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Pulmonary Vascular Engineering Using Induced Endothelial Progenitor-Like Cells
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A2676 - Pulmonary Vascular Engineering Using Induced Endothelial Progenitor-Like Cells
Author Block: T. Suzuki1, T. Waddell2, G. Karoubi2; 1Thoracic Surgery, Institute of development, aging and cancer, Sendai, Japan, 2Latner Thoracic Surgery Research Laboratories, University Health Network, Toronto, ON, Canada.
BACKGROUND A critical step in whole lung engineering is the achievement of a fully vascularized organ scaffold. While several cell sources have been investigated, vascular integrity after recellularization remains limited. Recent reports have identified partially reprogrammed cell populations en route to pluripotency. We’ve previously shown that timed expression of reprogramming factors OCT4, Klf-4, Sox2, and c-Myc (OKSM) in pulmonary epithelial cells results in generation of lung-specific induced progenitor cells (iPL) which demonstrate phenotypic reversal to their original cell state after withdrawal of reprogramming factors. Here we hypothesize that the transient expression of reprogramming factors will generate endothelial-specific iPLs with high proliferative potential and lineage restricted differentiation capacity, that will be suitable for organ specific vascular engineering. MATERIALS AND METHODS Primary lung endothelial cells were isolated from R26rtTA;Col1a14F2A mice and OKSM was induced via culture in the presence of doxycycline. After the induction period, the cells were subjected to our newly developed mouse-scale perfusion-based bioreactor system. RESULTS A 3-week induction period resulted in a 200-fold expansion of lung endothelial cells. Cells were able to grow after multiple freeze-thaw cycles in a transgene dependent manner. Endothelial-iPLs lost CD31 expression, which they recovered after being injected into decellularized lung scaffolds. Endothelial coverage achieved in 3-day bioreactor culture was 60.1% (n=4) of native lungs. Interestingly, the engineered vasculature recapitulated segment specific vascular marker expression. DISCUSSION Interrupted reprogramming significantly enhances endothelial cell growth while maintaining endothelial phenotype. The mouse-scale whole lung engineering platform developed in this study was useful to test multiple types of cells.
Author Block: T. Suzuki1, T. Waddell2, G. Karoubi2; 1Thoracic Surgery, Institute of development, aging and cancer, Sendai, Japan, 2Latner Thoracic Surgery Research Laboratories, University Health Network, Toronto, ON, Canada.
BACKGROUND A critical step in whole lung engineering is the achievement of a fully vascularized organ scaffold. While several cell sources have been investigated, vascular integrity after recellularization remains limited. Recent reports have identified partially reprogrammed cell populations en route to pluripotency. We’ve previously shown that timed expression of reprogramming factors OCT4, Klf-4, Sox2, and c-Myc (OKSM) in pulmonary epithelial cells results in generation of lung-specific induced progenitor cells (iPL) which demonstrate phenotypic reversal to their original cell state after withdrawal of reprogramming factors. Here we hypothesize that the transient expression of reprogramming factors will generate endothelial-specific iPLs with high proliferative potential and lineage restricted differentiation capacity, that will be suitable for organ specific vascular engineering. MATERIALS AND METHODS Primary lung endothelial cells were isolated from R26rtTA;Col1a14F2A mice and OKSM was induced via culture in the presence of doxycycline. After the induction period, the cells were subjected to our newly developed mouse-scale perfusion-based bioreactor system. RESULTS A 3-week induction period resulted in a 200-fold expansion of lung endothelial cells. Cells were able to grow after multiple freeze-thaw cycles in a transgene dependent manner. Endothelial-iPLs lost CD31 expression, which they recovered after being injected into decellularized lung scaffolds. Endothelial coverage achieved in 3-day bioreactor culture was 60.1% (n=4) of native lungs. Interestingly, the engineered vasculature recapitulated segment specific vascular marker expression. DISCUSSION Interrupted reprogramming significantly enhances endothelial cell growth while maintaining endothelial phenotype. The mouse-scale whole lung engineering platform developed in this study was useful to test multiple types of cells.
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