Session 6: Regenerative and Bioinspired Material I

Abstract

Direct stem cell encapsulation and cardiac differentiation within supporting biomaterial scaffolds are critical for reproducible and scalable production of the functional human tissues needed in regenerative medicine and drug-testing applications. Producing cardiac tissues directly from pluripotent stem cells rather than assembling tissues using pre-differentiated cells can eliminate multiple cell-handling steps that otherwise limit the potential for process automation and production scale-up. Previously having produced 3D cardiac tissues directly from human pluripotent stem cells (hPSCs) through hiPSC encapsulation and direct differentiation in PEG-fibrinogen (PEG-Fb), this study established a rapid, one-step hiPSC microsphere encapsulation system and used it to directly produce functional cardiac tissue microspheres in suspension culture.Following suspension of hiPSCs in PEG-Fb precursor solution, microspheres were rapidly formed (<1 sec) within a custom built microfluidic device through emulsification and photo-crosslinked using a full spectrum visible light source (day -3). Cell-laden microspheres were maintained three days in mTeSR-1 media and then differentiation was initiated (day 0). Microspheres were highly uniform and encapsulated hiPSCs remained viable and continued to divide, forming dense cell-laden spheroids. Following initiation of differentiation (day 0), microsphere diameter continued increasing as a result of cell growth. First isolated areas of contraction were typically seen on day 8 of differentiation, with microspheres establishing synchronous tissue-level contraction by day 14 and retaining their function for several months. Microspheres supported high efficiency cardiac differentiation with 71.6±8.4% cTnT+, 7.1±1.7% cTnT+/Ki67+, and 8.4±6.5% P4HB+ cells (day 20, n=3 separate batches). Microsphere cardiomyocytes (CMs) could be exogenously paced up to 6.0 Hz and responded appropriately to isoproterenol and propranolol. CMs stained positive for αSA with aligned and well-defined sarcomeres.In summary, rapid encapsulation of hiPSCs in PEG-Fb microspheres supported high-efficiency, direct differentiation to obtain 3D cardiac microspheres. Direct cardiac tissue formation through hPSC encapsulation in PEG-Fb provides the ability to guide cardiac differentiation, to produce functional CMs, and to obtain highly reproducible cell-laden microspheres in scalable suspension culture. Abstracts -Annual Meeting of the German Society for Biomaterials 2017 -Würzburg, November 09 -11 • DOI 10.1515/bmt-2017-6006 Biomed. Eng. -Biomed. Tech. 2017; 62(s2): p34-p39 • © by Walter de Gruyter • Berlin •