A three-dimensional hybrid pacemaker electrode seamlessly integrates into engineered, functional human cardiac tissue in vitro

We have developed an original scaffold-free tissue engineering approach, “cell sheet engineering”, and this technology has been already applied to regenerative medicine of various organs including heart. As the bioengineered three-dimensional cardiac tissue is expected to not only function for repairing the broad injured heart but also to be the practicable heart tissue models, we have developed the cell sheet-based perfusable bioengineered three-dimensional cardiac tissue. Recently we have also developed the unique suspension cultivation system for the high-efficient cardiac differentiation of human iPS cells. Fourteen-day culture with the serial treatments of suitable growth factors and a small compound in this stirring system with the suitable dissolved oxygen concentration produced robust embryoid bodies that showed the spontaneous beating and were mainly composed of cardiomyocytes (~80%). When these differentiated cells were cultured on temperature-responsive culture dishes after the enzymatic dissociation, the spontaneous and synchronous beating was observed accompanied with the intracellular calcium influx all over the area even after cell were detached from culture dishes as cell sheets by lowering the culture temperature. The cardiac cell sheets were mainly composed of cardiomyocytes (~80%) and partially mural cells (~20%). Furthermore, extracellular action potential propagation was observed between cell sheets when two cardiac cell sheets were partially overlaid, and this propagation was inhibited by the treatment with some anti-arrhythmic drugs. When the triple layered cardiac tissue was transplanted onto the subcutaneous tissue of nude rats, the spontaneous pulsation was observed over 2 months and engrafted cardiomyocytes were vascularized with the host tissue-derived endothelial cells. These findings suggest that cardiac cell sheets formed by hiPSC-derived cardiomyocytes might have sufficient properties for the creation of thickened cardiac tissue. Now we are developing the vascularized thickened human cardiac tissue by the repeated layering of cardiac cell sheets on the artificial vascular bed in vitro.

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Production of zebrafish cardiospheres and cardiac progenitor cells in vitro and three-dimensional culture of adult zebrafish cardiac tissue in scaffolds

Biotechnology and Bioengineering ◽

10.1002/bit.26331 ◽

2017 ◽

Vol 114 (9) ◽

pp. 2142-2148 ◽

Cited By ~ 3

Author(s):

Wendy R. Zeng ◽

Siew-Joo Beh ◽

Robert J. Bryson-Richardson ◽

Pauline M. Doran

Keyword(s):

Progenitor Cells ◽

Cardiac Tissue ◽

Three Dimensional ◽

Cardiac Progenitor Cells ◽

Adult Zebrafish ◽

Three Dimensional Culture

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Abstract 30: In vitro Fabrication of Human Cardiac Tissues With Porcine Perfusable Blood Vessels

Circulation Research ◽

10.1161/res.119.suppl_1.30 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Akitoshi Inui ◽

Hidekazu Sekine ◽

Kazunori Sano ◽

Izumi Dobashi ◽

Azumi Yoshida ◽

...

Keyword(s):

Blood Vessels ◽

Cardiac Tissue ◽

Severe Heart Failure ◽

Perfusion Culture ◽

Cardiac Cell ◽

Cell Sheets ◽

Cardiac Tissues ◽

Bioreactor System ◽

Human Cardiac Tissue

The definitive treatment of severe heart failure is heart transplantation; however the number of heart transplantation procedures performed in Japan per year ranges from 30-40 due to donor shortage. Therefore, recently other treatments such as ventricular assist device or regenerative therapy by human cardiac tissue engineering have been developed and are considered as appropriate alternatives. We have developed an original technology, which was named cell-sheet based tissue engineering to fabricate functional three-dimensional tissue by layering cell sheets. The utilization of this technique allowed us to successfully engineer thick rat cardiac tissue with perfusable blood vessels in vitro. Here, we demonstrate a technique to engineer human cardiac tissue with perfusable blood vessels using cardiac cell sheets derived from human induced pluripotent stem cells, and porcine small intestine as a vascular bed for perfusion culture. The small intestine was harvested from with a branch of the superior mesenteric artery and vein and underwent mucosal resection after harvested tissue was cut open. To engineer cardiac tissue with perfusable blood vessels, cardiac cell sheets co-cultured with endothelial cells, were triple-layered and then was overlaid on the vascular bed in the bioreactor system. One day after perfusion culture, overlaid cardiac tissues pulsated spontaneously and were synchronized. The cardiac tissue construct was viable tissue without any observable necrosis. Furthermore we examined the possibility of transplantation of the in vitro engineered human cardiac tissue with the connectable host artery and vein. Engineered cardiac tissue was removed from the bioreactor system after 4-day perfusion, and transplanted to another pig heart. The branch of the superior mesenteric artery and vein of the graft were then reconnected to the host internal thoracic artery and vein. When the cardiac tissue reperfused, it began to beat spontaneously after a few minutes. We believe that this method is useful to fabricate functional cardiac tissue and may become an appropriate treatment for severe heart failure.

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Three‐dimensional bioprinting human cardiac tissue chips of using a painting needle method

Biotechnology and Bioengineering ◽

10.1002/bit.27126 ◽

2019 ◽

Vol 116 (11) ◽

pp. 3136-3142 ◽

Cited By ~ 2

Author(s):

Shohei Chikae ◽

Akifumi Kubota ◽

Haruka Nakamura ◽

Atsushi Oda ◽

Akihiro Yamanaka ◽

...

Keyword(s):

Cardiac Tissue ◽

Three Dimensional ◽

Human Cardiac Tissue

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Strength-duration relationship as a tool to prioritize cardiac tissue properties that govern electrical excitability

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00161.2019 ◽

2019 ◽

Vol 317 (1) ◽

pp. H13-H25 ◽

Cited By ~ 3

Author(s):

Michael N. Sayegh ◽

Natasha Fernandez ◽

Hee Cheol Cho

Keyword(s):

Cardiac Tissue ◽

Three Dimensional ◽

Assessment Tools ◽

Electrical Strength ◽

Design Parameters ◽

Electrical Excitability ◽

Cell Alignment ◽

Tissue Properties ◽

Duration Relationship

Gaps exist in the availability of in vitro functional assessment tools that can emulate the integration of regenerative cells and tissues to the host myocardium. We use strength-duration relationships of electrically stimulated two- and three-dimensional myocardial constructs to study the effects of pacing frequency, culture dimensions, anisotropic cell alignment, fibroblast content, and pacemaker phenotype on electrical excitability. Our study delivers electrical strength-duration as a quantifiable parameter to evaluate design parameters of engineered cardiac tissue constructs.

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In Vitro Studies of Human Cardiac Tissue

Anesthesiology ◽

10.1097/00000542-197011000-00002 ◽

1970 ◽

Vol 33 (5) ◽

pp. 480-480

Author(s):

RICHARD A. JENSEN

Keyword(s):

Cardiac Tissue ◽

In Vitro Studies ◽

Human Cardiac Tissue

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Quantification of Myocyte Disarray in Human Cardiac Tissue

Frontiers in Physiology ◽

10.3389/fphys.2021.750364 ◽

2021 ◽

Vol 12 ◽

Author(s):

Francesco Giardini ◽

Erica Lazzeri ◽

Giulia Vitale ◽

Cecilia Ferrantini ◽

Irene Costantini ◽

...

Keyword(s):

Spatial Scale ◽

Cardiac Tissue ◽

Spatial Scales ◽

Three Dimensional ◽

Structural Data ◽

Multi Scale ◽

Tissue Organization ◽

Human Cardiac Tissue ◽

Microscopy Techniques ◽

Scale Of Analysis

Proper three-dimensional (3D)-cardiomyocyte orientation is important for an effective tension production in cardiac muscle. Cardiac diseases can cause severe remodeling processes in the heart, such as cellular misalignment, that can affect both the electrical and mechanical functions of the organ. To date, a proven methodology to map and quantify myocytes disarray in massive samples is missing. In this study, we present an experimental pipeline to reconstruct and analyze the 3D cardiomyocyte architecture in massive samples. We employed tissue clearing, staining, and advanced microscopy techniques to detect sarcomeres in relatively large human myocardial strips with micrometric resolution. Z-bands periodicity was exploited in a frequency analysis approach to extract the 3D myofilament orientation, providing an orientation map used to characterize the tissue organization at different spatial scales. As a proof-of-principle, we applied the proposed method to healthy and pathologically remodeled human cardiac tissue strips. Preliminary results suggest the reliability of the method: strips from a healthy donor are characterized by a well-organized tissue, where the local disarray is log-normally distributed and slightly depends on the spatial scale of analysis; on the contrary, pathological strips show pronounced tissue disorganization, characterized by local disarray significantly dependent on the spatial scale of analysis. A virtual sample generator is developed to link this multi-scale disarray analysis with the underlying cellular architecture. This approach allowed us to quantitatively assess tissue organization in terms of 3D myocyte angular dispersion and may pave the way for developing novel predictive models based on structural data at cellular resolution.

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Abstract 213: Evolving Challenges in Promoting Stem Cell Based Cardiovascular Repair and Regeneration

Circulation Research ◽

10.1161/res.119.suppl_1.213 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

Mani T Valarmathi ◽

Jiang Li

Keyword(s):

Stem Cells ◽

Stem Cell ◽

Cardiac Muscle ◽

Adult Stem Cells ◽

Cardiac Tissue ◽

Three Dimensional ◽

Induced Pluripotent Stem Cell ◽

Myocardial Damage ◽

Culture Conditions

Introduction: Use of adult stem cells in the stimulation of mammalian cardiac muscle regeneration is in its infancy, and to date, it has been difficult to determine the efficacy of the procedures that have been employed. The outstanding question remains whether stem cells derived from the bone-marrow or some other location within or outside of the heart can populate a region of myocardial damage and transform into tissue-specific cells, and also exhibit functional synchronization. As a result, this necessitates the development of an appropriate in vitro three-dimensional (3-D) model of cardiomyogenesis and prompts the development of a 3-D cardiac muscle construct for tissue engineering purposes, especially using the adult stem cells. Hypothesis: Functioning vascularized cardiac tissue can be generated by the interaction of human induced pluripotent stem cell-derived embryonic cardiac myocytes (hiPSC-ECMs) and human multipotent mesenchymal stem cells (hMSCs) on a 3-D prevascularized collagen cell carrier (CCC) scaffold. Methods and Results: In order to achieve the above aim, we have developed an in vitro 3-D functioning vascularized cardiac muscle construct using hiPSC-ECMs and hMSCs. First, to generate the prevascularized scaffold, human cardiac microvascular endothelial cells (hCMVECs) and hMSCs were co-cultured on 3-D CCCs for 7 days under vasculogenic culture conditions, hCMVECs/hMSCs underwent maturation, differentiation, and morphogenesis characteristic of micro vessels, and formed extensive plexuses of vascular networks. Next, the hiPSC-ECMs and hMSCs were co-cultured onto this generated prevascularized CCCs for further 7 or 14 days in myogenic culture conditions. Finally, the vascular and cardiac phenotypic inductions were analyzed at the morphological, immunological, biochemical, molecular, and functional levels. Expression and functional analyses of the differentiated cells revealed dramatic neo-angiogenesis and neo-cardiomyogenesis. Conclusions: Thus, our unique 3-D co-culture system provided us the apt in vitro functioning prevascularized 3-D cardiac patch that can be utilized for cellular cardiomyoplasty.

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SARS-CoV-2 infects and induces cytotoxic effects in human cardiomyocytes

10.1101/2020.06.01.127605 ◽

2020 ◽

Cited By ~ 4

Author(s):

Denisa Bojkova ◽

Julian Wagner ◽

Mariana Shumliakivska ◽

Galip Aslan ◽

Umber Saleem ◽

...

Keyword(s):

Cardiac Tissue ◽

Cardiac Injury ◽

Transcriptional Response ◽

Human Cardiomyocytes ◽

Global Pandemic ◽

Virus Rna ◽

Human Cardiac Tissue ◽

Induced Pluripotent ◽

Apoptotic Effects

Background: The coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and has emerged as global pandemic. SARS-CoV-2 infection can lead to elevated markers of cardiac injury associated with higher risk of mortality in COVID-19 patients. It is unclear whether cardiac injury may have been caused by direct infection of cardiomyocytes or is mainly secondary to lung injury and inflammation. Here we investigate whether human cardiomyocytes are permissive for SARS-CoV-2 infection. Methods: Infection was induced by two strains of SARS-CoV-2 (FFM1 and FFM2) in human induced pluripotent stem cells-derived cardiomyocytes (hiPS-CM) and in two models of human cardiac tissue. Results: We show that SARS-CoV-2 infects hiPS-CM as demonstrated by detection of intracellular double strand viral RNA and viral spike glycoprotein protein expression. Increasing concentrations of virus RNA are detected in supernatants of infected cardiomyocytes, which induced infections in CaCo-2 cell lines documenting productive infections. SARS-COV-2 infection induced cytotoxic and pro-apoptotic effects and abolished cardiomyocyte beating. RNA sequencing confirmed a transcriptional response to viral infection as demonstrated by the up-regulation of genes associated with pathways related to viral response and interferon signaling, apoptosis and reactive oxygen stress. SARS-CoV-2 infection and cardiotoxicity was confirmed in a iPS-derived human 3D cardiosphere tissue models. Importantly, viral spike protein and viral particles were detected in living human heart slices after infection with SARS-CoV-2. Conclusions: The demonstration that cardiomyocytes are permissive for SARS-CoV-2 infection in vitro warrants the further in depth monitoring of cardiotoxic effects in COVID-19 patients.

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