in lipid droplets consist of triacylglycerols (TAGs) and cholesterolesters. In HSCs the LDs contains in addition to these neutral lipids also retinyl esters. In fact, the surplus of retinol/vitamin A is mainly is mainly stored as retinyl esters in hepatic stellate cells. Lipid droplet dynamics during HSC activation To obtain more insight in the mechanism of LD loss and its role in HSC activation, we investigate the LD dynamics in HSCs. With a combined approach of confocal Raman microspectroscopy and mass spectrometric analysis of lipids (lipidomics) we recently demonstrated that in individual quiescent HSCs all LDs have a similar Raman spectrum, suggesting a similar lipid profile of retinylesters, cholesterol esters and triacylglycerols.

Figure: Freshly isolated HSCs were cultured and fixed at day 0 (quiescent state). Confocal Raman microscpectroscopy on LD enriched regions was performed and Cluster images (20 × 20 μm2) were constructed from Raman imaging data of the square area in the white light image. Each color represents a different cluster. The cluster averages show the average Raman spectra extracted from the black, pink, green and blue clusters displayed in the cluster image. * indicates (characteristic) RE peaks; # indicates characteristic acyl peak (Testerink et al (2012) PLoS One.

Together with lipidomic analysis, we were able to show a highly dynamic behavior of lipid droplets as well as a high lipid turnover within lipid droplets. The rapid replacement of retinyl esters by polyunsaturated fatty acids in LDs suggests a role for both retinoid and eicosanoid signaling during HSC activation (Testerink et al (2012) PLoS One).