Among the components of the VEGF signaling pathway, the three VEGF receptors and their coreceptor Nrp2 were shown to be strongly down-regulated in LSEC in vitro. Wnt-2, previously identified by us as a positive regulator of VegfR2, and its receptors were also drastically decreased upon culture.

Thus, defective Wnt signaling may enhance and synergize with defective Vegf receptor activity in cultured LSEC in a vicious circle. Furthermore, Sirolimus mw primary Vegf receptor deficiency in cultured LSEC may explain impaired LSEC proliferation in culture despite the presence of high concentrations of Vegf in LSEC culture media. As of now, a unique blood vascular EC-specific master regulator, as is Prox1 for lymphatic EC, has not be identified and specific gene expression in different blood vascular EC is thought to be mediated by combinations of otherwise

nonspecific transcriptional regulators. The LSEC-specific transcription factors Tfec, Gata4, Maf, and Lmo3 identified here may well represent such an EC subtype-specific combination of transcriptional regulators. Gata4 has been shown to be important in development of the liver and of cardiac myocytes. Although Gata2, 3, and 6 are expressed in different EC and transcriptionally target EC-specific genes such as vWF, VCAM-1, and Tie-2,22 Gata4 is not generally expressed in blood vascular selleck compound EC. Interestingly, endothelial Gata4 expression specifically induces the formation of the heart valves, a site where the sinusoidal endothelial marker proteins Stabilin-1 and -2 are also expressed.23 Thus, specific overexpression of Gata4 in LSEC versus LMEC-associated overexpression of Gata2, 3, and 5 renders Gata4 an attractive candidate for at least coregulating LSEC-specific gene transcription. Tfec, a bHLH transcription factor of the Mitf family contributes to IL-4-induced macrophage activation, suggesting a possible role in regulation of immune system processes in selleck LSEC; interestingly, the Mitf-family member Tfeb is involved in placental vascularization.24 Although the proto-oncogene c-Maf has been

shown to induce the angiogenic surface aminopeptidase N/CD13 in EC in vitro, our microarray analysis showed that CD13 expression was decreased in LSEC versus, LMEC indicating that MAF may target different genes in LSEC.25 Because Lmo family members have been shown to interact with Gata and bHLH transcription factors,26 Lmo3 could be involved in the regulation of LSEC-specific gene expression, possibly by interaction with Gata4 and/or Tfec. In this study we furthermore show that the LSEC-specific differentiation program comprises a novel, highly conserved 278 aa type-1 transmembrane protein selectively expressed in liver endothelium that was named liver endothelial differentiation-associated protein (Leda)-1.