In Vivo Engineering of Metabolically Active Hepatic Tissues in a Neovascularized Subcutaneous Cavity

Abstract

Recent success in clinical hepatocyte transplantation therapy has encouraged further investigation into bioengineering hepatic tissues in vivo. Engineering tissues in the subcutaneous space is an attractive method; however, hepatocyte survival has been transient due to insufficient vascular network formation. To establish a vascularized cavity, we created a polyethylene terephthalate mesh device coated with poly(vinylalcohol) that allowed for the gradual release of basic fibroblast growth factor (bFGF), a potent angiogenic factor. The efficacy of the bFGF-releasing device in inducing vascular network formation in the subcutaneous space was observed in mouse and rat studies. Isolated mouse hepatocytes transplanted into newly vascularized subcutaneous cavities allowed for persistent survival up to 120 days. In the absence of a vascularized compartment, the survival of the transplanted hepatocytes was markedly diminished. Functional maintenance of the engineered hepatic tissues was confirmed by high expression of liver-specific mRNAs and proteins. These engineered hepatic tissues have the ability to take up inoculated compounds and express strong induction of drug-metabolizing enzymes, demonstrating functional relevance as a metabolic tissue. In conclusion, we have created a novel technology to engineer functionally active hepatic tissues in the subcutaneous space, which will likely facilitate hepatocyte-based therapies.

Liver Tissue Engineering Utilizing Hepatocytes Propagated in Mouse Livers in Vivo

“…Prior to the liver tissue engineering procedure, the basic fibroblast growth factor (bFGF)-releasing device was inserted into the subcutaneous space on the back of FVB/N mice as described previously (17,30). Ten days after the device insertion, a highly vascularized subcutaneous platform was developed.…”

“…Although this approach has shown considerable modified two-step collagenase perfusion method as previously described (5,11,12,16,17,30). Briefly, the livers promise in recent years, there are several hurdles that need to be overcome for continual advancement to occur were primarily perfused with Hank's balanced salt solution (HBSS) (Sigma, St. Louis, MO) containing 0.09% using hepatocyte-based technologies.…”

“…First, the creation of a vascularized platform using a growth factor-releasing device at the target subcutaneous site prior Liver tissue engineering using primary hepatocytes presents the opportunity to create a de novo liver systo the transplantation of the donor hepatocytes (30). The prevascularization process plays a pivotal role in the surtem, which can lead to a new therapeutic modality towards the treatment of various liver diseases (4,10, vival of the transplanted hepatocytes by providing an adequate blood supply to these transplanted cells, due to 15,18,28) and may have the potential to obviate the need for organ liver transplantation.…”

“…hepatocyte sheets in the subcutaneous space (17). The confirmed functionality of the engineered liver system Hepatocyte Isolation and Purification includes liver-specific protein productions, chemical uptake and subsequent metabolizing enzyme expressions, Hepatocytes were isolated from hA1AT-TG mice or hA1AT hepatocyte-repopulated uPA/SCID mice using a and regenerative growth (17).Although this approach has shown considerable modified two-step collagenase perfusion method as previously described (5,11,12,16,17,30). Briefly, the livers promise in recent years, there are several hurdles that need to be overcome for continual advancement to occur were primarily perfused with Hank's balanced salt solution (HBSS) (Sigma, St. Louis, MO) containing 0.09% using hepatocyte-based technologies.…”

Humanized mice with ectopic artificial liver tissues

“…Use of partial hepatectomy with portal-caval shunt has been shown to improve hepatocyte engraftment and function in allogenic rat models (19,20), suggesting that transplanted hepatocytes depend on signals (hepatotrophic factors) circulating from the portal vein. Despite these findings, support of implanted hepatocytes by sequential delivery of seeded cells, growth factors, hormones, and/or proangiogenic molecules has had limited success (21)(22)(23)(24), and recent evidence showing promise for the survival of hepatocytes transplanted in the ectopic lymph node remains dependent on mouse liver injury (25). We hypothesized that an integrated system, based on hepatocyte encapsulation and the incorporation of juxtacrine and paracrine signals to stabilize the cell phenotype before, rather than during/after transplantation, could mitigate the requirement for portal venous flow or liver injury in vivo.…”