Pre-Conditioning Stem Cells in a Biomimetic Environment for Enhanced Cardiac Tissue Repair: In Vitro and In Vivo Analysis
Stem cell-based therapies represent a valid approach to restore cardiac function due to their beneficial effect in reducing scar area formation and promoting angiogenesis. However, their translation into the clinic is limited by the poor differentiation and inability to secrete sufficient therapeutic factors. To address this issue, several strategies such as genetic modification and biophysical pre-conditioning have been used to enhance the efficacy of stem cells for cardiac tissue repair.
In this study, a biomimetic approach was used to mimic the natural mechanical stimulation of the myocardium tissue. Specifically, human adipose-derived stem cells (hASCs) were cultured on a thin gelatin methacrylamide (GelMA) hydrogel disc and placed on top of a beating cardiomyocyte layer. qPCR studies and metatranscriptomic analysis of hASCs gene expression were investigated to confirm the correlation between mechanical stimuli and cardiomyogenic differentiation. In vivo intramyocardial delivery of pre-conditioned hASCs was carried out to evaluate their efficacy to restore cardiac function in mice hearts post-myocardial infarction.
The cyclic strain generated by cardiomyocytes significantly upregulated the expression of both mechanotransduction and cardiomyogenic genes in hASCs as compared to the static control group. The inherent angiogenic secretion profile of hASCs was not hindered by the mechanical stimulation provided by the designed biomimetic system. Finally, in vivo analysis confirmed the regenerative potential of the pre-conditioned hASCs by displaying a significant improvement in cardiac function and enhanced angiogenesis in the peri-infarct region.
Overall, these findings indicate that cyclic strain provided by the designed biomimetic system is an essential stimulant for hASCs cardiomyogenic differentiation, and therefore can be a potential solution to improve stem-cell based efficacy for cardiovascular repair.
KeywordsMechanical stimulation Myogenic differentiation Angiogenesis Cardiac repair
AP acknowledges an investigator grant provided by the Institutional Development Award (IDeA) from the National Institute of General Medical Sciences (NIGMS) of the NIH Award Number P20GM103638 and Umbilical Cord Matrix Project fund from State of Kansas. RPHA acknowledges the support from National Institute of Health (NIH) Grant 1R01HL-10690. AC acknowledges support from AHA 16GRNT31030030 and NIH GM102801. Research reported in this publication was made possible by the services of Dr. Erik Lundquist and Ms. Jennifer Hackett at the KU Genome Sequencing Core. The authors also acknowledge the services provided by Dr. Stuart Macdonald and Ms. Boryana S Koseva at the KU-INBRE Bioinformatics Core. This core lab is supported by an Institutional Development Award (IDeA) from the NIGMS (P20GM103418) from the NIH. We also gratefully thank Ms. Heather Shinogle of the University of Kansas Microscopy and Analytical Imaging Laboratory for her assistance with confocal fluorescence microscopy. We further acknowledge Ms. Dona Gréta Isai from the University of Kansas Medical Center for her help in the non-invasive image analysis of cardiomyocytes contractility.
Conflict of interest
Aparna R. Chakravarti, Settimio Pacelli, Perwez Alam, Samik Bagchi, Saman Modaresi, Andras Czirok, Rafeeq P.H. Ahmed, and Arghya Paul declare no conflict of interests.
All animal studies were carried out in accordance with the Guide for the Care and Use of Laboratory Animals (NIH publication No. 85-23 revised 1985) and approved by IACUC. No human studies were carried out by the authors for this article. Only commercially obtained cells were used.
Supplementary material 2 (WMV 12,122 kb)
Supplementary material 3 (WMV 60,309 kb)
Supplementary material 4 (WMV 41,200 kb)
- 3.Augiere, C., S. Megy, R. El Malti, A. Boland, L. El Zein, B. Verrier, A. Mégarbané, J.-F. Deleuze, and P. Bouvagnet. A novel alpha cardiac actin (ACTC1) mutation mapping to a domain in close contact with myosin heavy chain leads to a variety of congenital heart defects, arrhythmia and possibly midline defects. PLoS ONE 10:0127903, 2015.CrossRefGoogle Scholar
- 8.Dai, R., Z. Wang, R. Samanipour, K.-I. Koo, and K. Kim. Adipose-derived stem cells for tissue engineering and regenerative medicine applications. Stem Cells Int. 201:19, 2016.Google Scholar
- 13.Fischer, K. M., C. T. Cottage, W. Wu, S. Din, N. A. Gude, D. Avitable, P. Quijada, B. L. Collins, J. Fransioli, and M. A. Sussman. Enhancement of myocardial regeneration through genetic engineering of cardiac progenitor cells expressing Pim-1 kinase: Fischer: Pim-1 kinase enhances myocardial regeneration. Circulation 120:2077–2087, 2009.CrossRefGoogle Scholar
- 18.Haider, H., Y. J. Lee, S. Jiang, R. P. Ahmed, M. Ryon, and M. Ashraf. Phosphodiesterase inhibition with tadalafil provides longer and sustained protection of stem cells. Am. J. Physiol. 299:H1395–H1404, 2010.Google Scholar
- 21.Hu, X., S. P. Yu, J. L. Fraser, Z. Lu, M. E. Ogle, J. A. Wang, and L. Wei. Transplantation of hypoxia-preconditioned mesenchymal stem cells improves infarcted heart function via enhanced survival of implanted cells and angiogenesis. J. Thorac Cardiovasc. Surg. 135:799–808, 2008.CrossRefGoogle Scholar
- 24.Kabaeva, Z. T., A. Perrot, B. Wolter, R. Dietz, N. Cardim, J. M. Correia, H. D. Schulte, A. A. Aldashev, M. M. Mirrakhimov, and K. J. Osterziel. Systematic analysis of the regulatory and essential myosin light chain genes: genetic variants and mutations in hypertrophic cardiomyopathy. Eur. J. Hum. Genet. 10:741–748, 2002.CrossRefGoogle Scholar
- 29.Kubalak, S. W., W. C. Miller-Hance, T. X. O’Brien, E. Dyson, and K. R. Chien. Chamber specification of atrial myosin light chain-2 expression precedes septation during murine cardiogenesis. J. Biol. Chem. 269:16961–16970, 1994.Google Scholar
- 32.Li, W., N. Ma, L.-L. Ong, C. Nesselmann, C. Klopsch, Y. Ladilov, D. Furlani, C. Piechaczek, J. M. Moebius, K. Lützow, A. Lendlein, C. Stamm, R. K. Li, G. Steinhoff. Bcl-2 engineered MSCs inhibited apoptosis and improved heart function. Stem cells (Dayton, OH, U.S.), 25:2118-27, 2007.Google Scholar
- 33.Li, H., S. Zuo, Z. He, Y. Yang, Z. Pasha, Y. Wang, and M. Xu. Paracrine factors released by GATA-4 overexpressed mesenchymal stem cells increase angiogenesis and cell survival. Am. J. Physiol. 299:H1772–H1781, 2010.Google Scholar
- 35.Matsumoto, R., T. Omura, M. Yoshiyama, T. Hayashi, S. Inamoto, K. R. Koh, K. Ohta, Y. Izumi, Y. Nakamura, K. Akioka, K. Takeuchi, and J. Yoshikawa. Vascular endothelial growth factor-expressing mesenchymal stem cell transplantation for the treatment of acute myocardial infarction. Arterioscler. Thromb. Vasc. Biol. 25:1168–1173, 2005.CrossRefGoogle Scholar
- 40.Moulik, M., M. Vatta, S. H. Witt, A. M. Arola, R. T. Murphy, W. J. McKenna, A. M. Boriek, K. Oka, S. Labeit, N. E. Bowles, T. Arimura, A. Kimura, and J. A. Towbin. ANKRD1, the gene encoding cardiac ankyrin repeat protein, is a novel dilated cardiomyopathy gene. J. Am. Coll. Cardiol. 54:325–333, 2009.CrossRefGoogle Scholar
- 41.Nagao, K., N. Sowa, K. Inoue, M. Tokunaga, K. Fukuchi, K. Uchiyama, H. Ito, F. Hayashi, T. Makita, T. Inada, M. Tanaka, T. Kimura, and K. Ono. Myocardial expression level of neural cell adhesion molecule correlates with reduced left ventricular function in human cardiomyopathy. Circ. Heart Failure 7:351–358, 2014.CrossRefGoogle Scholar
- 42.Orr, N., R. Arnaout, L. J. Gula, D. A. Spears, P. Leong-Sit, Q. Li, W. Tarhuni, S. Reischauer, V. S. Chauhan, M. Borkovich, S. Uppal, A. Adler, S. R. Coughlin, D. Y. Stainier, and M. H. Gollob. A mutation in the atrial-specific myosin light chain gene (MYL4) causes familial atrial fibrillation. Nat. Commun. 7:11303, 2016.CrossRefGoogle Scholar
- 43.Ou, L., W. Li, Y. Zhang, W. Wang, J. Liu, H. Sorg, D. Furlani, R. Gäbel, P. Mark, C. Klopsch, L. Wang, K. Lützow, A. Lendlein, K. Wagner, D. Klee, A. Liebold, R. K. Li, D. Kong, G. Steinhoff, and N. Ma. Intracardiac injection of matrigel induces stem cell recruitment and improves cardiac functions in a rat myocardial infarction model. J. Cell. Mol. Med. 15:1310–1318, 2011.CrossRefGoogle Scholar
- 47.Pandey, R., S. Velasquez, S. Durrani, M. Jiang, M. Neiman, J. S. Crocker, J. B. Benoit, J. Rubinstein, A. Paul, and A. Rafeeq. MicroRNA-1825 induces proliferation of adult cardiomyocytes and promotes cardiac regeneration post ischemic injury. Am. J. Transl. Res. 9:3120–3137, 2017.Google Scholar
- 51.Paul, A., A. Hasan, H. A. Kindi, A. K. Gaharwar, V. T. S. Rao, M. Nikkhah, S. R. Shin, D. Krafft, M. R. Dokmeci, D. Shum-Tim, and A. Khademhosseini. Injectable graphene oxide/hydrogel-based angiogenic gene delivery system for vasculogenesis and cardiac repair. ACS Nano. 8:8050–8062, 2014.CrossRefGoogle Scholar
- 63.Tiso, N., M. Majetti, F. Stanchi, A. Rampazzo, R. Zimbello, A. Nava, and G. A. Danieli. Fine mapping and genomic structure of ACTN2, the human gene coding for the sarcomeric isoform of alpha-actinin-2, expressed in skeletal and cardiac muscle. Biochem. Biophys. Res. Commun. 265:256–259, 1999.CrossRefGoogle Scholar
- 65.Trapnell, C., B. A. Williams, G. Pertea, A. Mortazavi, G. Kwan, M. J. van Baren, S. L. Salzberg, B. J. World, L. Pachter. Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat. Biotechnol. 28:511–515, 2010.CrossRefGoogle Scholar
- 66.Turbay, D., S. B. Wechsler, K. M. Blanchard, and S. Izumo. Molecular cloning, chromosomal mapping, and characterization of the human cardiac-specific homeobox gene hCsx. Mol. Med. 2:86–96, 1996.Google Scholar
- 74.Youssef, A., D. Aboalola, and V. K. M. Han. The roles of insulin-like growth factors in mesenchymal stem cell niche. Stem Cells Int. 201:9453108, 2017.Google Scholar