Local Activation or Implantation of Cardiac Progenitor Cells Rescues Scarred Infarcted Myocardium Improving Cardiac Function

Rationale: Cardiac progenitor cells (CPCs) are multipotent, self-renewing cells that can regenerate the myocardium and improve cardiac function in animal models of myocardial infarction (MI). However, limited survival of stem/progenitor cells inhibits cardiac regeneration. Force dependent Notch activation promotes cardiac development and cardiac gene expression in many adult stem cells. As dysregulation of Notch signaling leads to embryonic lethal cardiovascular defects, activating this critical pathway during cell transplantation could improve efficacy of stem cell therapy. Objective: Investigate i) whether self-assembling peptide scaffolds can be used to activate Notch1 signaling in CPCs to promote cardiogenic differentiation and ii) the effect of scaffold stiffness on Notch1 activation and differentiation. Methods: Rat CPCs (c-kit + ) were cultured for 48h in 3D self-assembling scaffolds of varying stiffness (1% low, 2% high): empty scaffolds (RADA), scaffolds modified with peptide mimicking Notch1 ligand, Jagged1 (RJAG), or scaffolds modified with a scrambled peptide (RSCR) and cardiogenic gene expression measured by qRT-PCR. CHO cells expressing Notch1 responsive YFP were also cultured in the above scaffolds for 48h and YFP expression was determined. Results are mean ± SEM with p<0.05 considered significant by one or two-way ANOVA with appropriate post test. Results: In the Notch1 reporter cells, Notch1 activation increased significantly in presence of RJAG (p<0.01) and on increasing scaffold stiffness (p<0.01,n=6) indicating scaffold stiffness-dependent Notch1 activation. Culture of CPCs in RJAG containing 1% scaffolds (low stiffness) significantly increased early endothelial and smooth muscle but not cardiac gene expression while in 2% scaffolds (high stiffness) significantly increased only cardiac and not endothelial or smooth muscle gene expression (p<0.05, n≥4). Conclusions: Taken together, these data show that i) Notch1 activation in 3D is dependent on ligand density and scaffold stiffness and ii) stiffness dependent Notch1 activation differentially regulates cardiogenic gene expression in CPCs. Therefore, delivery of CPCs in JAG containing scaffolds could be used to improve cardiac function following MI.

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Abstract 212: Aggregation of Child Cardiac Progenitor Cells into 3D Spheres Improves Endothelial Lineage Commitment

Circulation Research ◽

10.1161/res.119.suppl_1.212 ◽

2016 ◽

Vol 119 (suppl_1) ◽

Author(s):

David Q Trac ◽

Chunhui Xu ◽

Michael E. Davis

Keyword(s):

Stem Cell ◽

Cardiac Function ◽

Notch Signaling ◽

Progenitor Cells ◽

Gene Array ◽

Lineage Commitment ◽

Cardiac Progenitor Cells ◽

Notch Ligand ◽

Monolayer Cultures ◽

Endothelial Lineage

Congenital heart disease is rarely cured by surgery and can lead to life-threatening, intractable right ventricular heart failure (HF). In particular, children with hypoplastic left heart syndrome have a 10 year transplant-free survival rate of 50-75% despite palliative surgical repair. Currently, no effective stem-cell based treatments are available for pediatric HF. Recent stem-cell based clinical trials have been limited by poor differentiation rates and low cell retention. Additionally, we have shown that human cardiac progenitor cells (hCPCs) have reduced regenerative potential as they age, starting as early as 1 year old. We propose the aggregation of CPCs into scaffold-free spheres to improve the differentiation of child CPCs into mature cardiac phenotypes by enhancing intercellular Notch signaling. Notch signaling activity has been implicated in the regulation of CPC fate decisions and prior research in our lab has shown that intramyocardial delivery of Notch-ligand containing hydrogels improves cardiac function. Child CPC spheres were produced at a size of 1500 cells per sphere using a microwell array and cultured in suspension. Using immunohistochemistry, we showed that aggregation of CPCs increased Notch1 expression compared to parallel monolayer cultures. This effect is not limited to CPCs and was recapitulated in spheres of Chinese hamster ovarian cells transfected with Notch1-YFP. Additionally, Notch signaling pathway gene array data showed increased expression of the Notch-cleaving metalloprotease ADAM10 (3.6-fold) and Notch ligand DLL1 (25.0-fold) in CPC spheres by 3 days in culture compared to monolayer cultures. By 14 days in culture, we showed that aggregation of CPCs robustly increases the expression of the GATA4, a cardiac transcription factor associated with angiogenesis, and VEGFR1, an early marker of endothelial lineage commitment. Based on our results, we hypothesize that aggregation of CPCs into spheroids increases endothelial differentiation via a Notch-dependent mechanism. Transplantation of CPC spheres may improve cardiac function in vivo compared to transplantation of single CPCs. The results from our project will facilitate the development of autologous stem-cell based therapies for pediatric HF.

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Extracellular vesicles from human cardiac progenitor cells inhibit cardiomyocyte apoptosis and improve cardiac function after myocardial infarction

Cardiovascular Research ◽

10.1093/cvr/cvu167 ◽

2014 ◽

Vol 103 (4) ◽

pp. 530-541 ◽

Cited By ~ 357

Author(s):

L. Barile ◽

V. Lionetti ◽

E. Cervio ◽

M. Matteucci ◽

M. Gherghiceanu ◽

...

Keyword(s):

Myocardial Infarction ◽

Cardiac Function ◽

Progenitor Cells ◽

Extracellular Vesicles ◽

Cardiomyocyte Apoptosis ◽

Cardiac Progenitor Cells

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Abstract 21: Stiffness-Dependent Notch1 Activation Regulates Cardiogenic Differentiation of Cardiac Progenitor Cells

Circulation Research ◽

10.1161/res.111.suppl_1.a21 ◽

2012 ◽

Vol 111 (suppl_1) ◽

Author(s):

Archana V Boopathy ◽

Khalid Salaita ◽

Michael E Davis

Keyword(s):

Gene Expression ◽

Amino Acid ◽

Cardiac Function ◽

Notch Signaling ◽

Progenitor Cells ◽

Cardiac Progenitor Cells ◽

Critical Pathway ◽

3D Scaffolds ◽

Self Assembling ◽

Cardiogenic Differentiation

Cardiac progenitor cells (CPCs) are multipotent, self-renewing cells that can regenerate the myocardium and improve cardiac function in animal models of MI by cardiogenic differentiation. However, limited survival of stem/progenitor cells, myocardial scarring and fibrosis inhibit cardiac regeneration. Notch signaling promotes early cardiac development, cardiomyocyte survival and cardiac gene expression in circulating endothelial progenitor cells, mesenchymal stem cells and CPCs. As misregulation of Notch signaling during development is lethal due to cardiovascular defects, activating this critical pathway during cell transplantation could improve the efficacy of stem cell therapy. We investigated whether self-assembling peptide nanofiber hydrogels can be used to activate Notch1 signaling. The 16 amino acid self-assembling scaffold (RAD) was modified with a 20 amino acid peptide mimicking the active site of Notch1 ligand, Jagged1 (RJAG) or with the corresponding scrambled peptide (RSCR). To determine whether scaffold stiffness regulates Notch1 activation, CHO cells with Notch1 responsive YFP expression were cultured in scaffolds of 1-3% w/v in presence of RSCR or RJAG at a 1:10 ligand: scaffold ratio in 3D. Presence of the RJAG peptide (p<0.01) and % concentration of the scaffold (p<0.01) increased Notch1 activation significantly (n=5) indicating that RJAG mediated Notch1 activation in 3D is scaffold stiffness-dependent. Therefore, CPCs were cultured within 3D scaffolds (1-3% w/v; empty, scaffold +RJAG or RSCR) and cardiogenic gene expression was determined by qPCR. An increase in expression of early endothelial (Flk1, Flt1, vWF) and smooth muscle (sm22α, sm αactin) genes was observed in CPCs cultured in 3D scaffolds containing RJAG but not when cultured in 2D. These data show that Notch1 activation is dependent on ligand density and scaffold stiffness. Delivery of CPCs in JAG1 containing self-assembling scaffolds could be used to enhance therapeutic angiogenesis and improve cardiac function following myocardial infarction.

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iPSC‐derived human cardiac progenitor cells improve ventricular remodelling via angiogenesis and interstitial networking of infarcted myocardium

Journal of Cellular and Molecular Medicine ◽

10.1111/jcmm.12725 ◽

2016 ◽

Vol 20 (2) ◽

pp. 323-332 ◽

Cited By ~ 16

Author(s):

KP Myu Mia Ja ◽

Qingfeng Miao ◽

Nicole Gui Zhen Tee ◽

Sze Yun Lim ◽

Manasi Nandihalli ◽

...

Keyword(s):

Progenitor Cells ◽

Cardiac Progenitor Cells ◽

Ventricular Remodelling ◽

Infarcted Myocardium

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Optimisation of conductive polymer biomaterials for cardiac progenitor cells

RSC Advances ◽

10.1039/c6ra11682e ◽

2016 ◽

Vol 6 (67) ◽

pp. 62270-62277 ◽

Cited By ~ 5

Author(s):

C. Puckert ◽

A. Gelmi ◽

M. K. Ljunggren ◽

M. Rafat ◽

E. W. H. Jager

Keyword(s):

Tissue Engineering ◽

Cardiac Function ◽

Progenitor Cells ◽

Cardiac Tissue ◽

Conductive Polymer ◽

Cardiac Tissue Engineering ◽

Cardiac Progenitor Cells ◽

Engineering Applications ◽

Effective Treatments

The characterisation of biomaterials for cardiac tissue engineering applications is vital for the development of effective treatments for the repair of cardiac function.

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