||Angiogenesis, the formation of new capillaries from pre-existing capillaries or venules, is essential to both health and disease and involved in many tissue-engineering applications. In addition to well-known angiogenic growth factors, mechanical factors, including cyclic stretch, have also been shown to affect angiogenesis, but how cyclic stretch regulates angiogenesis is not yet completely understood. Thus, the purpose of this work was to (1) identify stretch-mediated angiogenic control mechanisms and (2) elucidate how chemical and mechanical angiogenic regulators work in concert. We utilized a stretchable three-dimensional sprouting angiogenesis model, where endothelial cells were seeded on a collagen gel and could invade the gel while concomitantly being exposed to uniaxial cyclic stretch. We discovered that cyclic stretch alone is a strong angiogenic stimulus, and the effects of cyclic stretch on angiogenesis are both stretch magnitude and frequency dependent. We also discovered that cyclic stretch induced angiogenic sprouts to align perpendicular to the direction of stretch. Both the effects of cyclic stretch on the number and alignment of new sprouts were abolished by cytochalasin D (an inhibitor of actin polymerization) or Y27632 (an inhibitor of Rho associated kinase). In contrast, Sunitinib (an inhibitor of receptor tyrosine kinases) abolished cyclic stretch induced sprouting angiogenesis but not associated alignment, suggesting it is possible to separately manipulate the quantity and orientation of new sprouts. We also discovered that the combined effects of stretch and growth factors on angiogenesis varied with growth factors. Stretch and basic fibroblast growth factor (bFGF), but not vascular endothelial growth factor (VEGF), had an additive effect on angiogenesis, and such additive induction was abolished by cytochalasin D. Angiogenesis under the combination of stretch and VEGF was not sensitive to cytochalasin D, but was sensitive to Sunitinib, suggesting a change in molecular controls in the presence of different pro-angiogenic chemical factors. However, actin filaments were vital in stretch-mediated cell alignment, regardless of the presence of additional bFGF or VEGF. Overall, these results provide (1) an in-depth understanding of cyclic stretchmediated angiogenesis and (2) an in vitro experimental model for further experimentation within this research focus, driving toward future angiogenic therapies.