Technical developments in endothelial cell research
Technical developments in endothelial cell research
Blood vessels play an essential role in health and disease. Endothelial cells (ECs) residing in the microvasculature of tissues exhibit organ-specific molecular profiles that are influenced by their local microenvironment. Investigating these profiles requires methods to isolate ECs from the organs without compromising their molecular status. This can be achieved by Laser microdissection (LMD), enabling the study of ECs in human and animal tissues.
This thesis of Peter Zwiers aimed to develop methods for analysing ECs in vivo in both healthy and diseased conditions, with a particular focus on the distinct microvascular compartments of the kidney. Protocols combining LMD with RT-qPCR were optimized to ensure a linear correlation between dissected tissue area and quantification of microRNAs. Extending this workflow to integrate LMD with next-generation sequencing (NGS) allowed us to capture the complete transcriptome of ECs within the renal cortical microvasculature of healthy mice. Furthermore, we applied the LMD/NGS workflow to kidney biopsies of patients suffering from lupus nephritis (LN), identifying distinct gene expression profiles in glomeruli associated with disease activity.
Finally, we established an inducible EC-specific gene knockout model to investigate the function of EC-related molecules in adult mouse blood vessels. Gene knockout efficiency varied across organs and (micro)vessel types, altering EC responses to inflammation related molecules especially in microvascular regions with high knockout efficiency.
This research demonstrated the importance of isolating ECs from tissues by LMD to uncover the molecular basis of microvascular EC behaviour in organs and provides directions for future research in molecular pathology and drug development.