Angiogenesis: Steps and Regulation

• Angiogenesis, or new vessel formation, takes place during embryo development, in postnatal life, in the female genital system cyclically, and in wound repair.
• It is limited in time and depends on the balance between activator and inhibitor systems, maintaining microcirculation in a quiescent state with low endothelial cell proliferation.
• Quiescent endothelium rests on an extracellular matrix (ECM), specifically the basement membrane, which contains laminin and type IV collagen.
• Angiogenesis proceeds in five steps, regardless of the initiating stimulus:

Angiogenesis Steps:

• Basement membrane degradation using enzymes secreted by endothelial cells
• Migration of endothelial cells toward the inducing stimulus
• Proliferation of endothelial cells
• Canalization, branching, and formation of vascular loops to establish a circulatory network
• Perivascular apposition of pericytes and neosynthesis of basement membrane components by endothelial cells and pericytes
• New microvessels develop via:

Microvessel Development:

• Budding from preexisting vessels
• Participation of circulating endothelial progenitor cells
• Intussusception, where endothelial cells in preexisting vessels bridge the lumen

Angiogenesis Dependence

• All mechanisms involve loosening preexisting endothelial cells from junctions, this being maintained by proteins like vascular endothelial (VE)-cadherin and platelet-endothelial cell adhesion molecule (PECAM)
• Angiopoietin-1 (Ang-1) increases junctions with pericytes, while Ang-2 decreases junctions
• Integrins like alpha beta3 govern the attachment to basement membrane proteins
• Local proteinases and their inhibitors are active in all mechanisms.
• Dilation of microvessels, mediated by nitric oxide synthase (NOS), also aids in angiogenesis
• Angiogenesis relies on the balance between positive and negative modulators within the vascular microenvironment.

Tumor Angiogenesis

• Tumor angiogenesis, essential for tumor progression in growth, invasion, and metastasis, shares similar steps but is uncontrolled and has increased endothelial cell proliferation

• The avascular phase of tumors has been studied using tumor spheroids and implants Tumor cell mass grows with the cube of radius, while surface area increases with the square

• Tumor growth reaches a steady state when the area limits nutrient supply and metabolite removal

• Spheroids are nourished by diffusion and reach a steady state when cell loss equals cell gain, resulting in slow ("dormant") proliferation and no metastasis

• Conversely, tumors implanted onto angiogenic sites are rapidly vascularized, leading to exponential proliferation and high steady-state levels

• Mitotic index is inversely correlated with tumor cell distance from the vessel:

• Cells closest to capillaries have highest index

• O₂ diffuses to ~150 µm, allowing a capillary (~100 µm long, 20-100 endothelial cells) to support ~10⁴ viable tumor cells

• Capillary elongation (~800 µm/day) can support ~10⁴ tumor cells/day, indicating an amplification loop for tumor growth

• The vascular phase corresponds to invasive and metastatic tumors, which have intravasation and colonization conditions

• Tumor vessels are lined with discontinuous endothelium, favoring cell transmigration

• To colonize other organs and grow, tumor cells induce further angiogenesis or remain as dormant micrometastases.

• The vascular phase parallels neoplastic changeover which is the progression of neoplastic or preneoplastic cell capability to induce new vessels.

• Ultimately, transitioning from avascular to vascular phase allows escape from the "dormant" phase.

Tumor Blood Vessel Characteristics

• Aberrant phenotype and genetic profile
• Chaotic mixture of dilated and tortuous channels lacking features of arterioles, capillaries, or venules
• Arteriovenous shunts
• Excessive branching
• Uneven diameters
• Increased permeability
• Endothelium of tumor-associated cells proliferates rapidly, has discontinuities, and facilitates macromolecule permeability and tumor cell traffic

Basement Membrane and Pericytes

• May have extra layers lacking association with cells
• Pericytes are loosely associated, have abnormal shapes, extend cytoplasmic processes, and have extra layers
• Abnormal vessel structures coincide with altered gene and protein expression
• Findings indicate a specialized tumor endothelial cell transcription profile, which is genetically different from other angiogenesis linked activation patterns

Angiogenesis Role

• Angiogenesis facilitates tumor access to nutrients and oxygen and a way to spread to other parts of the organism
• Other mechanisms contribute to tumor vascularization:
• Recruitment of circulating endothelial precursor cells (EPCs)
• Co-option of pre-existing blood vessels -Vascular mimicry
• Marrow-derived EPCs incorporation into tumor blood vessels is limited in effect
• Glioblastoma multiforme exemplifies co-option, in which tumor cells form cuffs around brain blood vessels
• Ocular melanomas exemplify vascular mimicry, in which tumor cells line spaces filled with red blood cells
• Solid tumors grow in two phases: avascular and vascular
• Dependence on angiogenesis and angiogenic factor release is evidence of progression from neoplastic transformation in tumor growth and metastasis
• Angiogenesis is important for hematological malignancies; increased microvascular density (MVD) in bone marrow indicates the importance of said angiogenesis.