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A 3-D cardiac muscle construct for exploring adult marrow stem cell based myocardial regeneration.

ABSTRACT: Adult bone marrow stromal cells (BMSCs) are capable of differentiating into cardiomyocyte-like cells in vitro and contribute to myocardial regeneration in vivo. Consequently, BMSCs may potentially play a vital role in cardiac repair and regeneration. However, this concept has been limited by inadequate and inconsistent differentiation of BMSCs into cardiomyocytes along with poor survival and integration of neo-cardiomyocytes after implantation into ischemic myocardium. In order to overcome these barriers and to explore adult stem cell based myocardial regeneration, we have developed an in vitro model of three-dimensional (3-D) cardiac muscle using rat ventricular embryonic cardiomyocytes (ECMs) and BMSCs. When ECMs and BMSCs were seeded sequentially onto a 3-D tubular scaffold engineered from topographically aligned type I collagen-fibers and cultured in basal medium for 7, 14, 21, or 28 days, the maturation and co-differentiation into a cardiomyocyte lineage was observed. Phenotypic induction was characterized at morphological, immunological, biochemical and molecular levels. The observed expression of transcripts coding for cardiomyocyte phenotypic markers and the immunolocalization of cardiomyogenic lineage-associated proteins revealed typical expression patterns of neo-cardiomyogenesis. At the biochemical level differentiating cells exhibited appropriate metabolic activity and at the ultrastructural level myofibrillar and sarcomeric organization were indicative of an immature phenotype. Our 3-D co-culture system sustains the ECMs in vitro continuum of differentiation process and simultaneously induces the maturation and differentiation of BMSCs into cardiomyocyte-like cells. Thus, this novel 3-D co-culture system provides a useful in vitro model to investigate the functional role and interplay of developing ECMs and BMSCs during cardiomyogenic differentiation.

SUBMITTER: Valarmathi MT

PROVIDER: S-EPMC2887929 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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A 3-D cardiac muscle construct for exploring adult marrow stem cell based myocardial regeneration.

Valarmathi Mani T MT Goodwin Richard L RL Fuseler John W JW Davis Jeffrey M JM Yost Michael J MJ Potts Jay D JD

Biomaterials 20100202 12

Adult bone marrow stromal cells (BMSCs) are capable of differentiating into cardiomyocyte-like cells in vitro and contribute to myocardial regeneration in vivo. Consequently, BMSCs may potentially play a vital role in cardiac repair and regeneration. However, this concept has been limited by inadequate and inconsistent differentiation of BMSCs into cardiomyocytes along with poor survival and integration of neo-cardiomyocytes after implantation into ischemic myocardium. In order to overcome these ...[more]

PMID: 20129663

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Project description:An overdose of Isoproterenol (ISO) causes acute cardiomyocyte (CM) dropout and activates the resident cardiac c-kitpos stem/progenitor cells (CSCs) generating a burst of new CM formation that replaces those lost to ISO. Recently, unsuccessful attempts to reproduce these findings using c-kitCre knock-in (KI) mouse models were reported. We tested whether c-kit haploinsufficiency in c-kitCreKI mice was the cause of the discrepant results in response to ISO. Male C57BL/6J wild-type (wt) mice and c-kitCreKI mice were given a single dose of ISO (200 and/or 400?mg/Kg s.c.). CM formation was measured with different doses and duration of BrdU or EdU. We compared the myogenic and regenerative potential of the c-kitCreCSCs with wtCSCs. Acute ISO overdose causes LV dysfunction with dose-dependent CM death by necrosis and apoptosis, whose intensity follows a basal-apical and epicardium to sub-endocardium gradient, with the most severe damage confined to the apical sub-endocardium. The damage triggers significant new CM formation mainly in the apical sub-endocardial layer. c-kit haploinsufficiency caused by c-kitCreKIs severely affects CSCs myogenic potential. c-kitCreKI mice post-ISO fail to respond with CSC activation and show reduced CM formation and suffer chronic cardiac dysfunction. Transplantation of wtCSCs rescued the defective regenerative cardiac phenotype of c-kitCreKI mice. Furthermore, BAC-mediated transgenesis of a single c-kit gene copy normalized the functional diploid c-kit content of c-kitCreKI CSCs and fully restored their regenerative competence. Overall, these data show that c-kit haploinsufficiency impairs the endogenous cardioregenerative response after injury affecting CSC activation and CM replacement. Repopulation of c-kit haploinsufficient myocardial tissue with wtCSCs as well c-kit gene deficit correction of haploinsufficient CSCs restores CM replacement and functional cardiac repair. Thus, adult neo-cardiomyogenesis depends on and requires a diploid level of c-kit.

Dataset Information

A 3-D cardiac muscle construct for exploring adult marrow stem cell based myocardial regeneration.

Publications

A 3-D cardiac muscle construct for exploring adult marrow stem cell based myocardial regeneration.

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