Ryo Noguchi

Scaffold-Free Tubular Tissues Created by a Bio-3D Printer Undergo Remodeling and Endothelialization when Implanted in Rat Aortae

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.

Development of a three-dimensional pre-vascularized scaffold-free contractile cardiac patch for treating heart disease

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.

Surgical Strategy for Retrograde Type A Aortic Dissection Based on Long-Term Outcomes

BACKGROUND The optimal management of a retrograde type A aortic dissection (RAAD) is controversial, and few reports have discussed the long-term outcomes of surgical strategies. To determine the most appropriate strategy, we studied the early and late outcomes of RAAD cases. METHODS From 1998 to 2014, 44 patients with RAAD (mean age of 63 ± 11 years) underwent surgical repair. Ascending aortic replacement (AAR) was performed in 21 patients and ascending and total arch replacement (TAR) was performed in 23 patients. Eight of the patients who received TAR underwent complete resection of the primary tear in the distal arch or descending aorta (TAR-R[+]), whereas the remaining 15 patients received elephant trunk implantation as an alternative procedure for tear resection (TAR-R[-]). The early and late outcomes (mean follow-up, 86.5 months) were evaluated. RESULTS Hospital mortality occurred in 4 of the 44 (9.1%) patients, with no mortalities among the patients undergoing TAR-R[-]. There was a tendency toward a higher incidence of late aorta-related events in the AAR group, with a significantly higher patency rate of the false lumen in the proximal site of the residual aorta compared with the TAR group (p = 0.009). Furthermore, the 5-year rate of freedom from aortic growth greater than 50 mm was significantly lower after AAR than after TAR (p = 0.04). A multivariate analysis indicated that the initial ascending aortic diameter (odds ratio [OR], 1.5; p = 0.02) and AAR (OR, 29.1; p = 0.01) were independent predictors of late aortic expansion. CONCLUSIONS The surgical outcomes were acceptable in both the AAR and TAR groups. The long-term outcomes potentially support the aggressive adoption of TAR in relatively younger patients with significant ascending aortic enlargement at presentation.

The addition of human iPS cell-derived neural progenitors changes the contraction of human iPS cell-derived cardiac spheroids

BACKGROUND We havebeen attempting to use cardiac spheroids to construct three-dimensional contractilestructures for failed hearts. Recent studies have reported that neuralprogenitors (NPs) play significant roles in heart regeneration. However, theeffect of NPs on the cardiac spheroid has not yet been elucidated. OBJECTIVE This studyaims to demonstrate the influence of NPs on the function of cardiac spheroids. METHODS Thespheroids were constructed on a low-attachment-well plate by mixing humaninduced pluripotent stem (hiPS) cell-derived cardiomyocytes and hiPScell-derived NPs (hiPS-NPs). The ratio of hiPS-NPs was set at 0%, 10%, 20%,30%, and 40% of the total cell number of spheroids, which was 2500. The motionwas recorded, and the fractional shortening and the contraction velocity weremeasured. RESULTS Spheroidswere formed within 48 h after mixing the cells, except for the spheroidscontaining 0% hiPS-NPs. Observation at day 7 revealed significant differencesin the fractional shortening (analysis of variance; p = 0.01). The bestfractional shortening was observed with the spheroids containing 30% hiPS-NPs.Neuronal cells were detected morphologically within the spheroids under aconfocal microscope. CONCLUSION Theaddition of hiPS-NPs influenced the contractile function of the cardiacspheroids. Further studies are warranted to elucidate the underlying mechanism.

Haptoglobin Reduces Inflammatory Cytokine INF-γ and Facilitates Clot Formation in Acute Severe Burn Rat Model

Haptoglobin exerts renal protective function by scavenging free hemoglobin from the urine and blood stream in patients with hemolytic disorders. Recent studies elucidate the relationships between haptoglobin and inflammation. In addition, coagulopathy is often induced by systemic inflammation characterized by the presence of vascular endothelial damage. We hypothesize that haptoglobin might have an anti-inflammatory effect and affect hypercoagulability using rat burn model. Thirty anesthetized rats of six-weeks of age received over 30% full-thickness scald burn on the dorsal skin surface. All rats were injected with either haptoglobin (Hpt) or normal saline (NS) intraperitoneally. The rats were divided into three groups: 1) control group (NS 20 mL/kg); 2) low concentration of Hpt group, L-Hpt, (Hpt 4 mL (80 U) /kg+NS 16 mL/kg); and 3) high concentration of Hpt group, H-Hpt, (Hpt 20 mL (400 U) /kg). While under anesthesia, all rats were euthanized by exsanguination at 6 hours (N=5) and 24 hours (N=5). Inflammatory and anti-inflammatory cytokines were measured and whole-blood viscoelastic tests were performed by thromboelastometry (ROTEM). Haptoglobin significantly reduced free hemoglobin 24 hours after the injury. Improvement of hematuria was confirmed in the H-Hpt group. There were no differences in thrombin-antithrombin complex and plasmin-α2 plasmin inhibitor complex. The haptoglobin tended to decrease interferon-gamma (IFN-γ) in H-Hpt group. ROTEM findings of the L-Hpt group showed significantly higher clot firmness and shorter time to maximum clot formation velocity than the control group. Haptoglobin reduced INF-γ, and accelerated speed of clot formation in acute phase of severe burn.

Persistent Gastrointestinal Angiodysplasia in Heyde's Syndrome after Aortic Valve Replacement

A 67-year-old woman with recurrent gastrointestinal bleeding and aortic valve stenosis presented with spurting bleeding from angiodysplasia of the upper jejunum. As electrophoresis revealed decreased levels of high-molecular-weight (HMW) von Willebrand factor (VWF) multimers, she was diagnosed with Heydes syndrome. After aortic valve replacement, her HMW VWF levels quickly recovered to normal, and the gastrointestinal bleeding ceased. However, capsule endoscopy still revealed gastrointestinal angiodysplasia six months later. This case shows that minute analyses of VWF multimers enable the diagnosis and confirmation of the resolution of Heydes syndrome, and implies that gastrointestinal angiodysplasia can be attributed to unknown factors other than decreased VWF multimers.