Gq-activated fibroblasts induce cardiomyocyte action potential prolongation and automaticity in a three-dimensional microtissue environment

Cardiac fibroblasts (CFs) are known to regulate cardiomyocyte (CM) function in vivo and in two-dimensional in vitro cultures. This study examined the effect of CF activation on the regulation of CM electrical activity in a three-dimensional (3-D) microtissue environment. Using a scaffold-free 3-D platform with interspersed neonatal rat ventricular CMs and CFs, Gq-mediated signaling was selectively enhanced in CFs by Gαq adenoviral infection before coseeding with CMs in nonadhesive hydrogels. After 3 days, the microtissues were analyzed by signaling assay, histological staining, quantitative PCR, Western blots, optical mapping with voltage- or Ca2+-sensitive dyes, and microelectrode recordings of CF resting membrane potential (RMPCF). Enhanced Gq signaling in CFs increased microtissue size and profibrotic and prohypertrophic markers. Expression of constitutively active Gαq in CFs prolonged CM action potential duration (by 33%) and rise time (by 31%), prolonged Ca2+ transient duration (by 98%) and rise time (by 65%), and caused abnormal electrical activity based on depolarization-induced automaticity. Constitutive Gq activation in CFs also depolarized RMPCF from –33 to −20 mV and increased connexin 43 and connexin 45 expression. Computational modeling confers that elevated RMPCF and increased cell-cell coupling between CMs and CFs in a 3-D environment could lead to automaticity. In conclusion, our data demonstrate that CF activation alone is capable of altering action potential and Ca2+ transient characteristics of CMs, leading to proarrhythmic electrical activity. Our results also emphasize the importance of a 3-D environment where cell-cell interactions are prevalent, underscoring that CF activation in 3-D tissue plays a significant role in modulating CM electrophysiology and arrhythmias. NEW & NOTEWORTHY In a three-dimensional microtissue model, which lowers baseline activation of cardiac fibroblasts but enables cell-cell, paracrine, and cell-extracellular matrix interactions, we demonstrate that selective cardiac fibroblast activation by enhanced Gq signaling, a pathophysiological trigger in the diseased heart, modulates cardiomyocyte electrical activity, leading to proarrhythmogenic automaticity.

Download Full-text

Selective optogenetic stimulation of fibroblasts enables quantification of hetero-cellular coupling to cardiomyocytes in a three-dimensional model of heart tissue

EP Europace ◽

10.1093/europace/euaa128 ◽

2020 ◽

Vol 22 (10) ◽

pp. 1590-1599

Author(s):

Maximilian Funken ◽

Tobias Bruegmann ◽

Philipp Sasse

Keyword(s):

Membrane Potential ◽

Growth Factor ◽

Connexin 43 ◽

Transforming Growth Factor ◽

Three Dimensional ◽

Transforming Growth Factor Β1 ◽

Resting Membrane Potential ◽

Human Cardiomyocytes ◽

Tgf Β1

Abstract Aims Besides providing mechanical stability, fibroblasts in the heart could modulate the electrical properties of cardiomyocytes. Here, we aim to develop a three-dimensional hetero-cellular model to analyse the electric interaction between fibroblasts and human cardiomyocytes in vitro using selective optogenetic de- or hyperpolarization of fibroblasts. Methods and results NIH3T3 cell lines expressing the light-sensitive ion channel Channelrhodopsin2 or the light-induced proton pump Archaerhodopsin were generated for optogenetic depolarization or hyperpolarization, respectively, and characterized by patch clamp. Cardiac bodies consisting of 50% fibroblasts and 50% human pluripotent stem cell-derived cardiomyocytes were analysed by video microscopy and membrane potential was measured with sharp electrodes. Myofibroblast activation in cardiac bodies was enhanced by transforming growth factor-β1 (TGF-β1)-stimulation. Connexin-43 expression was analysed by qPCR and fluorescence recovery after photobleaching. Illumination of Channelrhodopsin2 or Archaerhodopsin expressing fibroblasts induced inward currents and depolarization or outward currents and hyperpolarization. Transforming growth factor-β1-stimulation elevated connexin-43 expression and increased cell–cell coupling between fibroblasts as well as increased basal beating frequency and cardiomyocyte resting membrane potential in cardiac bodies. Illumination of cardiac bodies generated with Channelrhodopsin2 fibroblasts accelerated spontaneous beating, especially after TGF-β1-stimulation. Illumination of cardiac bodies prepared with Archaerhodopsin expressing fibroblasts led to hyperpolarization of cardiomyocytes and complete block of spontaneous beating after TGF-β1-stimulation. Effects of light were significantly smaller without TGF-β1-stimulation. Conclusion Transforming growth factor-β1-stimulation leads to increased hetero-cellular coupling and optogenetic hyperpolarization of fibroblasts reduces TGF-β1 induced effects on cardiomyocyte spontaneous activity. Optogenetic membrane potential manipulation selectively in fibroblasts in a new hetero-cellular cardiac body model allows direct quantification of fibroblast–cardiomyocyte coupling in vitro.

Download Full-text

Development of electrical activity in cardiac myocyte aggregates derived from mouse embryonic stem cells

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01106.2001 ◽

2003 ◽

Vol 284 (6) ◽

pp. H2114-H2123 ◽

Cited By ~ 76

Author(s):

K. Banach ◽

M. D. Halbach ◽

P. Hu ◽

J. Hescheler ◽

U. Egert

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Action Potential ◽

Electrical Activity ◽

Cardiac Myocyte ◽

Three Dimensional ◽

Embryonic Stem ◽

Electrophysiological Properties ◽

Chronotropic Response

Embryonic stem cells differentiate into cardiac myocytes, repeating in vitro the structural and molecular changes associated with cardiac development. Currently, it is not clear whether the electrophysiological properties of the multicellular cardiac structure follow cardiac maturation as well. In long-term recordings of extracellular field potentials with microelectrode arrays consisting of 60 substrate-integrated electrodes, we examined the electrophysiological properties during the ongoing differentiation process. The beating frequency of the growing preparations increased from 1 to 5 Hz concomitant to a decrease of the action potential duration and action potential rise time. A developmental increase of the conduction velocity could be attributed to an increased expression of connexin43 gap junction channels. Whereas isoprenalin elicited a positive chronotropic response from the first day of spontaneous beating onward, a concentration-dependent negative chronotropic effect of carbachol only developed after ∼4 days. The in vitro development of the three-dimensional cardiac preparation thus closely follows the development described for the mouse embryonic heart, making it an ideal model to monitor the differentiation of electrical activity in embryonic cardiomyocytes.

Download Full-text

Altered contractility and [Ca2+]i homeostasis in phospholemman-deficient murine myocytes: role of Na+/Ca2+ exchange

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01181.2005 ◽

2006 ◽

Vol 291 (5) ◽

pp. H2199-H2209 ◽

Cited By ~ 35

Author(s):

Amy L. Tucker ◽

Jianliang Song ◽

Xue-Qian Zhang ◽

JuFang Wang ◽

Belinda A. Ahlers ◽

...

Keyword(s):

Protein Kinase ◽

Action Potential ◽

Cardiac Contractility ◽

Resting Membrane Potential ◽

Wild Type ◽

Cell Width ◽

Western Blots ◽

Whole Cell ◽

Major Function ◽

Adult Rat

Phospholemman (PLM) regulates contractility and Ca2+ homeostasis in cardiac myocytes. We characterized excitation-contraction coupling in myocytes isolated from PLM-deficient mice backbred to a pure congenic C57BL/6 background. Cell length, cell width, and whole cell capacitance were not different between wild-type and PLM-null myocytes. Compared with wild-type myocytes, Western blots indicated total absence of PLM but no changes in Na+/Ca2+ exchanger, sarcoplasmic reticulum (SR) Ca2+-ATPase, α1-subunit of Na+-K+-ATPase, and calsequestrin levels in PLM-null myocytes. At 5 mM extracellular Ca2+ concentration ([Ca2+]o), contraction and cytosolic [Ca2+] ([Ca2+]i) transient amplitudes and SR Ca2+ contents in PLM-null myocytes were significantly ( P < 0.0004) higher than wild-type myocytes, whereas the converse was true at 0.6 mM [Ca2+]o. This pattern of contractile and [Ca2+]i transient abnormalities in PLM-null myocytes mimics that observed in adult rat myocytes overexpressing the cardiac Na+/Ca2+ exchanger. Indeed, we have previously reported that Na+/Ca2+ exchange currents were higher in PLM-null myocytes. Activation of protein kinase A resulted in increased inotropy such that there were no longer any contractility differences between the stimulated wild-type and PLM-null myocytes. Protein kinase C stimulation resulted in decreased contractility in both wild-type and PLM-null myocytes. Resting membrane potential and action potential amplitudes were similar, but action potential duration was much prolonged ( P < 0.04) in PLM-null myocytes. Whole cell Ca2+ current densities were similar between wild-type and PLM-null myocytes, as were the fast- and slow-inactivation time constants. We conclude that a major function of PLM is regulation of cardiac contractility and Ca2+ fluxes, likely by modulating Na+/Ca2+ exchange activity.

Download Full-text

OptoGap is an optogenetics-enabled assay for quantification of cell–cell coupling in multicellular cardiac tissue

Scientific Reports ◽

10.1038/s41598-021-88573-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Patrick M. Boyle ◽

Jinzhu Yu ◽

Aleksandra Klimas ◽

John C. Williams ◽

Natalia A. Trayanova ◽

...

Keyword(s):

Cardiac Tissue ◽

Cardiac Fibroblasts ◽

Direct Method ◽

Electrical Coupling ◽

Three Dimensional ◽

Computational Modelling ◽

Cell Coupling ◽

Excitable Cells ◽

Potential Applicability ◽

Cell Cell

Download Full-text

Transient Receptor Potential Vanilloid 4 channel participates in mouse ventricular electrical activity

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00497.2020 ◽

2021 ◽

Author(s):

Sebastien Chaigne ◽

Guillaume Cardouat ◽

Julien Louradour ◽

Fanny Vaillant ◽

Sabine Charron ◽

...

Keyword(s):

Action Potential ◽

Electrical Activity ◽

Transient Receptor Potential ◽

Ventricular Myocytes ◽

Receptor Potential ◽

Left Ventricular ◽

Transient Receptor Potential Vanilloid ◽

Western Blots ◽

Transient Receptor ◽

Trpv4 Channel

Introduction: Transient Receptor Potential Vanilloid 4 (TRPV4) channel is a calcium permeable channel (PCa/PNa ~ 10). Its expression was reported in ventricular myocytes where it is involved in several cardiac pathological mechanisms. In this study, we investigated the implication of TRPV4 in ventricular electrical activity. Methods and Results: Left ventricular myocytes were isolated from trpv4+/+ and trpv4-/- mice. TRPV4 membrane expression and its colocalization with Cav1.2 was confirmed using western-blots biotinylation, immunoprecipitation and immunostaining experiments. Then, electrocardiograms (ECGs) and patch-clamp recordings showed shortened QTc and action potential (AP) duration in trpv4-/- compared to trpv4+/+ mice. Thus, TRPV4 activator GSK1016790A produced a transient and dose-dependent increase in AP duration at 90 % of repolarization (APD90) in trpv4+/+, but not in trpv4-/- myocytes or when combined with TRPV4 inhibitor GSK2193874 (100 nM). Hence, GSK1016790A increased CaT amplitude in trpv4+/+ but not in trpv4-/- myocytes, suggesting that TRPV4 carries an inward Ca2+ current in myocytes. Conversely, TRPV4 inhibitor GSK2193874 (100 nM) alone reduced APD90 in trpv4+/+ but not in trpv4-/- myocytes, suggesting that TRPV4 prolongs AP duration (APD) in basal condition. Finally, introducing TRPV4 parameters in a mathematical model predicted the development of an inward TRPV4 current during repolarization that increases AP duration and CaT amplitude, in accordance with what found experimentally. Conclusion: This study shows for the first time that TRPV4 modulates AP and QTc durations and constitutes thereby a good therapeutical target against long QT-mediated ventricular arrhythmias. Keywords: TRPV4 channel, action potential, QT interval, mathematical modeling, trpv4-/-, calcium transient.

Download Full-text

Abstract 542: Chronic Doxorubicin Cardiotoxicity Assessed in Engineered Cardiac Tissues Generated in Biowire™ II Platform

Circulation Research ◽

10.1161/res.127.suppl_1.542 ◽

2020 ◽

Vol 127 (Suppl_1) ◽

Author(s):

Roozbeh Aschar-sobbi ◽

Julia E Napolitano ◽

Danielle R Bogdanowicz ◽

Michael P Graziano

Keyword(s):

Action Potential ◽

Action Potential Duration ◽

Cardiac Fibroblasts ◽

Left Ventricular ◽

Left Ventricular Ejection ◽

Resting Membrane Potential ◽

Ventricular Ejection Fraction ◽

Cardiac Tissues

The anthracycline doxorubicin is an effective anti-tumor agent widely used in both adults and children. One major adverse effect of doxorubicin therapy is dose-dependent cardiotoxicity, ranging from asymptomatic reduction in left ventricular ejection fraction to more serious, potentially fatal symptoms including arrythmias and congestive heart failure. The exact mechanism of doxorubicin-induced cardiotoxicity remains unknown. Recently, human induced pluripotent stem cells (hiPSC) have emerged as a potential tool to model cardiac toxicity, but their fetal-like phenotype raises concerns about the translatability of in vitro data to in vivo cardiotoxicity. To overcome this limitation, Biowire™ II platform was used to generate 3D engineered cardiac tissues (ECTs) from hiPSC-derived cardiomyocytes and human cardiac fibroblasts. Using long-term electrical stimulation, ECTs with a phenotype approaching that of adult human myocardium were obtained. The ECTs were then exposed to 1 μM doxorubicin for 8 days followed by 7 days of washout. Measurements of contractile force amplitude at 1 Hz stimulation showed a transient increase in force within 24 hours of doxorubicin exposure followed by decrease in force after 2 days. Intracellular recordings of action potential (AP) showed a decrease in maximum upstroke velocity (dV/dt), AP amplitude (APA), and resting membrane potential (RMP) after 8 days of doxorubicin treatment. In addition, action potential duration (APD) at 30% (APD30) repolarization was increased in doxorubicin-treated ECTs, whereas APD50 and APD90 were decreased. Following 7 days of washout, no difference in force or AP parameters was found between doxorubicin and vehicle-treated ECTs with the exception of APD50 and APD90 which remained abbreviated. A global untargeted analysis of the conditioned media from doxorubicin-treated ECTs identified 204 analytes and revealed an upregulation of redox homeostasis, differential fatty acid metabolism, altered glycolysis and TCA cycle metabolites, and decreased nucleoside metabolism compared to vehicle-treated ECTs. These results show that doxorubicin not only increases oxidative stress, but also irreversibly affects action potential duration which may predispose to cardiac arrhythmias.

Download Full-text

Compromised Biomechanical Properties, Cell–Cell Adhesion and Nanotubes Communication in Cardiac Fibroblasts Carrying the Lamin A/C D192G Mutation

International Journal of Molecular Sciences ◽

10.3390/ijms22179193 ◽

2021 ◽

Vol 22 (17) ◽

pp. 9193

Author(s):

Veronique Lachaize ◽

Brisa Peña ◽

Catalin Ciubotaru ◽

Dan Cojoc ◽

Suet Nee Chen ◽

...

Keyword(s):

Cell Adhesion ◽

Optical Tweezers ◽

Cardiac Fibroblasts ◽

Biomechanical Properties ◽

Neonatal Rat ◽

Lamin A ◽

Clinical Effects ◽

Tunneling Nanotubes ◽

Adhesion Properties ◽

Cell Cell

Clinical effects induced by arrhythmogenic cardiomyopathy (ACM) originate from a large spectrum of genetic variations, including the missense mutation of the lamin A/C gene (LMNA), LMNA D192G. The aim of our study was to investigate the biophysical and biomechanical impact of the LMNA D192G mutation on neonatal rat ventricular fibroblasts (NRVF). The main findings in mutated NRVFs were: (i) cytoskeleton disorganization (actin and intermediate filaments); (ii) decreased elasticity of NRVFs; (iii) altered cell–cell adhesion properties, that highlighted a strong effect on cellular communication, in particular on tunneling nanotubes (TNTs). In mutant-expressing fibroblasts, these nanotubes were weakened with altered mechanical properties as shown by atomic force microscopy (AFM) and optical tweezers. These outcomes complement prior investigations on LMNA mutant cardiomyocytes and suggest that the LMNA D192G mutation impacts the biomechanical properties of both cardiomyocytes and cardiac fibroblasts. These observations could explain how this mutation influences cardiac biomechanical pathology and the severity of ACM in LMNA-cardiomyopathy.

Download Full-text

OptoGap: an optogenetics-enabled assay for quantification of cell-cell coupling in multicellular cardiac tissue

10.1101/171397 ◽

2017 ◽

Cited By ~ 2

Author(s):

Jinzhu Yu ◽

Patrick M. Boyle ◽

Aleksandra Klimas ◽

John C. Williams ◽

Natalia Trayanova ◽

...

Keyword(s):

Messenger Rna ◽

Quantitative Methods ◽

Cardiac Tissue ◽

De Novo ◽

Cardiac Fibroblasts ◽

Direct Method ◽

Electrical Coupling ◽

Cell Coupling ◽

Western Blots ◽

Cell Cell

AbstractIntercellular electrical coupling is an essential means of communication between cells. It is important to obtain quantitative knowledge of such coupling between cardiomyocytes and nonexcitable cells when, for example, pathological electrical coupling between myofibroblasts and cardiomyocytes yields increased arrhythmia risk or during the integration of donor (e.g. cardiac progenitor) cells with native cardiomyocytes in cell-therapy approaches. Currently, there is no direct method for assessing heterocellular coupling within multicellular tissue. Here we demonstrate experimentally and computationally a new contactless assay for electrical coupling, OptoGap, based on selective illumination of inexcitable cells that express optogenetic actuators and optical sensing of the response of coupled excitable cells, e.g. cardiomyocytes, that are light-insensitive. Cell-cell coupling is quantified by the energy required to elicit an action potential via junctional current from the light-stimulated cell(s). The proposed technique is experimentally validated against the standard indirect approach, GapFRAP, using light-sensitive cardiac fibroblasts and non-transformed cardiomyocytes in a two-dimensional setting. It’s potential applicability to the complex three-dimensional setting of the native heart is corroborated by computational modeling and proper calibration.Intercellular coupling is a fundamental form of communication between cells, essential for the synchronization of physiological processes in different organs. Pathologically altered coupling or the emergence of de novo coupling between native and donor cells are problems of interest in many cardiac applications, e.g. during cell delivery and cell integration for cardiac repair therapy1,2. In particular, interactions between cardiomyocytes and fibroblasts are of interest, especially the pro-arrhythmic increase in coupling as the latter transition to myofibroblasts3-6.Electrical coupling in cardiac tissue is mediated primarily by low-resistance paths formed by gap-junctional proteins (connexins), that can link cardiomyocytes (CMs) to each other and to non-cardiomyocytes (nCMs), such as fibroblasts. Qualitative and quantitative methods, e.g. immunofluorescence, messenger RNA and Western blots, are often used to assay connexin expression levels as a surrogate measure of coupling, but they do not provide functional information. A method for direct quantification of cell-cell coupling within the multicellular tissue context is highly desirable.

Download Full-text

Abstract P016: Human Mesenchymal Bone Marrow--Derived Stem Cells Require Connexin 43 to Form Beating 3-Dimensional Tubes When Cocultured with Neonatal Rat Cardiomyocytes

Circulation Research ◽

10.1161/res.109.suppl_1.ap016 ◽

2011 ◽

Vol 109 (suppl_1) ◽

Author(s):

Cristina Sanina ◽

Claudia O Rodrigues ◽

Ivonne H Schulman ◽

Irene Margitich ◽

Wayne Balkan ◽

...

Keyword(s):

Stem Cells ◽

Real Time ◽

Real Time Pcr ◽

Heart Development ◽

Connexin 43 ◽

Three Dimensional ◽

Neonatal Rat ◽

Control Group ◽

Cx43 Expression ◽

Rat Cardiomyocytes

Mesenchymal stem cells (MSCs) have begun to manifest themselves as a safe and beneficial therapy for restoring cardiac function in failing hearts. However, the mechanism underlying this function is unclear. The present study was initiated to investigate the role of Connexin 43 (Cx43) in hMSC integration, migration and differentiation in co-culture with neonatal rat cardiomyocyte (NRVMs) in vitro. Cx43 is a gap junction connexin which is required for proper heart development and heart electrophysiology. We generated lentiviral constructs for Cx43 knockdown and Cx43 overexpression and used them to alter the level of Cx43 in hMSCs. The effectiveness of these vectors was confirmed by assessing Cx43 levels in hMSCs by Western Blot analysis and by Real Time-PCR. These hMSCs were co-cultured with NRVMs for up to 1 month on poly-lysine and collagen I coated glass cover slips. Co-cultures containing MSCs overexpressing Cx43 exhibited coordinated beating and three-dimensional tube formation in 10 days, whereas 14 days were required for control cells. The hMSCs were integrated into these beating three dimensional tubes. No tubes were observed in co-cultures with Cx43 knockdown hMSCs, rather we observed the formation of small, unconnected beating spheres after 30 days in co-culture. Human MSC-Cx43 knockdown cells were integrated into these spheres and immunofluorescence staining demonstrated that Cx43 expression in hMSC-Cx43 knockdown remained reduced when compared to the control group. Real Time-PCR analysis using human specific primers showed significant upregulation of KDR, smooth muscle actin, Pecam1, CD34, CDH2, and CaCNA1C. Several genes, not initially seen in hMSCs, including Gata4, CDH5, SCN5A, SLC8A1, and KCNQ1, appeared in co-cultures with hMSCs with normal and elevated expression levels of Cx43. However, in co-culture of cardiomyocytes with Cx43 knockdown MSCs we observed downregulation of KDR, CHD2, SCN5A, SLC8A1, KCNQ1 compare to control. These results suggest a strong correlation between the presence of Cx43 and the ability of hMSCs to differentiate into cardiac and endothelial lineages.

Download Full-text

Functional scaffold-free 3-D cardiac microtissues: a novel model for the investigation of heart cells

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00743.2011 ◽

2012 ◽

Vol 302 (10) ◽

pp. H2031-H2042 ◽

Cited By ~ 52

Author(s):

B. R. Desroches ◽

P. Zhang ◽

B.-R. Choi ◽

M. E. King ◽

A. E. Maldonado ◽

...

Keyword(s):

Cell Culture ◽

Cardiac Fibroblasts ◽

Neonatal Rat ◽

Resting Membrane Potential ◽

Heart Cells ◽

Cell Type ◽

Adenoviral Gene Transfer ◽

Culture Model ◽

Cell Type Specific ◽

Novel Scaffold

To bridge the gap between two-dimensional cell culture and tissue, various three-dimensional (3-D) cell culture approaches have been developed for the investigation of cardiac myocytes (CMs) and cardiac fibroblasts (CFs). However, several limitations still exist. This study was designed to develop a cardiac 3-D culture model with a scaffold-free technology that can easily and inexpensively generate large numbers of microtissues with cellular distribution and functional behavior similar to cardiac tissue. Using micromolded nonadhesive agarose hydrogels containing 822 concave recesses (800 μm deep × 400 μm wide), we demonstrated that neonatal rat ventricular CMs and CFs alone or in combination self-assembled into viable (Live/Dead stain) spherical-shaped microtissues. Importantly, when seeded simultaneously or sequentially, CMs and CFs self-sorted to be interspersed, reminiscent of their myocardial distribution, as shown by cell type-specific CellTracker or antibody labeling. Microelectrode recordings and optical mapping revealed characteristic triangular action potentials (APs) with a resting membrane potential of −66 ± 7 mV ( n = 4) in spontaneously contracting CM microtissues. Under pacing, optically mapped AP duration at 90% repolarization and conduction velocity were 100 ± 30 ms and 18.0 ± 1.9 cm/s, respectively ( n = 5 each). The presence of CFs led to a twofold AP prolongation in heterogenous microtissues (CM-to-CF ratio of 1:1). Importantly, Ba2+-sensitive inward rectifier K+ currents and Ca2+-handling proteins, including sarco(endo)plasmic reticulum Ca2+-ATPase 2a, were detected in CM-containing microtissues. Furthermore, cell type-specific adenoviral gene transfer was achieved, with no impact on microtissue formation or cell viability. In conclusion, we developed a novel scaffold-free cardiac 3-D culture model with several advancements for the investigation of CM and CF function and cross-regulation.

Download Full-text