A Feasible Method for Quantifying Living Pancreatic Human Islets in Murine Livers Posttransplantation by Confocal Laser Scanning Microscopy

Abstract

Current histological methods cannot accurately determine the survival rate of human pancreatic islets following portal vein infusion. This is due, in part, to the low number of infused islets relative to the whole liver. In this study, we assessed the ability of confocal laser scanning microscopy (CLSM) to track human islets posttransplantation. Immunodeficient mice were transplanted with human islets. Following engraftment, animals were euthanized, livers procured, and human islet β cells immunofluorescently labeled with an insulin-specific antibody and evaluated by CLSM. A calibration curve comparing the area of insulin + hepatic islet β cells to the number of human islets collected was developed. Levels of human C-peptide were measured in transplant recipients to determine islet function. The short-term survival rate of islet transplants was defined as y = 0.0422x + 2.7008, in which x is human islet number and y is liver islet β cell area. Employing CLSM, human islets were detected in immunofluorescent labeled murine liver tissue sections posttransplantation. The β cell-relative area of human islets in 500 islet equivalent (IEQ) specimens was 20.21 ± 1.16 mm and in 1000 IEQ specimens 39.4 ± 2.23 mm posttransplantation. Human islet posttransplant survival rates were 82.9 ± 5.50% (500 IEQ group) and 86.9 ± 5.28% (1000 IEQ group). These data indicate that CLSM can be employed to quantify and characterize pancreatic human islets after transplantation to murine livers.