Identification of Transcription Factors that Regulate Liver Sinusoidal Endothelial Cell Specialization and Liver Regeneration

The liver has a high regenerative capacity, which is often lost in liver disease. Liver sinusoidal endothelial cells (LSECs) are crucial in liver development and regeneration since capillaries provide nutrients and oxygen and LSECs secrete growth factors that stimulate hepatocyte proliferation. LSECs are highly specialized to allow for selective transport to hepatocytes. We identified ~30 genes that are highly enriched in LSECs among which 7 transcription factors (TFs; including ZEB2). We aimed to: (1) investigate how these 7 TFs regulate the expression of LSEC markers and whether they may drive LSEC specialization; (2) determine the role these TFs (and that of ZEB2 in particular) during liver development, maintenance and disease. We generated lentiviruses to induce the expression of the 7 TFs in human umbilical vein ECs (HUVECs) and blood outgrowth ECs (BOECs). We identified c-MAF, GATA4 and MEIS2 as major regulators of LSEC marker expression, together being able to regulate about 50% of the LSEC signature genes. Using a lentivirus expressing ZEB2, we found an increase of LYVE1 (a hyaluronan scavenger receptor) and a downregulation of CD31 (the expression of which is low in LSECs in the healthy liver but upregulated in liver disease) and CD34 (the expression of which is low in mature LSECs). To unravel the role of ZEB2 in vivo, we generated conditional mouse models lacking or overexpressing Zeb2 specifically in the endothelium. Using flow cytometry, we found recombination in ~80% of liver ECs. Complementary to our in vitro results, endothelial Zeb2 deficiency resulted in lower Lyve1 expression in recombined liver ECs. We will use these genetic models to further study the role of endothelial Zeb2 in liver maintenance and disease. We conclude that c-MAF, MEIS2 and GATA4 are major regulators of the LSEC phenotype and that ZEB2 may play a role in LSECs by regulating LYVE1 expression.status: publishe