Carla Campana

Human hepatocytes and endothelial cells in organotypic membrane systems

The realization of organotypic liver model that exhibits stable phenotype is a major challenge in the field of liver tissue engineering. In this study we developed liver organotypic co-culture systems by using synthetic and biodegradable membranes with primary human hepatocytes and human umbilical vein endothelial cells (HUVEC). Synthetic membranes prepared by a polymeric blend constituted of modified polyetheretherketone (PEEK-WC) and polyurethane (PU) and biodegradable chitosan membranes were developed by phase inversion technique and used in homotypic and organotypic culture systems. The morphological and functional characteristics of cells in the organotypic co-culture membrane systems were evaluated in comparison with homotypic cultures and traditional systems. Hepatocytes in the organotypic co-culture systems exhibit compact polyhedral cells with round nuclei and well demarcated cell-cell borders like in vivo, as a result of heterotypic interaction with HUVECs. In addition HUVECs formed tube-like structures directly through the interactions with the membranes and hepatocytes and indirectly through the secretion of ECM proteins which secretion improved in the organotypic co-culture membrane systems. The heterotypic cell-cell contacts have beneficial effect on the hepatocyte albumin production, urea synthesis and drug biotransformation. The developed organotypic co-culture membrane systems elicit liver specific functions in vitro and could be applied for the realization of engineered liver tissues to be used in tissue engineering, drug metabolism studies and bioartificial liver devices.

Biodegradable and synthetic membranes for the expansion and functional differentiation of rat embryonic liver cells

The insufficient availability of donor organs for orthotopic liver transplantation worldwide has urgently increased the requirement for new therapies for acute and chronic liver disease. The creation of an unlimited source of donor cells for hepatocyte transplantation therapy and pharmaceutical applications may be the isolation and expansion of liver progenitor cells or stem cells. Here we report the expansion and functional differentiation of rat embryonic liver cells on biodegradable and synthetic polymeric membranes in comparison with traditional substrates, such as collagen and polystyrene culture dishes. Membranes prepared from chitosan and modified polyetheretherketone were used for the culture of liver progenitor cells derived from rat embryonic liver. Cells proliferated, with a significant increase in their number within 8-11 days. The cells displayed functional differentiation showing urea synthesis, albumin production and diazepam biotransformation on all substrates investigated. In particular, on a chitosan membrane liver-specific functions were expressed at significantly higher levels for prolonged times compared with other synthetic membranes, utilizing traditional substrates (collagen and PSCD) as references. These results demonstrate that chitosan membranes offer cells favourable conditions to promote the expansion and functional differentiation of embryonic liver cells that could be effectively used in liver tissue engineering and in pharmaceutical applications.

Flat and tubular membrane systems for the reconstruction of hippocampal neuronal network.

The selection of appropriate biomaterials that promote cellular adhesion and growth is particularly important for the in vitro reconstruction of neuronal network. This study focused on the development of new polymeric membranes in flat and tubular (hollow‐fibre) configurations as novel biomaterials for neuronal outgrowth. Two membrane systems constituted by modified polyetheretherketone (PEEK‐WC) and polyacrylonitrile (PAN) membranes were developed and used for the culture of hamster hippocampal neurons. We demonstrated that all investigated membranes supported the adhesion and growth of hippocampal neurons enhancing neuronal differentiation and neurite alignment. The differences in cell behaviours between cells cultured on flat and hollow‐fibre (HF) membranes were highlighted by the quantitative analysis of neuronal marker fluorescence intensity, morphometric analysis, RT–PCR analysis and also by metabolic activity measurements. In particular, the PAN HF membranes showed ideal growth culture conditions, guaranteeing adequate levels of metabolic features. Primary hippocampal cells cultured on PAN HF membranes were able to recreate in vitro a 3D neural tissue‐like structure that, mimicking the hippocampal tissue, could be used as a tool for the study of natural and pathological neurobiological events. Copyright

Membrane Approaches for Liver and Neuronal Tissue Engineering

Many advances have been made in the last century to replace and restore damaged organs and tissues due to the increase of the aging population and the shortage of suitable organs for transplantation. Tissue engineering approaches based on the combination of smart biomaterials with advanced cell therapy should closely mimic and replicate the physical and biochemical milieu of the natural tissues and organs. Many efforts have been made to improve the design of natural and synthetic, resorbable and nonbiodegradable biomaterials in supporting cell adhesion, differentiation, and proliferation toward the formation of a new tissue. Micro- and nanostructured membranes have a great potential to create a biomimetic environment for cells providing them the biochemical, physical, and mechanical cues, which are necessary for the tissue formation. Tailor-made membranes are designed according to well-defined engineering criteria, taking into account the chemical composition and the physicochemical, morphological, and transport properties. These systems on the basis of their characteristics compartmentalize cells in a well-controlled microenvironment at molecular level providing a selective mass transfer of nutrients and metabolites to and from cells ensuring the system homeostasis. In this article, advances achieved in the development of liver and neuronal membrane systems as bioartificial organs and tissues were reviewed focusing on their evaluation in both the preclinical and clinical systems and their validation as in vitro platforms.