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Isolation of Perivascular Multipotent Precursor Cell Populations from Human Cardiac Tissue
Published on: October 8, 2016
Concurrent Isolation of 3 Distinct Cardiac Stem Cell Populations From a Single Human Heart Biopsy
Megan M Monsanto1, Kevin S White1, Taeyong Kim1
1From the San Diego Heart Research Institute, San Diego State University, CA (M.M.M., K.S.W., T.K., B.J.W., K.F., K.I., F.G.K., A.C., K.B., S.M., M.A.S.); and Sharp Memorial Hospital, San Diego, CA (W.P.D.).
Insights
Researchers developed a new protocol to isolate three distinct cardiac stem cell populations from a single human heart biopsy. This method aids in understanding myocardial regeneration for heart failure treatment.
Area of Science:
- Cardiovascular Biology
- Stem Cell Research
- Regenerative Medicine
Background:
- The synergistic effects of distinct myocardial-derived stem cell populations are not well understood.
- Optimal stem cell choices for heart failure treatment are debated, necessitating new isolation methods.
Purpose of the Study:
- To establish a reliable protocol for isolating and culturing three unique stem cell populations from a single human heart biopsy.
- To gain insights into myocardial regeneration strategies.
Main Methods:
- Human heart tissue samples were minced and processed into a single-cell suspension.
- Magnetic-activated cell sorting (MACS) was used, initially based on c-Kit expression, followed by CD90/CD105 for mesenchymal stem cells and CD133 for endothelial progenitor cells.
- In vitro cell surface marker persistence was verified using flow cytometry and immunocytochemistry.
Main Results:
- Successfully isolated three distinct endogenous cardiac stem cell populations: c-Kit+, CD133+ endothelial progenitor cells, and c-Kit-, CD90+, CD105+ mesenchymal stem cells.
- The protocol effectively processed discarded heart tissue from left ventricular assist device implantations.
- Confirmed the phenotypic properties of isolated cells in vitro.
Conclusions:
- Concurrent isolation and expansion of cardiac progenitor cells, endothelial progenitor cells, and mesenchymal stem cells from a single human heart failure patient sample is feasible.
- The developed protocol yields distinct cell populations with specific phenotypic characteristics.
Rationale:
The relative actions and synergism between distinct myocardial-derived stem cell populations remain obscure. Ongoing debates on optimal cell population(s) for treatment of heart failure prompted implementation of a protocol for isolation of multiple stem cell populations from a single myocardial tissue sample to develop new insights for achieving myocardial regeneration.
Objective:
Establish a robust cardiac stem cell isolation and culture protocol to consistently generate 3 distinct stem cell populations from a single human heart biopsy.
Methods And Results:
Isolation of 3 endogenous cardiac stem cell populations was performed from human heart samples routinely discarded during implantation of a left ventricular assist device. Tissue explants were mechanically minced into 1 mm3 pieces to minimize time exposure to collagenase digestion and preserve cell viability. Centrifugation removes large cardiomyocytes and tissue debris producing a single cell suspension that is sorted using magnetic-activated cell sorting technology. Initial sorting is based on tyrosine-protein kinase Kit (c-Kit) expression that enriches for 2 c-Kit+ cell populations yielding a mixture of cardiac progenitor cells and endothelial progenitor cells. Flowthrough c-Kit- mesenchymal stem cells are positively selected by surface expression of markers CD90 and CD105. After 1 week of culture, the c-Kit+ population is further enriched by selection for a CD133+ endothelial progenitor cell population. Persistence of respective cell surface markers in vitro is confirmed both by flow cytometry and immunocytochemistry.
Conclusions:
Three distinct cardiac cell populations with individualized phenotypic properties consistent with cardiac progenitor cells, endothelial progenitor cells, and mesenchymal stem cells can be successfully concurrently isolated and expanded from a single tissue sample derived from human heart failure patients.
