Shinji Kawaguchi

Heterotopic transplantation of a decellularized and recellularized whole porcine heart

OBJECTIVES One of the final treatments for end-stage heart failure is heart transplantation. However, a shortage of donor hearts has created a long waiting list and limited benefits. Our ultimate goal is to create a whole beating heart fabricated on an organ scaffold for human heart transplantation. Here, we successfully performed the first transplantation using a decellularized whole porcine heart with mesenchymal stem cells. METHODS A porcine heart was harvested following cardiac arrest induced by a high-potassium solution and stored at -80°C for 24 h. The porcine heart was completely decellularized with 1% sodium dodecyl sulphate and 1% Triton X-100 under the control of perfusion pressure (100 mmHg) and maintained at 37°C. A decellularized whole-heart scaffold was sterilized with gamma irradiation. Cultured mesenchymal stem cells were collected and either infused into the ascending aorta or injected directly into the left ventricular wall. Finally, recellularized whole-heart scaffolds were transplanted into pigs under systemic anticoagulation treatment with heparin. Coronary artery angiography of the transplanted heart graft was performed. RESULTS In our decellularization method, all cellular components were removed, preserving the heart extracellular matrix. Heterotopic transplantations were successfully performed using a decellularized heart and a recellularized heart. The scaffolds were well perfused, without bleeding from the surface or anastomosis site. Coronary angiography revealed a patent coronary artery in both scaffolds. The transplanted decellularized heart was harvested on Day 3. Haematoxylin and eosin staining showed thrombosis in the coronary arteries and migrated inflammatory cells. Haematoxylin and eosin staining of the transplanted recellularized heart showed similar findings, with the exception of injected mesenchymal stem cells. CONCLUSIONS To the best of our knowledge, this is the first report of heterotopic transplantation of a decellularized whole porcine heart with mesenchymal stem cells. The scaffolds endured surgical procedures. We detected short-term coronary artery perfusion in the transplanted scaffolds by angiography. Future studies should analyse the histological features of transplanted decellularized scaffolds and optimize the system for recellularization to apply this unique technology clinically.

Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes

Summary Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%–87%. Approximately 6.2–7.0 × 108 cells (4-layer) and 1.5–2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future.

Construction of Routing Information Knowledge Base towards Wide Area Network Management

Nowadays each network management system (NMS) adopts different methods for collecting network information and different data structures. This makes NMS cooperation difficult, especially in multi-AS wide area network management. The current goal of the KANVAS (Knowledge base system in wide Area Networks with general Versatility, Availability and Scalability) project is to realize wide area network management by constructing a knowledge base system in networks. This paper discusses construction of a knowledge base system in a single AS (Autonomous System) towards the future wide area network management. This paper also shows the design and implementation of the knowledge base system that collects network configuration information from OSPF and SNMP, and stores it as instances of our proposed network ontology called Bonsai. Additionally, a prototype application that accesses the stored knowledge was implemented. The evaluation of basic performance and scalability of the system was carried out using a real AS topology and synthetic topologies and it was made sure that the prototype application works correctly. As a result, this paper shows that feasibility of the future wide area network management that adopts the knowledge base system.

Multidisciplinary approach to the treatment of cardiac AA amyloidosis and aortic stenosis due to Castleman's disease: a hybrid therapy with tocilizumab and aortic valve replacement.

A 67-year-old female patient with multicentric Castlemans disease(MCD)wasadmittedtoourhospitalwithsyncopeonexertion.Sheprevi-ously had episodes of chronic fatigue, weight loss, prolonged fever, andskin rash accompanied by anemia, polyclonal hypergammaglobulinemia,and elevated inflammatory markers. Computed tomography showedmultiple lymphadenopathies. The diagnosis of MCD with cutaneous in-volvement was made in 2010 based on the clinical manifestations andskinbiopsyhistopathological findings,whichshowedmassivein filtrationof polyclonal plasma cells in the dermis [1,2]. Comorbidities such as viralinfections, collagen diseases, and other malignancies were excluded. Shesimultaneously had renal impairment because of secondary AA amyloid-osisdue to MCD, whichwasdiagnosedb yrenal biopsybased on multipleAA-type amyloid glomerular deposits. Fortunately, following the ad-ministration of tocilizumab, a humanized monoclonal interleukin-6receptor antibody [3], the patient achieved a stable disease statusconcomitant with improved clinical and laboratory manifestations(decrease in C-reactive proteinfrom 10.0 mg/dL to 0.2 mg/dL andserum creatinine from 3.4 mg/dL to 2.0 mg/dL).Echocardiography on admission demonstrated a severely stenot-ic trileaflet aortic valve with concentric ventricular hypertrophy(Fig. 1A, B, C, and D), providing an evident reason for syncope onceother relevant cardiac and extracardiac abnormalities were exclud-ed. The prominent hypertrophicventricle with abnormal myocardialtexture and the extremely increased ventricular mass reconfirmedby cardiac magnetic resonance imaging (Fig. 1EandF)suggestedthe existence of not only cardiac hypertrophy induced by aortic ste-nosis (AS), but also infiltrative cardiomyopathy due to secondaryamyloidosis. As part of our preoperative evaluation, we performedcardiac catheterization including right ventricular endomyocardialbiopsy. Multiple AA-type amyloid infiltrations were identified inthe myocardium, suggesting cardiac involvement in secondary AAamyloidosis due to MCD (Fig. 2A, B, and C). Although the preciseoutcome of cardiac AA amyloidosis has not been fully elucidatedbecause of rarity, it seems to be comparatively preferred if the treat-ment of underlying inflammatory etiology is successful [3]. Curativesurgical therapy for symptomatic severe AS was therefore consid-ered in this case and we subsequently performed a successful surgi-cal aortic valve replacement (SAVR). Although within expectations,histopathological examination of the resected native aortic valveshowed severe calcific degeneration accompanied by amyloid pro-tein infiltration (Fig. 2D, E, and F). The patient has maintained agood clinical course since the surgery (decrease in brain natriureticpeptide from 1390 pg/mL to 614 pg/mL).MCD is a rare systemic lymphoproliferative disorder character-ized by abnormal proliferation of polyclonal plasma cells in the lym-phoidfollicles [1],withonlyafewcasesofsecondary AAamyloidosiscomplicating MCD reported [4].Thisisafirst report of secondary AAamyloidosis with cardiac involvement including the aortic valve dueto MCD. Although the long-term prognosis for MCD remains poor,especially for the plasma cell type of disease, recent advances inimmunological therapies such as tocilizumab have dramaticallyimproved the outcome both of MCD and subsequent amyloidosis

Thoracic Endovascular Aortic Repair in Patients with Prior Open Aortic Surgery

OBJECTIVE To review our experience of thoracic endovascular aortic repair (TEVAR) in patients with prior open aortic repair (OAR). MATERIALS AND METHODS Stent-grafts were deployed in the arch, descending thoracic and thoracoabdominal aortae of 39, 13 and 5 patients, respectively, and in a deteriorated extra-anatomical prosthesis in one. The access route was the femoral artery in 10 of 23 patients with, and in 30 of 35 patients without a prior abdominal prosthesis. Prior prostheses and elephant trunks comprised 57 of 116 landing zones and 23 proximal landing zones, respectively. RESULTS Three patients died before discharge. Type II endoleaks developed in six patients, and Types I and III developed in one patient each. Type I endoleaks were not found at landing zones comprising prosthetic grafts. The overall actuarial three-year survival rate including early mortality was 86.5%. CONCLUSION The clinical outcomes of TEVAR were excellent, even in patients with prior OAR. Prosthetic grafts, including elephant trunks, provided good landing zones for TEVAR. Prostheses with larger-caliber designs are recommended for iliac artery reconstruction in future TEVAR.

Kinked Graft and Anastomotic Stenosis-Induced Hemolytic Anemia Requiring Reoperation

We report a case of hemolytic anemia caused initially by narrowing of a proximal anastomotic site and subsequently by a kinked prosthetic graft after total aortic arch replacement. A 55-year-old man was diagnosed with acute type A aortic dissection by computed tomography (CT). After total aortic arch replacement, hemolytic anemia and renal dysfunction developed. CT detected narrowing of proximal anastomotic site of the graft. Removing the graft and ascending aortic replacement resolved the signs of hemolytic anemia. However, 50 days after the surgery, severe hemolytic anemia developed again. CT revealed a sharply kinked graft. Total arch replacement was again performed to resect the kinked graft. He was discharged on the 24th postoperative day without hemodialysis.