Efficient long-term survival of cell grafts after myocardial infarction with thick viable cardiac tissue entirely from pluripotent stem cells

Abstract

Poor engraftment of cells after transplantation to the heart is a common and unresolved problem in the cardiac cell therapies. We previously generated cardiovascular cell sheets entirely from pluripotent stem cells with cardiomyocytes, endothelial cells and vascular mural cells. Though sheet transplantation showed a better engraftment and improved cardiac function after myocardial infarction, stacking limitation (up to 3 sheets) by hypoxia hampered larger structure formation and long-term survival of the grafts. Here we report an efficient method to overcome the stacking limitation. Insertion of gelatin hydrogel microspheres (GHMs) between each cardiovascular cell sheet broke the viable limitation via appropriate spacing and fluid impregnation with GHMs. Fifteen sheets with GHMs (15-GHM construct; >1 mm thickness) were stacked within several hours and viable after 1 week in vitro. Transplantation of 5-GHM constructs (≈2 × 106 of total cells) to a rat myocardial infarction model showed rapid and sustained functional improvements. The grafts were efficiently engrafted as multiple layered cardiovascular cells accompanied by functional capillary networks. Large engrafted cardiac tissues (0.8 mm thickness with 40 cell layers) successfully survived 3 months after TX. We developed an efficient method to generate thicker viable tissue structures and achieve long-term survival of the cell graft to the heart.

Impact of Cell Composition and Geometry on Human Induced Pluripotent Stem Cells-Derived Engineered Cardiac Tissue

“…ECTs were incubated with staining solution (50 mL/L Ethidium Homodimer III and 50 mL/L Hoechst 33342, PromoCell GmbH, Heidelberg, Germany) in a pH-adjusted buffer for 60 minutes at room temperature and protected from light22. Fluorescent images were obtained using an Olympus DP72 optical microscope (Olympus, Tokyo, Japan).…”

“…Several ECT scale-up strategies have been described including pre-vascularization2021, stacking cell sheets22, scalable scaffolds23, and bioprinting24. Guided by initial works from the Bursac lab using Polydimethylsiloxane (PDMS) molds to form porous engineered tissues from neonatal rat skeletal myoblasts and CMs25, human ESC-derived CMs26, and mouse iPSCs27, we fabricated a range of mold geometries from 0.5mm thick PDMS sheets.…”

Pirfenidone controls the feedback loop of the AT1R/p38 MAPK/renin-angiotensin system axis by regulating liver X receptor-α in myocardial infarction-induced cardiac fibrosis

“…p38 MAPK, a major member of the MAPKs, has been shown to play a vital role in the development of cardiac fibrosis, MI, and cardiac hypertrophy4. Recent studies have suggested the involvement of the AT1R/p38 MAPK pathway in pancreatic fibrosis5, renal tubulointerstitial fibrosis6, and peritoneal fibrosis7.…”

Stem cell-derived cell sheet transplantation for heart tissue repair in myocardial infarction

“…Therefore, in vitro cell-dense tissues with the desired thickness can be fabricated by multiple cell sheet layering. Additionally, Matsuo et al inserted gelatin hydrogel microspheres (GHMs) between each cardiovascular cell sheet to break the viability limitation via appropriate spacing and fluid impregnation with GHMs [86]. Fifteen sheets with GHMs (> 1 mm thickness) were stacked within several hours and viable after 1 week in vitro.…”