scholarly journals Adult human cardiac stem cell supplementation effectively increases contractile function and maturation in human engineered cardiac tissues

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Jack F. Murphy ◽  
Joshua Mayourian ◽  
Francesca Stillitano ◽  
Sadek Munawar ◽  
Kathleen M. Broughton ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Contractile Function ◽  
Human Myocardium ◽  
Cardiac Stem Cells ◽  
Force Frequency ◽  
Cardiac Tissues ◽  
Adult Human ◽  
Beat Rate ◽  
Electrical Pacing

Abstract Background Delivery of stem cells to the failing heart is a promising therapeutic strategy. However, the improvement in cardiac function in animal studies has not fully translated to humans. To help bridge the gap between species, we investigated the effects of adult human cardiac stem cells (hCSCs) on contractile function of human engineered cardiac tissues (hECTs) as a species-specific model of the human myocardium. Methods Human induced pluripotent stem cell-derived cardiomyoctes (hCMs) were mixed with Collagen/Matrigel to fabricate control hECTs, with an experimental group of hCSC-supplemented hECT fabricated using a 9:1 ratio of hCM to hCSC. Functional testing was performed starting on culture day 6, under spontaneous conditions and also during electrical pacing from 0.25 to 1.0 Hz, measurements repeated at days 8 and 10. hECTs were then frozen and processed for gene analysis using a Nanostring assay with a cardiac targeted custom panel. Results The hCSC-supplemented hECTs displayed a twofold higher developed force vs. hCM-only controls by day 6, with approximately threefold higher developed stress and maximum rates of contraction and relaxation during pacing at 0.75 Hz. The spontaneous beat rate characteristics were similar between groups, and hCSC supplementation did not adversely impact beat rate variability. The increased contractility persisted through days 8 and 10, albeit with some decrease in the magnitude of the difference of the force by day 10, but with developed stress still significantly higher in hCSC-supplemented hECT; these findings were confirmed with multiple hCSC and hCM cell lines. The force-frequency relationship, while negative for both, control (− 0.687 Hz− 1; p = 0.013 vs. zero) and hCSC-supplemented (− 0.233 Hz− 1;p = 0.067 vs. zero) hECTs, showed a significant rectification in the regression slope in hCSC-supplemented hECT (p = 0.011 vs. control). Targeted gene exploration (59 genes) identified a total of 14 differentially expressed genes, with increases in the ratios of MYH7/MHY6, MYL2/MYL7, and TNNI3/TNNI1 in hCSC-supplemented hECT versus controls. Conclusions For the first time, hCSC supplementation was shown to significantly improve human cardiac tissue contractility in vitro, without evidence of proarrhythmic effects, and was associated with increased expression of markers of cardiac maturation. These findings provide new insights about adult cardiac stem cells as contributors to functional improvement of human myocardium.

scholarly journals Human Blood Serum Induces p38-MAPK- and Hsp27-Dependent Migration Dynamics of Adult Human Cardiac Stem Cells: Single-Cell Analysis via a Microfluidic-Based Cultivation Platform

Biology ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 708
Author(s):  
Anna L. Höving ◽  
Julian Schmitz ◽  
Kazuko E. Schmidt ◽  
Johannes F. W. Greiner ◽  
Cornelius Knabbe ◽  
...  
Keyword(s):  
Stem Cells ◽  
Stem Cell ◽  
Blood Serum ◽  
P38 Mapk ◽  
Human Blood ◽  
Human Blood Serum ◽  
Cardiac Stem Cells ◽  
Human Stem Cells ◽  
Adult Human ◽  
Migration Dynamics

Migratory capabilities of adult human stem cells are vital for assuring endogenous tissue regeneration and stem cell-based clinical applications. Although human blood serum has been shown to be beneficial for cell migration and proliferation, little is known about its impact on the migratory behavior of cardiac stem cells and underlying signaling pathways. Within this study, we investigated the effects of human blood serum on primary human cardiac stem cells (hCSCs) from the adult heart auricle. On a technical level, we took advantage of a microfluidic cultivation platform, which allowed us to characterize cell morphologies and track migration of single hCSCs via live cell imaging over a period of up to 48 h. Our findings showed a significantly increased migration distance and speed of hCSCs after treatment with human serum compared to control. Exposure of blood serum-stimulated hCSCs to the p38 mitogen-activated protein kinase (p38-MAPK) inhibitor SB239063 resulted in significantly decreased migration. Moreover, we revealed increased phosphorylation of heat shock protein 27 (Hsp27) upon serum treatment, which was diminished by p38-MAPK-inhibition. In summary, we demonstrate human blood serum as a strong inducer of adult human cardiac stem cell migration dependent on p38-MAPK/Hsp27-signalling. Our findings further emphasize the great potential of microfluidic cultivation devices for assessing spatio-temporal migration dynamics of adult human stem cells on a single-cell level.

scholarly journals Transcriptome Analysis Reveals High Similarities between Adult Human Cardiac Stem Cells and Neural Crest-Derived Stem Cells

Biology ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 435
Author(s):  
Anna L. Höving ◽  
Katharina Sielemann ◽  
Johannes F. W. Greiner ◽  
Barbara Kaltschmidt ◽  
Cornelius Knabbe ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
Stem Cell ◽  
Neural Crest ◽  
Global Gene Expression ◽  
Cell Populations ◽  
Cardiac Stem Cells ◽  
Human Stem Cell ◽  
Adult Human ◽  
Stem Cell Populations

For the identification of a stem cell population, the comparison of transcriptome data enables the simultaneous analysis of tens of thousands of molecular markers and thus enables the precise distinction of even closely related populations. Here, we utilized global gene expression profiling to compare two adult human stem cell populations, namely neural crest-derived inferior turbinate stem cells (ITSCs) of the nasal cavity and human cardiac stem cells (hCSCs) from the heart auricle. We detected high similarities between the transcriptomes of both stem cell populations, particularly including a range of neural crest-associated genes. However, global gene expression likewise reflected differences between the stem cell populations with regard to their niches of origin. In a broader analysis, we further identified clear similarities between ITSCs, hCSCs and other adherent stem cell populations compared to non-adherent hematopoietic progenitor cells. In summary, our observations reveal high similarities between adult human cardiac stem cells and neural crest-derived stem cells from the nasal cavity, which include a shared relation to the neural crest. The analyses provided here may help to understand underlying molecular regulators determining differences between adult human stem cell populations.

scholarly journals Blood Serum Stimulates p38-Mediated Proliferation and Changes in Global Gene Expression of Adult Human Cardiac Stem Cells

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1472 ◽  
Author(s):  
Anna L. Höving ◽  
Kazuko E. Schmidt ◽  
Madlen Merten ◽  
Jassin Hamidi ◽  
Ann-Katrin Rott ◽  
...  
Keyword(s):  
Gene Expression ◽  
Stem Cells ◽  
P38 Mapk ◽  
Regulatory Networks ◽  
Human Blood Plasma ◽  
Cardiac Stem Cells ◽  
Dependent Manner ◽  
Protective Effects ◽  
Adult Human ◽  
Age And Sex

During aging, senescent cells accumulate in various tissues accompanied by decreased regenerative capacities of quiescent stem cells, resulting in deteriorated organ function and overall degeneration. In this regard, the adult human heart with a generally low regenerative potential is of extreme interest as a target for rejuvenating strategies with blood borne factors that might be able to activate endogenous stem cell populations. Here, we investigated for the first time the effects of human blood plasma and serum on adult human cardiac stem cells (hCSCs) and showed significantly increased proliferation capacities and metabolism accompanied by a significant decrease of senescent cells, demonstrating a beneficial serum-mediated effect that seemed to be independent of age and sex. However, RNA-seq analysis of serum-treated hCSCs revealed profound effects on gene expression depending on the age and sex of the plasma donor. We further successfully identified key pathways that are affected by serum treatment with p38-MAPK playing a regulatory role in protection from senescence and in the promotion of proliferation in a serum-dependent manner. Inhibition of p38-MAPK resulted in a decline of these serum-mediated beneficial effects on hCSCs in terms of decreased proliferation and accelerated senescence. In summary, we provide new insights in the regulatory networks behind serum-mediated protective effects on adult human cardiac stem cells.

Annexin II Mediates Plasminogen-Dependant Matrix Invasion by Adult Human Mesenchymal Stem Cells.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2326-2326
Author(s):  
Paul B. Bolno ◽  
Doris A. Morgan ◽  
Mahesh Sharma ◽  
Martin Lazorik ◽  
Andrew S. Wechsler ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Extracellular Matrix ◽  
Mesenchymal Stem Cells ◽  
Stem Cell ◽  
Western Blot ◽  
Human Mesenchymal Stem Cells ◽  
Angiogenic Factors ◽  
Annexin Ii ◽  
Adult Human

Abstract Background: Annexin II (ANX2) is a fibrinolytic receptor that serves as a binding site for plasminogen and tissue plasminogen activator, facilitating the generation of plasmin. ANX2 is present on a wide variety of cells including vascular endothelial cells as well as macrophages. ANX2 has been shown to play a key role in extracellular matrix degradation, cellular migration, and invasion. This degradation of extracellular matrix may also cause the release of matrix-bound angiogenic factors such as VEGF and FGF. We hypothesized that adult human mesenchymal stem cells (hMSCs) express ANX2 and utilize this receptor for plasmin generation to facilitate basement membrane invasion. Methods: Primary hMSCs were isolated from the sternal bone marrow of patients undergoing median sternotomy. Stem cell surface markers were characterized via immuno-fluorescence. The presence of ANX2 protein by hMSCs was established via western blot. ANX2 mediated plasminogen activation and plasmin generation was quantified using chromozyme-P as a colorimetric substrate. Invasion assays were performed in dual-chamber culture wells containing matrigel inserts. hMSCs were plated into upper chambers containing: serum-free medium (SFM), SFM + Plasminogen, or SFM + Plasminogen + epsilon-aminocaproic acid (e-ACA inhibits binding of plasminogen to ANX2). After 24 hours, invasive cells were isolated and counted. Results: Sternal bone marrow derived hMSCs expressed the membrane phenotype CD34 (−), CD14 (−), CD44 (+), CD105 (+), CD106 (+). The presence of ANX2 was confirmed by western blot analysis. hMSCs generated 1.95 units of plasmin per milligram of protein. There was a 20% (p 0 .004) increase in hMSC invasion in the wells containing plasminogen as compared to SFM alone. When e-ACA was introduced there was a decrease in hMSC invasion back to control values. Conclusion: Our observations establish for the first time the presence and functional activity of ANX2 in hMSCs. These data suggest that mesenchymal stem cell expression of ANX2 facilitates plasminogen-mediated hMSC trans-endothelial invasion, migration and the release of pro-angiogenic factors from within the extracellular matrix, promoting stem cell directed repair and angiogenesis.

Bone Marrow-Derived Cells Regenerate Kit+ Cardiac Stem Cells (CSCs) in Damaged Heart.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 1681-1681
Author(s):  
Francesco Cerisoli ◽  
Lucio Barile ◽  
Roberto Gaetani ◽  
Letizia Cassinelli ◽  
Giacomo Frati ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Troponin I ◽  
Regulatory Elements ◽  
Cardiac Cells ◽  
Cardiac Differentiation ◽  
Pcr Analysis ◽  
Cardiac Stem Cells ◽  
Wild Type

Abstract A growing amount of data indicates that the heart harbours stem cells (CSCs) with regenerative potential, however the origin(s) of adult CSCs is still unknown. The expression of Kit a marker of several stem cell types, including hematopoietic and cardiac stem cells, suggests that Kit positive-CSCs may derive, at least in part, from extracardiac sources. In addition, it has been suggested that bone marrow (BM) cells may be mobilized, home into the heart and trans-differentiate into cardiomyocytes, following myocardial infarction. To investigate whether BM cells can contribute to repopulate the cardiac Kit+ stem cell pool, we transplanted BM cells from a mouse line expressing transgenic Green Fluorescent Protein (GFP) under the control of Kit regulatory elements, into wild type irradiated recipients. After hematological reconstution (4–5 months) and following cardiac infarction, cardiac cells were grown in vitro into typical “cardiospheres” (Messina et al., Circ. Res. 95,911;2004). The cardiospheres obtained, although not numerous, were all GFP fluorescent; this result was confirmed by PCR analysis of genomic DNA of individual CSs. At confocal microscopy, cells at the periphery of CSs showed coexistence of low GFP with cardiac markers, such as Troponin I and the transcription factor NKx2.5, consistent with the expected kit downregulation during cardiac differentiation. Our results show that cells of bone marrow origin can give rise, after homing into the heart, to cells with properties of Kit+ CSC. In contrast, CSCs isolated from kit/GFP transgenic mice are not able, upon transplantation, to repopulate the bone marrow of wild-type irradiated recipients. Thus, at least in pathological conditions, part of the Kit-positive CSCs population may be generated by BM-derived cells, capable of adopting in the heart the same function and features of cardiac stem cells.

scholarly journals Are MSCs Stem Cells? Demonstration of in Vitro and in Vivo Stem Cell Properties of Highly Enriched linneg/CD45neg/CD271pos/CD140alow/Neg Mesenchymal Stromal Cells from Adult Human Bone Marrow

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4374-4374
Author(s):  
Roshanak Ghazanfari ◽  
Hongzhe Li ◽  
Dimitra Zacharaki ◽  
Simón Méndez-Ferrer ◽  
Stefan Scheding
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Human Bone ◽  
Human Bone Marrow ◽  
Differentiation Potential ◽  
Self Renewal ◽  
Adult Human

Abstract Human bone marrow contains a rare population of non-hematopoietic mesenchymal stromal cells (BM-MSC) with multilineage differentiation capacity, which are essential constituents of the hematopoietic microenvironment. Self-renewal and differentiation are the two key properties of somatic stem cells, however, stem cell properties of human adult BM-MSC have not been demonstrated conclusively yet. We have previously shown that low/negative expression of PDGFRα on linneg/CD45neg/CD271pos cells identified a highly enriched population of primary BM-MSC in adult human bone marrow (Li et al. Blood, 2013, 122:3699). Based on this work, the current study aimed to investigate the in-vitro and in-vivo stem cell properties of this putative stromal stem cell population. The in-vitro clonogenic potential of freshly sorted human linneg/CD45neg/CD271pos/PDGFRlow/neg cells was evaluated by utilizing the CFU-F assay as well as the recently-developed mesensphere assay, which enables MSC amplification while preserving an immature phenotype (Isern et al, Cell Reports 2013, 30: 1714-24). Comparable colony frequencies were obtained with both assays (19.3 ± 2 and 17.5 ± 2.3 CFU-F and spheres per 100 plated cells, respectively, n=6, p=0.19). In order to test whether both assays identified the same population of clonogenic cells, colonies and spheres were replated under both conditions for up to three generations. The results showed comparable capacities of CFU-F and mesenspheres to form secondary and tertiary CFU-F and spheres. In-vitro self-renewal as indicated by increasing numbers of CFU-F and spheres (416.6 ± 431.7-fold and 49.5 ± 65.7-fold, respectively, n=3) was observed up to the third generation and decreased thereafter. The total number of generations was five (CFU-F) and six (spheres). In-vitro differentiation assays with both, CFU-F- and sphere-derived cells (tested until passage three) demonstrated tri-lineage differentiation potential (adipocytes, osteoblasts, chondrocytes). In addition, CFU-Fs and spheres had comparable surface marker profiles (CD73, CD90, CD105, and HLA-ABC positive; CD31, CD34 and HLA-DR negative), except for CD90, which was higher expressed on CFU-Fs. To investigate in-vivo self-renewal and differentiation potential of the putative stromal stem cells, linneg/CD45neg/CD271pos/PDGFRlow/neg -derived CFU-F and spheres were serially transplanted s.c into NSG mice. After 8 weeks, implants were harvested, human cells were FACS-isolated (CD90 and CD105 expression), and re-assayed under CFU-F and sphere conditions. Whereas in-vivo self-renewal of CFU-F could not be shown (111.5 ± 36 –fold decrease in total CFU-F numbers after primary transplantation, n=3), sphere self-renewal was clearly demonstrated by increased numbers of spheres after primary as well as secondary transplantation (1.13 ± 0.05 and 2.06 ± 0.26 –fold, respectively, n=3), which is remarkable given the fact that the number of recovered human cells is underestimated due to the isolation approach. Here, confirming GFP-marking experiments are ongoing. Finally, preliminary data indicate that linneg/CD45neg/CD271pos/PDGFRlow/neg –derived spheres display full in-vivo differentiation capacity in primary and secondary transplantations. Taken together, our data demonstrate - for the first time - that primary human linneg/CD45neg/CD271pos/PDGFRlow/neg cells meet stringent stem cell criteria, i.e. in-vitro and in-vivo self-renewal and differentiation. These findings answer the long-open question of the potential stem cell properties of adult human MSC and will enable to better understand the properties of native BM-MSC and their biological role in the bone marrow. Disclosures No relevant conflicts of interest to declare.

scholarly journals Stem Cell Studies in Cardiovascular Biology and Medicine: A Possible Key Role of Macrophages

Biology ◽  
2022 ◽  
Vol 11 (1) ◽  
pp. 122
Author(s):  
Nanako Kawaguchi ◽  
Toshio Nakanishi
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
Stromal Cells ◽  
Inflammatory Cells ◽  
Cardiac Stem Cells ◽  
Somatic Stem Cells ◽  
Micro Rnas ◽  
Cardiovascular Biology ◽  
Induced Pluripotent

Stem cells are used in cardiovascular biology and biomedicine, and research in this field is expanding. Two types of stem cells have been used in research: induced pluripotent and somatic stem cells. Stem cell research in cardiovascular medicine has developed rapidly following the discovery of different types of stem cells. Induced pluripotent stem cells (iPSCs) possess potent differentiation ability, unlike somatic stem cells, and have been postulated for a long time. However, differentiating into adult-type mature and functional cardiac myocytes (CMs) remains difficult. Bone marrow stem/stromal cells (BMSCs), adipose-derived stem cells (ASCs), and cardiac stem cells (CSCs) are somatic stem cells used for cardiac regeneration. Among somatic stem cells, bone marrow stem/stromal cells (BMSCs) were the first to be discovered and are relatively well-characterized. BMSCs were once thought to have differentiation ability in infarcted areas of the heart, but it has been identified that paracrine cytokines and micro-RNAs derived from BMSCs contributed to that effect. Moreover, vesicles and exosomes from these cells have similar effects and are effective in cardiac repair. The molecular signature of exosomes can also be used for diagnostics because exosomes have the characteristics of their origin cells. Cardiac stem cells (CSCs) differentiate into cardiomyocytes, smooth muscle cells, and endothelial cells, and supply cardiomyocytes during myocardial infarction by differentiating into newly formed cardiomyocytes. Stem cell niches and inflammatory cells play important roles in stem cell regulation and the recovery of damaged tissues. In particular, chemokines can contribute to the communication between inflammatory cells and stem cells. In this review, we present the current status of this exciting and promising research field.

Abstract 129: Human Mesenchymal Stem Cells Augment Contractile Function of Human Engineered Cardiac Tissues

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Timothy Cashman ◽  
Irene C Turnbull ◽  
Ioannis Karakikes ◽  
Jose Da Silva ◽  
Joshua M Hare ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Type I Collagen ◽  
High Efficiency ◽  
Contractile Function ◽  
Embryonic Stem ◽  
Type I ◽  
Cross Sectional ◽  
Cardiac Tissues ◽  
Twitch Force

Mesenchymal stem cells (MSC) have demonstrated efficacy for improving cardiomyocyte (CM) function in vitro, in vivo and in clinical trials, but the mechanism of this enhancement remains elusive. The objective of this study was to test the hypothesis that human engineered cardiac tissues (hECT) offer a viable model system to investigate the effects of human MSC on CM contractile function. Human CM (hCM) were produced from embryonic stem cells (hESC, H7 line) using a small-molecule based differentiation approach. Blebbistatin and BMP4 were added to hESC suspended in StemPro34 differentiation media for 24 h, followed by BMP4 and Activin A to day 4.5, followed by addition of IWR-1 Wnt inhibitor for at least 4 days. To create hECT, approximately 1 million hCM were mixed with 2.0 mg/ml bovine type I collagen and 0.9 mg/ml Matrigel, and pipetted into a mold fabricated from polydimethylsiloxane with integrated cantilever end-posts. To model hMSC cell therapy, two types of hECT were created: hCM-only control hECT, and hMSC-CM hybrid hECT containing hCM mixed with 5-10% of human bone marrow-derived MSC. Over several days in culture, the hECT self-assembled and started beating; end-post deflection was tracked in real time to compute twitch force using beam theory. Human CMs were produced with high efficiency (>70% cTnT+) with a predominantly ventricular phenotype (MLC2v+). Resulting hECTs exhibited spontaneous beating (1.3±0.4 Hz), cellular alignment, registered sarcomeres, and expression of cardiac specific genes cTnT, α-MHC, β-MHC and SERCA2a. After 11±2 days in culture, developed stress (force/area) was over 10-fold higher in hMSC-CM hybrid tissues (0.27±0.048 mN/mm 2 ) compared to hCM-only controls (0.02±0.006 mN/mm 2 ; p=0.04, n=5 per group). This reflected significantly greater twitch force (0.11±0.004 mN vs 0.033±0.016 mN, p=0.016) and smaller cross-sectional area (0.19±0.12 mm 2 vs 0.49±0.10 mm 2 ; p=0.003) in hMSC-CM hybrid vs hCM-only hECT. In conclusion, human ECT offer a novel system to study MSC-CM interactions. The findings suggest hMSC supplementation improves contractility compared to CM-only hECT. Investigating the mechanisms of hMSC-mediated enhancement of hECT function may yield insights into MSC-based therapies for cardiac regeneration.

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