CD8+ T cell-mediated rejection of allogenic human induced pluripotent stem cell-derived cardiomyocyte sheets in human PBMC-transferred NOG MHC double knockout mice.

BACKGROUND: Transplantation of human induced pluripotent stem cell-derived cardiomyocytes (hiPS-CMs) has emerged as a promising therapy to treat end-stage heart failure. However, immunogenicity of hiPS-CMs in transplanted patients has not been fully elucidated. Thus, in vivo models are required to estimate immune responses against hiPS-CMs in transplant recipients. METHODS: We transferred human peripheral blood mononuclear cells (hPBMCs) into NOD/Shi-scid IL2rgnull (NOG) MHC class I/II double knockout (NOG-ΔMHC) mice, which were reported to accept hPBMCs without xenogeneic-graft-versus-host disease (xeno-GVHD). Then, hiPS-CM sheets generated from the hiPS cell line 201B7 harboring a luciferase transgene were transplanted into the subcutaneous space of NOG-ΔMHC mice. Graft survival was monitored by bioluminescent images using a Xenogen In Vivo Imaging System. RESULTS: The human immune cells were engrafted for more than three months in NOG-ΔMHC mice without lethal xeno-GVHD. The hiPS-CMs expressed a moderate level of human leukocyte antigen (HLA)-class I, but not HLA-class II, molecules even after interferon-gamma (IFN-γ) stimulation. Consistently, the allogenic IFN-γ-treated hiPS-CMs induced weak CD8+ but not CD4+, T cell responses in vitro. hiPS-CM sheets disappeared approximately 17-24 days after transplantation in hPBMC-transferred NOG-ΔMHC mice, and CD8+ T cell depletion significantly prolonged graft survival, similar to what was observed following tacrolimus treatment. CONCLUSION: hiPS-CMs are less immunogenic in vitro but induce sufficient CD8+ T cell-mediated immune responses for graft rejection in vivo.

Pre-clinical evaluation of the efficacy and safety of human induced pluripotent stem cell-derived cardiomyocyte patch.

BACKGROUND: Cell- or tissue-based regenerative therapy is an attractive approach to treat heart failure. A tissue patch that can safely and effectively repair damaged heart muscle would greatly improve outcomes for patients with heart failure. In this study, we conducted a preclinical proof-of-concept analysis of the efficacy and safety of clinical-grade human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) patches. METHODS: A clinical-grade hiPSC line was established using peripheral blood mononuclear cells from a healthy volunteer that was homozygous for human leukocyte antigens. The hiPSCs were differentiated into cardiomyocytes. The obtained hiPSC-CMs were cultured on temperature-responsive culture dishes for patch fabrication. The cellular characteristics, safety, and efficacy of hiPSCs, hiPSC-CMs, and hiPSC-CM patches were analyzed. RESULTS: The hiPSC-CMs expressed cardiomyocyte-specific genes and proteins, and electrophysiological analyses revealed that hiPSC-CMs exhibit similar properties to human primary myocardial cells. In vitro and in vivo safety studies indicated that tumorigenic cells were absent. Moreover, whole-genome and exome sequencing revealed no genomic mutations. General toxicity tests also showed no adverse events posttransplantation. A porcine model of myocardial infarction demonstrated significantly improved cardiac function and angiogenesis in response to cytokine secretion from hiPSC-CM patches. No lethal arrhythmias were observed. CONCLUSIONS: hiPSC-CM patches are promising for future translational research and may have clinical application potential for the treatment of heart failure.

Human induced pluripotent stem cell-derived closed-loop cardiac tissue for drug assessment.

Human iPSC-derived cardiomyocytes (hiPSC-CMs) exhibit functional immaturity, potentially impacting their suitability for assessing drug proarrhythmic potential. We previously devised a traveling wave (TW) system to promote maturation in 3D cardiac tissue. To align with current drug assessment paradigms (CiPA and JiCSA), necessitating a 2D monolayer cardiac tissue, we integrated the TW system with a multi-electrode array. This gave rise to a hiPSC-derived closed-loop cardiac tissue (iCT), enabling spontaneous TW initiation and swift pacing of cardiomyocytes from various cell lines. The TW-paced cardiomyocytes demonstrated heightened sarcomeric and functional maturation, exhibiting enhanced response to isoproterenol. Moreover, these cells showcased diminished sensitivity to verapamil and maintained low arrhythmia rates with ranolazine-two drugs associated with a low risk of torsades de pointes (TdP). Notably, the TW group displayed increased arrhythmia rates with high and intermediate risk TdP drugs (quinidine and pimozide), underscoring the potential utility of this system in drug assessment applications.

Transplantation of Human Embryonic Stem Cell-Derived Pericyte-Like Cells Transduced with Basic Fibroblast Growth Factor Promotes Angiogenic Recovery in Mice with Severe Chronic Hindlimb Ischemia.

Critical limb ischemia (CLI) is a state of severe peripheral artery disease, with no effective treatment. Cell therapy has been investigated as a therapeutic tool for CLI, and pericytes are promising therapeutic candidates based on their angiogenic properties. We firstly generated highly proliferative and immunosuppressive pericyte-like cells from embryonic stem (ES) cells. In order to enhance the angiogenic potential, we transduced the basic fibroblast growth factor (bFGF) gene into the pericyte-like cells and found a significant enhancement of angiogenesis in a Matrigel plug assay. Furthermore, we evaluated the bFGF-expressing pericyte-like cells in the previously established chronic hindlimb ischemia model in which bone marrow-derived MSCs were not effective. As a result, bFGF-expressing pericyte-like cells significantly improved blood flow in both laser Doppler perfusion imaging (LDPI) and dynamic contrast-enhanced MRI (DCE-MRI). These findings suggest that bFGF-expressing pericyte-like cells differentiated from ES cells may be a therapeutic candidate for CLI.

Prevalence of bleeding events in real-world Japanese registry for Percutaneous Ventricular Assist Device.

PURPOSE: Bleeding complication is a critical risk factor for outcomes of acute heart failure patients requiring mechanical circulatory support (MCS), including percutaneous catheter-type heart pumps (Impella). The Japanese registry for Percutaneous Ventricular Assist Device (J-PVAD) is an ongoing, large-scale, real-world registry to characterize Japanese patients requiring Impella. Here we analyzed bleeding complication profiles in patients who received Impella. METHODS: All consecutive Japanese patients who received Impella from October 2017 to January 2020 were enrolled. The 30-day survival and bleeding complications were analyzed. RESULTS: A total of 1344 patients were included: 653 patients received Impella alone, 685 patients received a combination of veno-arterial extracorporeal membrane oxygenation and Impella (ECPELLA), and 6 patients had failed Impella delivery. Overall 30-day survival was 67.0%, with Impella alone at 81.9% and ECPELLA at 52.7%. Overall bleeding/hematoma adverse events with a relation or not-excluded relation to Impella was 6.92%. Among them, the rates of hematoma and bleeding from medical device access sites were 1.41% and 4.09%, respectively. There was no difference between etiologies for these events. CONCLUSION: This study represents the first 3-year survival and the safety profile focused on bleeding adverse events from the J-PVAD registry. The results show that the real-world frequency of bleeding adverse events for patients who received Impella was an expected range from previous reports, and future real-world studies should aim to expand this data set to improve outcomes and adverse events.

Pleiotropic effects of extracellular vesicles from induced pluripotent stem cell-derived cardiomyocytes on ischemic cardiomyopathy: A preclinical study.

BACKGROUND: Stem cell-secreted extracellular vesicles (EVs) play essential roles in intercellular communication and restore cardiac function in animal models of ischemic heart disease. However, few studies have used EVs derived from clinical-grade stem cells and their derivatives with stable quality. Moreover, there is little information on the mechanism and time course of the multifactorial effect of EV therapy from the acute to the chronic phase, the affected cells, and whether the effects are direct or indirect. METHODS: Induced pluripotent stem cell-derived cardiomyocytes (iPSCM) were produced using a clinical-grade differentiation induction system. EVs were isolated from the conditioned medium by ultracentrifugation and characterized in silico, in vitro, and in vivo. A rat model of myocardial infarction was established by left anterior descending artery ligation and treated with iPSCM-derived EVs. RESULTS: iPSCM-derived EVs contained microRNAs and proteins associated with angiogenesis, antifibrosis, promotion of M2 macrophage polarization, cell proliferation, and antiapoptosis. iPSCM-derived EV treatment improved left ventricular function and reduced mortality in the rat model by improving vascularization and suppressing fibrosis and chronic inflammation in the heart. EVs were uptaken by cardiomyocytes, endothelial cells, fibroblasts, and macrophages in the cardiac tissues. The pleiotropic effects occurred due to the direct effects of microRNAs and proteins encapsulated in EVs and indirect paracrine effects on M2 macrophages. CONCLUSIONS: Clinical-grade iPSCM-derived EVs improve cardiac function by regulating various genes and pathways in various cell types and may have clinical potential for treating ischemic heart disease.

Outcomes of Veno-Arterial Extracorporeal Membrane Oxygenation With Percutaneous Left Ventricular Unloading in Fulminant Myocarditis.

Fulminant myocarditis requiring peripheral veno-arterial extracorporeal membrane oxygenation (VA-ECMO) has a high mortality rate. We investigated clinical outcomes of combined use of VA-ECMO and percutaneous left ventricular assist device (VAD) (Impella) for fulminant myocarditis in 104 consecutive patients enrolled in the Japan Registry for Percutaneous VAD (J-pVAD) between October 2017 and January 2020. Patients were followed until hospital discharge and predictors of survival were analyzed with a Cox proportional hazards model. The median support duration of combined use of VA-ECMO and Impella (ECMO/Impella) was 6 days, and the median left ventricular ejection fraction improved from 15% to 52% during support ( p < 0.0001). Overall, 66 patients (63%) survived to discharge. Multivariate analysis revealed ECMO/Impella support at a transplant center as an independent predictor of survival ( p = 0.0231). Patients treated at transplant centers had better 60 days survival rates when compared to nontransplant centers (83% vs. 55%, p = 0.005). However, baseline characteristics and treatment strategies differed between the two groups. This real-world national registry database suggested the difference in survival after ECMO/Impella support for fulminant myocarditis between transplant and nontransplant centers, which may indicate hospital variations regarding patient management, although further controlled studies are needed.

Mid-term outcomes of surgical aortic valve replacement using a mosaic porcine bioprosthesis with concomitant mitral valve repair.

This study retrospectively evaluated the mid-term outcomes of surgical aortic valve replacement (SAVR) using a stented porcine aortic valve bioprosthesis (Mosaic; Medtronic Inc., Minneapolis, MN, USA) with concomitant mitral valve (MV) repair. From 1999 to 2014, 157 patients (median [interquartile range] age, 75 [70-79] years; 47% women) underwent SAVR with concomitant MV repair (SAVR + MV repair), and 1045 patients (median [interquartile range] age, 76 [70-80] years; 54% women) underwent SAVR only at 10 centers in Japan as part of the long-term multicenter Japan Mosaic valve (J-MOVE) study. The 5-year overall survival rate was 81.5% ± 4.1% in the SAVR + MV repair group and 85.1% ± 1.4% in the SAVR only group, and the 8-year overall survival rates were 75.2% ± 5.7% and 78.1% ± 2.1%, respectively. Cox proportional hazards analysis showed no significant difference in the survival rates between the two groups (hazard ratio, 0.87; 95% confidence interval, 0.54-1.40; P = 0.576). Among women with mild or moderate mitral regurgitation who were not receiving dialysis, those who underwent SAVR + MV repair, were aged > 75 years, and had a preoperative left ventricular ejection fraction of 30-75% tended to have a lower mortality risk. In conclusion, this subgroup analysis of the J-MOVE cohort showed relevant mid-term outcomes after SAVR + MV repair.

The Efficacy of Second-Stage Endovascular Repair Strategy for Acute Type A Aortic Dissection.

PURPOSE: To evaluate the usefulness of a hybrid treatment strategy for acute type A aortic dissection (AAAD). METHODS: We retrospectively evaluated the outcomes of 39 partial arch replacements (PAR; 26 male/13 female, mean age=67.9 years) in 62 patients with AAAD operated at our hospital from January 2019 to January 2023. The technique included PAR with graft-designed landing length and translocated the brachiocephalic artery inflow site during the initial surgery to minimize the invasiveness of the surgery. Thereafter, second-stage thoracic endovascular aortic repair (second TEVAR) for distal aortic events in the chronic phase was performed. RESULTS: There was 1 case of 30-day mortality (2.6%) and 2 cases of postprocedural cerebral infarction (5.1%). The cumulative survival rates were 97.4%/1 year and 97.4%/3 years. The cumulative freedom from aorta-related second-stage procedure for the distal aortic event after initial PAR, which was performed in 13 patients (33.3%), was 63.9%/1 year and 59.7%/3 years. All patients requiring re-intervention after initial PAR underwent a second TEVAR with a 100% success rate and no postoperative complications. CONCLUSION: Initial PAR for AAAD in anticipation of the second TEVAR is a valuable strategy for enabling minimally invasive additional treatment of aorta-related re-intervention for distal aortic events in the chronic phase. CLINICAL IMPACT: This study provides detailed information on the hybrid aortic repair strategy of the initial open partial arch repair and second staged endovascular repair for the acute type A aortic dissection. Based on this study, distal aortic re-intervention after initial open partial arch repair was necessary only in about 30% of cases, and no cases of SCI were observed in the initial treatment or in the second-stage endovascular repair and no cases of distal SINE were observed after the second staged endovascular repair. Overall, the results suggest that limiting the initial open partial arch repair can achieve good perioperative and early outcomes of initial surgery, and that second staged endovascular re-intervention for distal aortic events can be performed reliably, safely, and with minimal invasiveness.

Safety confirmation of induced pluripotent stem cell-derived cardiomyocyte patch transplantation for ischemic cardiomyopathy: first three case reports.

Introduction: With the expected increase in patients with heart failure and ischemic 15 cardiomyopathy, the development of myocardial regenerative medicine using cell transplantation as a novel treatment method is progressing. This first-in-human clinical trial aimed to confirm the safety of cardiomyocyte patch transplantation derived from allogeneic induced pluripotent stem (iPS) cells based on the results of several preclinical studies. Study design: The inclusion criteria were left ventricular ejection fraction of 35% or less; heart failure symptoms of New York Heart Association class III or higher despite existing therapies such as revascularization; and a 1-year observation period that included a 3-month immunosuppressive drug administration period after transplantation of iPS cell-derived cardiomyocyte patches to evaluate adverse events, cardiac function, myocardial blood flow, heart failure symptoms, and immune response. Results: In the first three cases of this trial, no transplanted cell-related adverse events were observed during the 1-year observation period, and improvement in heart failure symptoms was observed. In addition, improvements in left ventricular contractility and myocardial blood flow were observed in two of the three patients. Regarding immune response, an increase in transplant cell-specific antibody titer was observed in all three patients after immunosuppressive drug administration. In one patient with poor improvement in cardiac function and myocardial blood flow, an increase in antibody titer against HLA-DQ was observed even before cell transplantation. Conclusions: Our case findings demonstrate that the transplantation of iPS cell-derived cardiomyocyte patches for ischemic cardiomyopathy can be safely performed; however, further investigation of the therapeutic effect and its relationship with an immune response is needed by accumulating the number of patients through continued clinical trials.

Effect of morphological change on the maturation of human induced pluripotent stem cell-derived cardiac tissue in rotating flow culture.

Introduction: Understanding the critical factors for the maturation of human induced pluripotent stem cell (hiPSC)-derived cardiac tissue is important for further development of culture techniques. Rotating flow culture, where the tissues float in the culture medium by balancing its gravitational settling and the medium flow generated in rotating disk-shaped culture vessels, is one of culture systems used for tissue engineering. It has previously been demonstrated that rotating flow culture leads to the formation of matured cardiac tissue with higher levels of function and structure than the other culture systems. However, the detailed mechanisms underlying the maturation of cardiac tissue remain unclear. This study investigated the maturation process of hiPSC-derived cardiac tissue in rotating flow culture with a focus on morphological changes in the tissue, which is a trigger for maturation. Methods: The cardiac tissue, which consisted of cardiomyocytes derived from hiPSCs, was cultured on the 3D scaffold of poly (lactic-co-glycolic) acid (PLGA)-aligned nanofibers, in rotating flow culture for 5 days. During the culture, the time profile of projected area of tissue and formation of maturation marker proteins (ß-myosin heavy chain and Connexin-43), tissue structure, and formation of nuclear lamina proteins (Lamin A/C) were compared with that in static suspension culture. Results: The ratio of the projected area of tissue significantly decreased from Day 0 to Day 3 due to tissue shrinkage. In contrast, Western blot analysis revealed that maturation protein markers of cardiomyocytes significantly increased after Day 3. In addition, in rotating flow culture, flat-shaped nuclei and fiber-like cytoskeletal structures were distributed in the surface region of tissue where medium flow was continuously applied. Moreover, Lamin A/C, which are generally formed in differentiated cells owing to mechanical force across the cytoskeleton and critically affect the maturation of cardiomyocytes, were significantly formed in the tissue of rotating flow culture. Conclusions: In this study, we found that spatial heterogeneity of tissue structure and tissue shrinkage occurred in rotating flow culture, which was not observed in static suspension culture. Moreover, from the quantitative analysis, it was also suggested that tissue shrinkage in rotating flow culture contributed its following tissue maturation. These findings showed one of the important characteristics of rotating flow culture which was not revealed in previous studies.

3D puzzle-inspired construction of large and complex organ structures for tissue engineering.

3D printing as a powerful technology enables the fabrication of organ structures with a programmed geometry, but it is usually difficult to produce large-size tissues due to the limited working space of the 3D printer and the instability of bath or ink materials during long printing sessions. Moreover, most printing only allows preparation with a single ink, while a real organ generally consists of multiple materials. Inspired by the 3D puzzle toy, we developed a "building block-based printing" strategy, through which the preparation of 3D tissues can be realized by assembling 3D-printed "small and simple" bio-blocks into "large and complex" bioproducts. The structures that are difficult to print by conventional 3D printing such as a picture puzzle consisting of different materials and colors, a collagen "soccer" with a hollow yet closed structure, and even a full-size human heart model are successfully prepared. The 3D puzzle-inspired preparation strategy also allows for a reasonable combination of various cells in a specified order, facilitating investigation into the interaction between different kinds of cells. This strategy opens an alternative path for preparing organ structures with multiple materials, large size and complex geometry for tissue engineering applications.

Assessment of the Effectiveness of Zone 1-Landing Hybrid TEVAR by Comparing Its Outcomes with Those of Zone 2-Landing Hybrid TEVAR.

Background: Hybrid thoracic endovascular aortic repair (TEVAR) without median sternotomy is increasingly being performed in high-risk patients with aortic arch disease. The outcomes of hybrid TEVAR were reported to be worse with a more proximal landing zone. This study aims to clarify the effectiveness of zone 1-landing hybrid TEVAR by comparing the outcomes of zone 2-landing hybrid TEVAR. Methods: From April 2008 to October 2020, 213 patients (zone 1: zone 1-landing hybrid TEVAR, n = 82, 38.5%; zone 2: zone 2-landing hybrid TEVAR, n = 131, 61.5%) were enrolled (median age, 72 years; interquartile range [IQR], 65-78 years), with a median follow-up period of 6.0 years (IQR, 2.8-9.7 years). Results: The mean logistic EuroSCORE was 20.9 ± 14.8%: the logistic EuroSCORE of the zone 1 group (23.3 ± 16.1) was significantly higher than that of the zone 2 group (19.3 ± 12.4%, p = 0.045). The operative time and hospital stay of the zone 1 group were significantly longer than those of the zone 2 group. On the other hand, the in-hospital and late outcomes did not differ significantly between the two groups. There were no significant differences in cumulative survival (66.8% vs. 78.0% at 10 years, Log-rank p = 0.074), aorta-related death-free rates (97.6% vs. 99.2% at 10 years, Log-rank p = 0.312), and aortic event-free rates (81.4% vs. 87.9% at 10 years, Log-rank p = 0.257). Conclusions: Zone 1- and 2-landing hybrid TEVAR outcomes were satisfactory. Despite the high procedural difficulty and surgical risk, the outcomes of zone 1-landing hybrid TEVAR were equal to those of zone 2-landing hybrid TEVAR. If the surgical risk is high, zone 1-landing hybrid TEVAR should not be avoided.

Preoperative higher right ventricular stroke work index increases the risk of de novo aortic insufficiency after continuous-flow left ventricular assist device implantation.

During continuous-flow left ventricular assist device (CF-LVAD) support, hemodynamic shear stress causes a burden on aortic valve (AV) leaflets, leading to de novo aortic insufficiency (AI). This study investigated the influence of preoperative hemodynamic parameters on de novo AI in CF-LVAD recipients. We reviewed 125 patients who underwent CF-LVAD implantation without concomitant AV surgery between 2005 and 2018. De novo AI was defined as moderate or severe AI in those with none or trivial preoperative AI. During mean 30 ± 16 months of CF-LVAD support, de novo AI-free rate was 86% and 67% at 1 and 2 years, respectively. Multivariable analysis showed that higher right ventricular stroke work index (RVSWI) (hazard ratio, 1.12 /g/m2/beat; 95% confidence interval, 1.00-1.20; p = 0.047) and trivial grade AI (hazard ratio, 2.8; 95% confidence interval, 1.2-6.4; p = 0.020) were independent preoperative risk factors for de novo AI. The longitudinal analysis using generalized mixed effects model showed that higher RVSWI was associated with continuous AV closure after LVAD implantation (Odd ratio, 1.20/g/m2/beat; 95% confidence interval, 1.00-1.43 /g/m2/beat; p = 0.047). Right heart catheterization revealed that preoperative RVSWI was positively correlated with postoperative pump flow index in patients with continuously closed AV (r = 0.44, p = 0.04, n = 22). Preoperative higher RVSWI was a significant risk factor for de novo AI following CF-LVAD implantation. In patients with preserved right ventricular function, postoperative higher pump flow may affect AI development via hemodynamic stress on the AV.