Engineering Three-Dimensional Vascularized Cardiac Tissues

Abstract

Heart disease is one of the largest burdens to human health worldwide and has very limited therapeutic options. Engineered three-dimensional (3D) vascularized cardiac tissues have shown promise in rescuing cardiac function in diseased hearts and may serve as a whole organ replacement in the future. One of the major obstacles in reconstructing these thick myocardial tissues to a clinically applicable scale is the integration of functional vascular networks capable of providing oxygen and nutrients throughout whole engineered constructs. Without perfusion of oxygen and nutrient flow throughout the entire engineered tissue not only is tissue viability compromised, but also overall tissue functionality is lost. There are many supporting technologies and approaches that have been developed to create vascular networks such as 3D bioprinting, co-culturing hydrogels, and incorporation of soluble angiogenic factors. In this state-of-the-art review, we discuss some of the most current engineered vascular cardiac tissues reported in the literature and future directions in the field. Impact statement The field of cardiac tissue engineering is rapidly evolving and is now closer than ever to having engineered tissue models capable of predicting preclinical responses to therapeutics, modeling diseases, and being used as a means of rescuing cardiac function following injuries to the native myocardium. However, a major obstacle of engineering thick cardiac tissue remains to be the integration of functional vasculature. In this review, we highlight seminal and recently published works that have influenced and pushed the field of cardiac tissue engineering toward achieving vascularized functional tissues.

Keywords: 3D printed vasculature; angiogenesis; cardiac patch; engineered cardiac tissue; regenerative medicine; vascularized cardiac tissues.

FIG. 1.

Cardiac tissues can be cultured in numerous ways, which allow for their subsequent functionalization and transplantation. (A) Hydrogels with embedded cardiac-specific cell types and angiogenic growth factors can generate functional vascular cardiac patches that can be subsequently transplanted into diseased hearts. (B) An example of cardiac cell sheeting using thermosensitive gelatin for stacking several layers of cell sheets for mechanical stability and subsequent transplantation.

FIG. 2.

Cardiac patches demonstrate the ability to integrate with host vasculatures (A) 1-week post implantation in a rat MI model vasculature within the cardiac patch stains positive for human CD31⁺ endothelial networks. (B) Vessel-like structures within the cardiac patch contain leukocytes (white arrows) demonstrating functional integration into host vasculature. (C) H&E staining of the cardiac patch postimplantation reveals the presence of red blood cells within the patch (blue arrows). (D) Integration of host vasculature identified by the presence of cells staining positive for isolectin B4 (IB4) within the cardiac patch. (E) Implanted cardiac patch (hCMP) located to the right of the white dashed line was identified by visualization of engineered iPSC-CM expressing CD31 and cardiac troponin I (cTnT). (F) Maturing iPSC-CM with distinct sarcomeric structures were visualized by staining for coexpression of GFP and α-actinin. iPSC-CM, induced pluripotent stem cell-derived cardiomyocyte; MI, myocardial infarct.

FIG. 3.

Cell sheet engineering has been explored as a potential option for generating transplantable vascular cardiac tissues. (A) Cardiac cell sheets have been generated, layered, and cultured within vascular bioreactors to influence prevascularization before transplantation. (B, C) H&E and Azan staining show evidence of cardiac sheets containing perfusable vasculature capable (white arrows) of supporting the engineered tissue. (D) Transplanted cardiac cell sheet containing GFP expressing ECs stained with isolectin B4 demonstrates fused vasculature from cardiac cell sheet to host MI model. (E) Cardiac patch containing GFP expressing ECs stained for troponin T (red) shows transplanted cardiac cell sheet vascularizing host myocardium in rat MI model.

FIG. 4.

3D printing provides a platform for constructing detailed architectures of cardiac tissues (A) Bioinks composed of various cell types can be spatially patterned to give rise to functional cardiac tissues that can be subsequently evaluated both in vitro and in vivo. (B) Sacrificial inks are 3D printed into cardiac tissues and subsequently removed to create embedded luminal structures within engineered cardiac tissues that support thick tissue perfusion. (C) Support baths used 3D print soft materials with high fidelity that exhibit complex hierarchal structures of in vivo heart anatomy. 3D, three dimensional.

FIG. 5.

3D printing enables the generation of functional vascular cardiac tissues and complex cardiovascular structures. (A–C) Overview of bottom-up 3D printing process for the generation of a cardiac patch. (D) Image of excised omentum following 7 days of in vivo transplantation with white dashed lines highlighting the border of the cardiac patch. (E–G) Imaging of excised 3D printed cardiac patch following 7 days of in vivo transplantation for sarcomeric actinin (red) exhibiting a matured cardiac morphology at increasing magnification. (H, I) Cardiac organoid organ building block staining positive for cardiac specific markers cardiac troponin T (green) and α-actinin (red). (J, K) Viable SWIFT 3D printed perusable cardiac tissues imaged from top-down staining positive for cardiac troponin T and α-actinin following 24 h of perfusion. (L) SWIFT printed CAD rendering of left anterior descending artery into left ventricle-shaped mold. CAD, computer-aided designs; SWIFT, sacrificial writing into functional tissue.

FIG. 6.

3D printing into support material (A, B) CAD-generated renderings of cardiac structures used for FRESH v2.0 3D printing. (C, D) FRESH v2.0 printed neonatal scale human heart exhibiting complex cardiac morphology and adult size tri-leaflet heart valve, respectively. (E) FRESH v2.0 printed construct based on MRI-derived CAD rendering of the left anterior descending artery with computationally generated microvasculature capable of perfusion. (F) FRESH v2.0 printed full-size adult human heart. (G, H) Heart containing independent chambers, vasculature, and imbedded CMs and ECs printed into alginate microparticle support bath. FRESH, freeform reversible embedding of suspended hydrogels.