Human liver microtissue spheroids in hollow fiber membrane bioreactor.

The aim of this work was to create human liver microtissue spheroids metabolically active by using a hollow fiber membrane bioreactor whose design and structural features ensure a uniform microenvironment and adequate oxygenation. Human hepatocyte spheroids with uniform size and shape were formed through self-assembling and cultured into the bioreactor. Adjacent spheroids fused, giving rise to larger microstructures around the fibers forming liver-like tissue, which retained functional features in terms of urea synthesis, albumin production, and diazepam biotransformation up to 25days. The overall data strongly corroborates that within the bioreactor a proper oxygenation and supply of nutrients were provided to the cells ensuring a physiological amount even in the spheroids core. The oxygen uptake rate and the mathematical modelling of the mass transfer directly elucidated that liver microtissue spheroids are not exposed to any oxygen mass transfer limitation. The minimum oxygen concentration reached at the center of multiple spheroids with diameter of 200µm is significantly higher than the one of the perivenous zone in vivo, while for larger microtissues (400µm diameter) the oxygen concentration drops to values that are equal to the maximum concentration found in the liver periportal zone. Both experimental and modelling investigations led to the achievement of significant results in terms of liver cell performance. Indeed, the creation of a permissive microenvironment inside the bioreactor supported the formation and long-term maintenance of functional human liver microtissues.

3D liver membrane system by co-culturing human hepatocytes, sinusoidal endothelial and stellate cells.

In this study, a designed approach has been utilized for the development of a 3D liver system. This approach makes use of primary human sinusoidal endothelial cells, stellate cells and hepatocytes that are seeded sequentially on hollow fiber membranes (HF) in order to mimic the layers of cells found in vivo. To this purpose modified polyethersulfone (PES) HF membranes were used for the creation of a 3D human liver system in static and dynamic conditions. In order to verify the positive effect of non-parenchymal cells on the maintenance of hepatocyte viability and functions, homotypic cultures of hepatocytes alone on the HF membranes were further investigated. The membrane surface allowed the attachment and self-assembly of the cells, forming tissue-like structures around and between fibers. Sinusoidal cells formed tube-like structures that surrounded hepatocytes organized in cords within aggregates promoted by stellate cells. The co-culture of hepatocytes with sinusoidal endothelial and hepatic stellate cells preserved structural architecture of the construct and improved the liver-specific functions. Most importantly, cells co-cultured in a HF membrane bioreactor synthesized albumin and urea for 28 days. The liver membrane bioreactor also preserved the drug biotransformation activity with a continuous production of diazepam phase I metabolites for an extended period of time. Additionally, the cell oxygen uptake rates highlighted the maintenance of the actual oxygen concentration at a level compatible with their metabolic functions.

Recent Strategies Combining Biomaterials and Stem Cells for Bone, Liver and Skin Regeneration.

This review is focused on the combination of biomaterials with stem cells as a promising strategy for bone, liver and skin regeneration. At first, we describe stem cell-based constructs for bone tissue engineering with special attention to recent advanced approaches based on the use of biomaterial scaffolds with renewable stem cells that have been used for bone regeneration. We illustrate the strategies to improve liver regeneration by using liver stem cells and biomaterials and/or devices as therapeutic approaches. In particular, examples of biomaterials in combination with other technologies are presented since they allow the differentiation of stem cells in hepatocytes. After a description of the role and the benefit of MSCs in wound repair and in skin substitutes we highlight the suitability of biomaterials in guiding stem cell differentiation for skin regeneration and cutaneous repair in both chronic and acute wounds. Finally, an overview of the types of bioreactors that have been developed for the differentiation of stem cells and are currently in use, is also provided. The examples of engineered microenvironments reported in this review indicate that a detailed understanding of the various factors and mechanisms that control the behavior of stem cells in vivo has provided useful information for the development of advanced bioartificial systems able to control cell fate.