Abstract 209: Exosomes Derived From c-kit+ Cardiac Progenitor Cells are Essential for Myocardial Repair After Acute Myocardial Function

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Sudhish Sharma ◽  
Grace E Bigham ◽  
Rachana Mishra ◽  
Flaviu Gruia ◽  
Philip Z Brohawn ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Progenitor Cells ◽  
Left Ventricular ◽  
Biologically Active ◽  
Neonatal Rat ◽  
Right Atrial ◽  
Cardiac Progenitor Cells ◽  
Myocardial Repair ◽  
Paracrine Factors ◽  
Rat Cardiomyocytes

Background: Human cardiac progenitor cells (hCPCs), identified by ckit + /CD45 - , provide a promising therapeutic option following myocardial infarction (MI) as their clinical relevance has been validated in the S tem C ell I nfusion in P atients with I schemic Cardi o myopathy (SCIPIO) Phase I clinical trial. The mechanism for their functional recovery of the injured myocardium is unknown. Hypothesis: We hypothesized whether CPCs secrete biologically active exosomes and if these exosomes could provide cardioprotection after myocardial infarction (MI). Methods and results: Exosomes were isolated from cultured CPCs, generated from the biopsies of right atrial appendage (RAA) from neonatal (nCPCs) and adult (aCPCs) patients with normal functioning myocardium. TEM showed that both CPCs secrete microvesicles, which fall within the same size range as exosomes (80-170nM, diameter). FACS performed for canonical exosomal surface markers CD63, ALIX and CD9 confirmed the presence of exosomes in the secretome of CPCs. Quantification of exosomes by Nanosight NS300 showed that nCPCs produce more than twice the amount of exosomes as compared to aCPCs in 48 hours. Exosomes were internalized by cardiomyocytes, endothelial cells and fibroblasts, within the myocardium. CPCs derived exosomes enhanced angiogenesis as analyzed by HUVEC tube assay formation and proliferation of neonatal rat cardiomyocytes while inhibiting their apoptosis in the presence of oxidative stress and inflammation. Intra-myocardial injection of exosomes into rat myocardium after MI restored ejection fraction (CPCs 63.74±3.68% vs CPCs-exosomes 62 ± 2.97%), attenuated adverse left ventricular remodeling and reduced infarct size which were comparable to CSC therapy at 28 days post MI. CPC exosomes also contain distinctive cargo of miRs and proteins. Immunoblot analysis shows that CPC exosomes are enriched in the paracrine factors VEGFA, ANG1, SCF1 and HGF1, with cardioprotective roles. Conclusion: Our findings identify exosomes as the smallest functional unit and potential biomarkers of CPC therapy. CPCs derived exosomes can be utilized as an off the shelf cell-free therapy which eliminates several shortcomings of cell therapy, including cell retention, cell rejection and arrhythmia.

Abstract 855: Circulating Mesoangioblasts Differentiate Into Cardiac Myocytes And Improve Function After Acute Myocardial Infarction

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Masayoshi Iwasaki ◽  
Masamichi Koyanagi ◽  
Stefan Rapp ◽  
Corina Schuetz ◽  
Philipp Bushoven ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Cardiac Function ◽  
Troponin T ◽  
Marker Gene ◽  
Left Ventricular ◽  
Cardiac Differentiation ◽  
Neonatal Rat ◽  
Stem Cell Markers ◽  
Rt Pcr ◽  
Rat Cardiomyocytes

Mesoangioblasts (MAB) are vessel-associated cells identified during embryonic development. In contrast to hemangioblasts, MAB express mesenchymal (CD73) and endothelial marker, but lack the hematopoietic marker CD45. We recently identified circulating MAB in children. Children-derived MAB showed vigorous proliferation capacity and high telomerase activity. However, the potential of cardiac differentiation in these cells was not elucidated. Therefore, we tested the capacity of children-derived MAB to aquire a cardiomyogenic phenotype. MAB expressed several cardiac transcription factors such as Nkx2.5, GATA4 and MEF2C and the stem cell markers c-kit and islet-1. In order to assess cardiac differentiation capacity, we performed co-culture assays with neonatal rat cardiomyocytes (CM). Immunochemical analysis revealed that MAB expressed cardiac α-sarcomeric actinin 6 days after co-culture. Moreover, human troponin T (TnT) was expressed as demonstrated by human specific RT-PCR. To confirm these data, we examined TnT expression in MAB isolated of a 2 years old patient with a known mutation of TnT. Sequences of the cloned RT-PCR products were identical to human TnT except for the known mutation providing genetic proof of concept for cardiac differentiation. In order to exclude fusion between MAB and CM as a mechanism, we used paraformaldehyde-fixed CM as scaffold. In this assay, human TnT also was detected, indicating that differentiation is sufficient to induce cardiac marker gene expression. Next, we tested the effect of MAB to improve cardiac function. MAB were injected intramuscularly in nude mice after myocardial infarction. Functional analysis using Millar catheter 2 weeks after infarction demonstrated that cell therapy lowered filling pressure and preserved diastolic function when compared to the PBS injected group (LVEDP: −20.3%, tau: −20.6%, vs PBS injected heart). Furthermore, left ventricular volume was also decreased (LVEDV/weight −27.3%). In summary, children-derived MAB express cardiac-specific genes after co-culture with CM and improved cardiac function in vivo. Given that MAB can be easily isolated and expanded from peripheral blood, these cells might be suitable to augment cardiac repair in children with heart failure.

Abstract 4: Lymphatic Vessels Mediate the Mobilization of Cardiac Progenitor Cells after Myocardial Infarction

2014 ◽  
Vol 115 (suppl_1) ◽  
Author(s):  
Polina Goichberg ◽  
Maria Cimini ◽  
Antonio Cannata ◽  
Sergio Signore ◽  
Kanako Waight ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Progenitor Cells ◽  
Heart Diseases ◽  
Lymphatic Vessels ◽  
Mouse Heart ◽  
Cardiac Progenitor Cells ◽  
Myocardial Repair ◽  
Lymphatic Circulation ◽  
Lymphatic Vasculature

The delivery of adult cardiac progenitor cells (CPCs) or their activation in situ constitute an evolving approach for the treatment of heart failure. CPCs are endowed with regenerative capacity, producing differentiating myocytes and vascular structures in the course of homeostasis and upon injury. The regenerative function of CPCs is contingent to their ability to migrate to and engraft within the wounded area. Yet, the mechanisms governing CPC trafficking in the diseased myocardium are largely unknown. The lymphatic system is vital for tissue repair, and the role of the lymphatic vasculature in the trafficking of hematopoietic and cancer cells is well documented. We examined whether cardiac lymphatic vessels mediate the translocation of CPCs in the infarcted myocardium. By imaging of the heart from transgenic c-kit-GFP reporter mice, we found that as early as 4 hours after myocardial infarction (MI), uncommitted lineage-negative progenitors accumulated in the vicinity of the lymphatic vessels located in the region bordering the necrotic area. Histologically, extensive lymphangiogenesis was documented in the mouse heart in the acute (8-48 hours) and chronic (15-35 days) phases of infarct healing and scar formation. CPCs were detected traversing the wall of lymphatic vessels at different stages after MI, indicative of the functional role of the lymphatic circulation in the recruitment of primitive cells to the site of injury. Furthermore, isolated human CPCs exhibited chemotaxis and specific binding to the human lymphatic endothelial cells (LECs) in steady-state conditions and, increasingly, after exposure to an inflammatory cytokine, TNFα. CPCs performed trans-endothelial migration in vitro, and actively intravasated into the lumen of microvessels formed by LECs in three-dimensional matrices. Finally, our data suggest that sphingosine-1-phosphate (S1P)-stimulated signaling governs the interactions of CPCs with LECs. These findings on the direct role of lymphatic vasculature in CPC trafficking may contribute to the development of novel therapeutic modalities to increase mobilization of endogenous or transplanted CPCs, promoting myocardial repair in patients with ischemic heart diseases.

Abstract 19220: Potential of Human Cardiac Progenitor Cells from Young versus Old Donor Hearts in Cardiac Protection against Ischemic Left Ventricular Dysfunction and Late Remodeling

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Ann Sophie Walravens ◽  
Ellen Caluwé ◽  
Sander Trenson ◽  
Hilde Gillijns ◽  
Nina Vanden Driessche ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Left Ventricular Dysfunction ◽  
Ventricular Dysfunction ◽  
Left Ventricular ◽  
Border Zone ◽  
Coronary Artery Ligation ◽  
Cardiac Progenitor Cells ◽  
Myocardial Repair ◽  
Old Patients ◽  
Expression Levels

Introduction: Recent discovery of c-Kit + resident cardiac progenitor cells (CPC) has demonstrated cardiac regenerative capacity, which is lost upon increasing age in mice. Accordingly, we hypothesized that c-KIT + -selected CPC from young human hearts (yCPC) have a greater myocardial repair potential after myocardial infarction (MI) than CPC from old patients (oCPC). Methods: After Ethics Committee approval, cardiac biopsies from young (1-15y old) and old (58-80y old) patients, undergoing corrective or valve surgery were obtained. After collagenase-based digestion, cells were culture-expanded for 2 passages prior to c-KIT + selection using magnetic beads. Purity was confirmed using qRT-PCR for c-KIT, GATA4, CD90, CD105, DDR2, CD31 and MYH11 expression levels. To trace transplanted cells, CPC were lentivirally tagged with GFP. After permanent LAD coronary artery ligation in immunodeficient mice, 250,000 CPCs were injected in the infarct border zone (4x 62,500 cells/2.5 μl; yCPC, n =8; oCPC, n =6; or PBS, n =6). Cardiac ultrasound (VisualSonics, 3D 30 MHz probe) was performed at baseline, 1 and 35 days after MI. At 35 days post-MI, infarct remodeling and cell survival was evaluated using Sirius red and CPC engraftment by anti-GFP staining. Results: In contrast to low MYH11 and CD31 expression levels, c-KIT and GATA4 were markedly and equally expressed in yCPC and oCPC, whereas transcript levels of DDR2, CD105 and CD90 were 89, 109 and 56% higher in oCPC compared to yCPC. Systolic function was better preserved in yCPC than in oCPC and PBS-treated mice 24h after MI (LVEF from baseline - D1: 53±1 - 38±3 vs. 52±3 - 30±5 vs. 50±3 - 27±4%; ESVi from baseline - D1: 3.7±0.2 - 5.1±0.4 vs. 3.8±0.5 - 6.2±0.7 vs. 3.9±0.5 - 6.5±0.6ml/mm 2 ). Moreover, after 35 days, the dilatory response was reduced in yCPC compared to oCPC and PBS-treated mice (EDVi from D1 - D35: 8.3±0.5 - 13.5±1.3 vs. 8.8±0.5 - 16.7±1.8 vs. 8.5±0.5 - 16.6±1.3ml/mm 2 ). Improved remodeling was consistent with a markedly greater rim of surviving myocardium while anti-GFP staining showed only rarely retained CPC. Conclusions: In summary, these findings suggest enhanced protection by c-KIT + CPC isolated from young donor hearts against acute systolic left ventricular dysfunction and late remodeling after MI.

Abstract 5569: Paracrine Effects of Cardiomyocyte STAT3 Determine the Vasculogenic Differentiation Potential of Cardiac Progenitor Cells: Potential Role of Endogenous Erythropoietin

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Philipp Fischer ◽  
Melanie Hoch ◽  
Britta Stapel ◽  
Helmut Drexler ◽  
Denise Hilfiker-Kleiner
Keyword(s):  
Progenitor Cells ◽  
Adipocyte Differentiation ◽  
Left Ventricular ◽  
Prostaglandin E ◽  
Cardiac Progenitor Cells ◽  
Rt Pcr ◽  
Paracrine Factors ◽  
Differentiation Potential ◽  
Paracrine Effects

Mice with a cardiomyocyte-restricted knock out of STAT3 (αMHC-Cretg/+; STAT3flox/flox, STAT3-KO) show a continuous decrease of cardiac vascularization and develop heart failure beyond the age of 9 months. We investigated the role of cardiomyocyte STAT3-driven paracrine effects on Sca-1+ cardiac progenitor cells (CPC) in the mouse heart. CPC were immunomag-netically isolated from KO and wildtype (STAT3flox/flox, WT) hearts (age: 3 months). PCR and Western blot confirmed deletion of STAT3 in cardiomyocytes of KO mice, while CPC from KO showed normal expression of STAT3. The total number of CPC per heart was similar between WT and KO mice. FACS analysis revealed a reduced number of endothelial progenitor cells (defined by co-expression of Sca-1, CD31 and CD38, −25%, P<0.05) in CPC from KO compared to CPC from WT. In vitro culture for 4 weeks on fibronectin-coated plates of CPC from KO revealed reduced proliferation (−33%, p<0.01), impaired endothelial cell (EC) tubeformation (monitored with Tie2, eNOS and CD31 immunohistochemistry (IHC), p<0.01) and enhanced adipocyte differentiation (oil red staining and RT-PCR, p<0.05) compared with CPC from WT. Microarray of freshly isolated CPC reflected this differences in EC and adipocyte differentiation on the mRNA level (i.e. EC marker Prostaglandin E Rezeptor-3: 2.3-fold lower; adipocyte marker Lipocalin-2 2.7-fold higher in CPC from KO hearts). Microarray results from whole left ventricular tissue showed a decrease in gene expression of Erythropoietin (Epo) in KO hearts (-9,25-fold). ELISA, IHC and methylcellulose assay confirm expression of active EPO by cardiomyocytes. CPC express high levels of EPO receptor (IHC, RT-PCR). Epo enhanced tube formation and sprouting of EC and attenuated adipocyte differentiation of CPCs from KO. In vivo treatment with Epo rescued impaired proliferation, promoted EC differentiation and attenuated adipocyte differentiation of CPC from KO hearts. Conclusion: STAT3-dependent paracrine factors from cardiomyocytes regulate proliferation, differentiation and vasculogenic properties of CPCs. Cardiomyocyte derived EPO is an important paracrine mediator that promotes differentiation into EC and attenuates differentiation into adipocytes from CPCs in the adult heart.

scholarly journals LRRK2 Deficiency Protects Hearts from Myocardial Infarction Injury in Mice via the P53/HMGB1 Pathway

2021 ◽  
Author(s):  
Yuan Liu ◽  
Changgui Chen ◽  
Lu Chen ◽  
Xiaoxin Pei ◽  
Zekai Tao ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Left Ventricular ◽  
Neonatal Rat ◽  
Coronary Artery Ligation ◽  
Wild Type ◽  
Artery Ligation ◽  
Rat Cardiomyocytes ◽  
Ventricular Fibrosis ◽  
Deficient Mice

Abstract Purpose: LRRK2 is a Ser/Thr kinase with multiple functional domains. Current studies have shown that its mutations are closely related to hereditary Parkinson's disease. However, its role in cardiovascular disease, especially in myocardial infarction, is unclear. The aim of this study was to explore the functional role of LRRK2 in myocardial infarction. Methods: Wild-type and LRRK2 knockout mice were subjected to coronary artery ligation (left anterior descent) to establish a myocardial infarction mouse model. Neonatal rat cardiomyocytes were subjected to hypoxia to induce hypoxia injury in vitro. Results: We found increased LRRK2 expression levels in the infarct periphery of mouse hearts and hypoxic cardiomyocytes. LRRK2-deficient mice exhibited a decreased death rate and reduced infarction area compared to the wild-type controls 14 days after infarction. LRRK2-deficient mice showed reduced left ventricular fibrosis and inflammatory response, as well as improved cardiac function. In the in vitro study, LRRK2 silencing decreased the cleaved-caspase3 activity, reduced cardiomyocyte apoptosis, and diminished hypoxia-induced inflammation. However, LRRK2 overexpression enhanced the cleaved-caspase3 activity, increased the number of apoptotic cardiomyocytes, and caused remarkable hypoxia-induced inflammation. When exploring the related underlying mechanisms, we found that hypoxia induced an increase in HIFα expression, which enhanced LRRK2 expression. LRRK2 induced high expression of HMGB1 via P53. When blocking HMGB1 using the anti-HMGB1 antibody, the deteriorating effects caused by LRRK2 overexpression following hypoxia were inhibited in cardiomyocytes.Conclusions: In summary, LRRK2 deficiency protects hearts from myocardial infarction injury. The mechanism underlying this phenomenon involves the P53-HMGB1 pathway.

Abstract 1845: Adult Cardiac Progenitor Cells Promote Angiogenesis and Cardioprotection through Their Secreted Soluble Vcam-1

Circulation ◽  
2008 ◽  
Vol 118 (suppl_18) ◽  
Author(s):  
Katsuhisa Matsuura ◽  
Atsushi Honda ◽  
Toshio Nagai ◽  
Noritoshi Fukushima ◽  
Tatsuya Shimizu ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Cell Injury ◽  
Cell Sheet ◽  
Fold Increase ◽  
Neonatal Rat ◽  
Antibody Array ◽  
Cardiac Progenitor Cells ◽  
Protective Effects ◽  
Rat Cardiomyocytes ◽  
Promising Source

Although cardiac progenitor cells have been thought to be the promising source of cell therapy, the precise mechanisms of their paracrine action have not been fully elucidated. Since we observed that the transplantation of clonal expanded Sca-1 positive cardiac progenitor cells (cSca-1 cells) derived from adult murine heart by using cell sheet technique improved cardiac function of infarcted heart compared to adipose tissue derived mesenchymal cells (ATMC), we explored the secreted factors highly expressed in cSca-1 cells and identified that soluble VCAM-1 (sVCAM-1) was much abundant in cSca-1 cells compared to ATMC by using cytokine antibody array. cSca-1 cells-derived conditioned medium (CM) significantly enhanced endothelial migration and matrigel tube formation and these effects were abolished by knock down of VCAM-1 (Fold increase: control, 1.0; CM, 2.97 ± 0.21; siVCAM-1, 1.98 ± 0.09; siControl, 2.76 ± 0.05, p<0.01), suggesting that cSca-1 cells promote angiogenesis via their secreted sVCAM-1. We next examined whether sVCAM-1 conferred direct protective effects on cardiomyocytes. We exposed cardiomyocytes to 0.2 mM H 2 O 2 in the absence or presence of sVCAM-1 or CM and examined cardiomyocyte viability by MTT assay. The exposure of cardiomyocytes to H 2 O 2 significantly induced the cell injury. Interestingly when pretreated with sVCAM-1 or CM, the cell damages of cardiomyocytes by H 2 O 2 were significantly reduced. However when pretreated with anti-VLA4 antibody, a principal coreceptor of sVCAM-1, CM mediated cell protected effect was completely inhibited (Fold increase: control, 1.0; anti-VLA4, 0.89 ± 0.33; sVCAM-1, 1.69 ± 0.27; CM, 2.08 ± 0.28; CM+anti-VLA4, 1.07 ± 0.07, p<0.01), suggesting that a crucial role of the VLA4 in inducing survival of cardiomyocytes by CM. sVCAM-1 and CM induced phosphorylation of FAK, Akt, Erk and p38 MAPK in neonatal rat cardiomyocytes. When pretreated with wortmannin, SB203580 and PD98059, the cardioprotective effects of sVCAM-1 and CM significantly inhibited, suggesting that sVCAM-1 might protect cardiomyocytes from oxidative stress via integrated upregulation of Akt, Erk and p38MAPK. These findings suggest cardiac progenitor cells promote angiogenesis and cardioprotection through their secreted sVCAM-1.

scholarly journals Novel Identified Circular Transcript of RCAN2, circ-RCAN2, Shows Deviated Expression Pattern in Pig Reperfused Infarcted Myocardium and Hypoxic Porcine Cardiac Progenitor Cells In Vitro

2021 ◽  
Vol 22 (3) ◽  
pp. 1390
Author(s):  
Julia Mester-Tonczar ◽  
Patrick Einzinger ◽  
Johannes Winkler ◽  
Nina Kastner ◽  
Andreas Spannbauer ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Progenitor Cells ◽  
Regulatory Networks ◽  
Circular Rnas ◽  
Cardiac Progenitor Cells ◽  
Tissue Samples ◽  
Healthy Myocardium ◽  
Acute Mi

Circular RNAs (circRNAs) are crucial in gene regulatory networks and disease development, yet circRNA expression in myocardial infarction (MI) is poorly understood. Here, we harvested myocardium samples from domestic pigs 3 days after closed-chest reperfused MI or sham surgery. Cardiac circRNAs were identified by RNA-sequencing of rRNA-depleted RNA from infarcted and healthy myocardium tissue samples. Bioinformatics analysis was performed using the CIRIfull and KNIFE algorithms, and circRNAs identified with both algorithms were subjected to differential expression (DE) analysis and validation by qPCR. Circ-RCAN2 and circ-C12orf29 expressions were significantly downregulated in infarcted tissue compared to healthy pig heart. Sanger sequencing was performed to identify the backsplice junctions of circular transcripts. Finally, we compared the expressions of circ-C12orf29 and circ-RCAN2 between porcine cardiac progenitor cells (pCPCs) that were incubated in a hypoxia chamber for different time periods versus normoxic pCPCs. Circ-C12orf29 did not show significant DE in vitro, whereas circ-RCAN2 exhibited significant ischemia-time-dependent upregulation in hypoxic pCPCs. Overall, our results revealed novel cardiac circRNAs with DE patterns in pCPCs, and in infarcted and healthy myocardium. Circ-RCAN2 exhibited differential regulation by myocardial infarction in vivo and by hypoxia in vitro. These results will improve our understanding of circRNA regulation during acute MI.

scholarly journals Adropin-based dual treatment enhances the therapeutic potential of mesenchymal stem cells in rat myocardial infarction

2021 ◽  
Vol 12 (6) ◽  
Author(s):  
HuiYa Li ◽  
DanQing Hu ◽  
Guilin Chen ◽  
DeDong Zheng ◽  
ShuMei Li ◽  
...  
Keyword(s):  
Myocardial Infarction ◽  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Therapeutic Potential ◽  
Left Ventricular ◽  
Left Ventricular Ejection ◽  
Paracrine Factors ◽  
Dual Treatment

AbstractBoth weak survival ability of stem cells and hostile microenvironment are dual dilemma for cell therapy. Adropin, a bioactive substance, has been demonstrated to be cytoprotective. We therefore hypothesized that adropin may produce dual protective effects on the therapeutic potential of stem cells in myocardial infarction by employing an adropin-based dual treatment of promoting stem cell survival in vitro and modifying microenvironment in vivo. In the current study, adropin (25 ng/ml) in vitro reduced hydrogen peroxide-induced apoptosis in rat bone marrow mesenchymal stem cells (MSCs) and improved MSCs survival with increased phosphorylation of Akt and extracellular regulated protein kinases (ERK) l/2. Adropin-induced cytoprotection was blocked by the inhibitors of Akt and ERK1/2. The left main coronary artery of rats was ligated for 3 or 28 days to induce myocardial infarction. Bromodeoxyuridine (BrdU)-labeled MSCs, which were in vitro pretreated with adropin, were in vivo intramyocardially injected after ischemia, following an intravenous injection of 0.2 mg/kg adropin (dual treatment). Compared with MSCs transplantation alone, the dual treatment with adropin reported a higher level of interleukin-10, a lower level of tumor necrosis factor-α and interleukin-1β in plasma at day 3, and higher left ventricular ejection fraction and expression of paracrine factors at day 28, with less myocardial fibrosis and higher capillary density, and produced more surviving BrdU-positive cells at day 3 and 28. In conclusion, our data evidence that adropin-based dual treatment may enhance the therapeutic potential of MSCs to repair myocardium through paracrine mechanism via the pro-survival pathways.

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