Table 2.

Examples of 3D vascular drug screening models developed using different tissue engineering approaches including traditional tissue engineering, self-organization (3D bioprinting, cell sheets, and organoids), and self- assembly.

Approach	Cell type (s)	Scaffold/biomaterial	3D model	References
Traditional tissue engineering	Human coronary artery smooth muscle cells and HUVECs	Type I collagen, and aligned PLA nanofibers scaffold	Model (17, 18): Layer-by-layer assembly of a medial layer composed of human coronary artery smooth muscle cells in type I collagen, covered with an intimal layer composed of a HUVEC-seeded aligned PLA nanofibers scaffold. Application (17): Thrombosis model: Perfusion of human blood or platelets under physiological flow conditions using a parallel-plate flow chamber. Application (18): Real time monitoring of cytosolic Ca2+ in human platelets exposed to tissue engineered vessels to quantitatively compare the construct ability to promote or prevent platelet activation.	(17, 18)
	Endothelial and smooth muscle cells derived from human embryonic stem cells and iPSCs	Fibrin gels	Model: Cells were cultured into fibrin gels to induce 3D tissue formation. Application: The system was used to test a high-throughput screening strategy to assess chemical toxicity and drug efficacy.	(19)
	Human neonatal dermal fibroblasts or human bone marrow-derived MSCs	Rat-tail collagen I matrices	Model: Dense collagen gel matrices were developed by embedding human neonatal dermal fibroblasts or human bone marrow derived MSCs in rat-tail collagen I. The seeded matrix was then poured into a mandrel and allowed to gel. Application: Custom-made perfusion bioreactor chamber to test pharmacological and immunological responses of tissue engineered vascular grafts.	(20)
	Primary or iPSC-derived smooth muscle cells and EPCs	Collagen gel	Model: Medial cells (primary or iPSC-derived smooth muscle cells) embedded in a mixture of collagen gel and injected into molds to fabricate arteriole-scale human vessel grafts that are then endothelialized in the perfusion chamber.	(21)
	Vascular cells generated from PBMCs-derived iPSCs	PGA-P4HB starter matrices	Model: Vascular cells were used to seed tubular non-woven synthetic scaffolds and formulate small diameter vascular grafts under static and pulsatile flow conditions Application: Autologous PBMC derived iPSC-derived vascular constructs could be used for disease modeling and drug testing.	(22)
Self-organization (3D bioprinting, cell sheets, organoids)	Smooth muscle and endothelial cells derived from human PSCs	Fibrin matrix	Model: Induced self-organization of smooth muscle and endothelial cells derived from human PSCs in fibrin matrix using vascular endothelial growth factor to form microvasculature constructs. Application: 3D constructs arrayed in high throughput were used to screen a library of environmental and clinical vascular toxicants for immunological and toxicological responses.	(19)
	Human smooth muscle cells derived from pulmonary hypertension patients	–	Model: Culture of the media layer of blood vessel stimulating the thickening of a 3D media layer formed of human smooth muscle cells derived from pulmonary hypertension patients. Application: Effect of pulmonary hypertension drugs to suppress medial thickening.	(23)
	Human MSCs and EPCs	–	Model: Scaffoldless aligned human MSC sheets coated with human EPCs and cultured in a rotating wall bioreactor. Application: Tested the vascoactivity of the developed human cell-based endothelialized grafts in response to phenylephrine. This microphysiological system could be used for autologous drug screening.	(8)
	PSCs differentiated into endothelial cells and pericytes (24, 25) HUVECs, and smooth muscle cells derived from human ESCs and human iPSCs (26)	Matrigel/ collagen Methylcellulose-based hydrogel system (26)	Model (24, 25): Organoids model of diabetic vasculopathy. Model (26): Organoid co-culture model of smooth muscle and endothelial cells. Determined vascularization of organoids embedded in collagen/fibrinogen/fibronectin hydrogel. Application (26): in vitro co-culture model to study paracrine interactions between vascular cells. The system mimics physiological assembly of vessels and could be used for drug development and preclinical metabolic and toxicology studies.	(24, 25) (26)
	Endothelial cells	Polylactic acid for fused-filament 3D fabrication and PDMS for the cast	Model: 3D printing/microfluidics model of in vivo blood vessel network biology from healthy and diseased tissues. 3D printing of blood vessel images using fused-filament 3D fabrication by Polylactic acid. The 3D printout is cast in PDMS and dissolved, to produce the channels which are then lined with endothelial cells. Application: This model could be an effective tool to study drugs interactions with the endothelium under physiological flow conditions.	(27)
	Endothelial and smooth muscle cells	Nanoengineered hydrogel-based cell-laden bioinks	Model: 3D bioprinting of anatomically accurate, multi-cellular blood vessels using Nanoengineered hydrogel-based cell-laden bioinks. Application: Upon cytokine stimulation and blood perfusion, this 3D bioprinted vessel is able to recapitulate thromboinflammatory responses.	(28)
	HUVECs and MSCs	Gelatin-norbornene hydrogel cast	High throughput sample-agnostic bioreactor system, that was tested on vascular grafts made of HUVECs and MSCs encapsulated in gelatin-norbornene hydrogel cast into stereolithography 3D bioprinted well inserts.	(29)
Self-assembly	Smooth muscle cells	Pre-structured annular agarose well	Model: Smooth muscle cells were seeded into a pre-structured annular agarose well, which induced cell aggregation and self-assembly to develop tissue rings. Application: use the developed rings to formulate tissue tubes based on ring fusion, in presence of gelatin microspheres (30) that can deliver growth factors and influence cell phenotype (31).	(30, 31)
	Smooth muscle cells derived from human iPSCs	Agarose well systems	Development of vascular rings in agarose well systems using highly enriched functional smooth muscle cells derived from human induced pluripotent stem cells	(32)
	HUVECs and aortic smooth muscle cells	Agarose well systems	The use of agarose well systems in combination with cellularized microcarriers composed of gelatin microcarriers loaded with HUVECs and aortic smooth muscle cells to develop tubular structures.	(33)