Cell and tissue engineering for liver disease

Abstract

Despite the tremendous hurdles presented by the complexity of the liver's structure and function, advances in liver physiology, stem cell biology and reprogramming, and the engineering of tissues and devices are accelerating the development of cell-based therapies for treating liver disease and liver failure. This State of the Art Review discusses both the near- and long-term prospects for such cell-based therapies and the unique challenges for clinical translation.

Figure 1. Structure of the Liver

The liver is the largest internal organ in the body and performs over 500 functions, including numerous metabolic, synthetic, immunologic, and detoxification processes. (A) The liver exhibits a hierarchical structure consisting of repeated functional tissue units (liver lobules). Within a lobule, oxygenated blood enters through branches of the hepatic artery and portal vein, and flows in specialized sinusoidal vessels towards the central vein. Bile, that is produced and excreted by hepatocytes, flows in the counter direction towards the intrahepatic bile duct. (B) Hepatocytes are polarized epithelial cells that interact closely with a number of non-parenchymal cells types along the sinusoidal tracts of the liver lobule. Collectively, these cellular components and multiscale tissue structures contribute to the diverse functional roles of the liver.

Figure 2. The Liver in Health and Disease

Mechanisms that lead to hepatocyte damage and reduce liver function include drug-mediated toxicity, alcohol-induced and nonalcoholic fatty liver disease, hepatotrophic infections, and hereditary disorders. Fatty liver disease, resulting from both chronic alcohol exposure as well as nonalcoholic mechanisms, is increasingly common and leads to the chronic accumulation of fat droplets within the liver. Liver cirrhosis can be caused by hepatitis virus infection, autoimmune processes, chronic alcohol abuse, as well as chronic inflammation and fat accumulation. Cirrhosis is characterized by alterations in the sinusoidal structure and function of the liver and the accumulation of extracellular matrix, which is commonly referred to as scarring. These alterations lead to a reduction in hepatic function that can progress to hepatic failure and increased risk of hepatocellular carcinoma.

Figure 3. Organ transplantation and cell-based therapies

Currently, liver transplantation is the primary treatment for liver failure and the only therapy shown to improve survival in patients with liver failure. Due to the limited number of livers suitable for transplantation, advanced surgical procedures including split liver and partial donor transplants have been pursued clinically. Additionally, a diverse range of cell-based therapies are currently being explored to treat liver disease and liver failure. These include the transplantation of various adult and stem cell-derived cell populations, the development of extracorporeal bioartificial liver devices, and the implantation of engineered tissues.

Figure 4. Extracorporeal bioartificial liver devices

Extracorporeal bioartificial liver devices incorporate functional liver cells and aim to provide an array of important liver functions (detoxification, metabolic, synthetic) for a patient by processing the patient’s blood/plasma outside of the body. This approach could serve as a temporary support and bridge until a liver becomes available for transplantation. Currently, such devices are either in the clinical trial or exploratory research stage. Liver cell-based bioreactor designs primarily fall into four general categories based on device configuration. These include hollow fiber devices, packed beds, flat plate systems, and encapsulation-based reactors. The majority of current clinical trials utilize a hollow fiber design in which cells are positioned outside the fibers and the patient’s blood/plasma is perfused through the fiber lumen. Cell sources include three categories; currently existing tumor cell lines, porcine and human primary hepatocytes, and hepatic cells derived from embryonic stem cells (ESC), induced pluripotent stem cells (iPSC), or reprogrammed from other cell types. Due to their increased availability compared to primary human hepatocytes, primary porcine hepatocytes are the most common cellular component of current bioartificial liver devices. Device design characteristics have been shown to affect the functional stability of the cellular components. Many design challenges exist including the balanced delivery of oxygen and nutrients to the cells, preventing mechanical shear forces from damaging the cells, and clinically relevant scale-up.

Figure 5. In vitro culture systems for hepatocytes

Improved in vitro systems have been developed for culturing primary human hepatocytes. Elucidating the roles of many microenvironmental signals in governing hepatocellular processes has enabled optimization of in vitro culture systems. These optimized systems include co-cultures to provide specific cell-cell interactions as well as culture conditions with defined concentrations of soluble factors and extracellular matrix molecules (left). The application of microfabrication technologies to in vitro hepatic tissue engineering has facilitated the control of culture platforms down to the microscale, such as the patterning of co-culture configurations (middle). Further, natural and synthetic biomaterial systems have been applied towards the optimization of three-dimensional in vitro culture platforms (right). Collectively, these engineering approaches have further advanced the understanding of how combinations of microenvironmental cues influence cell functions, and provided important insights into the temporal and spatial dynamics of hepatic cell and tissue function.

Figure 6. Sources of Hepatocytes

Obtaining appropriate sources of hepatocytes is a major limitation for developing cell-based therapies for treating liver disease. Many different approaches are under investigation including methods for improving the expansion of primary human hepatocytes in vitro, the directed differentiation of pluripotent stem cells (both embryonic stem cells and induced pluripotent stem cells), the differentiation of either intrahepatic or extrahepatic adult progenitor cells, as well as new methods for the direct reprogramming of hepatic cells from adult somatic cells.

Figure 7. Liver cell and tissue engineering

Progress in the field of liver cell and tissue engineering serves a bidirectional role as both a means for establishing robust model systems for investigating the human liver in health and disease, as well as the foundation for the development of new cell-based therapies. Consequently, applications exist on a continuum ranging from fundamental in vitro studies (left), to engineered approaches for interfacing with animal models (middle), and finally to translational clinical applications (right). In order to further understand human liver function and disease processes, engineered culture platforms can serve to complement animal models. Concurrently, the foundation of novel cell-based therapies is based on advances in cell and tissue engineering and the progression of these technologies from relevant animal disease models to clinical settings.