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. 2021 Mar;1(3):e62.
doi: 10.1002/cpz1.62.

Cardio Phenotypic Potential of Mesenchymal Stem Cells

Karen M Peterson  1 , Federico Franchi  1 , M Olthoff  1 , Ramasamy Paulmurugan  2   3 , Martin Rodriguez-Porcel  1
Affiliations

Cardio Phenotypic Potential of Mesenchymal Stem Cells

Karen M Peterson et al. Curr Protoc. 2021 Mar.

Erratum in

Abstract

Cell therapy is being investigated as a powerful intervention to ameliorate the consequences of coronary artery disease. Among the different stem cell options, mesenchymal stem cells (MSCs) are particularly attractive due to their high availability, as well as immune-privileged status. However, it is still unclear whether mesenchymal stem cells can acquire cardiomyogenic characteristics after they are transplanted to the myocardium. In this article, we outline protocols that illustrate the plasticity of MSCs and their ability to acquire cardiogenic characteristics when they are in an ischemic-like environment, as typically encountered after transplantation into the ischemic heart. © 2021 Wiley Periodicals LLC. Basic Protocol 1: Isolation of mesenchymal stem cells (MSCs) Support Protocol 1: Characterization of MSCs by flow cytometry Basic Protocol 2: Isolation of neonatal cardiomyoctes (NCMs) Support Protocol 2: Characterization of NCMs Basic Protocol 3: Cardiogenic plasticity of MSCs under ischemic-like conditions Support Protocol 3: Characterization of the cardiomyogenic potential of MSCs.

Keywords: cardiogenic; luciferase; mesenchymal stem cells; neonatal cardiomyocytes.

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Figures

Figure 1.
Figure 1.
Cartoon depicting the process for isolation of mesenchymal stem cells (MSCs) from bone marrow.
Figure 2.
Figure 2.
Characterization of isolated MSCs. Top panels, show the negative controls (IgG) of the CD29, CD45, CD11b/c and CD90. Bottom panels show that isolated MSCs were positive for CD29 (99.4%) and CD90 (99.98%) and negative for CD45 (0.32%) and CD11b/c (0.93%).
Figure 3.
Figure 3.
Cartoon depicting the process for isolation of neonatal cardiomyocytes (NCM).
Figure 4.
Figure 4.
Characterization of NCM. A and B show that isolated NCM are strongly positive for MHC (Texas Red) and Trop2 (FITC). Panel C depicts the RT-qPCR with increased expression of cardiac proteins of Myl2, TropT, Nkx2.5, Mef2C, and Gja-1, compared to total RNA.
Figure 5.
Figure 5.
Step-by-step process for the induction of MSCs towards a cardio phenotype.
Figure 6.
Figure 6.
Real Time Quantitative PCR showing the increased mRNA expression of Mef2c, Myl2, Gja1 (early) and TropT (late) markers in cardio MSCs compared to naïve MSCs. Data is expressed as fold change compared to naïve MSCs. P<0.05 vs. control for all genes.
Figure 7.
Figure 7.
Immunofluorescence staining for cardiac proteins. NCM (left), naïve MSCs (middle) and cardio MSCs (right column) were stained for (A) MHC (Texas Red), (B) TropT (FITC) and (C) Cx43 (Texas Red) and counterstained with DAPI. Cardio MSCs showed increased expression of cardiac proteins, indicating a shift to a cell with cardiomyogenic characteristics. Slides were photographed on a laser scanning confocal microscope at 20x magnification.
Figure 8.
Figure 8.
MSCs, carrying the cTnT-TSTA-Fluc reporter gene, were exposed to NCM-conditioned medium in a time dependent fashion, and their phenotype assessed using bioluminescence. Data is expressed as fold change compared to (non-treated) control MSCs. Both groups of cardio MSCs (increasing exposure to ischemic-like conditioned media) were significantly different (p<0.05) compared to naïve MSCs.
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