Arnar Ingi Vilhjálmsson and Óttar Rolfsson
Patients suffering from shock-induced endotheliopathy (SHINE) share phenotypic features of endothelial dysfunction and sustained high levels of circulating catecholamines. These phenotypic features correlate with poor disease outcome and are statistically associated with development of organ failure and death. Using an in vitro endothelial cell culture model treated with catecholamines, we aim to further understand the molecular response in SHINE. Alterations in the transcriptome are observed upon high levels of catecholamines, significantly increasing GRAMD1B transcription which is involved in intracellular cholesterol transport. The role of GRAMD1B in the response to catecholamines is yet to be clarified.
Using CRISPR-Cas9, a knock-out cell line was established to investigate the role of GRAMD1B in relation to SHINE. Seahorse assays were used to assess changes in cellular oxygen consumption rate and extracellular acidification rate. Liquid chromatography-mass spectrometry (LC-MS) was utilized to measure changes within the proteome, metabolome and lipidome following catecholamine stimulation.
We observed changes in the energy demand of endothelial cells treated with catecholamines, where oxygen consumption and glycolysis were increased. Additionally, changes observed in the proteome, metabolome and lipidome further support the transcriptomic data. Comparing the response of the knock-out cell line, the absence of GRAMD1B leads to impaired mitochondrial function and distinct changes in the lipidome, effecting cholesteryl ester and fatty acid metabolism.
GRAMD1B thus plays an important role in mitochondrial function, effecting lipid metabolism in response to catecholamine stimulation.
