**3. Endothelial cells**

Endothelial cells are crucial in the promotion of cancer cell migration, invasion, and metastasis. During tumorigenesis, gaseous exchange and nutrient transport occur by passive diffusion however an increase in the volume of the tumours (1–2 mm3 ) leads to insufficient oxygen and a build-up of metabolic waste in the tumour microenvironment [6, 7, 16, 23]. This makes the tumour microenvironment to become hypoxic and acidic thereby highlighting a need for the tumours to develop their own blood supply to overcome this.

The vascular endothelium, a thin layer of endothelial cells, aids in orchestrating the separation of circulating blood from tissues, delivery of water, oxygen and nutrients, movement and adhesion of leucocytes, and formation of blood vessels within the tumour microenvironment [6, 7, 23]. The vascular endothelium is highly organised and hierarchical in structure, and this enables the interaction between stromal and non-stromal cells to provide support and stability for the blood vessels.

Tumours co-opt existing blood vessels and induce growth of new blood vessels by a mechanism known as vessel sprouting [7, 23]. Abnormal sprouts are characteristic of the tumour vasculature along with intercellular gaps and no hierarchical arrangement. These vascular endothelial cells within the tumour microenvironment interact with tumour cells and other stromal cells to promote tumorigenesis and metastasis.

The hypoxic tumour microenvironment leads to expression and activation of hypoxia-inducible factors (HIFs) that co-ordinate cellular response to low oxygen levels. These HIFs then instruct the endothelial cells to secrete and release proangiogenic factors including vascular endothelial growth factor (VEGF), platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) thereby initiating vessel sprouting [6, 7, 23, 24]. These proangiogenic factors, especially VEGF then promote vascular permeability and angiogenesis by stimulating the migration of endothelial cells to form new blood vessel lumen in an autocrine and paracrine fashion. Activation of VEGF receptors (VEGFR) on endothelial cells also activate several downstream signalling pathways including the mitogen-activated protein kinases and extracellular signal-regulated kinases (MAPK/ERK) and phosphatidylinositol-3 kinases (PI3K/ Akt) pathways involved in the regulation of cell survival, cell cycle progression, cell growth and angiogenesis [23].

The endothelial cells secrete proteins to form new basement membranes, which are often immature and fail to reach final stages of maturation thus resulting in a leaky vasculature [6, 7, 23]. Endothelial cells communicate with the basement membrane and the ECM through integrin proteins (collagen, elastin, fibronectin and fibrillin) and proteoglycans for mechanical and physical support [25]. The basement membrane degrades to activate stroma thus allowing activated stroma to have a direct contact with tumour cells during tumorigenesis [26]. This induces alterations such as enhanced vascularity and increased ECM production, which are all essential for invasion.

During tumour metastasis, following cell detachment, tumour cells first undergo intravasation by first escaping the primary tumour site and enter the vasculature [6, 7, 23, 27]. Upon entering the vasculature, the tumour cells then adhere to endothelial cells during intravasation thereby changing the endothelial barrier, which allows the tumour cells to migrate between two endothelial cells. This signifies that the interaction between the endothelial cells and tumour cells is reciprocal. The tumour cells can differentiate into endothelial cells within the tumour microenvironment to support and sustain tumour growth. Endothelial cells can also change cell fate and often undergo endothelial-mesenchymal transition (EMT), organised by TGF-β, EGF and bone morphogenetic protein (BMP), to cancer-associated fibroblasts (CAFs) during tumour progression [6, 24, 28]. This leads to a loss in cell-to-cell connection, detachment and elongation, enhanced migration, and a loss of endothelial properties. Cell adhesion establishes a tight a tight connection between cells as well as between cells and ECM thus activating cell proliferation and survival pathways [7]. Therefore, this loss in cell-to-cell connections enables cancer cells to transverse the vasculature (extravasation) and interact with pre-metastatic niches that permits cell proliferation and colonisation at the secondary sites [7, 24].

Furthermore, vascularization and endothelial cell expansion enhance tumour initiation and self-renewal properties of cancer stem cells within the tumour microenvironment. Endothelial cells secrete soluble factors that aid in the maintenance of stem cell properties in neural stem cells and activate cancer cells thereby promoting tumour growth [6]. Endothelial cells also secrete cytokines such as IL-8, which promote characteristics of cancer stem cells in glioblastoma including their migration and invasion abilities [6]. In a positive feedback loop of IL-8 mediated signalling, glioblastoma cells induce endothelial cell migration toward the tumour bulk thereby promoting brain tumour growth. In oesophageal cancer, epiregulin (EREG) overexpression is induced by endothelial cells and this leads to an increase in actin rearrangement, spheroid formation and enrichment of cancer stem cells [6].
