Abstract 1115: Impaired Angiogenic Potential of Cardiac Progenitor Cells in Mice with a Cardiomyocyte-Restricted Knock Out of STAT3

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Philipp Fischer ◽  
Ewa Missol-Kolka ◽  
Nils-Holger Zschemisch ◽  
Christian Templin ◽  
Helmut Drexler ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Adipocyte Differentiation ◽  
Prostaglandin E ◽  
Mouse Heart ◽  
Cardiac Progenitor Cells ◽  
Rt Pcr ◽  
Differentiation Potential ◽  
Intramyocardial Injection ◽  
Knock Out

Mice with a cardiomyocyte-restricted knock out of STAT3 (KO: alpha-MHC-Cre tg/+; STAT3 flox/flox ) show a continuous decrease of the cardiac capillary density and develop heart failure beyond the age of 9 months. We sought to determine the paracrine influence of cardiomyocyte STAT3 on the endothelial differentiation potential of cardiac progenitor cells (CPC) of the adult mouse heart. Sca-1 + CPC were isolated from male mice hearts by MACS separation. STAT3 was entirely deleted in cardiomyocytes of KO mice, while CPC from KO showed normal expression of STAT3 (confirmed by PCR and Western blot). No difference in the total number of CPC per heart was observed between wildtype (WT: STAT3 flox/flox ) and KO mice. FACS analysis revealed a reduced number of endothelial progenitor cells (as defined by coexpression of Sca-1, CD31 and CD38, −25%, P<0.05) among CPC from KO compared to CPC from WT. The differentiation potential of CPC from WT and KO was analyzed during in vitro culture on fibronectin-coated plates. After 4 weeks of culture RT-PCR for CD31 and immunohistochemistry (IHC) for endothelial cell (EC) marker tie2 and isolectin B4 was performed. CPC from WT showed markedly more efficient EC differentiation and tube formation compared to CPC from KO (p<0.01). In contrast, adipocyte differentiation was enhanced in CPC from KO (p<0.05, oil red staining and RT-PCR). Proliferation capacity of CPC from KO was reduced by 33% (p<0.01) as compared to CPC from WT. Microarray results of freshly isolated CPC were consistent with the differences in EC and adipocyte differentiation (i.e. prostaglandin E receptor 3 up 2.3-fold in CPC from WT, Lipocalin-2 up 2.7-fold in CPC from KO). We did not observe cardiomyocyte differentiation (IHC for alpha-sarcomeric actinin; RT-PCR for Nkx 2.5, alpha-MHC, or alpha-skeletal actin) of CPC from both genotypes, neither in vitro by addition of oxytocin, 5-AZA, DMSO, nor following intramyocardial injection of CPC in vivo. Conclusion: STAT3-dependent paracrine mediators released from cardiomyocytes are determinants of differentiation and vasculogenic properties of new EC derived from cardiac progenitor cells. The identification of these factors may offer new approaches to enforce the endogenous vasculogenic repair potential of the adult heart.

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.

Abstract P014: Oxidative Stress Mediated Regulation of Antioxidants in Cardiac Progenitor Cells

2011 ◽  
Vol 109 (suppl_1) ◽  
Author(s):  
Gokulakrishnan Iyer ◽  
Michael E Davis
Keyword(s):  
Oxidative Stress ◽  
Xanthine Oxidase ◽  
Progenitor Cells ◽  
Cardiac Progenitor Cells ◽  
Dependent Manner ◽  
Akt Inhibitor ◽  
Differentiation Potential ◽  
Lineage Differentiation ◽  
Phosphorylated Akt

Cardiac diseases are the leading causes of death throughout the world and transplantation of endogenous myocardial progenitor population with robust cardiovascular lineage differentiation potential is a promising therapeutic strategy. Therefore, in vitro expansion and transplantation of cardiac progenitor cells (CPCs) is currently in early clinical testing as a potential treatment for severe cardiac dysfunction. However, poor survival and engraftment of cells is one of the major limitations of cell transplantation therapy. Oxidative stress is increased in the ischemic myocardium and indirect inferences suggest the vulnerability of CPCs to oxidative stress. In this study, we show that in vitro, resident c-kit positive CPCs isolated from rat myocardium are significantly (p<0.05) resistant to superoxide-induced apoptosis compared to cardiomyocytes as analyzed by the number of sub-G1 population following xanthine/xanthine oxidase treatment. Interestingly, CPCs have two to four fold higher basal SOD1 and SOD2 activities (p<0.01) compared to cardiomyocytes and endothelial cells. Superoxide treatment increased expression of SOD1 (p<0.01), SOD2 (p<0.01), and glutathione peroxidase (p<0.05) mRNAs within 6 h of treatment compared to control cells. Recent studies suggest the involvement of AKT in controlling cell death, survival and also expression of SOD enzymes. Therefore, we investigated the involvement of AKT in CPCs subjected to oxidative stress. Western blot analysis revealed that the amount of phosphorylated AKT increased significantly within 10 minutes of xanthine/xanthine oxidase treatment. In addition, treatment with LY294002 - a PI3 kinase/AKT inhibitor, increased apoptosis in CPCs treated with superoxide. Our studies demonstrate a novel finding in which resident progenitor cells are protected from oxidative injury by containing higher basal levels of antioxidants as compared to myocytes. Moreover, under oxidant challenge antioxidant levels are regulated, possibly in an AKT-dependent manner. Further elucidation of this pathway may lead to novel therapeutic opportunities.

Abstract 217: Long Non-coding RNA Myocardial Infarction-associated Transcript (MIAT) Protects Cardiac Progenitor Cells From Oxidative Stress-induced Cell Death

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Liu Yang ◽  
Yang Yu ◽  
Baron Arnone ◽  
Chan Boriboun ◽  
Jiawei Shi ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Myocardial Infarction ◽  
Cell Death ◽  
Progenitor Cells ◽  
Cardiac Cells ◽  
Border Zone ◽  
Mouse Heart ◽  
Cardiac Progenitor Cells

Background: Long non-coding RNAs (lncRNAs) are an emerging class of RNAs with no or limited protein-coding capacity; a few of which have recently been shown to regulate critical biological processes. Myocardial infarction-associated transcript (MIAT) is a conserved mammalian lncRNA, and single nucleotide polymorphisms (SNPs) in 6 loci of this gene have been identified to be strongly associated with the incidence and severity of human myocardial infarction (MI). However, whether and how MIAT impacts on the pathogenesis of MI is unknown. Methods & Results: Quantitative RT-PCR analyses revealed that MIAT is expressed in neonatal mouse heart and to a lesser extent in adult heart. After surgical induction of MI in adult mice, MIAT starts to increase in 2 hours, peaks at 6 hours in atria and 12 hours in ventricles, and decreases to baseline at 24 hours. Fluorescent in situ hybridization (FISH) revealed a slight increase in the number of MIAT-expressing cells in the infarct border zone at 12 hours post-MI. Moreover, qRT-PCR analyses of isolated cardiac cells revealed that MIAT is predominantly expressed in cardiosphere-derived cardiac progenitor cells (CPCs). Treatment of CPCs with H 2 O 2 led to a marked upregulation of MIAT, while knockdown (KD) of MIAT resulted in a significantly impaired cell survival in vitro with H 2 O 2 treatment and in vivo after administered in the ischemic/reperfused heart. Notably, bioinformatics prediction and RNA immunoprecipitation identified FUS (fused in sarcoma) as a novel MIAT-interacting protein. FUS-KD CPCs displayed reduced cell viability and increased apoptosis under oxidative stress. Furthermore, MIAT overexpression enhanced survival of WT CPCs but not FUS-KD CPCs, suggesting that the protective role of MIAT is mediated by FUS. Conclusions: MIAT interacts with FUS to protect CPCs from oxidative stress-induced cell death.

Abstract 16189: Lineage Tracing of Postnatal Cardiomyogenic Progenitor Cells

Circulation ◽  
2014 ◽  
Vol 130 (suppl_2) ◽  
Author(s):  
Yuan-Hung Liu ◽  
Shih-Yun Huang ◽  
Yi-Shuan Lin ◽  
Hsing-Yu Huang
Keyword(s):  
Progenitor Cells ◽  
Heart Development ◽  
Lineage Tracing ◽  
Mouse Heart ◽  
Differentiation Potential ◽  
Genetic Ablation ◽  
Epicardial Cells ◽  
Cardiomyogenic Differentiation

Recent studies report that postnatal mammalian hearts undergo cardiomyocyte refreshment. While the exact origin of the cells involved in postnatal cardiomyogenesis remains unclear. Here, we identified a pool of Nkx2.5 enhancer expressing cells in the postnatal mouse heart with cardiomyogenic differentiation potential in vitro. We tracked the expression of a cardiac-specific enhancer of Nkx2.5 using inducible Nkx2.5 enhancer-Cre mice from embryonic development to adulthood and post-myocardial infarction (MI) and documented the Nkx2.5 enhancer expressing cells directly contribute to postnatal cardiomyogenesis in vivo. Upon genetic ablation of these activated progenitors after myocardial injury, the cardiac function deteriorated. Transcriptomic analysis of Nkx2.5 enhancer expressing cells showed high expression of heart development genes. To trace the developmental origin of the activated Nkx2.5 cardiomyogenic progenitor cells, we created different lineage-Cre/Nkx2.5 enh-eGFP/ROSA26 reporter triple transgenic mice. Post-MI Nkx2.5 cardiomyogenic progenitor cells originated from the embryonic epicardial cells, not from the pre-existing cardiomyocytes, endothelial cells, cardiac neural crest cells, or perinatal/postnatal epicardial cells. Together, this study confirmed that cardiac lineage-specific progenitor cells, which originate from embryonic epicardium-derived cells, contribute to postnatal mammalian cardiomyogenesis.

scholarly journals Increased prostaglandin-D2 in male but not female STAT3-deficient hearts shifts cardiac progenitor cells from endothelial to white adipocyte differentiation

2020 ◽  
Author(s):  
Elisabeth Stelling ◽  
Melanie Ricke-Hoch ◽  
Sergej Erschow ◽  
Steve Hoffmann ◽  
Anke Katharina Bergmann ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Adipocyte Differentiation ◽  
Young Male ◽  
Capillary Density ◽  
Prostaglandin D2 ◽  
Cardiac Progenitor Cells ◽  
Differentiation Potential ◽  
White Adipocyte ◽  
Female Mice ◽  
Age Related

AbstractCardiac levels of the signal transducer and activator of transcription factor-3 (STAT3) decline with age, and male but not female mice with a cardiomyocyte-specific STAT3 deficiency (CKO) display premature age-related heart failure associated with reduced cardiac capillary density. In the present study isolated male and female CKO-cardiomyocytes exhibit increased prostaglandin (PG)-generating cyclooxygenase-2 (COX-2) expression. The PG-degrading hydroxyprostaglandin-dehydrogenase-15 (HPGD) expression is only reduced in male cardiomyocytes, which is associated with increased PGD2 secretion from isolated male but not female CKO-cardiomyocytes. Reduced HPGD expression in male cardiomyocytes derive from impaired androgen-receptor-(AR)-signaling due to loss of its co-factor STAT3. Elevated PGD2 secretion in males is associated with increased white adipocyte accumulation in aged male but not female hearts. Adipocyte differentiation is enhanced in isolated SCA-1+-cardiac-progenitor-cells (CPC) from young male CKO-mice compared to the adipocyte differentiation of male wildtype (WT)-CPC and CPC isolated from female mice. Epigenetic analysis in freshly isolated male CKO-CPC display hypermethylation in pro-angiogenic genes (Fgfr2, Epas1) and hypomethylation in the white adipocyte differentiation gene Zfp423 associated with upregulated ZFP423 expression and a shift from endothelial to white adipocyte differentiation compared to WT-CPC. The expression of the histone-methyltransferase EZH2 is reduced in male CKO-CPC compared to male WT-CPC whereas no differences in the EZH2 expression in female CPC were observed. Clonally expanded CPC can differentiate into endothelial cells or into adipocytes depending on the differentiation conditions. ZFP423 overexpression is sufficient to induce white adipocyte differentiation of clonal CPC. In isolated WT-CPC, PGD2 stimulation reduces the expression of EZH2 thereby upregulating ZFP423 expression and promoting white adipocyte differentiation.Thus, cardiomyocyte STAT3-deficiency leads to age-related and sex-specific cardiac remodeling and failure in part due to sex-specific alterations in PGD2 secretion and subsequent epigenetic impairment of the differentiation potential of CPC. Causally involved is the impaired AR signaling in absence of STAT3, which reduces the expression of the PG degrading enzyme HPGD.

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.

Abstract 18845: Induced Cardiac Progenitor Cells Differentiate Into Cardiac Lineage Cells in vivo and Improve Survival in Mice post Myocardial Infarction

Circulation ◽  
2015 ◽  
Vol 132 (suppl_3) ◽  
Author(s):  
Pratik A Lalit ◽  
Max R Salick ◽  
Daryl O Nelson ◽  
Jayne M Squirrell ◽  
Christina M Shafer ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Disease Modeling ◽  
Ex Vivo ◽  
Cardiac Progenitor Cells ◽  
Heart Tube ◽  
Whole Embryo Culture ◽  
Cardiac Actin ◽  
Cardiac Lineage

Several studies have reported reprogramming of fibroblasts (Fibs) to induced cardiomyocytes, and we have recently reprogrammed mouse Fibs to induced cardiac progenitor cells (iCPCs), which may be more favorable for cardiac repair because of their expandability and multipotency. Adult cardiac (AC), lung and tail-tip Fibs from an Nkx2.5-EYFP reporter mouse were reprogrammed using a combination of five defined factors into iCPCs. Transcriptome and immunocytochemistry analysis revealed that iCPCs were cardiac mesoderm-restricted progenitors that expressed CPC markers including Nkx2.5, Gata4, Irx4, Tbx5, Cxcr4, Flk1 etc. iCPCs could be extensively expanded (over 30 passages) while maintaining multipotency to differentiate in vitro into cardiac lineage cells including cardiomyocytes (CMs), smooth muscle cells and endothelial cells. iCPC derived CMs upon co-culture with mESC-derived CMs formed intercellular gap junctions, exhibited calcium transients, and contractions. The purpose of this study was to determine the in vivo potency of iCPCs. Given that the Nkx2.5-EYFP reporter identifies embryonic CPCs, we first tested the embryonic potency of iCPCs using an ex vivo whole embryo culture model injecting cells into the cardiac crescent (CC) of E8.5 mouse embryos and culturing for 24 to 48 hours. GFP labeled AC Fibs were first tested and live imaging revealed that after 24 hours these cells were rejected from the embryo proper and localized to the ecto-placental cone. In contrast, iCPCs reprogrammed from AC Fibs when injected into the CC localized to the developing heart tube and differentiated into MLC2v, αMHC and cardiac actin expressing CMs. Further we injected iCPCs into infarcted adult mouse hearts and determined their regenerative potential after 1-4 wks. The iCPCs significantly improved survival (p<0.01 Mantel-Cox test) in treated animals (75%) as compared to control (11%). Immunohistochemistry revealed that injected iCPCs localized to the scar area and differentiated into cardiac lineage cells including CMs (cardiac actin). These results indicate that lineage reprogramming of adult somatic cells into iCPCs provides a scalable cell source for cardiac regenerative therapy as well as drug discovery and disease modeling.

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