No abstract

Whole organ engineering has emerged as a promising alternative avenue to fill the gap of donor organ shortage in organ transplantation. Recent breakthroughs in the decellularization of solid organs and repopulation with desired cell populations have generated neo-organ constructs with promising functional outcomes. The realization of this goal requires engineering advancement in the perfusion-based bioreactors to (i) efficiently deliver decellularization agents, followed by (ii) its reconstruction with relevant cell types and (iii) maintenance of viability and function of the repopulated organ. In this study, we report the development and assembly of a perfusion bioreactor with the potential to enable regenerative reconstruction of pancreas. The assembled bioreactor is versatile to efficiently decellularize multiple organs, as demonstrated by complete decellularization of pancreas, liver and heart in the same set-up. Further, the same system is amenable to support organ repopulation with diverse cell types. Using our in-house bioreactor system, we demonstrate pancreas repopulation with both immortalized MIN-6 beta cells and differentiating human pluripotent stem cells. Importantly, we show the significant advantage of perfusion culture over static culture in enhancing cell engraftment, viability and phenotypic maintenance of the repopulated pancreas. In addition, this study is a significant step forward for whole organ engineering as it will facilitate cost-effective and easy assembly of perfusion bioreactors to enable rapid advancement in regenerative organ reconstruction.

Pluripotent stem cells are promising source of cells for tissue engineering, regenerative medicine and drug discovery applications. The process of stem cell differentiation is regulated by multi-parametric cues from the surrounding microenvironment, one of the critical one being cell interaction with extracellular matrix (ECM). The ECM is a complex tissue-specific structure which is an important physiological regulator of stem cell function and fate. Recapitulating this native ECM microenvironment niche is best facilitated by decellularized tissue/organ derived ECM, which can faithfully reproduce the physiological environment with high fidelity to in vivo condition and promote tissue-specific cellular development and maturation. Recognizing the need for organ specific ECM in a 3D culture environment in driving phenotypic differentiation and maturation of hPSCs, we fabricated an ECM array platform using native-mimicry ECM from decellularized organs (namely pancreas, liver and heart), which allows cell-ECM interactions in both 2D and 3D configuration. The ECM array was integrated with rapid quantitative imaging for a systematic investigation of matrix protein profiles and sensitive measurement of cell-ECM interaction during hPSC differentiation. We tested our platform by elucidating the role of the three different organ-specific ECM in supporting induced pancreatic differentiation of hPSCs. While the focus of this report is on pancreatic differentiation, the developed platform is versatile to be applied to characterize any lineage specific differentiation.