Health issues associated with liver diseases afflict millions of individuals and account for over 70,000 deaths annually in the United States. Due in part to an aging population, liver diseases are expected to rise significantly over the next two decades, increasing the need for more effective treatment therapies and increased success rates with transplants. Unfortunately, there are no effective treatments to curb the pathology and there remains a shortage of available livers for transplantation. This challenge is further compounded with alloreactive responses leading to transplant rejection. However, a viable solution is the use of a model liver systems that accurately mimic the biomechanical and biochemical functioning of in vivo liver tissue. Additionally, alternative methods to expand recipient autologous hepatic cells while maintaining function would serve as efficient methods to generate liver systems for transplantation. However, while liver models for in vivo use have been attempted, none have yet successfully expanded autologous hepatic cells in vitro followed by successful implantation to alleviate liver failure in recipients using an in vivo model system. My laboratory has recently demonstrated success in this approach, where we have established an effective in vitro 3D hepatocyte culture system for rapid expansion. Furthermore, our preliminary work shows great promise in applying the system for in vivo adoptive implantation using our innovative in-house designed 3D scaffold system. Therefore, this proposal's objective is to develop a method for rapid expansion of hepatic cells in a novel 3D printed bioscaffold for assembly of a liver organoid for in vivo tissue restoration and ex vivo drug screening. The central hypothesis is that primary hepatic cells seeded in a novel biomaterial scaffold will display similar metabolic function, structure, and biomechanical properties to that of the original liver tissues. The success of this approach will restore liver function following transplantation in a liver-damaged mouse model. The innovative combination of rheological biomaterial tuning, 3D bioprinting, and culture methods that utilize a novel bioscaffold will be applied in pursuit of two specific aims: 1) Engineering an ex vivo model for screening therapeutic drugs targeting hepatocytes through 3D printed bioscaffolds and 2) Development of an implantable hepatic organoid for in vivo tissue restoration to alleviate liver failure in a mouse model. Dedicated equipment for high resolution bulk rheological measurements will support 1) characterization of liver viscoelasticity through bulk shear rheology 2) evaluation of complex moduli of hepatocytes growing in a novel biomaterial, and 3) correlation of multiscale structural information to bulk data. These investigations will establish a platform for novel mechanically tuned 3D culture systems for both rigorous in vitro diagnostic screening and for in vivo adoptive transfer approaches to physiol...