The epicardium plays an essential role in cardiogenesis by providing cardiac cell types and paracrine cues to the developing myocardium. The human adult epicardium is quiescent, but recapitulation... Show moreThe epicardium plays an essential role in cardiogenesis by providing cardiac cell types and paracrine cues to the developing myocardium. The human adult epicardium is quiescent, but recapitulation of developmental features may contribute to adult cardiac repair. The cell fate of epicardial cells is proposed to be determined by the developmental persistence of specific subpopulations. Reports on this epicardial heterogeneity have been inconsistent, and data regarding the human developing epicardium are scarce. Here we specifically isolated human fetal epicardium and used single-cell RNA sequencing to define its composition and to identify regulators of developmental processes. Few specific subpopulations were observed, but a clear distinction between epithelial and mesenchymal cells was present, resulting in novel population-specific markers. Additionally, we identified CRIP1 as a previously unknown regulator involved in epicardial epithelial-to-mesenchymal transition. Overall, our human fetal epicardial cell-enriched dataset provides an excellent platform to study the developing epicardium in great detail. Show less
Dronkers, E.; Herwaarden, T. van; Brakel, T.J. van; Sanchez Duffhues, G.; Goumans, M.J.; Smits, A.M. 2021
The epicardium, the mesothelial layer covering the heart, is a crucial cell source for cardiac development and repair. It provides cells and biochemical signals to the heart to facilitate... Show moreThe epicardium, the mesothelial layer covering the heart, is a crucial cell source for cardiac development and repair. It provides cells and biochemical signals to the heart to facilitate vascularization and myocardial growth. An essential element of epicardial behavior is epicardial epithelial to mesenchymal transition (epiMT), which is the initial step for epicardial cells to become motile and invade the myocardium. To identify targets to optimize epicardium-driven repair of the heart, it is vital to understand which pathways are involved in the regulation of epiMT. Therefore, we established a cell culture model for human primary adult and fetal epiMT, which allows for parallel testing of inhibitors and stimulants of specific pathways. Using this approach, we reveal Activin A and ALK4 signaling as novel regulators of epiMT, independent of the commonly accepted EMT inducer TGF beta. Importantly, Activin A was able to induce epicardial invasion in cultured embryonic mouse hearts. Our results identify Activin A/ALK4 signaling as a modulator of epicardial plasticity which may be exploitable in cardiac regenerative medicine. Show less
Patients with a congenital bicuspid aortic valve (BAV), a valve with two instead of three aortic leaflets, have an increased risk of developing thoracic aneurysms and aortic dissection. The... Show morePatients with a congenital bicuspid aortic valve (BAV), a valve with two instead of three aortic leaflets, have an increased risk of developing thoracic aneurysms and aortic dissection. The mechanisms underlying BAV-associated aortopathy are poorly understood. This study examined BAV-associated aortopathy in Nos3(-/-) mice, a model with congenital BAV formation. A combination of histological examination and in vivo ultrasound imaging was used to investigate aortic dilation and dissections in Nos3(-/-) mice. Moreover, cell lineage analysis and single-cell RNA sequencing were used to observe the molecular anomalies within vascular smooth muscle cells (VSMCs) of Nos3(-/-) mice. Spontaneous aortic dissections were found in ascending aortas located at the sinotubular junction in similar to 13% of Nos3(-/-) mice. Moreover, Nos3(-/-) mice were prone to developing aortic dilations in the proximal and distal ascending aorta during early adulthood. Lower volumes of elastic fibres were found within vessel walls of the ascending aortas of Nos3(-/-) mice, as well as incomplete coverage of the aortic inner media by neural crest cell (NCC)-derived VSMCs. VSMCs of Nos3(-/-) mice showed downregulation of 15 genes, of which seven were associated with aortic aneurysms and dissections in the human population. Elastin mRNA was most markedly downregulated, followed by fibulin-5 expression, both primary components of elastic fibres. This study demonstrates that, in addition to congenital BAV formation, disrupted endothelial-mediated nitric oxide (NO) signalling in Nos3(-/-) mice also causes aortic dilation and dissection, as a consequence of inhibited elastic fibre formation in VSMCs within the ascending aorta. Show less
Qiao, X.H.; Zanden, S.Y. van der; Wander, D.P.A.; Borras, D.M.; Song, J.Y.; Li, X.Y.; ... ; Neefjes, J. 2020
The anthracycline doxorubicin (Doxo) and its analogs daunorubicin (Daun), epirubicin (Epi), and idarubicin (Ida) have been cornerstones of anticancer therapy for nearly five decades. However, their... Show moreThe anthracycline doxorubicin (Doxo) and its analogs daunorubicin (Daun), epirubicin (Epi), and idarubicin (Ida) have been cornerstones of anticancer therapy for nearly five decades. However, their clinical application is limited by severe side effects, especially dose-dependent irreversible cardiotoxicity. Other detrimental side effects of anthracyclines include therapy-related malignancies and infertility. It is unclear whether these side effects are coupled to the chemotherapeutic efficacy. Doxo, Daun, Epi, and Ida execute two cellular activities: DNA damage, causing double-strand breaks (DSBs) following poisoning of topoisomerase II (Topo II), and chromatin damage, mediated through histone eviction at selected sites in the genome. Here we report that anthracycline-induced cardiotoxicity requires the combination of both cellular activities. Topo II poisons with either one of the activities fail to induce cardiotoxicity in mice and human cardiac microtissues, as observed for aclarubicin (Acla) and etoposide (Etop). Further, we show that Doxo can be detoxified by chemically separating these two activities. Anthracycline variants that induce chromatin damage without causing DSBs maintain similar anticancer potency in cell lines, mice, and human acute myeloid leukemia patients, implying that chromatin damage constitutes a major cytotoxic mechanismof anthracyclines. With these anthracyclines abstained from cardiotoxicity and therapy-related tumors, we thus uncoupled the side effects from anticancer efficacy. These results suggest that anthracycline variants acting primarily via chromatin damage may allow prolonged treatment of cancer patients and will improve the quality of life of cancer survivors. Show less
The anthracycline doxorubicin (Doxo) and its analogs daunorubicin (Daun), epirubicin (Epi), and idarubicin (Ida) have been cornerstones of anticancer therapy for nearly five decades. However, their... Show moreThe anthracycline doxorubicin (Doxo) and its analogs daunorubicin (Daun), epirubicin (Epi), and idarubicin (Ida) have been cornerstones of anticancer therapy for nearly five decades. However, their clinical application is limited by severe side effects, especially dose-dependent irreversible cardiotoxicity. Other detrimental side effects of anthracyclines include therapy-related malignancies and infertility. It is unclear whether these side effects are coupled to the chemotherapeutic efficacy. Doxo, Daun, Epi, and Ida execute two cellular activities: DNA damage, causing double-strand breaks (DSBs) following poisoning of topoisomerase II (Topo II), and chromatin damage, mediated through histone eviction at selected sites in the genome. Here we report that anthracycline-induced cardiotoxicity requires the combination of both cellular activities. Topo II poisons with either one of the activities fail to induce cardiotoxicity in mice and human cardiac microtissues, as observed for aclarubicin (Acla) and etoposide (Etop). Further, we show that Doxo can be detoxified by chemically separating these two activities. Anthracycline variants that induce chromatin damage without causing DSBs maintain similar anticancer potency in cell lines, mice, and human acute myeloid leukemia patients, implying that chromatin damage constitutes a major cytotoxic mechanism of anthracyclines. With these anthracyclines abstained from cardiotoxicity and therapy-related tumors, we thus uncoupled the side effects from anticancer efficacy. These results suggest that anthracycline variants acting primarily via chromatin damage may allow prolonged treatment of cancer patients and will improve the quality of life of cancer survivors. Show less
Balbi, C.; Lodder, K.; Costa, A.; Moimas, S.; Moccia, F.; Herwaarden, T. van; ... ; Bollini, S. 2019
The data and information presented here refer to the research article entitled: "Reactivating endogenous mechanisms of cardiac regeneration via paracrine boosting with the human amniotic fluid stem... Show moreThe data and information presented here refer to the research article entitled: "Reactivating endogenous mechanisms of cardiac regeneration via paracrine boosting with the human amniotic fluid stem cell secretome" (Balbi et al., 2019, Apr 04). This dataset illustrates the in vitro paracrine effect exerted by the human amniotic fluid stem cell secretome on rodent neonatal cardiomyocytes, human endothelial progenitors and different subsets of cardiac progenitor cells. Cytokine/chemokine profiling of the human amniotic fluid stem cell secretome is provided as well. This data can provide useful insights in regenerative medicine as demonstrating the in vitro cardioprotective and proliferative secretory paracrine potential of human fetal stem cells. (c) 2019 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). Show less
Balbi, C.; Lodder, K.; Costa, A.; Moimas, S.; Moccia, F.; Herwaarden, T. van; ... ; Bollini, S. 2019
Background: The adult mammalian heart retains residual regenerative capability via endogenous cardiac progenitor cell (CPC) activation and cardiomyocyte proliferation. We previously reported the... Show moreBackground: The adult mammalian heart retains residual regenerative capability via endogenous cardiac progenitor cell (CPC) activation and cardiomyocyte proliferation. We previously reported the paracrine cardioprotective capacity of human amniotic fluid-derived stem cells (hAFS) following ischemia or cardiotoxicity. Here we analyse the potential of hAFS secretome fractions for cardiac regeneration and future clinical translation.Methods: hAFS were isolated from amniotic fluid leftover samples from prenatal screening. hAFS conditioned medium (hAFS-CM) was obtained following hypoxic preconditioning. Anti-apoptotic, angiogenic and proliferative effects were evaluated on rodent neonatal cardiomyocytes (r/mNVCM), human endothelial colony forming cells (hECFC) and human CPC. Mice undergoing myocardial infarction (MI) were treated with hAFS-CM, hAFS-extracellular vesicles (hAFS-EV), or EV-depleted hAFS-CM(hAFS-DM) by single intra-myocardial administration and evaluated in the short and long term.Results: hAFS-CM improved mNVCM survival under oxidative and hypoxic damage, induced Ca2+-dependent angiogenesis in hECFC and triggered hCPC and rNVCM proliferation. hAFS-CM treatment after MI counteracted scarring, supported cardiac function, angiogenesis and cardiomyocyte cell cycle progression in the long term. hAFS-DM had no effect. hAFS-CM and hAFS-EV equally induced epicardium WT1+CPC reactivation. Although no CPC cardiovascular differentiation was observed, our data suggests contribution to local angiogenesis by paracrine modulation. hAFS-EV alone were able to recapitulate all the beneficial effects exerted by hAFS-CM, except for stimulation of vessel formation.Conclusions: hAFS-CM and hAFS-EV can improve cardiac repair and trigger cardiac regeneration via paracrine modulation of endogenous mechanisms. While both formulations are effective in sustaining myocardial renewal, hAFS-CM retains higher pro-angiogenic potential, while hAFS-EV particularly enhances cardiac function. (c) 2019 Elsevier B.V. All rights reserved. Show less
