# Pre-Conditioning Methods and Novel Approaches with Mesenchymal Stem Cells Therapy in Cardiovascular Disease

Anthony G Matta, Vanessa Nader, Marine Lebrin, Fabian Gross, Anne-Catherine Prats, Daniel Cussac, Michel Galinier, Jérôme Roncalli

► To cite this version:

Anthony G Matta, Vanessa Nader, Marine Lebrin, Fabian Gross, Anne-Catherine Prats, et al. Pre-Conditioning Methods and Novel Approaches with Mesenchymal Stem Cells Therapy in Cardiovascular Disease. Cells, 2022, 11 (10), pp.1620. 10.3390/cells11101620 . hal-04880325

HAL Id: hal-04880325

https://hal.science/hal-04880325v1

Submitted on 10 Jan 2025

HAL is a multi-disciplinary open access
archive for the deposit and dissemination of sci-
entific research documents, whether they are pub-
lished or not. The documents may come from
teaching and research institutions in France or
abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est
destinée au dépôt et à la diffusion de documents
scientifiques de niveau recherche, publiés ou non,
émanant des établissements d'enseignement et de
recherche français ou étrangers, des laboratoires
publics ou privés.

<figure><img src="image_1.png" /></figure>
Review

# Pre-Conditioning Methods and Novel Approaches with Mesenchymal Stem Cells Therapy in Cardiovascular Disease

Anthony Matta<sup>1,2,3</sup>, Vanessa Nader<sup>1,4</sup>, Marine Lebrin<sup>1,5</sup>, Fabian Gross<sup>1,5 ID</sup>, Anne-Catherine Prats<sup>6 ID</sup>, Daniel Cussac<sup>6 ID</sup>, Michel Galinier<sup>1</sup> and Jerome Roncalli<sup>1,5,6,* ID</sup>

**Abstract:** Transplantation of mesenchymal stem cells (MSCs) in the setting of cardiovascular disease, such as heart failure, cardiomyopathy and ischemic heart disease, has been associated with good clinical outcomes in several trials. A reduction in left ventricular remodeling, myocardial fibrosis and scar size, an improvement in endothelial dysfunction and prolonged cardiomyocytes survival were reported. The regenerative capacity, in addition to the pro-angiogenic, anti-apoptotic and anti-inflammatory effects represent the main target properties of these cells. Herein, we review the different preconditioning methods of MSCs (hypoxia, chemical and pharmacological agents) and the novel approaches (genetically modified MSCs, MSC-derived exosomes and engineered cardiac patches) suggested to optimize the efficacy of MSC therapy.

**Keywords:** mesenchymal stem cells; preconditioning; exosome; engineered cardiac patches

<figure><img src="image_2.png" /></figure>

check for
updates

**Citation:** Matta, A.; Nader, V.; Lebrin, M.; Gross, F.; Prats, A.-C.; Cussac, D.; Galinier, M.; Roncalli, J. Pre-Conditioning Methods and Novel Approaches with Mesenchymal Stem Cells Therapy in Cardiovascular Disease. *Cells* **2022**, *11*, 1620. https://doi.org/10.3390/cells11101620

Academic Editor: Joni H. Ylostalo

Received: 13 April 2022

Accepted: 10 May 2022

Published: 12 May 2022

**Publisher's Note:** MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations.

<figure><img src="image_3.png" /></figure>

**Copyright:** © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).

## 1. Introduction

Several clinical trials have established the safety of mesenchymal stem cell (MSC) therapy and have shown promising results in the setting of cardiovascular disease over the past decades [1,2]. In ischemic heart disease, the role of existing conventional therapy, including percutaneous coronary intervention, coronary artery bypass graft and medical treatment, is limited to prevent future ischemic events and further expansion of myocardial damage [3]. Unlike MSC transplantation, there are no effects on myocardial repair, lost myocardial tissue and cardiomyocytes regeneration. Data from the literature showed a reduction in scar burden, myocardial fibrosis and infarct size, a reversion of left ventricular remodeling and an improvement in cardiac function after MSC therapy [1,4,5].

MSCs are undifferentiated, multipotent and self-renewable cells recognized for their potential of differentiation [6,7] and paracrine activity [2,8–10]. MSCs secrete diverse biological active cytokines, growth factors, chemokines and miRNA, resulting in anti-fibrotic, anti-inflammatory, regenerative, proliferative, immunomodulatory and angiogenic effects [11–14]. Neovascularization, angiogenesis, cardiomyocytes apoptosis inhibition, myocardial repair enhancement and dead cardiomyocytes replacement are the major targets of MSC therapy within the context of myocardial infarction [2]. MSCs are present in different human organs, but usually isolated from the following three main sources: umbilical cord, adipose tissue and bone marrow [2]. The latter is commonly used, despite the fact that it provides a mixture of non-purified miscellaneous cells [15]. After injection, MSCs are

<footnote n="1">¹ Department of Cardiology, Institute CARDIOMET, University Hospital of Toulouse, 31059 Toulouse, France; dr.anthonymatta@hotmail.com (A.M.); nader.e.vanessa@gmail.com (V.N.); lebrin.m@chu-toulouse.fr (M.L.); gross.f@chu-toulouse.fr (F.G.); galinier.m@chu-toulouse.fr (M.G.)</footnote>

<footnote n="2">² Faculty of Medicine, Holy Spirit University of Kaslik, Kaslik 446, Lebanon</footnote>

<footnote n="3">³ Department of Cardiology, Intercommunal Hospital Centre Castres-Mazamet, 81100 Castres, France</footnote>

<footnote n="4">⁴ Faculty of Pharmacy, Lebanese University, Beirut 6573/14, Lebanon</footnote>

<footnote n="5">5 CIC-Biotherapies, University Hospital of Toulouse, 31059 Toulouse, France</footnote>

<footnote n="6">6 INSERM I2MC—UMR1297, 31432 Toulouse, France; anne-catherine.prats@inserm.fr (A.-C.P.); daniel.cussac@inserm.fr (D.C.)</footnote>

<footnote>* Correspondence: roncalli.j@chu-toulouse.fr; Tel.: +33-56-132-3334; Fax: +33-56-132-2246</footnote>
able to home, accumulate and engraft with the adjacent cellular components of the injured tissue and, subsequently, recruit additional progenitor cells [15,16]. However, hypoxia and increased free radical concentration in the context of myocardial infarction generate a detrimental microenvironment for transplanted MSCs [17]. Thus, preconditioning of MSCs with hypoxia or pharmacological or chemical agents in addition to novel strategies, such as exosome-mediated MSCs, genetically modified MSCs and engineered cardiac patches, were performed for improving the overall efficacy of MSC transplantation (Figure 1). All these techniques promote MSC survival and their capacity to form a regenerative and proliferative environment. Herein, we review the different preconditioning methods and novel approaches with MSCs in the setting of ischemic cardiac disease.

<figure><img src="image_4.png" /><figcaption>**Figure 1.** Figure illustrating mesenchymal stem cell (MSC) preconditioning methods, novel approaches and their main impacts.</figcaption></figure>

## 2. Preconditioning Methods

### 2.1. Hypoxia-Preconditioned MSCs

The purpose of hypoxic preconditioning is to prolong the short survival time of grafted MSCs in the ischemic area, a major limitation of the therapeutic potential of stem cell therapy [18–20]. Indeed, hypoxic preconditioning increases the expression of protective factors against future hypoxic insult (hypoxia inducible factor-1 α (HIF-1α)), angiogenic factors (vascular epithelial growth factor, angiopoietin-1 and erythropoietin), pro-survival proteins (P65, P50 and P105) and anti-apoptotic proteins (Bcl-xl et Bcl-2) [21]. Previous study results showed that 24 h hypoxia exposure could dramatically amplify MSC proliferation and reduce their apoptosis by mainly activating the HIF-1α/Apelin/APJ axis [22]. First, HIF-1α modulates oxygen homeostasis and promotes cell function and tolerance in a
hypoxic microenvironment [23]. It plays a crucial role in cardiomyocytes protection against ischemia-reperfusion injury by regulating mitochondrial reactive oxygen species [24] and heme oxgenase-1 [25]. Then, the inhibition of inflammatory reaction and apoptosis, up-regulation of collagen matrix and glycolysis, stimulation of angiogenesis and improvement of oxygen delivery are mediated by HIF-1α [26]. Second, the stimulation of Apelin/APJ enhances MSC survival and differentiation [27]. Moreover, hypoxia preconditioning activates other pathways, such as SDF-1α/CXCR4 axis implicated in MSC migration, detention and homing [28,29], PI3K/Akt signaling pathway that blocks cell death [30] and GRP78 that interferes in angiogenic cytokine secretion and cell migration [31]. A recent study revealed that extracellular vesicles from hypoxia-preconditioned MSCs may partly alleviate myocardial injury by targeting the thioredoxin-interacting protein-mediated HIF-1α pathway [32]. The evidence suggests that transplantation of hypoxia-preconditioned MSCs in the setting of myocardial infarction results in better cardiovascular outcomes by enhancing MSC engraftment, proliferation, differentiation, survival and paracrine activity [33,34]. Furthermore, it has shown that hypoxic preconditioning enhances survival and proangiogenic capacity of human first trimester chorionic villus-derived MSCs for fetal tissue engineering [35]. Lastly, we spotlight that different percentages of hypoxia have different outcomes. For example, 1% hypoxia extends MSC lifespan and maintains their proliferation rate [36,37]. In addition, 2% and 5 % hypoxia increased MSC number and viability [34]. Upregulation of stemness-related genes was observed with 3% hypoxia [38,39]. In other words, severe hypoxia (<1%) activates glycolytic metabolism and induces MSC quiescence, whereas moderate hypoxia (3–5%) stimulates MSC proliferation [40–42]. Although, short duration exposure to hypoxia (24 h) yields a better result than that of longer duration (72 h).

## 2.2. Preconditioning with Pharmacological and Chemical Agents

Numerous growth factors, drugs and pharmacological and chemical substances have been used for MSC preconditioning (Table 1). For example, the treatment of MSCs with IGF-1 showed a positive impact on survival, detrimental infarct consequences (infarct size, ventricular remodeling and fibrosis) and pro-inflammatory cytokines [43]. HGF promotes MSC differentiation into cardiomyocytes, whereas the effect of IGF-1 on MSC potential of differentiation remains uncertain [44,45]. On the other hand, pretreatment with bFGF improves stem cells’ homing ability to the infarct zone and angiogenesis [46]. The pretreatment of MSCs with growth factor combinations (FGF-2, IGF-1 and BMP-2) leads to stronger engraftment, better viability in hypoxic situations, enhanced cell to cell communication and greater cytoprotective effects [47]. The results of a recent study showed superior cardiac function recovery and vasculogenesis in the infarcted myocardium 6 weeks after an injection of treated MSCs with SDF-1α in a rat model [48]. Beyond growth factors, variant biological active substances have been tested to improve the therapeutic efficacy of MSC therapy. Indeed, MSC pretreatment with angiotensin II potentiates the paracrine activity, angiogenesis, gap junction formation and global clinical outcome, by up-regulating the expression of VEGF, Cx43 with no effects on the differentiation mechanisms [49]. In addition, the left ventricular cardiac function and cardiomyogenic transdifferentiation have been significantly improved after transplantation of pioglitazone pretreated MSCs [50]. Thus, it seems a promising preconditioning method to predict cardiomyogenesis. Furthermore, pretreatment of MSCs with atorvastatin significantly improved cardiac function, reduced infarct size, decreased serum marker level of inflammation and fibrosis, inhibited apoptosis and enhanced survival of implanted MSCs, via activating the subtype eNOS of nitric oxide synthase [51]. Atorvastatin also improved the migration capacity of MSCs by increasing the expression of CXCR4 [52]. Benefits on MSC survival and differentiation have been observed with simvastatin pretreated MSCs [53]. Statin pretreatment positive outcomes have been also observed after transplantation of sevoflurane-preconditioned MSCs, which increase the expression of HIF-1α, HIF-2α, VEGF and p/Akt/Akt [54]. Transplantation of LPS-(lipopolysaccharide) preconditioned MSCs in the setting of myocardial infarction improves their biological and functional characteristics by up-regulating VEGF, phosphorylated Akt
and TLR4 pathway [55]. Thereby, longer survival of transplanted cells, intense neovascularization and greater amelioration of left ventricular ejection fraction have been reported [55]. Vitamine E decreases oxidative stress and H₂O₂-related senescence by up-regulating the expression of VEGF, TGF-β and LDH [56]. The proliferation ability of MSCs has been promoted with astragaloside IV by inhibiting the translocation of NF-κBp65 [57], apple ethanol extract by inducing the phosphorylation of eIF4E, p44, p70S6K, MAPK, eIF48, p44/42, mTOR and S6RP [58], oxytocin by activating the Akt/ERK1/2 axis [59], LL-37 by activating the MAPK pathway [60] and migration inhibitory factor by releasing VEGF, BFGF, HGF and IGF [61]. Although, the migration and homing abilities of MSCs have been improved with deferoxamine by expressing HIF-1α, CXCR4, CCR2, MMP-2 and MMP-9 [62], IL-1β by producing different cytokines, chemokines and adhesions molecules [63] and TGF-β1 by triggering the canonical SMADs [64]. In addition, the improvement of cardiovascular stem cell therapeutic outcomes has been associated with transplantation of 2,4-dinitrophenol [65], oxytocin [66] and dimethyloxyglycine [67] pretreated MSCs. Finally, our group has shown that melatonin (pineal hormone to protect tissue from oxidative damage) pretreated MSCs modulate survival, differentiation and antifibrotic activity of cardiac fibroblasts [68]. Our results showed that MSCs significantly improved morphological and functional cardiac parameters two weeks after injection. However, the partial recovery of ventricular ejection fraction was maintained up to two months only when MSC survival was increased by melatonin treatment. These data indicate that the increased number of viable cells is critical for the amplification of the beneficial effects of MSCs on injured myocardium and ventricular function recovery. These properties of MSCs opened new perspective for understanding the mechanisms of action of MSCs and anticipated their potential therapeutic effects.

<table><caption>**Table 1.** MSC preconditioning with pharmacological and chemical agents.</caption><thead><tr><td>Agents</td><td>Effects on</td><td>References</td></tr></thead><tbody><tr><td>IGF-1</td><td>survival, infarct consequences, pro-inflammatory cytokines</td><td>[43]</td></tr><tr><td>HGF</td><td>differentiation into cardiomyocytes</td><td>[44,45]</td></tr><tr><td>bFGF</td><td>stem cells homing and angiogenesis</td><td>[46]</td></tr><tr><td>FGF-2, IGF-1 and BMP-2 combination</td><td>engraftment, viability, cell to cell communication, cytoprotective effect</td><td>[47]</td></tr><tr><td>SDF-1α</td><td>cardiac function recovery and vasculogenesis</td><td>[48]</td></tr><tr><td>Angiotensin II</td><td>paracrine activity, angiogenesis and gap junction formation</td><td>[49]</td></tr><tr><td>Pioglitazone</td><td>cardiac function and cardiomyogenic trans differentiation</td><td>[50]</td></tr><tr><td>Atorvastatin</td><td>cardiac function, infarct size, serum markers level of inflammation and fibrosis, apoptosis, migration capacity and survival of implanted MSCs</td><td>[51,52]</td></tr><tr><td>Simvastatin</td><td>MSC survival and differentiation</td><td>[53]</td></tr><tr><td>Sevoflurane</td><td>homing, survival and differentiation</td><td>[54]</td></tr><tr><td>LPS (lipopolysaccharide)</td><td>biological and functional characteristics of MSCs</td><td>[55]</td></tr><tr><td>Vitamine E</td><td>decreases oxidative stress and H2O2-related senescence</td><td>[56]</td></tr><tr><td>Astragaloside</td><td>proliferation ability of MSCs</td><td>[57]</td></tr><tr><td>Apple ethanol</td><td>proliferation ability of MSCs</td><td>[58]</td></tr><tr><td>Oxytocin</td><td>proliferation ability of MSCs</td><td>[59]</td></tr><tr><td>LL-37</td><td>proliferation ability of MSCs</td><td>[60]</td></tr><tr><td>Deferoxamine</td><td>migration and homing abilities of MSC</td><td>[62]</td></tr></tbody></table>
<table><caption>**Table 1.** *Cont.*</caption><thead><tr><td>Agents</td><td>Effects on</td><td>References</td></tr></thead><tbody><tr><td>IL-1β</td><td>migration and homing abilities of MSCs</td><td>[63]</td></tr><tr><td>TGF-β1</td><td>migration and homing abilities of MSCs</td><td>[64]</td></tr><tr><td>2,4-dinitrophenol</td><td>cardiovascular stem cell therapeutic outcomes</td><td>[65]</td></tr><tr><td>Oxytocin</td><td>cardiovascular stem cell therapeutic outcomes</td><td>[66]</td></tr><tr><td>Dimethyloxalyglycine</td><td>cardiovascular stem cell therapeutic outcomes</td><td>[67]</td></tr><tr><td>Melatonin</td><td>survival, differentiation and antifibrotic activity</td><td>[68]</td></tr></tbody></table>

### 3. Novel Approaches

#### 3.1. Genetic Modification of MSCs

Genetic modification of MSCs up-regulates the expression of specific genes implicated in MSC migration, adhesion, survival and premature senescence (Table 2). To begin, the migratory ability of MSCs has been promoted by overexpressing nuclear receptors (Nur1, Nur77) [69,70], integrin subunit-α4 [71], aquaporin-1 [72] and CXCR4/CXCR7 that serve as receptors for major cellular migratory process chemokine (SDF-1) [73,74]. Then, the overexpression of α(1,3)fucosyltransferase [75], focal adhesion kinase [76], integrin-linked kinase [77] and miR-9-5p [78] have been linked to stronger MSC adhesion and engraftment. However prolonged survival of transplanted MSCs has been demonstrated with overexpression of integrin-linked kinase that activates AKT, mTOR, JAK2/STAT3 signaling pathways [79,80], protein kinase Cε [81], Trkβ [82] and Gremlin1 [83]. The up-regulation of Sox2 and Oct4 genes accelerates cell transition from phase G1 into phase S, enhancing MSC proliferation, differentiation and anti-inflammatory effect [84,85]. EphB2 overexpression reduced premature senescence by suppressing mitochondrial reactive oxygen species accumulation, which triggers MSC senescence [86]. Transplantation of Kallikrein-1 genetically modified MSCs attenuates cardiac inflammation, cardiomyocytes apoptosis and myocardial fibrosis via VEGF, GSK-3β and NO signaling pathways activation [87–91]. Thus, pleiotropic, angiogenic proteolytic and cardioprotective effects have been attributed to Kallikrein-1 [91]. In the context of acute myocardial infarction, several clinical trials have demonstrated the therapeutic benefits of transplantation of genetically modified MSCs in animal models. For example, the target outcomes of MSC therapy were maintained for longer durations with transplanted Akt or angiopoietin1-MSCs [92]. Although, an injection of Bcl-2 or SDF-1α-or TNFR gene modified MSCs or miR-377 depleted MSCs potentiates the required efficacy of vascular density, cardiac function, infarct size and myocardial fibrosis [93–98]. Genetic modification of MSCs is applied using viral vectors, such as adenoviral, lentiviral and retroviral vectors for nucleic acid delivery [99], non-viral delivery systems, such as plasmid DNA, polymers, nanoplasmids, liposomes and DNA minicircles [100–102] and the novel gene-editing technology, clustered regularly interspaced short palindromic repeats (CRISPR/Cas9) [103]. This last technique allows one to insert a new sequence in the genome via homology-directed repair, which could rectify an acquired gene mutation or provoke a knock-in or knock-out mutation or suppress a specific gene expression [103].
<table><caption>**Table 2.** Outcomes of genetic modifications of MSCs.</caption><thead><tr><th>Function</th><th>Up-Regulating Genes</th><th>References</th></tr></thead><tbody><tr><td rowspan="4">Improved MSC migration</td><td>Nur1, Nur7</td><td>[69,70]</td></tr><tr><td>Integrin subunit- α4</td><td>[71]</td></tr><tr><td>Aquaporin-1</td><td>[72]</td></tr><tr><td>CXCR4/VXCR7</td><td>[73,74]</td></tr><tr><td rowspan="4">Improved MSC adhesion and engraftment</td><td>α(1,3)fucosyltransferase</td><td>[75]</td></tr><tr><td>Focal adhesin kinase</td><td>76]</td></tr><tr><td>Integrin-linked kinase</td><td>[77]</td></tr><tr><td>miR-9-5-p</td><td>[78]</td></tr><tr><td rowspan="4">Prolonged MSC survival</td><td>Integrin-linked kinase</td><td>[79,80]</td></tr><tr><td>Protein kinase Cε</td><td>[81]</td></tr><tr><td>Trkβ</td><td>[82]</td></tr><tr><td>Gremlin 1</td><td>[83]</td></tr><tr><td>Enhanced MSC proliferation and differentiation</td><td>Sox2 and Oct4</td><td>[84,85]</td></tr><tr><td>Reduced premature senescence</td><td>EphB2</td><td>[86]</td></tr><tr><td>Sustained therapeutic efficacy</td><td>AktAngiopoietin 1</td><td>[92]</td></tr><tr><td rowspan="4">Better outcomes in setting of acute myocardial infarction</td><td>Bcl-2</td><td>[93]</td></tr><tr><td>SDF-1α</td><td>[95]</td></tr><tr><td>TNFR</td><td>[97]</td></tr><tr><td>miR-377</td><td>[98]</td></tr></tbody></table>

## 3.2. MSCs *Derived-Exosomes*

Exosomes are classified as extracellular vesicles that are continuously produced and released by various hematopoietic and non-hematopoietic cells [104–106]. Exosomes interfere in variant cell to cell interaction pathways that are implicated in different physiological and pathological patterns [107]. Endocytosis, membrane fusion and membrane receptors represent the three exosomal mechanisms to regulate cell to cell communication [108]. Exosomes are mainly isolated for therapeutic application, either by ultrafiltration or ultracentrifugation-based methods [107]. Preclinical experimental animal models have demonstrated the therapeutic benefits of MSC-derived exosomes in the setting of myocardial infarction. An injection of MSC-derived miRNA-enriched exosomes have showed remarkable outcomes, such as reduction in infarct size and myocardial fibrosis with miR-22 via acting on MECP2 [109], enhancement of anti-apoptotic and cardioprotective effects with miR-221 by inhibiting PUMA expression [110], promotion of cardiac function recovery with miR-19a by suppressing PTEN and activating ERK pathways, respectively [111], and improvement in angiogenesis with miR-210 [112]. Overall, the transplantation of exosomes-derived MSCs leads to stronger cardioprotective effects [113] and reduction in the risk of tumorigenicity [114] than MSC-based therapies.

## 3.3. Engineered Cardiac Patches

Cell sheets and cell containing scaffolds represent the two forms of engineered cardiac patches [115]. Multiple cell types, such as endothelial cells, cardiac fibroblasts, pluripotent stem cells, cardiomyocytes, progenitor cells and smooth muscle cells have been incorporated into engineered cardiac patches [116–119]. Consequently, the replacement of damaged cardiomyocytes with functional cardiac cells is the ultimate target of engineered cardiac tissue transplantation. Promising results with evidence of remuscularization of the fibrotic myocardium have been demonstrated in numerous pre-clinical studies [120–125]. Indeed,
cardiac function recovery has been observed in rats and minipigs after 4 weeks of transplantation of cell-free patches in the setting of acute anterior myocardial infarction [126]. Furthermore, the implantation of a bioengineered cardiac patch has shown superior therapeutic efficacy compared to that of decellularized placenta and human-induced pluripotent stem cells for myocardial repair, mediated by growth and pro-angiogenic factors that promote engraftment, neovascularization and paracrine function [127]. However, the need for a huge quantity of exogenous cardiac cells to refill the injured myocardium and stable electromechanical coupling between the transplanted cardiac patches and host tissue for long-term engraftment are the main challenges for this novel cardiac approach [128]. Thereby, larger and thicker vascularized cardiac patches that are synchronized with the circulatory and electromechanical systems of the native myocardium are required to overcome these limitations. The safety concern of cardiac patch therapy was limited to arrhythmias, which were generally transient and non-fatal [129–131]. It is noteworthy that a recently published study has revealed the efficacy of upscaled engineered heart tissue to improve left ventricular function and reduce the infarct size in the context of ischemic myocardial disease without documenting a significant difference in arrhythmogenicity, compared to a cell-free patch group in a rabbit model [132].

Overall, the therapeutic benefits of MSCs have been demonstrated in the treatment of ischemic cardiomyopathies [133]; however, the limited engraftment and poor survival of MSCs injected into an ischemic heart hindered the efficacy of the treatment. The use of scaffolds and polymeric supports to provide transplanted cells anchorage, a straightforward approach to circumvent this limitation, has already been tested [134]. Indeed, a robust therapeutic benefit of ADSCs when transplanted with a collagen scaffold in a preclinical porcine model of myocardial infarction, compared with cells without a collagen scaffold, has been successfully demonstrated. The functional improvement in cardiac function and myocardial remodeling after ADSC-collagen scaffold transplantation was associated with increased cell engraftment [135]. The positive preclinical results obtained using different biomaterials and cell types invited researchers to test whether these experimental procedures could be translated into the clinical setting. Thus, the phase I MAGNUM clinical trial was designed with the purpose of comparing the effects exerted by bone-marrow mononucleated cells-seeded cellularized collagen matrices with those exerted by cells alone, in patients presenting left ventricular post-ischemic myocardial scars. The results were promising because no treatment-related serious adverse events were reported during the follow-up period and heart functionality and mechanical parameters improved significantly in patients who received the cellularized patches. In other words, clinically, this procedure seems to be safe, feasible, and effective [136]. We mention that one of the first clinical trials on engineered heart muscle in patients with terminal heart failure is ongoing, BioVAT-HF (ClinicalTrials.gov: NCT04396899). However, a recent report of in-human transplantation of an allogenic-induced pluripotent stem cell-derived cardiomyocytes patch into the epicardium of the anterior and lateral walls via the fourth intercostal space in a patient with ischemic cardiomyopathy has been currently published [137]. This report signals the safety and efficacy of these patches on NYHA class, left ventricular end systolic volume and Vo2 peak at the 1-year follow-up after transplantation [137]. Moreover, the ESCORT trial on six patients referred to cardiac surgery has also demonstrated the technical feasibility of producing clinical-grade human embryonic stem cell-derived cardiovascular progenitors delivered in a fibrin epicardial patch, and supported their short- and medium-term safety, thereby, setting the grounds for adequately powered efficacy studies [138]. Finally, the translation of preclinical findings to the first clinical results requires the creation of cardiac scaffolds following all the GMP regulatory and quality requirements in order to test their safety as potential therapeutic products. The CARDIOPATCH Interreg Sudoe program aims to create a 2.0 version patch (v2.0) with growth factors and genetically improved mesenchymal cells and iPS-derived cardiac cells that improve cell survival of both the implanted cells and the ischemic cardiac tissue, as well as their pro-angiogenic capacity.
## 4. MSCs Perspectives

As is known for most new therapies, the progression of MSC therapy has been hard, slow and punctuated by difficulties. The available evidence proves the safety of MSC transplantation, which represents a new, hopeful strategy for the management of cardiovascular disease, particularly ischemic and non-ischemic heart failure [139–141]. Up to date, numerous Phase I and Phase II trials have demonstrated promising results with regenerative medicine in the setting of heart failure and myocardial infarction [2]. The findings from these trials are divergent. However, several important points have not yet been defined, such as the preferred cell source, preparation method, appropriate dose and recommended manner of administration. Defining these parameters constitutes an important step towards establishing a standard approach with MSC therapy and ensuring result reproducibility. The results from pivotal phase III trials are required to support the clinical application of MSC therapy in the cardiovascular field. Recently, stem cell therapy was approved for the management of complex perianal fistulas in Crohn's disease [142]. We emphasize that pre-conditioning methods have contributed to overcome numerous hurdles, such as injected cell migration, engraftment, proliferation, differentiation and survival, resulting in stronger efficacy and better outcomes. Furthermore, recent studies have proved the benefits of mechanical stimulation on MSCs and the surrounding microenvironment and showed the interest of its application for bone regeneration therapy [143]. Lastly, engineered cardiac patch technology represents a revolution in stem cell therapy for cardiovascular disease, but manufacturing larger and thicker constructs that are suitably vascularized and incorporated with the electromechanical and circulatory systems of the vernacular myocardium is necessary for the clinical translation step.

## 5. Conclusions

To conclude, transplantation of pre-conditioned MSCs results in better therapeutic efficacy in the setting of cardiovascular disease, especially with moderate hypoxia pre-conditioning. In parallel, the available novel techniques are able to overcome the limitations (MSCs homing ability, engraftment and survival) of this regenerative medicine, promoting stronger cardiovascular outcomes. Starting translational engineered cardiac patch practice from pre-clinical trials in animal models to in-human trials may change our future management of heart failure.

**Funding:** This research was partly supported by CARDIOPATCH (SOE4/P1/E1063) project co-funded by the Interreg Sudoe program.

**Institutional Review Board Statement:** Not applicable.

**Informed Consent Statement:** Not applicable.

**Data Availability Statement:** Not applicable.

**Conflicts of Interest:** The authors declare no conflict of interest.

## References

1. Guijarro, D.; Lebrin, M.; Lairez, O.; Bourin, P.; Piriou, N.; Pozzo, J.; Lande, G.; Berry, M.; Le Tourneau, T.; Cussac, D.; et al. Intramyocardial transplantation of mesenchymal stromal cells for chronic myocardial ischemia and impaired left ventricular function: Results of the MESAMI 1 pilot trial. *Int. J. Cardiol.* **2016**, *209*, 258–265. [CrossRef] [PubMed]

2. Razeghian-Jahromi, I.; Matta, A.G.; Canitrot, R.; Zibaeenezhad, M.J.; Razmkhah, M.; Safari, A.; Nader, V.; Roncalli, J. Surfing the clinical trials of mesenchymal stem cell therapy in ischemic cardiomyopathy. *Stem Cell Res.* **2021**, *12*, 361. [CrossRef] [PubMed]

3. Awada, H.K.; Hwang, M.P.; Wang, Y. Towards comprehensive cardiac repair and regeneration after myocardial infarction: Aspects to consider and proteins to deliver. *Biomaterials* **2016**, *82*, 94–112. [CrossRef] [PubMed]

4. Hare, J.M.; Fishman, J.E.; Gerstenblith, G.; DiFede Velazquez, D.L.; Zambrano, J.P.; Suncion, V.Y.; Tracy, M.; Ghersin, E.; Johnston, P.V.; Brinker, J.A.; et al. Comparison of allogeneic vs autologous bone marrow-derived mesenchymal stem cells delivered by transendocardial injection in patients with ischemic cardiomyopathy: The POSEIDON randomized trial. *JAMA* **2012**, *308*, 2369–2379. [CrossRef]
5. Butler, J.; Hamo, C.E.; Udelson, J.E.; O'Connor, C.; Sabbah, H.N.; Metra, M.; Shah, S.J.; Kitzman, D.W.; Teerlink, J.R.; Bernstein, H.S.; et al. Reassessing Phase II Heart Failure Clinical Trials: Consensus Recommendations. *Circ. Heart Fail.* **2017**, *10*, e003800. [CrossRef]

6. Pittenger, M.F. Multilineage potential of adult human mesenchymal stem cells. *Science* **1999**, *284*, 143–147. [CrossRef]

7. Chamberlain, G.; Fox, J.; Ashton, B.; Middleton, J. Concise review: Mesenchymal stem cells: Their phenotype, differentiation capacity, immunological features, and potential for homing. *Stem Cells* **2009**, *25*, 2739–2749. [CrossRef]

8. Caplan, A.I. Adult mesenchymal stem cells for tissue engineering versus regenerative medicine. *J. Cell Physiol.* **2007**, *213*, 341–347. [CrossRef]

9. Wei, X.; Yang, X.; Han, Z.P.; Qu, F.F.; Shao, L.; Shi, Y.F. Mesenchymal stem cells: A new trend for cell therapy. *Acta Pharm. Sin.* **2013**, *34*, 747–754. [CrossRef]

10. Castro-Manreza, M.E.; Montesions, J.J. Immunoregulation by msenchymal stem cells: Biological aspects and clinical applications. *J. Imminol. Res.* **2015**, *2015*, 394917.

11. Kuraitis, D.; Giordano, C.; Ruel, M.; Musaro, A.; Suuronen, E.J. Exploiting extracellular matrix-stem cell interactions: A review of natural materials for therapeutic muscle regeneration. *Biomaterials* **2012**, *33*, 428–443. [CrossRef] [PubMed]

12. Khubutiya, M.S.; Vagabov, A.V.; Temmov, A.A.; Sklifas, A.N. Paracrine mechanisms of proliferative, anti-apoptotic and anti-inflammatory effects of mesenchymal stromal cells in models of acute organ injury. *Cryotherapy* **2014**, *16*, 579–585. [CrossRef] [PubMed]

13. Gnecchi, M.; Zhang, Z.; Ni, A.; Dzau, V.J. Paracrine mechanisms in adult stem cell signaling and therapy. *Circ. Res.* **2008**, *103*, 1204–1219. [CrossRef] [PubMed]

14. Williams, A.R.; Hare, J.M. Mesenchymal stem cells: Biology, pathophysiology, translational findings, and therapeutic implications for cardiac disease. *Circ. Res.* **2011**, *109*, 923–940. [CrossRef]

15. Quevedo, H.C.; Hatzistergos, K.E.; Oskouei, B.N.; Feigenbaum, G.S.; Rodriguez, J.E.; Valdes, D.; Pattany, P.M.; Zambrano, J.P.; Hu, Q.; McNiece, I.; et al. Allogeneic mesenchymal stem cells restore cardiac function in chronic ischemic cardiomyopathy via trilineage differentiating capacity. *Proc. Natl. Acad. Sci. USA* **2009**, *106*, 14022–14027. [CrossRef]

16. Barbash, I.M.; Chouraqui, P.; Baron, J.; Feinberg, M.S.; Etzion, S.; Tessone, A.; Miller, L.; Guetta, E.; Zipori, D.; Kedes, L.H.; et al. Systemic delivery of bone marrow-derived mesenchymal stem cells to the infarcted myocardium: Feasibility, cell migration, and body distribution. *Circulation* **2003**, *108*, 863–868. [CrossRef]

17. Brodarac, A.; Šarić, T.; Oberwallner, B.; Mahmoodzadeh, S.; Neef, K.; Albrecht, J. Susceptibility of murine induced pluripotent stem cell-derived cardiomyocytes to hypoxia and nutrient deprivation. *Stem Cell Res.* **2015**, *6*, 83. [CrossRef]

18. Zhang, M.; Methot, D.; Poppa, V.; Fujio, Y.; Walsh, K.; Murry, C.E. Cardiomyocyte grafting for cardiac repair: Graft cell death and anti-death strategies. *J. Mol. Cell Cardiol.* **2001**, *33*, 907–921. [CrossRef]

19. Reinecke, H.; Murry, C.E. Cell grafting for cardiac repair. *Methods Mol. Biol.* **2003**, *219*, 97–112.

20. Toma, C.; Pittenger, M.F.; Cahill, K.S.; Byrne, B.J.; Kessler, P.D. Human mesenchymal stem cells differentiate to a cardiomyocyte phenotype in the adult murine heart. *Circulation* **2002**, *105*, 93–98. [CrossRef]

21. Hu, X.; Yu, S.P.; Fraser, J.L.; Lu, Z.; Ogle, M.E.; Wang, J.A.; Wei, L. Transplantation of hypoxia-preconditioned of mesenchymal stem cells improves infarcted heart function via enhanced survival of implanted cells and angiogenesis. *J. Thorac. Cardiovasc. Surg.* **2008**, *135*, 799–808. [CrossRef]

22. Hou, J.; Wang, L.; Long, H.; Wu, H.; Wu, Q.; Zhong, T.; Chen, X.; Zhou, C.; Guo, T.; Wang, T. Hypoxia preconditioning promotes cardiac stem cell survival and cardiogenic differentiation in vitro involving activation of the HIF-1α/Apelin/APJ axis. *Stem Cell Res.* **2017**, *8*, 215. [CrossRef] [PubMed]

23. Chen, J.; Kang, J.G.; Keyvanfar, K.; Youn, N.S.; Hwang, P.M. Long-term adaptation to hypoxia preserves hematopoietic stem cell function. *Exp. Hematol.* **2016**, *44*, 866–873. [CrossRef] [PubMed]

24. Cai, Z.; Zhong, H.; Bosch-Marce, M.; Fox-Talbot, K.; Wang, L.; Wei, C.; Trush, M.A.; Semenza, G.L. Complete loss of ischaemic preconditioning-induced cardioprotection in mice with partial deficiency of HIF-1 alpha. *Cardiovasc. Res.* **2008**, *77*, 463–470. [CrossRef] [PubMed]

25. Ockaili, R.; Natarajan, R.; Salloum, F.; Fisher, B.J.; Jones, D.; Fowler III, A.A.; Kukreja, R.C. HIF-1 activation attenuates postischemic myocardial injury: Role for heme oxygenase-1 in modulating microvascular chemokine generation. *Am. J. Physiol. Heart Circ. Physiol.* **2005**, *289*, H542–H548. [CrossRef]

26. Loor, G.; Schumacker, P.T. Role of hypoxia-inducible factor in cell survival during myocardial ischemia-reperfusion. *Cell Death Differ.* **2008**, *15*, 686–690. [CrossRef] [PubMed]

27. Zhang, N.K.; Cao, Y.; Zhu, Z.M.; Zheng, N.; Wang, L.; Xu, X.H.; Gao, L.R. Activation of endogenous cardiac stem cells by apelin-13 in infarcted rat heart. *Cell Transpl.* **2016**, *2*, 1645–1652. [CrossRef]

28. Tang, Y.L.; Zhu, W.; Cheng, M.; Chen, L.; Zhang, J.; Sun, T.; Kishore, R.; Phillips, M.I.; Losordo, D.W.; Qin, G. Hypoxic preconditioning enhances the benefit of cardiac progenitor cell therapy for treatment of myocardial infarction by inducing CXCR4 expression. *Circ. Res.* **2009**, *104*, 1209–1216. [CrossRef]

29. Kucia, M.; Reca, R.; Miekus, K.; Wanzeck, J.; Wojakowski, W.; Janowska-Wieczorek, A.; Ratajczak, J.; Ratajczak, M.Z. Trafficking of normal stem cells and metastasis of cancer stem cells involve similar mechanisms: Pivotal role of the SDF-1-CXCR4 axis. *Stem Cells* **2005**, *23*, 879–894. [CrossRef]
30. Alvarez-Tejado, M.; Naranjo-Suarez, S.; Jiménez, C.; Carrera, A.C.; Landázuri, M.O.; del Peso, L. Hypoxia induces the activation of the phosphatidylinositol 3-kinase/Akt cell survival pathway in PC12 cells: Protective role in apoptosis. *J. Biol. Chem.* **2001**, *276*, 22368-22374. [CrossRef]

31. Lee, J.H.; Yoon, Y.M.; Lee, S.H. Hypoxic preconditioning promotes the bioactivities of mesenchymal stem cells via the HIF-1α-GRP78-Akt axis. *Int. J. Mol. Sci.* **2017**, *18*, 1320. [CrossRef] [PubMed]

32. Mao, C.Y.; Zhang, T.T.; Li, D.J.; Zhou, E.; Fan, Y.Q.; He, Q.; Wang, C.Q.; Zhang, J.F. Extracellular vesicles from hypoxia-preconditioned mesenchymal stem cells alleviates myocardial injury by targeting thioredoxin-interacting protein-mediated hypoxia-inducible factor-1α pathway. *World J. Stem Cells* **2022**, *14*, 183-199. [CrossRef] [PubMed]

33. Raziyeva, K.; Smagulova, A.; Kim, Y.; Smagul, S.; Nurkesh, A.; Saparov, A. Preconditioned and genetically modified stem cells for myocardial infarction treatment. *Int. J. Mol. Sci.* **2020**, *21*, 7301. [CrossRef] [PubMed]

34. Bahir, B.; Choudhery, M.S.; Hussain, I. Hypoxic preconditioning as a strategy to maintain the regenerative potential of mesenchymal stem cells. In *Regenerative Medicine*, Choudhery, M., Ed.; IntechOpen: London, UK, 2020.

35. Hao, D.; Chuanchao, H.; Ma, B.; Lankford, L.; Reynaga, L.; Farmer, D.L.; Guo, F.; Wang, A. Hypoxic preconditioning enhances survival and proangiogenic capacity of human first trimester chorionic villus-derived mesenchymal stem cells for fetal tissue engineering. *Stem Cells Int.* **2019**, *2019*, 9695239. [CrossRef]

36. Lee, C.W.; Kang, D.; Kim, A.K.; Kim, D.Y.; Kim, D.I. Improvement of cell cycle lifespan and genetic damage susceptibility of human mesenchymal stem cells by hypoxic priming. *Int. J. Stem Cells* **2018**, *11*, 61. [CrossRef]

37. Suryawan, I.G.R.; Pikir, B.S.; Rantam, F.A.; Ratri, A.K.; Nugraha, R.A. Hypoxic preconditioning promotes survival of human adipocyte mesenchymal stem cell via expression of prosurvival and proangiogenic biomarkers. *F1000Research* **2021**, *10*, 843. [CrossRef]

38. Safwani, W.K.; Choi, J.R.; Yong, K.W.; Ting, I.; Adenan, N.A.; Pingguan-Murphy, B. Hypoxia enhances the viability, growth and chondrogenic potential of cryopreserved human adipose-derived stem cells. *Cryobiology* **2017**, *75*, 91-99. [CrossRef]

39. Werle, S.B.; Chagastelles, P.; Pranke, P.; Casagrande, L. Hypoxia upregulates the expression of the pluripotency markers in the stem cells from human deciduous teeth. *Clin. Oral Investig.* **2019**, *23*, 199-207. [CrossRef]

40. Obradovic, H.; Krstic, J.; Trivanovic, D.; Mojsilovic, S.; Okic, I.; Kukolj, T.; Ilic, V.; Jaukovic, A.; Terzic, M.; Bugarski, D. Improving Stemness and Functional Features of Mesenchymal Stem Cells from Wharton's Jelly of a Human Umbilical Cord by Mimicking the Native, Low Oxygen Stem Cell Niche. *Placenta* **2019**, *82*, 25-34. [CrossRef]

41. Park, S.E.; Kim, H.; Kwon, S.; Choi, S.; Oh, S.; Ryu, G.H.; Jeon, H.B.; Chang, J.W. Pressure Stimuli Improve the Proliferation of Wharton's Jelly-Derived Mesenchymal Stem Cells under Hypoxic Culture Conditions. *Int. J. Mol. Sci.* **2020**, *21*, 7092. [CrossRef]

42. Moniz, I.; Ramalho-Santos, J.; Branco, A.F. Differential oxygen exposure modulates mesenchymal stem cell metabolism and proliferation through mTOR signaling. *Int. J. Mol. Sci.* **2022**, *23*, 3749. [CrossRef] [PubMed]

43. Guo, J.; Zheng, D.; Li, W.F.; Li, H.R.; Zhang, A.D.; Li, Z.C. Insulin-like growth factor 1 treatment of MSCs attenuates inflammation and cardiac dysfunction following MI. *Inflammation* **2014**, *37*, 2156-2163. [CrossRef] [PubMed]

44. Zhang, G.W.; Gu, T.X.; Guan, X.Y.; Sun, X.J.; Qi, X.; Li, X.Y.; Wang, X.B.; Lv, F.; Yu, L.; Jiang, D.Q.; et al. HGF and IGF-1 promote protective effects of allogeneic BMSC transplantation in rabbit model of acute myocardial infarction. *Cell Prolif.* **2015**, *48*, 661-670. [CrossRef]

45. Farzaneh, M.; Rahimi, F.; Alishahi, M.; Khoshnam, S.E. Paracrine Mechanisms Involved in Mesenchymal Stem Cell Differentiation into Cardiomyocytes. *Curr. Stem Cell Res.* **2019**, *14*, 9-13. [CrossRef]

46. Ling, L.; Gu, S.; Cheng, Y.; Ding, L. bFGF promotes Sca-1+ cardiac stem cell migration through activation of the PI3K/Akt pathway. *Mol. Med. Rep.* **2018**, *17*, 2349-2356. [CrossRef] [PubMed]

47. Hahn, J.Y.; Cho, H.J.; Kang, H.J.; Kim, T.S.; Kim, M.H.; Chung, J.H.; Bae, J.W.; Oh, B.H.; Park, Y.B.; Kim, H.S. Pre-treatment of mesenchymal stem cells with a combination of growth factors enhances gap junction formation, cytoprotective effect on cardiomyocytes, and therapeutic efficacy for myocardial infarction. *J. Am. Coll Cardiol.* **2008**, *51*, 933-943. [CrossRef]

48. Esmaeili, R.; Darbandi-Azar, A.; Sadeghpour, A.; Majidzadeh, K.; Eini, L.; Jafarbeik-Iravani, N.; Hoseinpour, P.; Vajhi, A.; Oghabi Bakhshaiesh, T.; Masoudkabir, F.; et al. Mesenchymal stem cells pretreatment with stromal-derived factor-1 alpha augments cardiac function and angiogenesis in infarcted myocardium. *Am. J. Med. Sci.* **2021**, *361*, 765-775. [CrossRef]

49. Liu, C.; Fan, Y.; Zhou, L.; Zhu, H.Y.; Song, Y.C.; Hu, L.; Wang, Y.; Li, Q.P. Pretreatment of mesenchymal stem cells with angiotensin II enhances paracrine effects, angiogenesis, gap junction formation and therapeutic efficacy for myocardial infarction. *Int. J. Cardiol.* **2015**, *188*, 22-32. [CrossRef]

50. Shinmura, D.; Togashi, I.; Miyoshi, S.; Nishiyama, N.; Hida, N.; Tsuji, H.; Tsuruta, H.; Segawa, K.; Tsukada, Y.; Ogawa, S.; et al. Pretreatment of human mesenchymal stem cells with pioglitazone improved efficiency of cardiomyogenic transdifferentiation and cardiac function. *Stem Cells* **2011**, *29*, 357-366. [CrossRef]

51. Song, L.; Yang, Y.J.; Dong, Q.T.; Qian, H.Y.; Gao, R.L.; Qiao, S.B.; Shen, R.; He, Z.X.; Lu, M.J.; Zhao, S.H.; et al. Atorvastatin enhance efficacy of mesenchymal stem cells treatment for swine myocardial infarction via activation of nitric oxide synthase. *PLoS ONE* **2013**, *8*, e65702. [CrossRef]

52. Li, N.; Yang, Y.J.; Qian, H.Y.; Li, Q.; Zhang, Q.; Li, X.D.; Dong, Q.T.; Xu, H.; Song, L.; Zhang, H. Intravenous administration of atorvastatin-pretreated mesenchymal stem cells improves cardiac performance after acute myocardial infarction: Role of CXCR4. *Am. J. Transl. Res.* **2015**, *7*, 1058-1070. [PubMed]
53. Yang, Y.J.; Qian, H.Y.; Huang, J.; Li, J.J.; Gao, R.L.; Dou, K.F.; Yang, G.S.; Willerson, J.T.; Geng, Y.J. Combined therapy with simvastatin and bone-marrow derived mesenchymal stem cells increases benefits in infarcted swine hearts. *Arter. Thromb Vasc. Biol.* **2009**, *29*, 2076–2082. [CrossRef] [PubMed]

54. Sun, X.; Fang, B.; Zhao, X.; Zhang, G.; Ma, H. Preconditioning of mesenchymal stem cells by sevoflurane to improve their therapeutic potential. *PLoS ONE* **2014**, *9*, e90667.

55. Yao, Y.; Zhang, F.; Wang, L.; Zhang, G.; Wang, Z.; Chen, J.; Gao, X. Lipopolysaccharide preconditioning enhances the efficacy of mesenchymal stem cells transplantation in a rat model of acute myocardial infarction. *J. Biomed. Sci.* **2009**, *16*, 74. [CrossRef]

56. Bhatti, F.U.; Mehmood, A.; Latief, N.; Zahra, S.; Cho, H.; Khan, S.N.; Riazuddin, S. Vitamin E protects rat mesenchymal stem cells against hydrogen peroxide-induced oxidative stress *in vitro* and improves their therapeutic potential in surgically-induced rat model of osteoarthritis. *Osteoarthr. Cartil.* **2017**, *25*, 321–331. [CrossRef]

57. Li, M.; Yu, L.; She, T.; Gan, Y.; Liu, F.; Hu, Z.; Chen, Y.; Li, S.; Xia, H.; Xia, H. Astragaloside IV attenuates Toll-like receptor 4 expression via NF-kappaB pathway under high glucose condition in mesenchymal stem cells. *Eur. J. Pharm. Sci.* **2012**, *696*, 203–209. [CrossRef]

58. Lee, J.; Shin, M.S.; Kim, M.O.; Jang, S.; Oh, S.W.; Kang, M.; Jung, K.; Park, Y.S.; Lee, J. Apple ethanol extract promotes proliferation of human adult stem cells, which involves the regenerative potential of stem cells. *Nutr. Res.* **2016**, *36*, 925–936. [CrossRef]

59. Noiseux, N.; Borie, M.; Desnoyers, A.; Menaouar, A.; Stevens, L.M.; Mansour, S.; Danalache, B.A.; Roy, D.C.; Jankowski, M.; Gutkowska, J. Preconditioning of stem cells by oxytocin to improve their therapeutic potential. *Endocrinology* **2012**, *153*, 5361–5372. [CrossRef]

60. Yang, Y.; Choi, H.; Seon, M.; Cho, D.; Bang, S.I. LL-37 stimulates the functions of adipose-derived stromal/stem cells via early growth response 1 and the MAPK pathway. *Stem Cell Res.* **2016**, *7*, 58. [CrossRef]

61. Xia, W.; Zhang, F.; Xie, C.; Jiang, M.; Hou, M. Macrophage migration inhibitory factor confers resistance to senescence through CD74-dependent AMPK-FOXO3a signaling in mesenchymal stem cells. *Stem Cell Res.* **2015**, *6*, 82. [CrossRef]

62. Najafi, R.; Sharifi, A.M. Deferoxamine preconditioning potentiates mesenchymal stem cell homing *in vitro* and in streptozotocin-diabetic rats. *Expert Opin. Biol.* **2013**, *13*, 959–972. [CrossRef] [PubMed]

63. Carrero, R.; Cerrada, I.; Lledo, E.; Dopazo, J.; García-García, F.; Rubio, M.P.; Trigueros, C.; Dorronsoro, A.; Ruiz-Sauri, A.; Montero, J.A.; et al. IL1beta induces mesenchymal stem cells migration and leucocyte chemotaxis through NF-kappaB. *Stem Cell Rev.* **2012**, *8*, 905–916. [CrossRef] [PubMed]

64. Dubon, M.J.; Yu, J.; Choi, S.; Park, K.S. Transforming growth factor beta induces bone marrow mesenchymal stem cell migration via noncanonical signals and N-cadherin. *J. Cell Physiol.* **2017**, *233*, 201–213. [CrossRef] [PubMed]

65. Khan, I.; Ali, A.; Akhter, M.A.; Naeem, N.; Chotani, M.A.; Mustafa, T.; Salim, A. Preconditioning of mesenchymal stem cells with 2,4-dinitrophenol improves cardiac function in infarcted rats. *Life Sci.* **2016**, *162*, 60–69. [CrossRef]

66. Kim, Y.S.; Kwon, J.S.; Hong, M.H.; Kang, W.S.; Jeong, H.Y.; Kang, H.J.; Jeong, M.h.; Ahn, Y. Restoration of angiogenic capacity of diabetes-insulted mesenchymal stem cells by oxytocin. *BMC Cell Biol.* **2013**, *14*, 38. [CrossRef]

67. Liu, X.B.; Wang, J.A.; Ji, X.Y.; Yu, S.P.; Wei, L. Preconditioning of bone marrow mesenchymal stem cells by prolyl hydroxylase inhibition enhances cell survival and angiogenesis *in vitro* and after transplantation into the ischemic heart of rats. *Stem Cell Res.* **2014**, *5*, 111. [CrossRef]

68. Mias, C.; Lairez, O.; Trouche, E.; Roncalli, J.; Calise, D.; Seguelas, M.H.; Ordener, C.; Piercecchi-Marti, M.D.; Auge, N.; Salvayre, A.N.; et al. Mesenchymal stem cells promote matrix metalloproteinase secretion by cardiac fibroblasts and reduce cardiac ventricular fibrosis after myocardial infarction. *Stem Cells* **2009**, *27*, 2734–2743. [CrossRef]

69. Cornelissen, A.S.; Maijenburg, M.W.; Nolte, M.A.; Voermans, C. Organ-specific migration of mesenchymal stromal cells: Who, when, where and why? *Immunol. Lett.* **2015**, *168*, 159–169. [CrossRef]

70. Maijenburg, M.W.; Gilissen, C.; Melief, S.M.; Kleijer, M.; Weijer, K.; Ten Brinke, A.; Roelofs, H.; Van Tiel, C.M.; Veltman, J.A.; de Vries, C.J.; et al. Nuclear receptors Nur77 and Nurr1 modulate mesenchymal stromal cell migration. *Stem Cells Dev.* **2012**, *21*, 228–238. [CrossRef]

71. Andrzejewska, A.; Nowakowski, A.; Janowski, M.; Koniusz, S.; Walczak, P.; Grygorowicz, T.; Jablonska, A.; Lukomska, B. In vitro and in vivo functional studies on itga4 overexpressing human bone marrow mesenchymal stem cells. *J Regen Med.* **2015**, *4*, 4.

72. Pelagalli, A.; Nardelli, A.; Lucarelli, E.; Zannetti, A.; Brunetti, A. Autocrine signals increase ovine mesenchymal stem cells migration through aquaporin-1 and CXCR4 overexpression. *J. Cell Physiol.* **2018**, *233*, 6241–6249. [CrossRef] [PubMed]

73. De Becker, A.; Van Riet, I. Homing and migration of mesenchymal stromal cells: How to improve the efficacy of cell therapy? *World J. Stem Cells* **2016**, *8*, 73–87. [CrossRef] [PubMed]

74. Ullah, M.; Liu, D.D.; Thakor, A.S. Mesenchymal stromal cell homing: Mechanisms and strategies for improvement. *iScience* **2019**, *15*, 421–438. [CrossRef] [PubMed]

75. Lo, C.Y.; Weil, B.R.; Palka, B.A.; Momeni, A.; Canty, J.M.; Neelamegham, S. Cell surface glycoengineering improves selectin-mediated adhesion of mesenchymal stem cells (MSCs) and cardiosphere-derived cells (CDCs): Pilot validation in porcine ischemia-reperfusion model. *Biomaterials* **2016**, *74*, 19–30. [CrossRef] [PubMed]

76. Wang, H.; Wang, X.; Qu, J.; Yue, Q.; Hu, Y.; Zhang, H. VEGF enhances the migration of MSCs in neural differentiation by regulating focal adhesion turnover. *J. Cell Physiol.* **2015**, *230*, 2728–2742. [CrossRef] [PubMed]
77. Song, S.W.; Chang, W.; Song, B.W.; Song, H.; Lim, S.; Kim, H.J.; Cha, M.J.; Choi, E.; Im, S.H.; Chang, B.C.; et al. Integrin-linked kinase is required in hypoxic mesenchymal stem cells for strengthening cell adhesion to ischemic myocardium. *Stem Cells* **2009**, *27*, 1358-1365. [CrossRef]

78. Li, X.; He, L.; Yue, Q.; Lu, J.; Kang, N.; Xu, X.; Wang, H.; Zhang, H. MiR-9-5p promotes MSC migration by activating β-catenin signaling pathway. *Am. J. Physiol. Cell Physiol.* **2017**, *313*, C80–C93. [CrossRef]

79. Zeng, B.; Liu, L.; Wang, S.; Dai, Z. ILK regulates MSCs survival and angiogenesis partially through AKT and mTOR signaling pathways. *Acta HistoChem.* **2017**, *119*, 400–406. [CrossRef]

80. Mao, Q.; Liang, X.L.; Wu, Y.F.; Pang, Y.H.; Zhao, X.J.; Lu, Y.X. ILK promotes survival and self-renewal of hypoxic MSCs via the activation of lncTCF7-Wnt pathway induced by IL-6/STAT3 signaling. *Gene* **2019**, *26*, 165–176. [CrossRef]

81. He, H.; Zhao, Z.H.; Han, F.S.; Liu, X.H.; Wang, R.; Zeng, Y.J. Overexpression of protein kinase C ε improves retention and survival of transplanted mesenchymal stem cells in rat acute myocardial infarction. *Cell Death Dis.* **2016**, *7*, e2056. [CrossRef]

82. Heo, H.; Yoo, M.; Han, D.; Cho, Y.; Joung, I.; Kwon, Y.K. Upregulation of TrkB by forskolin facilitated survival of MSC and functional recovery of memory deficient model rats. *BioChem. Biophys. Res. Commun.* **2013**, *431*, 796–801. [CrossRef] [PubMed]

83. Xiang, Q.; Hong, D.; Liao, Y.; Cao, Y.; Liu, M.; Pang, J.; Zhou, J.; Wang, G.; Yang, R.; Wang, M.; et al. Overexpression of gremlin1 in mesenchymal stem cells improves hindlimb ischemia in mice by enhancing cell survival. *J. Cell Physiol.* **2017**, *232*, 996–1007. [CrossRef] [PubMed]

84. Han, S.M.; Han, S.H.; Coh, Y.R.; Jang, G.; Chan Ra, J.; Kang, S.K.; Lee, H.W.; Youn, H.Y. Enhanced proliferation and differentiation of Oct4- and Sox2-overexpressing human adipose tissue mesenchymal stem cells. *Exp. Mol. Med.* **2014**, *46*, e101. [CrossRef]

85. Li, Q.; Han, S.M.; Song, W.J.; Park, S.C.; Ryu, M.O.; Youn, H.Y. Anti-inflammatory effects of Oct4/Sox2-overexpressing human adipose tissue-derived mesenchymal stem cells. *Vivo* **2017**, *31*, 349–356. [CrossRef] [PubMed]

86. Jung, Y.H.; Lee, H.J.; Kim, J.S.; Lee, S.J.; Han, H.J. EphB2 signaling-mediated Sirt3 expression reduces MSC senescence by maintaining mitochondrial ROS homeostasis. *Free Radic. Biol. Med.* **2017**, *110*, 368–380. [CrossRef]

87. Agata, J.; Chao, L.; Chao, J. Kallikrein gene delivery improves cardiac reserve and attenuates remodeling after myocardial infarction. *Hypertension* **2002**, *40*, 653–659. [CrossRef]

88. Yao, Y.Y.; Yin, H.; Shen, B.; Smith, R.S.; Liu, Y.; Gao, L.; Chao, L.; Chao, J. Tissue kallikrein promotes neovascularization and improves cardiac function by the Akt-glycogen synthase kinase-3beta pathway. *Cardiovasc. Res.* **2008**, *80*, 354–364. [CrossRef]

89. Bledsoe, G.; Chao, L.; Chao, J. Kallikrein gene delivery attenuates cardiac remodeling and promotes neovascularization in spontaneously hypertensive rats. *Am. J. Physiol. Heart Circ. Physiol.* **2003**, *285*, H1479–H1488. [CrossRef]

90. Yao, Y.Y.; Yin, H.; Shen, B.; Chao, L.; Chao, J. Tissue kallikrein infusion prevents cardiomyocyte apoptosis, inflammation and ventricular remodeling after myocardial infarction. *Regul. Pept.* **2007**, *140*, 12–20. [CrossRef]

91. Devetzi, M.; Goulielmaki, M.; Khoury, N.; Spandidos, D.A.; Sotiropoulo, G.; Christodoulo, I.; Zoumpourlis, V. Genetically-modified stem cells in treatment of human diseases: Tissue kallikrein (KLK1)-based targeted therapy. *Int. J. Mol. Med.* **2018**, *41*, 1177–1186. [CrossRef]

92. Shujia, J.; Haider, H.K.; Idris, N.M.; Lu, G.; Ashraf, M. Stable therapeutic effects of mesenchymal stem cell-based multiple gene delivery for cardiac repair. *Cardiovasc. Res.* **2008**, *77*, 525–533. [CrossRef] [PubMed]

93. Li, W.; Ma, N.; Ong, L.L.; Nesselmann, C.; Klopsch, C.; Ladilov, Y.; Furlani, D.; Piechaczek, C.; Moebius, J.M.; Lützow, K.; et al. Bcl-2 engineered MSCs inhibited apoptosis and improved heart function. *Stem Cells* **2007**, *25*, 2118–2127. [CrossRef] [PubMed]

94. Grunewald, M.; Avraham, I.; Dor, Y.; Bachar-Lustig, E.; Itin, A.; Jung, S.; Chimenti, S.; Landsman, L.; Abramovitch, R.; Keshet, E. VEGF-induced adult neovascularization: Recruitment, retention, and role of accessory cells. *Cell* **2006**, *124*, 175–189. [CrossRef] [PubMed]

95. Tang, J.; Wang, J.; Yang, J.; Kong, X.; Zheng, F.; Guo, L.; Zhang, L.; Huang, Y. Mesenchymal stem cells over-expressing SDF-1 promote angiogenesis and improve heart function in experimental myocardial infarction in rats. *Eur. J. CardioThorac. Surg.* **2009**, *36*, 644–650. [CrossRef] [PubMed]

96. Tang, J.; Wang, J.; Zheng, F.; Kong, X.; Guo, L.; Yang, J.; Zhang, L.; Huang, Y. Combination of chemokine and angiogenic factor genes and mesenchymal stem cells could enhance angiogenesis and improve cardiac function after acute myocardial infarction in rats. *Mol. Cell BioChem.* **2010**, *339*, 107–118. [CrossRef] [PubMed]

97. Bao, C.; Guo, J.; Lin, G.; Hu, M.; Hu, Z. TNFR gene-modified mesenchymal stem cells attenuate inflammation and cardiac dysfunction following MI. *Scand. Cardiovasc. J.* **2008**, *42*, 56–62. [CrossRef]

98. Wen, Z.; Huang, W.; Feng, Y.; Cai, W.; Wang, Y.; Wang, X.; Liang, J.; Wani, M.; Chen, J.; Zhu, P.; et al. MicroRNA-377 regulates mesenchymal stem cell-induced angiogenesis in ischemic hearts by targeting VEGF. *PLoS ONE* **2014**, *9*, e104666.

99. Lundstrom, K. Viral vectors in gene therapy. *Diseases* **2018**, *6*, 42. [CrossRef]

100. Yin, H.; Kanasty, R.L.; Eltoukhy, A.A.; Vegas, A.J.; Dorkin, J.R.; Anderson, D.G. Non-viral vectors for gene-based therapy. *Nat. Rev. Genet.* **2014**, *15*, 541–555. [CrossRef]

101. Hardee, C.L.; Arévalo-Soliz, L.M.; Hornstein, B.D.; Zechiedrich, L. Advances in non-viral DNA vectors for gene therapy. *Genes* **2017**, *8*, 65. [CrossRef]

102. Suschak, J.J.; Williams, J.A.; Schmaljohn, C.S. Advancements in DNA vaccine vectors, non-mechanical delivery methods, and molecular adjuvants to increase immunogenicity. *Hum. Vaccin. Immunother.* **2017**, *13*, 2837–2848. [CrossRef] [PubMed]

103. Zhang, J.H.; Adikaram, P.; Pandey, M.; Genis, A.; Simonds, W.F. Optimization of genome editing through CRISPR-Cas9 engineering. *Bioengineered* **2016**, *7*, 166–174. [CrossRef] [PubMed]
104. Trams, E.G.; Lauter, C.J.; Salem, N.; Heine, U. Exfoliation of membrane ecto-enzymes in the form of micro-vesicles. *Biochim. Biophys. Acta* **1981**, *645*, 63–70. [CrossRef]

105. Potolicchio, I.; Carven, G.J.; Xu, X.; Stipp, C.; Riese, R.J.; Stern, L.J.; Santambrogio, L. Proteomic analysis of microglia-derived exosomes: Metabolic role of the aminopeptidase CD13 in neuropeptide catabolism. *J. Immunol.* **2005**, *175*, 2237–2243. [CrossRef]

106. Février, B.; Raposo, G. Exosomes: Endosomal-derived vesicles shipping extracellular messages. *Curr. Opin. Cell Biol.* **2004**, *16*, 415–421. [CrossRef]

107. Nikfarjam, S.; Rezaie, J.; Zolbanine, N.M.; Jafari, R. Mesenchymal stem cell derived-exosomes: A modern approach in translational medicine. *J. Transl. Med.* **2020**, *18*, 449. [CrossRef]

108. Loyer, X.; Vion, A.C.; Tedgui, A.; Boulanger, M. Microvesicles as cell-cell messengers in cardiovascular diseases. *Circ. Res.* **2014**, *114*, 345–353. [CrossRef]

109. Feng, Y.; Huang, W.; Wani, M.; Yu, X.; Ashraf, M. Ischemic preconditioning potentiates the protective effect of stem cells through secretion of exosomes by targeting Mecp2 via miR-22. *PLoS ONE* **2014**, *9*, e88685. [CrossRef]

110. Yu, B.; Gong, M.; Wang, Y.; Millard, R.W.; Pasha, Z.; Yang, Y.; Ashraf, M.; Xu, M. Cardiomyocyte protection by GATA-4 gene engineered mesenchymal stem cells is partially mediated by translocation of miR-221 in microvesicles. *PLoS ONE* **2013**, *8*, e73304. [CrossRef]

111. Yu, B.; Kim, H.W.; Gong, M.; Wang, J.; Millard, R.W.; Wang, Y.; Ashraf, M.; Xu, M. Exosomes secreted from GATA-4 overexpressing mesenchymal stem cells serve as a reservoir of anti-apoptotic microRNAs for cardioprotection. *Int. J. Cardiol.* **2015**, *182*, 349–360. [CrossRef]

112. Wang, N.; Chen, C.; Yang, D.; Liao, Q.; Luo, H.; Wang, X.; Zhou, F.; Yang, X.; Yang, J.; Zeng, C.; et al. Mesenchymal stem cells-derived extracellular vesicles, via miR-210, improve infarcted cardiac function by promotion of angiogenesis. *Biochim. Biophys. Acta Mol. Basis Dis.* **2017**, *1863*, 2085–2092. [CrossRef] [PubMed]

113. De Abreu, R.C.; Fernandes, H.; Da Costa Martins, P.A.; Sahoo, S.; Emanueli, C.; Ferreira, L. Native and bioengineered extracellular vesicles for cardiovascular therapeutics. *Nat. Rev. Cardiol.* **2020**, *17*, 685–697. [CrossRef] [PubMed]

114. Barkholt, L.; Flory, E.; Jekerle, V.; Lucas-Samuel, S.; Ahnert, P.; Bisset, L.; Büscher, D.; Fibbe, W.; Foussat, A.; Kwa, M.; et al. Risk of tumorigenicity in mesenchymal stromal-cell based therapies-bridging scientific observations and regulatory viewpoints. *Cytotherapy* **2013**, *15*, 753–759. [CrossRef] [PubMed]

115. Zhang, J. Engineered tissue patch for cardiac cell therapy. *Curr. Treat. Options Cardiovasc.* **2015**, *17*, 399. [CrossRef]

116. Zhang, J.; Zhu, W.; Radisic, M.; Vunjak-Novakovic, G. Can we engineer a human cardiac patch for therapy? *Circ. Res.* **2018**, *123*, 244–265. [CrossRef]

117. Zhou, P.; Pu, W.T. Recounting cardiac cellular composition. *Circ. Res.* **2016**, *118*, 368–370. [CrossRef]

118. Fleischer, S.; Vunjak-Novakovic, G. Cardiac tissue engineering: From repairing to modeling the human heart. In *Encyclopedia of Tissue Engineering and Regenerative Medicine*, 1st ed.; Academic Press: Oxford, UK, 2019; pp. 131–144.

119. Chong, J.J.; Yang, X.; Don, C.W.; Minami, E.; Liu, Y.W.; Weyers, J.J.; Mahoney, W.M.; Van Biber, B.; Cook, S.M.; Palpant, N.J.; et al. Human embryonic-stem-cell-derived cardiomyocytes regenerate non-human primate hearts. *Nature* **2014**, *510*, 273–277. [CrossRef]

120. Ye, L.; Chang, Y.H.; Xiong, Q.; Zhang, P.; Zhang, L.; Somasundaram, P.; Lepley, M.; Swingen, C.; Su, L.; Wendel, J.S.; et al. Cardiac repair in a porcine model of acute myocardial infarction with human induced pluripotent stem cell-derived cardiovascular cells. *Cell Stem Cell* **2014**, *15*, 750–761. [CrossRef]

121. Weinberger, F.; Breckwoldt, K.; Pecha, S.; Kelly, A.; Geertz, B.; Starbatty, J.; Yorgan, T.; Cheng, K.H.; Lessmann, K.; Stolen, T.; et al. Cardiac repair in guinea pigs with human engineered heart tissue from induced pluripotent stem cells. *Sci. Transl. Med.* **2016**, *8*, 363. [CrossRef]

122. Shiba, Y.; Fernandes, S.; Zhu, W.Z.; Filice, D.; Muskheli, V.; Kim, J.; Palpant, N.J.; Gantz, J.; Moyes, K.W.; Reinecke, H.; et al. Human ES-cell-derived cardiomyocytes electrically couple and suppress arrhythmias in injured hearts. *Nature* **2012**, *489*, 322–325. [CrossRef]

123. Wei, K.; Serpooshan, V.; Hurtado, C.; Diez-Cunado, M.; Zhao, M.; Maruyama, S.; Zhu, W.; Fajardo, G.; Noseda, M.; Nakamura, K.; et al. Epicardial FSTL1 reconstitution regenerates the adult mammalian heart. *Nature* **2015**, *525*, 479–485. [CrossRef] [PubMed]

124. Serpooshan, V.; Zhao, M.; Metzler, S.A.; Wei, K.; Shah, P.B.; Wang, A.; Mahmoudi, M.; Malkovskiy, A.V.; Rajadas, J.; Butte, M.J.; et al. The effect of bioengineered acellular collagen patch on cardiac remodeling and ventricular function post myocardial infarction. *Biomaterials* **2013**, *34*, 9048–9055. [CrossRef] [PubMed]

125. Masumoto, H.; Nakane, T.; Tinney, J.P.; Yuan, F.; Ye, F.; Kowalski, W.J.; Minakata, K.; Sakata, R.; Yamashita, J.K.; Keller, B.B. The myocardial regenerative potential of three-dimensional engineered cardiac tissues composed of multiple human iPS cell-derived cardiovascular cell lineages. *Sci. Rep.* **2016**, *6*, 29933. [CrossRef] [PubMed]

126. Wang, L.; Liu, Y.; Ye, G.; He, Y.; Li, B.; Guan, Y.; Gong, B.; Mequanint, K.; Xing, M.M.Q.; Qiu, X. Injectable and conductive cardiac patches repair infarcted myocardium in rats and minipigs. *Nat. Biomed. Eng.* **2021**, *5*, 1157–1173. [CrossRef]

127. Jiang, Y.; Sun, S.J.; Zhen, Z.; Wei, R.; Zhang, N.; Liao, S.Y.; Tse, H.F. Myocardial repair of bioengineered cardiac patches with decellularized placental scaffold and human-induced pluripotent stem cells in a rat model of myocardial infarction. *Stem Cell Res.* **2021**, *12*, 13. [CrossRef]

128. Wang, L.; Serpooshan, V.; Zhang, J. Engineering human cardiac muscle patch constructs for prevention of post-infarction LV remodeling. *Front. Cardiovasc. Med.* **2021**, *8*, 621781. [CrossRef]
129. Liu, Y.W.; Chen, B.; Yang, X.; Fugate, J.A.; Kalucki, F.A.; Futakuchi-Tsuchida, A.; Couture, L.; Vogel, K.W.; Astley, C.A.; Baldessari, A.; et al. Human embryonic stem cell-derived cardiomyocytes restore function in infarcted hearts of non-human primates. *Nat. Biotechnol.* **2018**, *36*, 597–605. [CrossRef]

130. Shiba, Y.; Gomibuchi, T.; Seto, T.; Wada, Y.; Ichimura, H.; Tanaka, Y.; Ogasawara, T.; Okada, K.; Shiba, N.; Sakamoto, K.; et al. Allogeneic transplantation of iPS cell-derived cardiomyocytes regenerates primate hearts. *Nature* **2016**, *538*, 388–391. [CrossRef]

131. Romagnuolo, R.; Masoudpour, H.; Porta-Sánchez, A.; Qiang, B.; Barry, J.; Laskary, A.; Qi, X.; Massé, S.; Magtibay, K.; Kawajiri, H.; et al. Human embryonic stem cell-derived cardiomyocytes regenerate the infarcted pig heart but induce ventricular tachyarrhythmias. *Stem Cell Rep.* **2019**, *12*, 967–981. [CrossRef]

132. Jabbour, R.J.; Owen, T.J.; Pandey, P.; Reinsch, M.; Wang, B.; King, O.; Couch, L.S.; Pantou, D.; Pitcher, D.S.; Chowdhury, R.A.; et al. In vivo grafting of large engineered heart tissue patches for cardiac repair. *JCI Insight* **2021**, *6*, e144068. [CrossRef]

133. Guo, Y.; Yu, Y.; Hu, S.; Chen, Y.; Shen, Z. The therapeutic potential of mesenchymal stem cells for cardiovascular diseases. *Cell Death Dis.* **2020**, *11*, 349. [CrossRef] [PubMed]

134. Perez-Estenaga, I.; Prosper, F.; Pelacho, B. Allogeneic Mesenchymal Stem Cells and Biomaterials: The Perfect Match for Cardiac Repair? *Int. J. Mol. Sci.* **2018**, *19*, 3236. [CrossRef] [PubMed]

135. Araña, M.; Gavira, J.J.; Peña, E.; Gonzalez, A.; Abizanda, G.; Cilla, M.; Pérez, M.M.; Albiasu, E.; Aguado, N.; Casado, M.; et al. Epicardial delivery of collagen patches with adipose-derived stem cells in rat and minipig models of chronic myocardial infarction. *Biomaterials* **2014**, *35*, 143–151. [CrossRef] [PubMed]

136. Chachques, J.C.; Trainini, J.C.; Lago, N.; Cortes-Morichetti, M.; Schussler, O.; Carpentier, A. Myocardial assistance by grafting a new bioartificial upgraded myocardium (MAGNUM trial): Clinical feasibility study. *Ann. Thorac. Sur.* **2008**, *85*, 901–908. [CrossRef]

137. Miyagawa, S.; Kainuma, S.; Kawamura, T.; Suzuki, K.; Ito, Y.; Iseoka, H.; Ito, E.; Takeda, M.; Sasai, M.; Mochizuki-Oda, N.; et al. Transplantation of IPSC-derived cardiomyocyte patches for ischemic cardiomyopathy. *BMJ* **2022**, *2021*, 12. [CrossRef]

138. Menasché, P.; Vanneaux, V.; Hagège, A.; Bel, A.; Cholley, B.; Parouchev, A.; Cacciapuoti, I.; Al-Daccak, R.; Benhamouda, N.; Blons, H.; et al. Transplantation of Human Embryonic Stem Cell-Derived Cardiovascular Progenitors for Severe Ischemic Left Ventricular Dysfunction. *J. Am. Coll Cardiol.* **2018**, *71*, 429–438. [CrossRef]

139. Bolli, R.; Solankhi, M.; Tang, X.L.; Kahlon, A. Cell therapy in patients with heart failure: A comprehensive review and emerging concepts. *Cardiovasc. Res.* **2022**, *118*, 951–976. [CrossRef]

140. Arjmand, B.; Abedi, M.; Arabi, M.; Alavi-Moghadam, S.; Rezaei-Tavirani, M.; Hadavandkhani, M.; Tayanloo-Beik, A.; Kordi, R.; Roudsari, P.P.; Larijani, B. Regenerative medicine for the treatment of ischemic heart disease; status and future perspectives. *Front. Cell Dev. Biol.* **2021**, *9*, 704903. [CrossRef]

141. Chang, D.; Fan, T.; Gao, S.; Jin, Y.; Zhang, M.; Ono, M. Application of mesenchymal stem cell sheet to treatment of ischemic heart disease. *Stem Cell Res.* **2021**, *12*, 384. [CrossRef]

142. Panès, J.; Garcia-Olmo, D.; Assche, G.V.; Colombel, J.F.; Reinisch, W.; Baumgart, D.C.; Dignass, A.; Nachury, M.; Ferrante, M.; Kazemi-Shirazi, L.; et al. ADMIRE CD Study Group Collaborators. Long-term efficacy and safety of stem cell therapy (Cx601) for complex perianal fistulas in patients with Crohn's disease. *Gastroenterology* **2018**, *154*, 1334–1342. [CrossRef]

143. Sun, Y.; Wan, B.; Wang, R.; Zhang, B.; Luo, P.; Wang, D.; Nie, J.J.; Chen, D.; Wu, X. Mechanical stimulation on mesenchymal stem cells and surrounding microenvironments in bone regeneration: Regulations and applications. *Front. Cell Dev. Biol.* **2022**, *10*, 808303. [CrossRef] [PubMed]