Pacemaker cell characteristics of differentiated and HCN4-transduced human mesenchymal stem cells

Darche, Fabrice F.; Rivinius, Rasmus; Köllensperger, Eva; Leimer, Uwe; Germann, Günter; Seckinger, Anja; Hose, Dirk; Schröter, Julian; Bruehl, Claus; Draguhn, Andreas; Rinaldi, Gabriel; Schmidt, Manfred; Koenen, Michael; Thomas, Dierk; Katus, Hugo A.; Schweizer, Patrick

doi:10.1016/j.lfs.2019.116620

Cited by 11 publications

(27 citation statements)

References 51 publications

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“…Immunohistochemical analyses revealed specific fluorescence signals for the pacemaker-specific markers HCN1, Ca v 1.2, Cx31.9 and Cx45 (Figure 8B ) in transplanted dhaMSC at injections sites, while anti-HCN2 antibodies showed only low signals[ 20 ] (Figure 8B ). It is noteworthy that dhaMSC engrafted within pig hearts for 6 weeks showed considerable anti-HCN4 fluorescence signals (Figure 8B ), which were not detected at dhaMSC membranes, when cultured in vitro [ 17 ]. Furthermore, nhaMSC transplanted in pig hearts for 6 wk according to the similar protocol lacked HCN4 or any pacemaker-typical markers (Figure 9B ).…”

Section: Resultsmentioning

confidence: 99%

“…Isolation and handling of haMSC were approved by the local ethical committee (University of Heidelberg ethical committee number: S-462/2010). Cells were cultured in hMSC expansion medium as previously described[ 17 ].…”

Section: Methodsmentioning

confidence: 99%

“…Native haMSC (nhaMSC) were cultured for 4 wk in serum-free differentiation medium[ 17 ] free from teratogenic substances. In detail, differentiation medium comprised RPMI 1640 (Thermo Fisher Scientific, Waltham, MA, United States), 10 mL 50x B27 supplement (Thermo Fisher Scientific), 5 mL 10 5 UI penicillin (Thermo Fisher Scientific), 5 mL 10 5 µg/mL streptomycin (Thermo Fisher Scientific) and 10 ng/mL human recombinant BMP4 (PeproTech, Rocky Hill, NY, United States)[ 18 ].…”

Section: Methodsmentioning

confidence: 99%

“…According to this approach, MSC primarily served as a "vector shuttle" to guide the depolarizing gene HCN2 to the heart. Recently, we reported the differentiation of human MSC (hMSC) derived from adipose tissue into cells with cardiac pacemaker characteristics[ 17 ]. Pacemaker cell differentiation was achieved by cell culture using a customized cell medium, omitting the need for genetic manipulation.…”

Section: Introductionmentioning

confidence: 99%

“…Here, we asked whether differentiated hMSC derived from adipose tissue (dhaMSC), according to our customized protocol[ 17 ] may elicit biological pacemaker function after transplantation into porcine myocardium. To this end, we performed intramyocardial implantation of dhaMSC in pigs with experimental total AV block to elucidate the capacity of dhaMSC to integrate within the myocardium and to generate escape pacemaker activity.…”

Section: Introductionmentioning

confidence: 99%

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In vivo cardiac pacemaker function of differentiated human mesenchymal stem cells from adipose tissue transplanted into porcine hearts

Darche

Rivinius

Rahm

et al. 2020

WJSC

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BACKGROUND Mesenchymal stem cells (MSC) modified by gene transfer to express cardiac pacemaker channels such as HCN2 or HCN4 were shown to elicit pacemaker function after intracardiac transplantation in experimental animal models. Human MSC derived from adipose tissue (haMSC) differentiate into cells with pacemaker properties in vitro , but little is known about their behavior after intracardiac transplantation. AIM To investigate whether haMSC elicit biological pacemaker function in vivo after transplantation into pig hearts. METHODS haMSC under native conditions (nhaMSC) or after pre-conditioning by medium differentiation (dhaMSC) ( n = 6 pigs each, 5 × 10 6 cells/animal) were injected into the porcine left ventricular free wall. Animals receiving PBS injection served as controls ( n = 6). Four weeks later, total atrioventricular (AV)-block was induced by radiofrequency catheter ablation, and electronic pacemaker devices were implanted for backup stimulation and heart rate monitoring. Ventricular rate and rhythm of pigs were evaluated during a follow-up of 15 d post ablation by 12-lead-ECG with heart rate assessment, 24-h continuous rate monitoring recorded by electronic pacemaker, assessment of escape recovery time, and pharmacological challenge to address catecholaminergic rate response. Finally, hearts were analyzed by histological and immunohistochemical investigations. RESULTS In vivo transplantation of dhaMSC into the left ventricular free wall of pigs elicited spontaneous and regular rhythms that were pace-mapped to ventricular injection sites (mean heart rate 72.2 ± 3.6 bpm; n = 6) after experimental total AV block. Ventricular rhythms were stably detected over a 15-d period and were sensitive to catecholaminergic stimulation (mean maximum heart rate 131.0 ± 6.2 bpm; n = 6; P < 0.001). Pigs, which received nhaMSC or PBS presented significantly lower ventricular rates (mean heart rates 47.2 ± 2.5 bpm and 37.4 ± 3.2 bpm, respectively; n = 6 each; P < 0.001) and exhibited little sensitivity towards catecholaminergic stimulation (mean maximum heart rates 76.4 ± 3.1 bpm and 60.5 ± 3.1 bpm, respectively; n = 6 each; P < 0.05). Histological and immunohistochemical evaluation of hearts treated with dhaMSC revealed local clusters of transplanted cells at the injection sites that lacked macrophage or lymphocyte infiltrations or tumor formation. Intense fluorescence signals at these sites indicated membrane expression of HCN4 and other pacemaker-specific proteins involved in cardiac automaticity and impulse propagation. CONCLUSION dhaMSC transplanted ...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

In vivo cardiac pacemaker function of differentiated human mesenchymal stem cells from adipose tissue transplanted into porcine hearts

Darche

Rivinius

Rahm

et al. 2020

WJSC

Self Cite

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Disrupting methyl‐CpG‐binding protein 2 expression induces the transformation of induced pluripotent stem cell cardiomyocytes into pacemaker‐like cells by insulin gene enhancer binding protein 1

Zhang

Shi

et al. 2023

The Journal of Gene Medicine

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Background The experiment will explore whether interfering with the expression of methyl‐CpG‐binding protein 2 (MecP2) can enhance the ability of insulin gene enhancer binding protein 1 (ISL1) to induce iPSC‐CMs to differentiate into pacemaker‐like cells. Methods Differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes (CMs) can be induced via the regulation of the Wnt signaling pathway. Real‐time quantitative PCR (qPCR), western blotting, immunofluorescence staining, and patch‐clamp technique were used to analyze the ability of ISL1 to induce the transformation of iPSC‐CMs into pacemaker‐like cells. Calcium spark, patch‐clamp technique, and real‐time qPCR were used to verify whether disrupting the expression of MeCP2 enhanced this ability of ISL1 to induce the differentiation of iPSC‐CMs into pacemaker cells. Transplant pacemaker‐like cardiomyocytes into the myocardium of mice to observe whether the pacemaker cells can survive in the tissue for a long time. Results RT‐qPCR and patch‐clamp analyses showed that overexpression of ISL1 induced the successful differentiation of iPSC‐CMs into pacemaker cells. ISL1‐overexpressing pacemaker‐like cells possessed typical characteristics of pacemaker morphology, including action potential and If inward current. Chromatin immunoprecipitation results showed that MeCP2 bound to the promoter region of HCN4. Following disruption of MeCP2 expression, the gene expression of sinoatrial node‐specific transcription factors, If inward current, and cardiac rhythm changes in iPSC‐CMs resembled those of sinoatrial node pacemaker cells. Therefore, ISL1 induced the differentiation of iPSC‐CMs into pacemaker‐like cells, and knockdown of MeCP2 increased this effect. Frozen section results showed that surviving pacemaker‐like cells could still be observed in myocardial tissue after 45 days. Conclusions Experiments have found that interfering with the expression of MeCP2 can increase the ability of ISL1 to induce iPSC‐CM cells to differentiate into pacemaker‐like cells. And the pacemaker‐like cells obtained in this experiment can survive in myocardial tissue for a long time.

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Histone demethylase KDM5B catalyzed H3K4me3 demethylation to promote differentiation of bone marrow mesenchymal stem cells into cardiomyocytes

Wang

Zhong

2022

Mol Biol Rep

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Background Studies have shown that histone H3 methylation is involved in regulating the differentiation of Bone Marrow Mesenchymal Stem Cells (BMSCs). KDM5B can specifically reduce the level of histone 3 lysine 4 trimethylation (H3K4me3), thereby activating the expression of related genes and participating in biological processes such as cell differentiation, embryonic development and tumor formation. Whether KDM5B is involved in the regulation of BMSCs differentiation into cardiomyocytes through the above manner has not been reported. Objective To investigate the effect of KDM5B on the induction and differentiation of swine BMSCs into myocardial cells in vitro. Methods Swine bone marrow BMSCs were isolated and cultured, and the overexpression, interference expression and blank vector of KMD5B were constructed and transfected by lentivirus. BMSCs was induced to differentiate into cardiomyocytes by 5-azacytidine (5-AZA) in vitro, and the differentiation efficiency was compared by immunofluorescence, RT-PCR, Western Blot and whole-cell patch clamp detection. Result Compared with the control group, the expression levels of histone H3K4me3 and pluripotency gene Nanog in KDM5B overexpression group were significantly decreased, while the expression level of key myocardial gene HCN4 and myocardial marker gene α-Actin and cTNT were significantly increased, and the Na+ current density on the surface of differentiated myocardial cell membrane was significantly increased. Meanwhile, the corresponding results of the KDM5B silent expression group were just opposite. Conclusions It indicated that enhanced KDM5B expression could promote the differentiation of BMSCs into cardiomyocytes and improve the differentiation efficiency by controlling H3K4 methylation levels.

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Pacemaker cell characteristics of differentiated and HCN4-transduced human mesenchymal stem cells

Cited by 11 publications

References 51 publications

In vivo cardiac pacemaker function of differentiated human mesenchymal stem cells from adipose tissue transplanted into porcine hearts

In vivo cardiac pacemaker function of differentiated human mesenchymal stem cells from adipose tissue transplanted into porcine hearts

Disrupting methyl‐CpG‐binding protein 2 expression induces the transformation of induced pluripotent stem cell cardiomyocytes into pacemaker‐like cells by insulin gene enhancer binding protein 1

Histone demethylase KDM5B catalyzed H3K4me3 demethylation to promote differentiation of bone marrow mesenchymal stem cells into cardiomyocytes

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