In the mouse embryo, the first definitive haematopoietic stem cells (HSCs), capable of repopulating adult irradiated mice, emerge at mid-gestation by embryonic day E11. At this stage, the aorta-gonad-mesonephros (AGM) region is able to initiate and expand HSCs. Recently, it has been shown that the development of HSC in the AGM region results from the maturation of haematopoietic precursors called pre-HSCs. Mounting evidence points at an endothelial origin for these cells, the haematogenic endothelium. Analysis of VEGFs mutants, a critical pathway for endothelial developement, suggested that it also plays a role during early haematopoiesis. The main receptor of the pathway, FLK-1 (also known as VEGRR2 or KDR), is expressed in early hematopoietic and endothelial cells in the mouse embryo. Knock-out mutants for Flk-1 showed a decrease of endothelial and intra-embryonic haematopoietic progenitors. Although Flk-1 has been identified as an essential gene for HSC emergence, its exact point of action in HSC development remains unknown. In this thesis, I investigated the role of FLK-1 signalling in haematopoietic development and defined precise stages and cell types during HSC emergence in which FLK-1 is critically involved. by using a reporter line and antibody staining, I demonstrated that FLK-1 is expressed in the pre-HSCs/HSC lineage. Germ-line Flk-1 knockout results in embryonic lethality at around E9.0, before HSC emergence, mainly due to defects in vasculogenesis. Since arterial specification precedes HSC formation, it has never been elucidated whether the haematopoietic defects found in the knockouts are a secondary effect of the loss of vasculature or it FLK-1 is directly involved in haematopoietic specification. Therefore, to determine the role of the receptor in HSC development, I used a conditional inducible mutagenesis approach that allowed the deletion of Flk-1 precisely when pre-HSCs mature into HSCs at E10.5 and E11.5. My data showed that Flk-1 deletion at these stages affects both endothelial and haematopoietic progenitors, as well as HSCs. This suggests that the VEGF pathway is not only essential in early stages of haematopoietic development, as previously demonstrated, but it may be also involved in the maturation of pre HSC into HSCs at later stages.

A new concept quickly gaining ground in the field of cancer research is that the inflammatory process plays a key role in cancer development and metastasis; however, the molecular mechanisms of such an involvement in cancer progression remain largely unspecified. YKL-40, also known as human cartilage glycoprotein 39, is a secreted heparin-binding protein with ties to both cancers and inflammatory disease. In these diseases, YKL-40 has been suggested to play a role in regulating tissue and extracellular matrix remodeling. It has been found that in certain cancers, including breast, colorectal and brain, that high YKL-40 serum levels correlate with poor outcome, and consequently it may serve as a biomarker. Our recent study has shown that tumor-derived YKL-40 acts as an angiogenic factor due to its ability to up-regulate vessel formation and metastasis during tumor development. However, blockade of the function of YKL-40, which implicates therapeutic value, has not been explored yet. The goal of this project was to better understand the importance of tumor-derived YKL-40 in angiogenesis through both functional and structural studies. By establishing a monoclonal YKL-40 antibody for blocking YKL-40, the function of tumor-derived YKL-40 in inducing endothelial cell angiogenesis and tumor cell survival was uncovered, confirming YKL-40's importance in tumor signaling as well as offering evidence in the benefit of its neutralization. Additionally, a postulated heparin-binding domain on YKL-40 was mutated in hopes of revealing the relevance of this binding ability on YKL-40's function and whether this could serve as a target in inhibiting YKL-40 signaling.

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