Stem cells are regarded with great promise in the next decades for treatment of congenital disease. Well known for their capacity to differentiate into a broad spectrum of cell types (pluripotency), embryonic stem (ES) cells also secrete corrective factors that prevent lethal congenital heart defects from occurring. Using a mouse model of the """"""""thin myocardial syndrome"""""""" (Id knockout mice), we have shown that secreted factors from ES cells normalize gene expression profiles in neighbor cells. The ES cells can rescue congenital heart defects not only when injected into Id KO blastocysts but also into mouse females that will bear Id KO embryos. The ES cells can also rescue muscular dystrophy when injected into mdx (a mouse model of Duchenne muscular dystrophy, DMD) blastocysts. Rather than factor secretion, the main mechanism of the rescue in this case is spreading of ES-derived dystrophin (the protein absent in DMD) throughout most of the musculature to stabilize the muscle. The potential use of ES cells to treat human disease is clouded by ethical concerns surrounding the destruction of human oocytes or fertilized embryos. The scientific community is looking for alternatives that would not use embryos as starting material. Recent experiments demonstrated that murine somatic cells de-differentiate to an embryonic stem cell-like status by the incorporation of transcription factors. These cells were named induced pluripotent stem cells (iPS cells). The iPS cells are similar to the ES cells. Remarkably, the generation of iPS cells does not require destruction of embryos, therefore there are no ethical concerns. In addition, a recent proof-of-principle experiment showed that iPS cells can correct disease (sickle cell anemia) in mice. In this application we would like to characterize the murine iPS cells in their role to rescue cardiac and skeletal muscle disease, exemplified by the Id knockout mice and the mdx mice. To this end, we will inject murine iPS cells into murine blastocysts (Id KO and mdx), intraperitoneally into female mice that will harbor mutant embryos (Id KO), and also intraperitoneally into mice predisposed to develop dilated cardiomyopathy (Id conditional KO). We hypothesize that iPS cells will rescue the cardiac phenotype of the Id KO embryos as well as muscular dystrophy in mdx mice. Survival of the Id KO embryos will be evaluated. Corrections will be evaluated at the histological (immunohistochemistry, H&E, immunofluorescence) and functional (echocardiography, treadmill) level. Secretion of potential rescue molecules in the thin myocardial syndrome (Id) rescue and spreading of dystrophin in the muscular dystrophy (mdx) rescue will be determined. These experiments will help elucidate the mechanisms that the iPS cells may utilize to effect corrections in muscle and will broaden the therapeutic applicability of the iPS cells.

Public Health Relevance

ES cells have the unique capacity to differentiate into all cell types and to emit healing factors. This feature places them at the center of the regenerative medicine arena. Technical and ethical issues compromise the enthusiasm for the use of ES cells in a clinical setting. Therefore it becomes imperative to find alternatives that will render cells equally potent. Recently the induced pluripotent stem (iPS) cells emerged as the greatest alternative. The iPS cells phenocopy the ES cells in the most rigorous assays that are used to characterize the ES cells, including the capacity to form mouse chimeras. As the generation of iPS cells does not involve the use of embryos, the ethical concerns are eliminated. To assess the therapeutic potential of the iPS cells, it will be extremely important to test the role that the iPS cells play in correcting a variety of genetic diseases. In these application we propose to challenge the murine iPS cells in two muscular diseases (heart and skeletal muscle) that we showed can be corrected by ES cell treatment. The mechanism of the correction of both diseases by the ES cells is different - in one case secretion of factors and in another spreading of a structural protein. We plan to inject iPS cells into early embryos that are predisposed to develop these diseases. We also plan to inject the iPS cells into female mice before conception. Finally, we plan to inject the iPS cells into mice predisposed to develop dilated cardiomyopathy. In all cases we hope the iPS cells will prevent pathology from occurring. These experiments will help elucidate the molecular mechanisms that the iPS cells may utilize to effect corrections in cardiac and skeletal muscle and will broaden the therapeutic applicability of the iPS cells.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Exploratory/Developmental Grants (R21)
Project #
1R21HL094905-01A1
Application #
7739137
Study Section
Cardiovascular Differentiation and Development Study Section (CDD)
Program Officer
Schramm, Charlene A
Project Start
2009-08-01
Project End
2011-07-31
Budget Start
2009-08-01
Budget End
2010-07-31
Support Year
1
Fiscal Year
2009
Total Cost
$195,000
Indirect Cost
Name
University of Medicine & Dentistry of NJ
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
623946217
City
Newark
State
NJ
Country
United States
Zip Code
07107
Zhao, Qingshi; Chang, Corey; Gonzalez, J Patrick et al. (2016) Combined Id1 and Id3 Deletion Leads to Severe Erythropoietic Disturbances. PLoS One 11:e0154480
Ziegler, Amber N; Schneider, Joel S; Qin, Mei et al. (2012) IGF-II promotes stemness of neural restricted precursors. Stem Cells 30:1265-76
Vitale, Joseph M; Schneider, Joel S; Beck, Amanda J et al. (2012) Dystrophin-compromised sarcoglycan-ýý-knockout diaphragm requires full wild-type embryonic stem cell reconstitution for correction. J Cell Sci 125:1807-13
Beck, Amanda J; Vitale, Joseph M; Zhao, Qingshi et al. (2011) Differential requirement for utrophin in the induced pluripotent stem cell correction of muscle versus fat in muscular dystrophy mice. PLoS One 6:e20065
Zhao, Qingshi; Beck, Amanda J; Vitale, Joseph M et al. (2011) Developmental ablation of Id1 and Id3 genes in the vasculature leads to postnatal cardiac phenotypes. Dev Biol 349:53-64
Zhao, Qingshi; Beck, Amanda; Vitale, Joseph M et al. (2010) Rescue of developmental defects by blastocyst stem cell injection: towards elucidation of neomorphic corrective pathways. J Cardiovasc Transl Res 3:66