Mesenteric Parametrial Fat Pad Surgery for in vivo Implantation of Hepatocytes in Nude Mice

Abstract

Cell-based liver therapies utilizing functionally stabilized engineered hepatic tissue hold promise in improving host liver functions and are emerging as a potential alternative to whole-organ transplantation. Owing to the ability to accommodate a large ex vivo engineered hepatocyte mass and dense vascularization, the mesenteric parametrial fat pad in female nude mice forms an ideal anatomic microenvironment for ectopic hepatocyte transplantation. However, the lack of any reported protocol detailing the presurgical preparation and construction of the engineered hepatic hydrogel, fat pad surgery, and postsurgical care and bioluminescence imaging to confirm in vivo hepatocyte implantation makes it challenging to reliably perform and test engraftment and integration with the host. In this report, we provide a step-by-step protocol for in vivo hepatocyte implantation, including preparation of hepatic tissue for implantation, the surgery process, and bioluminescence imaging to assess survival of functional hepatocytes. This will be a valuable protocol for researchers in the fields of tissue engineering, transplantation, and regenerative medicine.Key features• Primary human hepatocytes transduced ex vivo with a lentiviral vector carrying firefly luciferase are surgically implanted onto the fat pad.• Bioluminescence helps monitor survival of transplanted hepatic tissue over time.• Applicable for assessment of graft survival, graft-host integration, and liver regeneration.Graphical overview