Modeling Heart Diseases on a Chip: Advantages and Future Opportunities.

Heart disease is a significant burden on global health care systems and is a leading cause of death each year. To improve our understanding of heart disease, high quality disease models are needed. These will facilitate the discovery and development of new treatments for heart disease. Traditionally, researchers have relied on 2D monolayer systems or animal models of heart disease to elucidate pathophysiology and drug responses. Heart-on-a-chip (HOC) technology is an emerging field where cardiomyocytes among other cell types in the heart can be used to generate functional, beating cardiac microtissues that recapitulate many features of the human heart. HOC models are showing great promise as disease modeling platforms and are poised to serve as important tools in the drug development pipeline. By leveraging advances in human pluripotent stem cell-derived cardiomyocyte biology and microfabrication technology, diseased HOCs are highly tuneable and can be generated via different approaches such as: using cells with defined genetic backgrounds (patient-derived cells), adding small molecules, modifying the cells' environment, altering cell ratio/composition of microtissues, among others. HOCs have been used to faithfully model aspects of arrhythmia, fibrosis, infection, cardiomyopathies, and ischemia, to name a few. In this review, we highlight recent advances in disease modeling using HOC systems, describing instances where these models outperformed other models in terms of reproducing disease phenotypes and/or led to drug development.

Spike Protein Impairs Mitochondrial Function in Human Cardiomyocytes: Mechanisms Underlying Cardiac Injury in COVID-19.

BACKGROUND: COVID-19 has a major impact on cardiovascular diseases and may lead to myocarditis or cardiac failure. The clove-like spike (S) protein of SARS-CoV-2 facilitates its transmission and pathogenesis. Cardiac mitochondria produce energy for key heart functions. We hypothesized that S1 would directly impair the functions of cardiomyocyte mitochondria, thus causing cardiac dysfunction. METHODS: Through the Seahorse Mito Stress Test and real-time ATP rate assays, we explored the mitochondrial bioenergetics in human cardiomyocytes (AC16). The cells were treated without (control) or with S1 (1 nM) for 24, 48, and 72 h and we observed the mitochondrial morphology using transmission electron microscopy and confocal fluorescence microscopy. Western blotting, XRhod-1, and MitoSOX Red staining were performed to evaluate the expression of proteins related to energetic metabolism and relevant signaling cascades, mitochondrial Ca2+ levels, and ROS production. RESULTS: The 24 h S1 treatment increased ATP production and mitochondrial respiration by increasing the expression of fatty-acid-transporting regulators and inducing more negative mitochondrial membrane potential (Δψm). The 72 h S1 treatment decreased mitochondrial respiration rates and Δψm, but increased levels of reactive oxygen species (ROS), mCa2+, and intracellular Ca2+. Electron microscopy revealed increased mitochondrial fragmentation/fission in AC16 cells treated for 72 h. The effects of S1 on ATP production were completely blocked by neutralizing ACE2 but not CD147 antibodies, and were partly attenuated by Mitotempo (1 µM). CONCLUSION: S1 might impair mitochondrial function in human cardiomyocytes by altering Δψm, mCa2+ overload, ROS accumulation, and mitochondrial dynamics via ACE2.

SARS-CoV-2 spike protein-mediated cardiomyocyte fusion may contribute to increased arrhythmic risk in COVID-19.

BACKGROUND: SARS-CoV-2-mediated COVID-19 may cause sudden cardiac death (SCD). Factors contributing to this increased risk of potentially fatal arrhythmias include thrombosis, exaggerated immune response, and treatment with QT-prolonging drugs. However, the intrinsic arrhythmic potential of direct SARS-CoV-2 infection of the heart remains unknown. OBJECTIVE: To assess the cellular and electrophysiological effects of direct SARS-CoV-2 infection of the heart using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). METHODS: hiPSC-CMs were transfected with recombinant SARS-CoV-2 spike protein (CoV-2 S) or CoV-2 S fused to a modified Emerald fluorescence protein (CoV-2 S-mEm). Cell morphology was visualized using immunofluorescence microscopy. Action potential duration (APD) and cellular arrhythmias were measured by whole cell patch-clamp. Calcium handling was assessed using the Fluo-4 Ca2+ indicator. RESULTS: Transfection of hiPSC-CMs with CoV-2 S-mEm produced multinucleated giant cells (syncytia) displaying increased cellular capacitance (75±7 pF, n = 10 vs. 26±3 pF, n = 10; P<0.0001) consistent with increased cell size. The APD90 was prolonged significantly from 419±26 ms (n = 10) in untransfected hiPSC-CMs to 590±67 ms (n = 10; P<0.05) in CoV-2 S-mEm-transfected hiPSC-CMs. CoV-2 S-induced syncytia displayed delayed afterdepolarizations, erratic beating frequency, and calcium handling abnormalities including calcium sparks, large "tsunami"-like waves, and increased calcium transient amplitude. After furin protease inhibitor treatment or mutating the CoV-2 S furin cleavage site, cell-cell fusion was no longer evident and Ca2+ handling returned to normal. CONCLUSION: The SARS-CoV-2 spike protein can directly perturb both the cardiomyocyte's repolarization reserve and intracellular calcium handling that may confer the intrinsic, mechanistic substrate for the increased risk of SCD observed during this COVID-19 pandemic.

TRPC3-Nox2 Protein Complex Formation Increases the Risk of SARS-CoV-2 Spike Protein-Induced Cardiomyocyte Dysfunction through ACE2 Upregulation.

Myocardial damage caused by the newly emerged coronavirus (SARS-CoV-2) infection is one of the key determinants of COVID-19 severity and mortality. SARS-CoV-2 entry to host cells is initiated by binding with its receptor, angiotensin-converting enzyme (ACE) 2, and the ACE2 abundance is thought to reflect the susceptibility to infection. Here, we report that ibudilast, which we previously identified as a potent inhibitor of protein complex between transient receptor potential canonical (TRPC) 3 and NADPH oxidase (Nox) 2, attenuates the SARS-CoV-2 spike glycoprotein pseudovirus-evoked contractile and metabolic dysfunctions of neonatal rat cardiomyocytes (NRCMs). Epidemiologically reported risk factors of severe COVID-19, including cigarette sidestream smoke (CSS) and anti-cancer drug treatment, commonly upregulate ACE2 expression level, and these were suppressed by inhibiting TRPC3-Nox2 complex formation. Exposure of NRCMs to SARS-CoV-2 pseudovirus, as well as CSS and doxorubicin (Dox), induces ATP release through pannexin-1 hemi-channels, and this ATP release potentiates pseudovirus entry to NRCMs and human iPS cell-derived cardiomyocytes (hiPS-CMs). As the pseudovirus entry followed by production of reactive oxygen species was attenuated by inhibiting TRPC3-Nox2 complex in hiPS-CMs, we suggest that TRPC3-Nox2 complex formation triggered by panexin1-mediated ATP release participates in exacerbation of myocardial damage by amplifying ACE2-dependent SARS-CoV-2 entry.

SARS-CoV-2 Nsp6 damages Drosophila heart and mouse cardiomyocytes through MGA/MAX complex-mediated increased glycolysis.

SARS-CoV-2 infection causes COVID-19, a severe acute respiratory disease associated with cardiovascular complications including long-term outcomes. The presence of virus in cardiac tissue of patients with COVID-19 suggests this is a direct, rather than secondary, effect of infection. Here, by expressing individual SARS-CoV-2 proteins in the Drosophila heart, we demonstrate interaction of virus Nsp6 with host proteins of the MGA/MAX complex (MGA, PCGF6 and TFDP1). Complementing transcriptomic data from the fly heart reveal that this interaction blocks the antagonistic MGA/MAX complex, which shifts the balance towards MYC/MAX and activates glycolysis-with similar findings in mouse cardiomyocytes. Further, the Nsp6-induced glycolysis disrupts cardiac mitochondrial function, known to increase reactive oxygen species (ROS) in heart failure; this could explain COVID-19-associated cardiac pathology. Inhibiting the glycolysis pathway by 2-deoxy-D-glucose (2DG) treatment attenuates the Nsp6-induced cardiac phenotype in flies and mice. These findings point to glycolysis as a potential pharmacological target for treating COVID-19-associated heart failure.

Investigating and Resolving Cardiotoxicity Induced by COVID-19 Treatments using Human Pluripotent Stem Cell-Derived Cardiomyocytes and Engineered Heart Tissues.

Coronavirus disease 2019 continues to spread worldwide. Given the urgent need for effective treatments, many clinical trials are ongoing through repurposing approved drugs. However, clinical data regarding the cardiotoxicity of these drugs are limited. Human pluripotent stem cell-derived cardiomyocytes (hCMs) represent a powerful tool for assessing drug-induced cardiotoxicity. Here, by using hCMs, it is demonstrated that four antiviral drugs, namely, apilimod, remdesivir, ritonavir, and lopinavir, exhibit cardiotoxicity in terms of inducing cell death, sarcomere disarray, and dysregulation of calcium handling and contraction, at clinically relevant concentrations. Human engineered heart tissue (hEHT) model is used to further evaluate the cardiotoxic effects of these drugs and it is found that they weaken hEHT contractile function. RNA-seq analysis reveals that the expression of genes that regulate cardiomyocyte function, such as sarcomere organization (TNNT2, MYH6) and ion homeostasis (ATP2A2, HCN4), is significantly altered after drug treatments. Using high-throughput screening of approved drugs, it is found that ceftiofur hydrochloride, astaxanthin, and quetiapine fumarate can ameliorate the cardiotoxicity of remdesivir, with astaxanthin being the most prominent one. These results warrant caution and careful monitoring when prescribing these therapies in patients and provide drug candidates to limit remdesivir-induced cardiotoxicity.

Elucidating the role of the L-type calcium channel in excitability and energetics in the heart: The ISHR 2020 Research Achievement Award Lecture.

Cardiovascular disease continues to be the leading health burden worldwide and with the rising rates in obesity and type II diabetes and ongoing effects of long COVID, it is anticipated that the burden of cardiovascular morbidity and mortality will increase. Calcium is essential to cardiac excitation and contraction. The main route for Ca2+ influx is the L-type Ca2+ channel (Cav1.2) and embryos that are homozygous null for the Cav1.2 gene are lethal at day 14 postcoitum. Acute changes in Ca2+ influx through the channel contribute to arrhythmia and sudden death, and chronic increases in intracellular Ca2+ contribute to pathological hypertrophy and heart failure. We use a multidisciplinary approach to study the regulation of the channel from the molecular level through to in vivo CRISPR mutant animal models. Here we describe some examples of our work from over 2 decades studying the role of the channel under physiological and pathological conditions. Our single channel analysis of purified human Cav1.2 protein in proteoliposomes has contributed to understanding direct molecular regulation of the channel including identifying the critical serine involved in the "fight or flight" response. Using the same approach we identified the cysteine responsible for altered function during oxidative stress. Chronic activation of the L-type Ca2+ channel during oxidative stress occurs as a result of persistent glutathionylation of the channel that contributes to the development of hypertrophy. We describe for the first time that activation of the channel alters mitochondrial function (and energetics) on a beat-to-beat basis via movement of cytoskeletal proteins. In translational studies we have used this response to "report" mitochondrial function in models of cardiomyopathy and to test efficacy of novel therapies to prevent cardiomyopathy.

Development and diseases of the coronary microvasculature and its communication with the myocardium.

We review the current understanding of formation and development of the coronary microvasculature which supplies oxygen and nutrients to the heart myocardium and removes waste. We emphasize the close relationship, mutual development, and communication between microvasculature endothelial cells and surrounding cardiomyocytes. The first part of the review is focused on formation of microvasculature during embryonic development. We summarize knowledge about establishing the heart microvasculature density based on diffusion distance. Then signaling mechanisms which are involved in forming the microvasculature are discussed. This includes details of cardiomyocyte-endothelial cell interactions involving hypoxia, VEGF, NOTCH, angiopoietin, PDGF, and other signaling factors. The microvasculature is understudied due to difficulties in its visualization. Therefore, currently available imaging methods to delineate the coronary microvasculature in development and in adults are discussed. The second part of the review is dedicated to the importance of the coronary vasculature in disease. Coronary microvasculature pathologies are present in many congenital heart diseases (CHD), especially in pulmonary atresia, and worsen outcomes. In CHDs, where the development of the myocardium is impaired, microvasculature is also affected. In adult patients coronary microvascular disease is one of the main causes of sudden cardiac death, especially in women. Coronary microvasculature pathologies affect myocardial ischemia and vice versa; myocardial pathologies such as cardiomyopathies are closely connected with coronary microvasculature dysfunction. Microvasculature inflammation also worsens the outcomes of COVID-19 disease. Our review stresses the importance of coronary microvasculature and provides an overview of its formation and signaling mechanisms and the importance of coronary vasculature pathologies in CHDs and adult diseases. This article is categorized under: Cardiovascular Diseases > Stem Cells and Development Congenital Diseases > Molecular and Cellular Physiology Cardiovascular Diseases > Molecular and Cellular Physiology.

Sars-Cov-2 Spike Protein-Induced Damage of hiPSC-Derived Cardiomyocytes.

Sars-Cov-2 may trigger molecular and functional alterations of cardiomyocytes (CMs) of the heart due to the presence of receptor angiotensin-converting enzyme 2 (ACE2) of the host cells. While the endocytic itinerary of the virus via cleavage of the spike protein of Sars-Cov-2 is well understood, the role of the remaining part of the spike protein subunit and ACE2 complex is still elusive. Herein, the possible effects of this complex are investigated by using synthetic spike proteins of Sars-Cov-2, human-induced pluripotent stem cells (hiPSC), and a culture device made of an arrayed monolayer of cross-linked nanofibers. hiPSCs are first differentiated into CMs that form cardiac tissue-like constructs with regular beating and expression of both ACE2 and gap junction protein Connexin 43. When incubated with the spike proteins, the hiPSC-CMs undergo a rhythmic fluctuation with overstretched sarcomere structures and dispersed gap junction proteins. When incubated with the spike proteins and supplementary angiotensin II, the damage of the spike protein on hiPSC-CMs is enhanced due to downregulated ACE2, chromatin margination, altered Connexin 43 expression, sarcomere disruption, and beating break. This discovery may imply latent effects of the spike proteins on the heart.

SARS-CoV-2 Infection Induces Ferroptosis of Sinoatrial Node Pacemaker Cells.

BACKGROUND: Increasing evidence suggests that cardiac arrhythmias are frequent clinical features of coronavirus disease 2019 (COVID-19). Sinus node damage may lead to bradycardia. However, it is challenging to explore human sinoatrial node (SAN) pathophysiology due to difficulty in isolating and culturing human SAN cells. Embryonic stem cells (ESCs) can be a source to derive human SAN-like pacemaker cells for disease modeling. METHODS: We used both a hamster model and human ESC (hESC)-derived SAN-like pacemaker cells to explore the impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection on the pacemaker cells of the heart. In the hamster model, quantitative real-time polymerase chain reaction and immunostaining were used to detect viral RNA and protein, respectively. We then created a dual knock-in SHOX2:GFP;MYH6:mCherry hESC reporter line to establish a highly efficient strategy to derive functional human SAN-like pacemaker cells, which was further characterized by single-cell RNA sequencing. Following exposure to SARS-CoV-2, quantitative real-time polymerase chain reaction, immunostaining, and RNA sequencing were used to confirm infection and determine the host response of hESC-SAN-like pacemaker cells. Finally, a high content chemical screen was performed to identify drugs that can inhibit SARS-CoV-2 infection, and block SARS-CoV-2-induced ferroptosis. RESULTS: Viral RNA and spike protein were detected in SAN cells in the hearts of infected hamsters. We established an efficient strategy to derive from hESCs functional human SAN-like pacemaker cells, which express pacemaker markers and display SAN-like action potentials. Furthermore, SARS-CoV-2 infection causes dysfunction of human SAN-like pacemaker cells and induces ferroptosis. Two drug candidates, deferoxamine and imatinib, were identified from the high content screen, able to block SARS-CoV-2 infection and infection-associated ferroptosis. CONCLUSIONS: Using a hamster model, we showed that primary pacemaker cells in the heart can be infected by SARS-CoV-2. Infection of hESC-derived functional SAN-like pacemaker cells demonstrates ferroptosis as a potential mechanism for causing cardiac arrhythmias in patients with COVID-19. Finally, we identified candidate drugs that can protect the SAN cells from SARS-CoV-2 infection.

Genome-wide analyses reveal the detrimental impacts of SARS-CoV-2 viral gene Orf9c on human pluripotent stem cell-derived cardiomyocytes.

Patients with coronavirus disease 2019 (COVID-19) commonly have manifestations of heart disease. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genome encodes 27 proteins. Currently, SARS-CoV-2 gene-induced abnormalities of human heart muscle cells remain elusive. Here, we comprehensively characterized the detrimental effects of a SARS-CoV-2 gene, Orf9c, on human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) by preforming multi-omic analyses. Transcriptomic analyses of hPSC-CMs infected by SARS-CoV-2 with Orf9c overexpression (Orf9cOE) identified concordantly up-regulated genes enriched into stress-related apoptosis and inflammation signaling pathways, and down-regulated CM functional genes. Proteomic analysis revealed enhanced expressions of apoptotic factors, whereas reduced protein factors for ATP synthesis by Orf9cOE. Orf9cOE significantly reduced cellular ATP level, induced apoptosis, and caused electrical dysfunctions of hPSC-CMs. Finally, drugs approved by the U.S. Food and Drug Administration, namely, ivermectin and meclizine, restored ATP levels and ameliorated CM death and functional abnormalities of Orf9cOE hPSC-CMs. Overall, we defined the molecular mechanisms underlying the detrimental impacts of Orf9c on hPSC-CMs and explored potentially therapeutic approaches to ameliorate Orf9c-induced cardiac injury and abnormalities.

Overexpression of the SARS-CoV-2 receptor angiotensin converting enzyme 2 in cardiomyocytes of failing hearts.

Hospitalized patients who die from Covid-19 often have pre-existing heart disease. The SARS-CoV-2 virus is dependent on the ACE2 receptor to be able to infect cells. It is possible that the strong link between cardiovascular comorbidities and a poor outcome following a SARS-CoV-2 infection is sometimes due to viral myocarditis. The aim was to examine the expression of ACE2 in normal hearts and hearts from patients with terminal heart failure. The ACE2 expression was measured by global quantitative proteomics and RT-qPCR in left ventricular (LV) tissue from explanted hearts. Immunohistochemistry was used to examine ACE2 expression in cardiomyocytes, fibroblasts and endothelial cells. In total, tissue from 14 organ donors and 11 patients with terminal heart failure were included. ACE2 expression was 2.6 times higher in 4 hearts from patients with terminal heart failure compared with 6 healthy donor hearts. The results were confirmed by immunohistochemistry where more than half of cardiomyocytes or fibroblasts showed expression of ACE2 in hearts from patients with terminal heart failure. In healthy donor hearts ACE2 was not expressed or found in few fibroblasts. A small subpopulation of endothelial cells expressed ACE2 in both groups. Upregulated ACE2 expression in cardiomyocytes may increase the risk of SARS-CoV-2 myocarditis in patients with heart failure.

Interaction of SARS-CoV-2 with cardiomyocytes: Insight into the underlying molecular mechanisms of cardiac injury and pharmacotherapy.

SARS-CoV-2 causes respiratory illness with a spectrum of systemic complications. However, the mechanism for cardiac infection and cardiomyocyte injury in COVID-19 patients remains unclear. The current literature supports the notion that SARS-CoV-2 particles access the heart either by the circulating blood cells or by extracellular vesicles, originating from the inflamed lungs, and encapsulating the virus along with its receptor (ACE2). Both cardiomyocytes and pericytes (coronary arteries) express the necessary accessory proteins for access of SARS-CoV-2 particles (i.e. ACE2, NRP-1, TMPRSS2, CD147, integrin α5ß1, and CTSB/L). These proteins facilitate the SARS-CoV-2 interaction and entry into the pericytes and cardiomyocytes thus leading to cardiac manifestations. Subsequently, various signaling pathways are altered in the infected cardiomyocytes (i.e. increased ROS production, reduced contraction, impaired calcium homeostasis), causing cardiac dysfunction. The currently adopted pharmacotherapy in severe COVID-19 subjects exhibited side effects on the heart, often manifested by electrical abnormalities. Nonetheless, cardiovascular adverse repercussions have been associated with the advent of some of the SARS-CoV-2 vaccines with no clear mechanisms underlining these complications. We provide herein an overview of the pathways involved with cardiomyocyte in COVID-19 subjects to help promoting pharmacotherapies that can protect against SARS-CoV-2-induced cardiac injuries.

Remdesivir induces persistent mitochondrial and structural damage in human induced pluripotent stem cell-derived cardiomyocytes.

AIMS: Remdesivir is a prodrug of an adenosine triphosphate analogue and is currently the only drug formally approved for the treatment of hospitalized coronavirus disease of 2019 (COVID-19) patients. Nucleoside/nucleotide analogues have been shown to induce mitochondrial damage and cardiotoxicity, and this may be exacerbated by hypoxia, which frequently occurs in severe COVID-19 patients. Although there have been few reports of adverse cardiovascular events associated with remdesivir, clinical data are limited. Here, we investigated whether remdesivir induced cardiotoxicity using an in vitro human cardiac model. METHODS AND RESULTS: Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were exposed to remdesivir under normoxic and hypoxic conditions to simulate mild and severe COVID-19, respectively. Remdesivir induced mitochondrial fragmentation, reduced redox potential, and suppressed mitochondrial respiration at levels below the estimated plasma concentration under both normoxic and hypoxic conditions. Non-mitochondrial damage such as electrophysiological alterations and sarcomere disarray were also observed. Importantly, some of these changes persisted after the cessation of treatment, culminating in increased cell death. Mechanistically, we found that inhibition of DRP1, a regulator of mitochondrial fission, ameliorated the cardiotoxic effects of remdesivir, showing that remdesivir-induced cardiotoxicity was preventable and excessive mitochondrial fission might contribute to this phenotype. CONCLUSIONS: Using an in vitro model, we demonstrated that remdesivir can induce cardiotoxicity in hiPSC-CMs at clinically relevant concentrations. These results reveal previously unknown potential side-effects of remdesivir and highlight the importance of further investigations with in vivo animal models and active clinical monitoring to prevent lasting cardiac damage to patients.

Tumor Necrosis Factor-Alpha Exacerbates Viral Entry in SARS-CoV2-Infected iPSC-Derived Cardiomyocytes.

The coronavirus disease 2019 (COVID-19) pandemic with high infectivity and mortality has caused severe social and economic impacts worldwide. Growing reports of COVID-19 patients with multi-organ damage indicated that severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) may also disturb the cardiovascular system. Herein, we used human induced pluripotent stem cell (iPSC)-derived cardiomyocytes (iCMs) as the in vitro platform to examine the consequence of SARS-CoV2 infection on iCMs. Differentiated iCMs expressed the primary SARS-CoV2 receptor angiotensin-converting enzyme-II (ACE2) and the transmembrane protease serine type 2 (TMPRSS2) receptor suggesting the susceptibility of iCMs to SARS-CoV2. Following the infection of iCMs with SARS-CoV2, the viral nucleocapsid (N) protein was detected in the host cells, demonstrating the successful infection. Bioinformatics analysis revealed that the SARS-CoV2 infection upregulates several inflammation-related genes, including the proinflammatory cytokine tumor necrosis factor-α (TNF-α). The pretreatment of iCMs with TNF-α for 24 h, significantly increased the expression of ACE2 and TMPRSS2, SASR-CoV2 entry receptors. The TNF-α pretreatment enhanced the entry of GFP-expressing SARS-CoV2 pseudovirus into iCMs, and the neutralization of TNF-α ameliorated the TNF-α-enhanced viral entry. Collectively, SARS-CoV2 elevated TNF-α expression, which in turn enhanced the SARS-CoV2 viral entry. Our findings suggest that, TNF-α may participate in the cytokine storm and aggravate the myocardial damage in COVID-19 patients.

Antihypertensive drug treatment and susceptibility to SARS-CoV-2 infection in human PSC-derived cardiomyocytes and primary endothelial cells.

The pathogenicity of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has been attributed to its ability to enter through the membrane-bound angiotensin-converting enzyme 2 (ACE2) receptor. Therefore, it has been heavily speculated that angiotensin-converting enzyme inhibitor (ACEI) or angiotensin receptor blocker (ARB) therapy may modulate SARS-CoV-2 infection. In this study, exposure of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) and human endothelial cells (hECs) to SARS-CoV-2 identified significant differences in protein coding genes involved in immunity, viral response, and cardiomyocyte/endothelial structure. Specifically, transcriptome changes were identified in the tumor necrosis factor (TNF), interferon α/ß, and mitogen-activated protein kinase (MAPK) (hPSC-CMs) as well as nuclear factor kappa-B (NF-κB) (hECs) signaling pathways. However, pre-treatment of hPSC-CMs or hECs with two widely prescribed antihypertensive medications, losartan and lisinopril, did not affect the susceptibility of either cell type to SARS-CoV-2 infection. These findings demonstrate the toxic effects of SARS-CoV-2 in hPSC-CMs/hECs and, taken together with newly emerging multicenter trials, suggest that antihypertensive drug treatment alone does not alter SARS-CoV-2 infection.

COVID19-associated cardiomyocyte dysfunction, arrhythmias and the effect of Canakinumab.

BACKGROUND: Cardiac injury associated with cytokine release frequently occurs in SARS-CoV-2 mediated coronavirus disease (COVID19) and mortality is particularly high in these patients. The mechanistic role of the COVID19 associated cytokine-storm for the concomitant cardiac dysfunction and associated arrhythmias is unclear. Moreover, the role of anti-inflammatory therapy to mitigate cardiac dysfunction remains elusive. AIMS AND METHODS: We investigated the effects of COVID19-associated inflammatory response on cardiac cellular function as well as its cardiac arrhythmogenic potential in rat and induced pluripotent stem cell derived cardiomyocytes (iPS-CM). In addition, we evaluated the therapeutic potential of the IL-1ß antagonist Canakinumab using state of the art in-vitro confocal and ratiometric high-throughput microscopy. RESULTS: Isolated rat ventricular cardiomyocytes were exposed to control or COVID19 serum from intensive care unit (ICU) patients with severe ARDS and impaired cardiac function (LVEF 41±5%; 1/3 of patients on veno-venous extracorporeal membrane oxygenation; CK 154±43 U/l). Rat cardiomyocytes showed an early increase of myofilament sensitivity, a decrease of Ca2+ transient amplitudes and altered baseline [Ca2+] upon exposure to patient serum. In addition, we used iPS-CM to explore the long-term effect of patient serum on cardiac electrical and mechanical function. In iPS-CM, spontaneous Ca2+ release events were more likely to occur upon incubation with COVID19 serum and nuclear as well as cytosolic Ca2+ release were altered. Co-incubation with Canakinumab had no effect on pro-arrhythmogenic Ca2+ release or Ca2+ signaling during excitation-contraction coupling, nor significantly influenced cellular automaticity. CONCLUSION: Serum derived from COVID19 patients exerts acute cardio-depressant and chronic pro-arrhythmogenic effects in rat and iPS-derived cardiomyocytes. Canakinumab had no beneficial effect on cellular Ca2+ signaling during excitation-contraction coupling. The presented method utilizing iPS-CM and in-vitro Ca2+ imaging might serve as a novel tool for precision medicine. It allows to investigate cytokine related cardiac dysfunction and pharmacological approaches useful therein.

Cardiomyocytes recruit monocytes upon SARS-CoV-2 infection by secreting CCL2.

Heart injury has been reported in up to 20% of COVID-19 patients, yet the cause of myocardial histopathology remains unknown. Here, using an established in vivo hamster model, we demonstrate that SARS-CoV-2 can be detected in cardiomyocytes of infected animals. Furthermore, we found damaged cardiomyocytes in hamsters and COVID-19 autopsy samples. To explore the mechanism, we show that both human pluripotent stem cell-derived cardiomyocytes (hPSC-derived CMs) and adult cardiomyocytes (CMs) can be productively infected by SARS-CoV-2, leading to secretion of the monocyte chemoattractant cytokine CCL2 and subsequent monocyte recruitment. Increased CCL2 expression and monocyte infiltration was also observed in the hearts of infected hamsters. Although infected CMs suffer damage, we find that the presence of macrophages significantly reduces SARS-CoV-2-infected CMs. Overall, our study provides direct evidence that SARS-CoV-2 infects CMs in vivo and suggests a mechanism of immune cell infiltration and histopathology in heart tissues of COVID-19 patients.