Synthetic prostacyclin ONO-1301 Enhance Survival of Human Pluripotent Stem Cell-Derived Cardiomyocytes Tissue Transplantation in a Rat Myocardial Infarction Model
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE205622 | GEO | 2023/03/26
REPOSITORIES: GEO
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| S-EPMC7822665 | biostudies-literature
Project description:Acute occlusion of a coronary artery results in swift tissue necrosis. Bordering areas of the infarcted myocardium may also experience impaired blood supply and reduced oxygen delivery leading to altered metabolic and mechanical processes. While transcriptional changes in hypoxic cardiomyocytes are well-studied, little is known about the proteins that are actively secreted from these cells. We established a novel secretome analysis of cardiomyocytes by combining stable isotope labeling and click chemistry with subsequent mass spectrometry analysis. Further functional validation experiments included ELISA measurement of human samples, murine LAD ligation and adeno-associated virus (AAV) 9-mediated in vivo overexpression in mice. The presented approach is feasible for the analysis of the secretome of primary cardiomyocytes without serum starvation. 1026 proteins were identified to be secreted within 24 hours, indicating a 5-fold increase in detection compared to former approaches. Among them, a variety of proteins have so far not been explored in the context of cardiovascular pathologies. One of the most strongly upregulated secreted factors upon hypoxia was proprotein convertase subtilisin/kexin type 6 (PCSK6). Validation experiments revealed an increase of PCSK6 on mRNA and protein level in hypoxic cardiomyocytes. PCSK6 expression was elevated in hearts of mice following 3 days of ligation of the left anterior descending artery, a finding confirmed by immunohistochemistry. ELISA measurements in human serum also indicate distinct kinetics for PCSK6 in patients suffering from acute myocardial infarction, with a peak on day 3 post-infarction. Transfer of PCSK6-depleted cardiomyocyte secretome resulted in decreased expression of collagen I and III in fibroblasts compared to control treated cells, and siRNA mediated knockdown of PCSK6 in cardiomyocytes impacted transforming growth factor-β activation and mothers against decapentaplegic homolog 3 (SMAD3) translocation in fibroblasts. An Adeno-associated virus (AAV) 9-mediated, cardiomyocyte-specific overexpression of PCSK6 in mice resulted in increased collagen expression and cardiac fibrosis as well as decreased left ventricular function after myocardial infarction. In conclusion, a novel mass spectrometry-based approach allows the investigation of the secretome of primary cardiomyocytes. Analysis of hypoxia-induced secretion led to the identification of PCSK6 to be crucially involved in cardiac remodeling after acute myocardial infarction. Secretome analysis was performed on neonatal rat ventricular cardiomyocytes (NRVCMs) which were incubated under hypoxic conditions (1.5% O2, 5% CO2, 93.5% N2) for 12 (Hypoxia 0-12h), 24 (Hypoxia 0-24h) and 30 (Hypoxia 24-30h) hours. Furthermore, knockdown (KD) of PCSK6 in vitro mediated by small interfering RNA (siRNA) was performed to investigate changes in the secretome of cardiomyocytes with PCSK6 KD vs. control (control siRNA) during 24 hours of hypoxia (PCSK6 KD 0-24h). Cells were pulse-labeled with AHA (L-azidohomoalanine) and SILAC (stable isotope labeling with amino acids in cell culture) for 12 (Hypoxia 0-12h), 24 (Hypoxia 0-24h/PCSK6 KD 0-24h) and 6 hours (Hypoxia 24-30h). For Hypoxia 0-12h 3 replicates, Hypoxia 0-24h 6 replicates, PCSK6 KD 0-24h 3 Replicates and Hypoxia 24-30h 2 replicates were performed with label-swap.
2020-03-05 | PXD016178 | Pride
Project description: Not available
| S-EPMC6341224 | biostudies-literature
Project description:Myocardial infarction (MI) contributes to cardiac mortality and morbidity. After myocardial infarction the innate immune response is pivotal in clearing of tissue debris as well as scar formation, but exaggerated cytokine and chemokine secretion with subsequent leukocyte infiltration also leads to further tissue damage. Post-translational regulation of cytokine / chemokine signaling occurs via ectodomain shedding through a disintegrin and metalloproteases (ADAMs). Here, we address the value of targeting a previously unknown ADAM10 / CX3CL1 axis in the regulation of neutrophil recruitment early after MI. We show that myocardial ADAM10 is distinctly upregulated in biopsies from patients with ischemia-driven cardiomyopathy and correlates with heart failure progression. Intriguingly, upon MI in mice, pharmacological treatment with the ADAM10 inhibitionor GI254023X as well as genetic cardiomycyte-specific ADAM10 deletion improves survival with markedly enhanced heart function and reduced scar size. Mechanistically, this is driven by abolished ADAM10-mediated CX3CL1 ectodomain shedding followed by diminished IL-1β-dependent inflammation, reduced neutrophil bone marrow egress as well as myocardial tissue infiltration. Genetic cardiomycyte-specific ADAM10 deletion confirmes the small-molecule data and leads to improved cardiac function and reduction in inflammatory markers after ischemic myocardial injury. Thus, our data shows a conceptual insight into how acute MI induces chemotactic signaling via ectodomain shedding in cardiomyocytes.
2022-11-10 | GSE217268 | GEO
Project description:Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-cardiomyocytes) are promising sources for the regenerative therapy to treat heart failure. To realize this therapy, the engraftment potential of hiPSC-cardiomyocytes following the injection into the host heart should be improved. Here, we established the efficient method to analyze the cell cycle activity of hiPSC-cardiomyocytes using Fluorescence Ubiquitination-based Cell Cycle Indicator (FUCCI) system. High-throughput screening analysis using FUCCI-expressing hiPSC-cardiomyocytes identified a retinoic acid receptor (RAR) agonist, Am80, as an effective cell cycle activator in hiPSC-cardiomyocytes. The transplantation of hiPSC-cardiomyocytes treated by Am80 prior to the injection significantly enhanced the engraftment into the damaged mouse heart for 6 months. RNA sequencing analysis in Am80-treated iPSC-cardiomyocytes revealed that both RARA and RARB played important roles in the Am80-mediated cell cycle activation in hiPSC-cardiomyocytes. Collectively, this study highlights the effectiveness of FUCCI system to easily analyze the cell cycle status in hiPSC-cardiomyocytes and the cell cycle activation by Am80 is the possible strategy to increase the graft size following the cell transplantation into the damaged heart.
2023-07-12 | GSE176303 | GEO
Project description:Background: Bone marrow-derived multipotent progenitor cell (MPC) transplantation leads to short term functional and bioenergetic improvement in a porcine model of postinfarction Left Ventricular (LV) remodeling despite a low engraftment rate. However, the long term outcome after MPC transplantation is unknown. Methods and Results: The objectives of this study were to determine the long term functional outcome after MPC transplantation in a porcine model of postinfarction LV remodeling and to elucidate its mechanisms. Myocardial infarction (MI) was created by ligating the first and second diagonal branches of the Left anterior descending artery after open thoracotomy. Intramyocardial injection of 50 million lacZ labeled MPC was performed in the periscar region (Cell, n=7) with 5 equal injections immediately after MI, while in control animals (CONT, n=7; only 6 were profiled by microarray) saline was injected. Outcome was assessed temporally for 4 months with MRI and P-31 MRS. Engraftment was studied on histology and gene chip (Affymetrix) array analysis was used to study differential expression of genes in the two groups at 4 months. MPC treatment resulted in improvement of ejection fraction as early as 10 days after MI (32.2±5.5 vs. 43.4±5.1 in CONT and Cell respectively, p <0.05). This improvement was seen each month and persisted up to 4 months (35.7±5.0 vs. 51.2±4.8 in CONT and Cell respectively, p<0.005). PCr/ATP ratio improved with MPC transplantation (1.88±0.11 vs. 2.13±0.13 in CONT and Cell respectively, p<0.05). Under increased workload with inotropic stimulation, the bioenergetic differences were even more pronounced. There was no significant difference in scar size (scar/LV area*100) at 10 days post infarction (9.7±2.3 vs. 8.3±1.5 in CONT and Cell respectively). However, at 4 months there was a significant decrease in scar size in the MPC treated animals (8.6±2.4 vs. 4.6±1.0 in CONT and Cell respectively, p <0.05). No significant engraftment of MPC was observed on histology at 4 months. Gene chip array analysis identified several genes which were differentially expressed in the two groups. MPC transplantation was associated with a downregulation of mitochondrial oxidative enzymes, increased levels of MEF2a (Myocyte Enhancer Factor 2a) and ZFP91 (Zinc finger protein 91). Conclusion: MPC transplantation results in long term improvement in ventricular function and myocardial energetics that is associated with a decreased scar size and differential expression of genes. 13 swine were randomized after myocardial infarction to receive either 50 million stem cells (n=7) or saline only (n=6). The pigs were followed for a total of 4 months after transplantation with MRI occurring at baseline, 10 days, 1 month and then subsequently each month for 4 months. At the end of the study open-chest NMR spectroscopy was performed and then after sacrifice histology and microarray analysis was done on the tissue samples.
2010-04-12 | E-GEOD-14643 | biostudies-arrayexpress
Project description: Not available
| S-EPMC4055118 | biostudies-literature
Project description:We designed microRNA-responsive, modified mRNA switches (miR-switches) that quantitatively control their own translation level by determining the miRNA activities. We found that the three miR-switches (i.e., miR-1-, miR-208-, and miR-499-switches) enable to purify cardiomyocytes differentiated from hPSCs with high efficiency, accuracy, and safety. The purified cardiomyocytes were engrafted in mouse heart and did not form tumor. Simultaneous purification of two different cell types, cardiomyocytes and endothelial cells, was also performed by transfecting the mRNA responding to the both miR-208 and miR-126, into heterogeneous cell population derived from hPSCs. In addition, selective induction of apoptosis in noncardiomyocytes triggered by miR-1/208-Bim switch enriched cardiomyocytes without cell sorting. The mRNA switch could purify desired cell types and control cell death pathways by monitoring the miRNA expression dynamics during cell fate conversion. MYH6-EIP4 iPSCs for mRNA, N=3 MYH6-EIP4 day18 cardiomyocytes (before transfection) for mRNA, N=3 MYH6-EIP4 day19 cardiomyocytes (miR-1-switch day1) for mRNA, N=3 MYH6-EIP4 day19 cardiomyocytes (without transfection) for mRNA, N=3 MYH6-EIP4 day25 cardiomyocytes (miR-1-switch day7) for mRNA, N=3 MYH6-EIP4 day25 cardiomyocytes (without transfection) for mRNA, N=3 201B7 d22 miR-208a-BFP sorted cells for mRNA, N=1 201B7 d22 miR-208a-BFP negative fraction sorted cells for mRNA, N=1 201B7 d22 SIRPA+LIN- sorted cells for mRNA, N=1 201B7 d22 SIRPA+LIN- negative fraction sorted cells for mRNA, N=1 201B7 d22 VCAM1+ sorted cells for mRNA, N=1 201B7 d22 VCAM1+negative fraction sorted cells for mRNA, N=1 MYH6-EIP4 day19 cardiomyocytes (miR-208a-Bim-switch day1) for mRNA, N=1 MYH6-EIP4 day19 cardiomyocytes (without transfection) for mRNA, N=1 MYH6-EIP4 day21 cardiomyocytes (miR-208a-Bim-switch day7) for mRNA, N=1 MYH6-EIP4 day21 cardiomyocytes (without transfection) for mRNA, N=1 MYH6-EIP4 day25 cardiomyocytes (miR-208a-Bim-switch day7) for mRNA, N=1 MYH6-EIP4 day25 cardiomyocytes (without transfection) for mRNA, N=1
2015-05-20 | E-GEOD-60632 | biostudies-arrayexpress
Project description:Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) were transplanted into rats for one month to determine the change in morphology, structure, and function. Single cell RNA seq analysis was performed to compared the transcriptome of in vivo matured PSC-CMs to adult rat cardiomyocytes and in vitro cultured PSC-CMs.
2016-12-13 | GSE92247 | GEO