Figure 3
![Immunostaining for cardiomyocyte markers in porcine and human cardiac fibroblasts. A, Representative immunofluorescence staining for DAPI ([blue] nuclear marker), GFP ([green] lentivirus infection marker) and the (red) cardiomyocyte markers cTnT (top) and a-actinin (bottom) in porcine cardiac fibroblasts transduced with GMT, GMT plus miR-590 or GMT plus H/My. GMT plus miR-590 and GMT plus H/My induced abundant cTnT and a-actinin expression 4 weeks after transduction (n=3). B, Immunofluorescence staining for DAPI ([blue] nuclear marker), GFP ([green] lentivirus infection marker) and the (red) cardiomyocyte markers cTnT (top) and a-actinin (bottom) in human cardiac fibroblasts transduced with GMT, GMT plus miR-590 or GMT plus H/My. GMT plus miR-590 and GMT plus H/My induced abundant cTnT and a](profile/Deepthi_Sanagasetti/publication/310389739/figure/fig2/AS:544079769083904@1506730133794/Immunostaining-for-cardiomyocyte-markers-in-porcine-and-human-cardiac-fibroblasts-A.png)
Immunostaining for cardiomyocyte markers in porcine and human cardiac fibroblasts. A, Representative immunofluorescence staining for DAPI ([blue] nuclear marker), GFP ([green] lentivirus infection marker) and the (red) cardiomyocyte markers cTnT (top) and a-actinin (bottom) in porcine cardiac fibroblasts transduced with GMT, GMT plus miR-590 or GMT plus H/My. GMT plus miR-590 and GMT plus H/My induced abundant cTnT and a-actinin expression 4 weeks after transduction (n=3). B, Immunofluorescence staining for DAPI ([blue] nuclear marker), GFP ([green] lentivirus infection marker) and the (red) cardiomyocyte markers cTnT (top) and a-actinin (bottom) in human cardiac fibroblasts transduced with GMT, GMT plus miR-590 or GMT plus H/My. GMT plus miR-590 and GMT plus H/My induced abundant cTnT and a
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Background
Reprogramming of cardiac fibroblasts into induced cardiomyocyte‐like cells represents a promising potential new therapy for treating heart disease, inducing significant improvements in postinfarct ventricular function in rodent models. Because reprogramming factors effective in transdifferentiating rodent cells are not sufficient to repr...
Contexts in source publication
Context 1
... studies likewise demonstrated that GMT plus miR-590 induced expression of the cardiomyocyte marker, a-sarcomeric actinin, as well as cTnT expression in porcine and human cardiac fibroblasts to an extent similar to that observed after administration of GMT plus H/My (Figure 3). ...
Context 2
... order to better discern the mechanism of action of miR- 590, we used an miRNA target prediction program (Tar- getscan 7.0), which identified Sp1 as a potential target of miR- 590 ( Figure S3A). Confirming these data, luciferase reporter assays showed the direct binding of miR-590 to the 3 0 -UTR of Sp1 ( Figure S3B). ...
Context 3
... order to better discern the mechanism of action of miR- 590, we used an miRNA target prediction program (Tar- getscan 7.0), which identified Sp1 as a potential target of miR- 590 ( Figure S3A). Confirming these data, luciferase reporter assays showed the direct binding of miR-590 to the 3 0 -UTR of Sp1 ( Figure S3B). ...
Citations
... Singh et al. demonstrated that lentiviral delivery of GHMT TFs coupled with Myocardin or miR-590 produced iCMs from rat, porcine, and human CFs in vitro. miR-590 promoted suppression of Sp1, a zinc-finger protein, which upregulated numerous genes linked with CM phenotype and downregulated various fibroblast-related genes [66]. Years later, Zhou et al. reported that GMT combined with miR−133 (GMT133) has the ability to convert human CFs into CMs [57]. ...
Cardiac diseases are the foremost cause of morbidity and mortality worldwide. The heart has limited regenerative potential; therefore, lost cardiac tissue cannot be replenished after cardiac injury. Conventional therapies are unable to restore functional cardiac tissue. In recent decades, much attention has been paid to regenerative medicine to overcome this issue. Direct reprogramming is a promising therapeutic approach in regenerative cardiac medicine that has the potential to provide in situ cardiac regeneration. It consists of direct cell fate conversion of one cell type into another, avoiding transition through an intermediary pluripotent state. In injured cardiac tissue, this strategy directs transdifferentiation of resident non-myocyte cells (NMCs) into mature functional cardiac cells that help to restore the native tissue. Over the years, developments in reprogramming methods have suggested that regulation of several intrinsic factors in NMCs can help to achieve in situ direct cardiac reprogramming. Among NMCs, endogenous cardiac fibroblasts have been studied for their potential to be directly reprogrammed into both induced cardiomyocytes and induced cardiac progenitor cells, while pericytes can transdifferentiate towards endothelial cells and smooth muscle cells. This strategy has been indicated to improve heart function and reduce fibrosis after cardiac injury in preclinical models. This review summarizes the recent updates and progress in direct cardiac reprogramming of resident NMCs for in situ cardiac regeneration.
... MiR-133 enhances generation of functional iCMs through direct repression of Snail, a master regulator of epithelial-to-mesenchymal transition. In addition, miRNA-590 can replace Hand2 and Myocd during conversion of human and porcine fibroblasts to iCMs (Eulalio et al., 2012;Singh et al., 2016). ...
Limited regenerative capacity of adult cardiomyocytes precludes heart repair and regeneration after cardiac injury. Direct cardiac reprograming that converts scar-forming cardiac fibroblasts (CFs) into functional induced-cardiomyocytes (iCMs) offers promising potential to restore heart structure and heart function. Significant advances have been achieved in iCM reprogramming using genetic and epigenetic regulators, small molecules, and delivery strategies. Recent researches on the heterogeneity and reprogramming trajectories elucidated novel mechanisms of iCM reprogramming at single cell level. Here, we review recent progress in iCM reprogramming with a focus on multi-omics (transcriptomic, epigenomic and proteomic) researches to investigate the cellular and molecular machinery governing cell fate conversion. We also highlight the future potential using multi-omics approaches to dissect iCMs conversion for clinal applications.
... Direct cellular reprogramming represents a novel strategy whereby resident cardiac fibroblasts in areas of myocardial infarction or fibrosis can be transdifferentiated into functional cardiomyocyte-like cells (iCMs) that can in turn enhance myocardial contractile function 4-7 . We and others have shown that different combinations of cardio-differentiating factors can transdifferentiate rodent and human cardiac fibroblasts into iCMs [8][9][10][11][12][13][14] . Human cells nevertheless appear resistant to reprogramming compared to rodent cells, likely due to epigenetic downregulation of cell plasticity [15][16][17] . ...
... Adult human cardiac fibroblasts were isolated using standard isolation techniques from ventricular myocardial tissue obtained from explants of heart failure patients undergoing mechanical assist device placement or cardiac transplantation at Baylor St. Luke's Medical Center 9,10 . A written informed consent was obtained from all the subjects and/or their legal guardian(s) prior to obtaining the tissue. ...
... Cell reprogramming. Lentivirus vectors each encoding Gata4, Mef2, or Tbx5 (GMT), Hand2/Myocardin (H/M), non-targeting (NT) shRNA, p63 short hairpin RNA (Origene, Rockville, MD), p63-transactivation inhibitory domain (Vectorbuilder, Chicago, IL) tagged with green fluorescent protein (GFP) or GFP control vectors were prepared from relevant plasmids by the Baylor College Of Medicine Gene Vector Core, as previously described 9,10,27,28 . ...
Abstract Direct cell reprogramming represents a promising new myocardial regeneration strategy involving in situ transdifferentiation of cardiac fibroblasts into induced cardiomyocytes. Adult human cells are relatively resistant to reprogramming, however, likely because of epigenetic restraints on reprogramming gene activation. We hypothesized that modulation of the epigenetic regulator gene p63 could improve the efficiency of human cell cardio-differentiation. qRT-PCR analysis demonstrated significantly increased expression of a panel of cardiomyocyte marker genes in neonatal rat and adult rat and human cardiac fibroblasts treated with p63 shRNA (shp63) and the cardio-differentiation factors Hand2/Myocardin (H/M) versus treatment with Gata4, Mef2c and Tbx5 (GMT) with or without shp63 (p
... Around the same time, HAND2 was found to help GMTMeMyEZ to facilitate the early progress of reprogramming of HDFs and H9Fs (Bektik et al., 2017), and MiR-590 could promote reprogramming of HCFs (Singh et al., 2016). However, the reprogramming efficiency and maturity in HDFderived induced cardiomyocyte-like cells (iCMs) were poorer than those in HCF-derived iCMs (Fu et al., 2013;Nam et al., 2013). ...
... In this study, GMTMeMy (5F)-induced HDFs were cultured with modified reprogramming medium containing TGF-b inhibitor, WNT inhibitors, and Vitamin C, which were previously reported to enhance cardiac reprogramming (Mohamed et al., 2017;Talkhabi et al., 2015). Medium was replaced by maintenance medium 2 weeks after reprogramming (Fig. 1A). ...
Reprogramming of human dermal fibroblasts (HDFs) into induced cardiomyocyte-like cells (iCMs) represents a promising strategy for human cardiac regeneration. Different cocktails of cardiac transcription factors can convert HDFs into iCMs, although with low efficiency and immature phenotype. Here, GATA4, MEF2C, TBX5, MESP1, and MYOCD (GMTMeMy for short) were used to reprogram HDFs by retrovirus infection. We found that the exogenous expression stoichiometry of GATA4 (GATA4 stoichiometry) significantly affected reprogramming efficiency. When 1/8 dosage of GATA4 virus (GATA4 dosage) plus MTMeMy was used, the reprogramming efficiency was obviously improved compared with average pooled virus encoding each factor, which measured, by the expression level of cardiac genes, the percentage of cardiac troponin T and alpha-cardiac myosin heavy-chain immunopositive cells and the numbers of iCMs showing calcium oscillation or beating synchronously in co-culture with mouse CMs. In addition, we prepared conditioned maintenance medium (CMM) by CM differentiation of H9 human embryonic stem cell line. We found that compared with traditional maintenance medium (TMM), CMM made iCMs show well-organized sarcomere formation and characteristic calcium oscillation wave earlier. These findings demonstrated that appropriate GATA4 stoichiometry was essential for cardiac reprogramming and some components in CMM were important for maturation of iCMs.
... Another proposed strategy for remuscularization of the diseased heart is the reprogramming of cardiac fibroblasts into cardiomyocyte-like cells [33]. Successful transdifferentiation of porcine cardiac fibroblasts into cardiomyocyte-like cells has been demonstrated in vitro utilizing lentiviral vectors expressing cardio-reprogramming factors (Gata4, Mef2c, Tbx5) alongside miR-590 [34]. However, whether this strategy will provide postinfarct recovery in vivo in swine has not been tested. ...
Swine are popular large mammals for cardiac preclinical testing due to their similarities with humans in terms of organ size and physiology. Recent studies indicate an early neonatal regenerative capacity for swine hearts similar to small mammal laboratory models such as rodents, inspiring exciting possibilities for studying cardiac regeneration with the goal of improved clinical translation to humans. However, while swine hearts are anatomically similar to humans, fundamental differences exist in growth mechanisms, nucleation, and the maturation of pig cardiomyocytes, which could present difficulties for the translation of preclinical findings in swine to human therapeutics. In this review, we discuss the maturational dynamics of pig cardiomyocytes and their capacity for proliferative cardiac regeneration during early neonatal development to provide a perspective on swine as a preclinical model for developing cardiac gene- and cell-based regenerative therapeutics.
... GMT, in combination with miR-590, induced the expression of the cardiomyocyte marker cardiac troponin T (cTnT) in~5% of human or porcine CFs [111], and this efficiency was further increased, combining the dual inhibition of HDACs and WNT with retinoic acid administration alongside GMT plus hsa-miR-590 transduction [112]. The use of Sendai virus to actively deliver GMTMM + mmu-miR-133a to human CFs further demonstrated the potential efficacy of miRNA and TF combinations in inducing direct cardiac reprogramming [113], although it has been recently reported that the administration of a polycistronic vector ensuring miRcombo delivery at a stoichiometric ratio can be more effective compared to other viral methods in inducing direct cardiac reprogramming [114]. ...
... Twf1, Col16a1, and Ezh2 [102] mmu-miR-208b-3p + ascorbic acid/BMP4 This combination (MAB) induces transdifferentiation to iCMs. Gata4 [105] mmu-miR-133a + GMT/GMTMM cocktails Increases the efficiency of iCM generation Snai1 [107] mmu-miR-1/mmu-mmu-miR-133a + GHMT cocktail Increases the efficiency of iCM generation Twf1 and Snai1 [109] hsa-miR-590 + GMT cocktail Increases the maturation of iCMs Sp1 [111] 4.3. Challenges and Opportunities of miRNA-Mediated Cardiac Reprogramming as a Therapeutic Strategy to Treat Heart Failure Patients ...
Cell reprogramming is a groundbreaking technology that, in few decades, generated a new paradigm in biomedical science. To date we can use cell reprogramming to potentially generate every cell type by converting somatic cells and suitably modulating the expression of key transcription factors. This approach can be used to convert skin fibroblasts into pluripotent stem cells as well as into a variety of differentiated and medically relevant cell types, including cardiomyocytes and neural cells. The molecular mechanisms underlying such striking cell phenotypes are still largely unknown, but in the last decade it has been proven that cell reprogramming approaches are significantly influenced by non-coding RNAs. Specifically, this review will focus on the role of microRNAs in the reprogramming processes that lead to the generation of pluripotent stem cells, neurons, and cardiomyocytes. As highlighted here, non-coding RNA-forced expression can be sufficient to support some cell reprogramming processes, and, therefore, we will also discuss how these molecular determinants could be used in the future for biomedical purposes.
... Recently, mouse cardiac and dermal fibroblasts could be efficiently reprogrammed into pounding iCM in vitro or vivo by defined factors Gata4, Mef2c and Tbx5 (GMT) [13][14][15][16]. Addition of another factor like HAND2, MESP1, MYOCD, ESRRG and ZFPM2 into GMT cocktail could also convert human cardiac or dermal fibroblasts into iCM (F-iCM) which was proved to replenish mouse diseased heart after transplantation in vivo [17][18][19]. Despite more complex reprogramming factor cocktails, human iCM generation is achieved at lower efficiency and requires longer reprogramming time. ...
... Despite more complex reprogramming factor cocktails, human iCM generation is achieved at lower efficiency and requires longer reprogramming time. Furthermore, different from mouse, human iCMs by defined factors don't contract spontaneously alone, suggesting biological function of human iCM is incomplete or immature [18]. Therefore, protocols for human iCM generation need further improvement. ...
... We first optimized F-iCM generation to help develop an effective protocol for U-iCM generation. In consideration of important roles of GATA4, MEF2C, TBX5, MYOCD, MESP1, ZFPM2 and ESRRG in human cardiac direct reprogramming of fibroblasts [17,18,25], we first generated retroviruses to overexpress these seven core cardiac transcription factors and transduced them together into dermal fibroblasts BJ purchased from ATCC which is widely used for human iPSCs generation [32,33] but not applied to cardiac reprogramming up to now. We found that these seven factors upregulated cardiac specific marker genes MYH6 and TNNT2 at one week. ...
Despite direct reprogramming of human cardiac fibroblasts into induced cardiomyocytes (iCM) holds great potential for heart regeneration, the mechanisms are poorly understood. Whether other human somatic cells could be reprogrammed into cardiomyocytes is also unknown. Here, we report human urine cells (hUCs) could be converted into CM-like cells from different donors and the related chromatin accessibility dynamics (CAD) by assay for transposase accessible chromatin(ATAC)-seq. hUCs transduced by MEF2C, TBX5, MESP1 and MYOCD but without GATA4 expressed multiple cardiac specific genes, exhibited Ca²⁺ oscillation potential and sarcomeric structures, and contracted synchronously in coculture with mouse CM. Additionally, we found that MYOCD is required for both closing and opening critical loci, mainly by hindering the opening of loci enriched with motifs for the TEAD and AP1 family and promoting the closing of loci enriched with ETS motifs. These changes differ partially from CAD observed during iCM induction from human fibroblasts. Collectively, our study offers one practical platform for iCM generation and insights into mechanisms for iCM fate determination.
... 3,4 We and others have shown that a wide variety of cardiac reprogramming cocktails based upon the core cardiac transcription factors Gata4, Mef2c and Tbx5 (GMT) can convert fibroblasts into iCMs, albeit with low efficiency in inducing contractile cells in vitro, especially as applied to human cells. [5][6][7][8][9][10][11][12][13][14][15][16] The TEAD (transcriptional enhance associate domain) transcription factor family is a component of the Hippo signaling pathway, the inactivation of which has been implicated in cell replication, growth, and development. [17][18][19][20] The role of TEAD in the Hippo pathway is Singh et al Tead1 Enhances Cardiac Cellular Reprogramming broadly believed to be regulated by the presence or the absence of nuclear YAP (yes-associated protein) and TAZ (transcriptional activator binding domain). ...
... 8 Adult human cardiac fibroblasts were harvested from ventricular tissue obtained from heart failure pateints undergoing cardiac transplantation at Baylor St. Luke's Medical Center and under a protocol approved by the Baylor College of Medicine Institutional Review Board (IRB H-33421). 9 Human tissues were manually sharply miced and placed in fibroblast growth medium (Dulbecco modified eagle medium (DMEM), 10% fetal bovine serum (FBS), and 1% penicillin/streptomycin), as previously described. 10 Cell Reprogramming Lentivirus vectors encoding Gata4 (G), Mef2c (M), Tbx5 (T), Tead1 (Td), YAP, TAZ, PGC (peroxisome proliferatoractivated receptor-γ coactivator)-1A, PGC-1B and GFP (green fluorescent protein) were prepared from relevant plasmids by the Baylor College of Medicine Gene Vector Core, as previously described. ...
... 10 Cell Reprogramming Lentivirus vectors encoding Gata4 (G), Mef2c (M), Tbx5 (T), Tead1 (Td), YAP, TAZ, PGC (peroxisome proliferatoractivated receptor-γ coactivator)-1A, PGC-1B and GFP (green fluorescent protein) were prepared from relevant plasmids by the Baylor College of Medicine Gene Vector Core, as previously described. 8,9,15 A combination of 4 factors (GMTTd) was used for reprogramming rat and human cardiac fibroblasts reprogramming. Rat and human cardiac fibroblasts were seeded at a density of 5×10 4 cells per well of a 24-well plate in Iscove modified dulbecco media medium containing 20% FBS, and 1% penicillin/streptomycin. Lentivirus vectors with a multiplicity of infection: 20 was added with polybrene (5 μg/ μL) after 24 hours. ...
Background
The conversion of fibroblasts into induced cardiomyocytes may regenerate myocardial tissue from cardiac scar through in situ cell transdifferentiation. The efficiency transdifferentiation is low, especially for human cells. We explored the leveraging of Hippo pathway intermediates to enhance induced cardiomyocyte generation.
Methods and Results
We screened Hippo effectors Yap (yes‐associated protein), Taz (transcriptional activator binding domain), and Tead1 (TEA domain transcription factor 1; Td) for their reprogramming efficacy with cardio‐differentiating factors Gata4, Mef2C, and Tbx5 (GMT). Td induced nearly 3‐fold increased expression of cardiomyocyte marker cTnT (cardiac troponin T) by mouse embryonic and adult rat fibroblasts versus GMT administration alone ( P <0.0001), while Yap and Taz failed to enhance cTnT expression. Serial substitution demonstrated that Td replacement of TBX5 induced the greatest cTnT expression enhancement and sarcomere organization in rat fibroblasts treated with all GMT substitutions (GMTd versus GMT: 17±1.2% versus 5.4±0.3%, P <0.0001). Cell contractility (beating) was seen in 6% of GMTd‐treated cells by 4 weeks after treatment, whereas no beating GMT‐treated cells were observed. Human cardiac fibroblasts likewise demonstrated increased cTnT expression with GMTd versus GMT treatment (7.5±0.3% versus 3.0±0.3%, P <0.01). Mechanistically, GMTd administration increased expression of the trimethylated lysine 4 of histone 3 (H3K4me3) mark at the promoter regions of cardio‐differentiation genes and mitochondrial biogenesis regulator genes in rat and human fibroblast, compared with GMT.
Conclusions
These data suggest that the Hippo pathway intermediate Tead1 is an important regulator of cardiac reprogramming that increases the efficiency of maturate induced cardiomyocytes generation and may be a vital component of human cardiodifferentiation strategies.
... fibroblasts into "induced cardiomyocytes" (iCMs) [1][2][3][4][5][6][7][8][9] . Despite encouraging observations that cardiac transdifferentiation improves post-infarct cardiac function in small animal models, limits on the efficiency of this process, especially in human cells, has catalyzed the search for more effective cardiac reprograming strategies [10][11][12][13][14] . ...
... Lentivirus vectors encoding ETV2 tagged with yellow fluorescent protein (Venus), or encoding Gata4, Mef2c orTbx5 tagged with green fluorescent protein (GFP), or GFP alone were prepared from relevant plasmids by the Baylor College of Medicine Gene Vector Core, as previously described 4,5,10 . Plasmids for human ETV2 and reverse tetracycline-controlled transactivator (rtTA) were gifts from Dr. Rinpei Morita, Department of Microbiology and Immunology, Keio University, Tokyo, Japan. ...
... Fluorescence-activated cell sorting (FACS) analysis. For FACS analysis, cells were washed, trypsinized and fixed with fixation buffer (BD Biosciences, San Jose, CA) as previously described 5,10 . Fixed cells were permeabilized with Perm/Wash buffer (BD Biosciences, San Jose, CA). ...
Fibroblast reprogramming offers the potential for myocardial regeneration via in situ cell transdifferentiation. We explored a novel strategy leveraging endothelial cell plasticity to enhance reprogramming efficiency. Rat cardiac endothelial cells and fibroblasts were treated with Gata4, Mef2c, and Tbx5 (GMT) to assess the cardio-differentiation potential of these cells. The endothelial cell transdifferentiation factor ETV2 was transiently over-expressed in fibroblasts followed by GMT treatment to assess “trans-endothelial” cardio-differentiation. Endothelial cells treated with GMT generated more cTnT ⁺ cells than did cardiac fibroblasts (13% ± 2% vs 4% ± 0.5%, p < 0.01). Cardiac fibroblasts treated with ETV2 demonstrated increased endothelial cell markers, and when then treated with GMT yielded greater prevalence of cells expressing cardiomyocyte markers including cTnT than did fibroblasts treated with GMT or ETV2 (10.3% ± 0.2% vs 1.7% ± 0.06% and 0.6 ± 0.03, p < 0.01). Rat cardiac fibroblasts treated with GMT + ETV2 demonstrated calcium transients upon electrical stimulation and contractility synchronous with surrounding neonatal cardiomyocytes, whereas cells treated with GMT or ETV2 alone failed to contract in co-culture experiments. Human cardiac fibroblasts treated with ETV2 and then GMT likewise demonstrated greater prevalence of cTnT expression than did cells treated with GMT alone (2.8-fold increase, p < 0.05). Cardiac fibroblast transitioning through a trans-endothelial state appears to enhance cardio-differentiation by enhancing fibroblast plasticity.
... Importantly, in vitro studies with mouse cells also provide specie-specific outcomes with limited relevance and predictivity for humans. As an exception, pig CFs might be employed considering their close features to human CFs (53). However, investigation on AHCFs is preferred in the perspective of future clinical translation of the approach. ...
Ischemic heart disease is the major cause of mortality worldwide. Despite the most recent pharmacological progresses, cardiac regeneration is yet not possible, and heart transplantation is the only therapeutic option for end-stage heart failure. Traditional cardiac regenerative medicine approaches, such as cell therapies and tissue engineering, have failed in the obtainment of human functional cardiac tissue, mainly due to unavailability of high quantities of autologous functional cardiomyocytes (CMs), low grafting efficiency, and/or arrhythmic events. Direct reprogramming (DR) of fibroblasts into induced CMs (iCMs) has emerged as a new promising approach for myocardial regeneration by in situ transdifferentiation or providing additional CM source for cell therapy. Among available DR methods, non-viral transfection with microRNAs (miRcombo: miR-1, miR-133, miR-208, and miR-499) appears promising for future clinical translation. MiRcombo transfection of fibroblasts could be significantly improved by the development of safe nanocarriers, efficiently delivering their cargo to target cells at the required stoichiometric ratio and overall dose in due times. Newly designed in vitro 3D culture microenvironments, providing biomimetic biophysical and biochemical stimuli to miRcombo-transfected cells, significantly increase the yield of fibroblast transdifferentiation into iCMs, enhancing CM gene expression. Epigenetic regulation of gene expression programs, critical to cell lineage commitment, can also be promoted by the administration of specific anti-inflammatory and anti-fibrotic soluble factors, helping in suppressing fibroblast signature. The aim of this mini-review is to introduce the readers to a relatively unknown field of cardiac research integrating bioengineering tools as relevant for the progress of miRNA-mediated cardiac DR.
