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Background Small caliber vascular prostheses are not clinically available because synthetic vascular prostheses lack endothelial cells which modulate platelet activation, leukocyte adhesion, thrombosis, and the regulation of vasomotor tone by the production of vasoactive substances. We developed a novel method to create scaffold-free tubular tissue from multicellular spheroids (MCS) using a “Bio-3D printer”-based system. This system enables the creation of pre-designed three-dimensional structures using a computer controlled robotics system. With this system, we created a tubular structure and studied its biological features. Methods and Results Using a “Bio-3D printer,” we made scaffold-free tubular tissues (inner diameter of 1.5 mm) from a total of 500 MCSs (2.5× 104 cells per one MCS) composed of human umbilical vein endothelial cells (40%), human aortic smooth muscle cells (10%), and normal human dermal fibroblasts (50%). The tubular tissues were cultured in a perfusion system and implanted into the abdominal aortas of F344 nude rats. We assessed the flow by ultrasonography and performed histological examinations on the second (n = 5) and fifth (n = 5) day after implantation. All grafts were patent and remodeling of the tubular tissues (enlargement of the lumen area and thinning of the wall) was observed. A layer of endothelial cells was confirmed five days after implantation. Conclusions The scaffold-free tubular tissues made of MCS using a Bio-3D printer underwent remodeling and endothelialization. Further studies are warranted to elucidate the underlying mechanism of endothelialization and its function, as well as the long-term results.

Objective:The optimalorientationof a bileaflet mechanical valvefor tricuspidvalvereplacement (TVR) hasnot yetbeen determined. Theaim of this study was to use fiberoptic cardioscopy to evaluate the effect of orientation of a mechanical valve implanted in the tricuspid position on bileaflet mechanical valve behavior. Methods: Twelve pigs (50—59 kg) underwent TVR with a St. Jude Mechanical Heart Valve (25 mm standard cuff model) after cardioplegic arrest. The mechanical valve was implanted horizontally in six pigs (Group H), and vertically in another six pigs (Group V). The heart was perfused with pellucid Krebs—Henseleit solution in situ and the mechanical valve behavior was observed with a fiberoptic endoscope during different heart rates (HRs) induced by ventricular pacing (60, 90, 120, 150 min 1 ). All images were recorded on a high-speed videosystem every 4 ms. The closingtime lag (CTL) betweenthe valve leaflets was calculated and compared betweenthe two groups. Results: In Group H, the lower valve leaflet tended to open incompletely and close earlier than the upper leaflet. The calculated CTL was 303 60 ms, 65 48 ms, 40 9 ms, and 40 26 ms at pacing HRs of 60, 90, 120, and 150 min 1 , respectively. In contrast to Group H, there was little difference in CTL between the right and left leaflets in Group V. The calculated CTL was 9 12 ms, 11 10 ms, 1 3 ms, and 6 7m s at pacing HRs of 60, 90, 120, and 150 min 1 , respectively. There were significant differences in CLT between the two groups at each ventricular pacingrate(P < 0.01).Conclusions:Orientationofan implanted bileaflet valveinthetricuspidpositionsignificantlyinfluencedleafletmotion.In a horizontal orientation, the lower valve leaflet opened incompletely and closed earlier than the upper leaflet. These results suggest that the gravity might affect leaflet motion and that bileaflet mechanical valves should be implanted vertically in TVR to prevent abnormal leaflet motion and thrombus formation.

Background The aim of our study was to develop a completely scaffold-free, viable, contractile cardiac tissue capable of being grafted into the damaged native heart. Methods Our technology is based on the fundamental characteristics of the self-assembling nature of cells. We created contractile cardiac spheroids by plating a mixture of rat neonatal ventricular cardiomyocytes, human dermal fibroblasts, and human coronary microartery endothelial cells in ultralow attachment plates. First, the optimal cell ratios for the 3 cell sources were determined. Next, approximately 1 × 10 4 optimal spheroids were fused into a patch-like construct, and the morphologic characteristics and mechanical functions of these patches were evaluated. Finally, the cardiac patches were grafted into the hearts of F344 nude rats, and histologic studies were performed after transplantation. Results Synchronous beating of the cardiac patch was confirmed electrophysiologically and mechanically. A micronetwork of endothelial cells was also demonstrated in the construct, and the histologic study performed 5 days after transplantation showed the grafts to be viable, with functioning microvascular structures inside the graft tissue. Conclusions We consider the application of our scaffold-free 3-dimensional tissue engineering technology to cardiac regeneration therapy is feasible and expect that this technology will become a promising tool for the treatment of end-stage heart failure.

Introduction: Small caliber synthetic vascular grafts are not clinically available. We developed a novel method to create scaffold-free tubular tissue from multicellular spheroids (MCS) using a “Bio-3D printer”-based system, which enables the creation of various three-dimensional structures pre-designed using a computer system. With this system, we created a tubular structure (Fig. 1), and studied its biological features. Methods: We made 1.5 mm in diameter scaffold-free tubular tissues from MCS (1.25 x 10[[Unable to Display Character: ⁷]] cells) composed of human umbilical vein endothelial cells (40%), human aortic smooth muscle cells (10%) and normal human dermal fibroblasts (50%) using a Bio-3D printer. The vessels were cultured in a perfusion system. We implanted grafts into the abdominal aortas of F344 nude rats, and assessed the flow by ultrasonography and performed histological examinations on the second (N=5) and fifth (N=5) days after implantation. Results: All grafts were patent. Remodeling of the vessel (enlargement of the lumen area and thinning of the wall) was observed (Fig. 2). A layer of endothelial cells was developed after implantation of the graft (Fig. 3). Conclusions: The scaffold-free vascular grafts made of MCS using a Bio-3D printer showed biological features comparable to native vessels. Further studies are warranted toward the clinical application of this novel technology.

Purpose The aim of our study is to develop completely scaffold-free, viable, contractile cardiac tissue which can be grafted to the damaged native heart. Methods and Materials Our technology is based on the fundamental characteristics of self-assembling nature of cells. We called these self-assembled spherical cell aggregates as “spheroids”. We created contractile cardiac spheroids by plating a mixture of 800 cells of rat neonate ventricular cardiomyocytes (RNVCM), 100 cells of human normal dermal fibroblasts (HNDFB), and 100 cells of human coronary micro-artery endothelial cells (HCMEC) in ultra-low-attachment plate. After 24 hr cultivation, prevascularized contractile cardiac spheroids were fabricated. Next, 10000 spheroids were fused into patch like construct with a maximum diameter of 1cm with 200μm in thickness. We evaluated morphological characteristics, and electro- and mechanical function. We also grafted the cardiac patch onto the heart of F344 nude rats, and performed histological study 3 weeks after transplantation. Results We confirmed synchronous beating of the cardiac patch electro-physiologically, and mechanically (video-motion analysis). Expression of connexin 43 (a component of gap junction) and micro network of endothelial cell in the construct were demonstrated. Histological study performed 3 weeks after transplantation showed that the grafts were viable with functioning micro-vascular structure inside the graft. [ figure 1 ] Conclusions We consider that applying our scaffold-free 3-Dimensinal tissue engineering technology to cardiac regeneration therapy is feasible. We expect that this technology will become a promising tool to treat end-stage heart failure.