Alfredo J. Fabrega

Transplantation of Hepatocytes: An In-Vitro and In-Vivo Study in Canines

We isolated and transplanted hepatocytes in the canine, large animal model to evaluate hepatocyte yield and purity as well as the optimal site for hepatocyte engraftment (i.e., the spleen or the portal bed). We obtained viable, pure, single hepatocyte suspensions that were readily preserved at 4°C in University of Wisconsin (UW) solution for up to 3 days. Both intrasplenic and portal vein injection were well tolerated, with minimal recipient morbidity and mortality when 1-2 × 109 hepatocytes were injected into immunosuppressed allogeneic hosts. We noted the embolization of hepatocytes into the parenchyma of the native liver within 7 days of intrasplenic transplantation that produced a mild reversible derangement of liver function and histology. These results warrant consideration prior to clinical trials of hepatocyte transplantation in man.

Hepatitis C infection in potential recipients with normal liver biochemistry does not preclude renal transplantation

The hepatitis C virus (HCV) may be an important cause of chronic liver disease in renal transplant recipients. We investigated retrospectively the incidence and outcome of HCV infection in long-term renal transplant recipients and patients on hemodialysis. Stored, pretransplant sera of transplant recipients with normal liver biochemistry at surgery were tested for hepatitis C by a second-generation enzyme immunoassay. Hemodialysis patients were tested by a first-generation enzyme-linked immunosorbent assay (ELISA) against c100-3. We studied 252 renal transplant recipients and 58 hemodialysis patients followed for 65±10 months and 26±6 months, respectively. Fifteen percent (38/252) of the transplant recipients were HCV positive as were 3/58 (5%) of the hemodialysis patients. Over liver disease occurred in 22/252 (8.7%) transplant recipients and none in the hemodialysis group. Thirty-six percent (8/22) of transplant recipients with overt liver disease were HCV positive. No HCV-positive patients died of liver failure. Of six biopsies in the HCV-positive transplant group, two had histological evidence of CAH. CAH was seen in six of eight biopsies in the HCV-negative transplants and two of these latter patients progressed to cirrhosis. No hemodialysis patients had clinical or histological evidence of chronic liver disease. Two HCV-negative transplant patients died of liver failure, while no deaths related to liver disease occurred in hemodialysis patients regardless of HCV status. We conclude that hepatitis C may cause chronic hepatitis in renal transplant patients. However, chronic liver disease in HCV-positive renal transplant recipients appears to be a clinically and histologically benign entity. HCV-positive potential renal allograft recipients with normal liver biochemistry should not be excluded from renal transplantation.

The morphology and function of rabbit hepatocytes isolated using ethylenediaminetetraacetate.

The enzyme digestion technique using collagenase is used in most studies that describe hepatocyte isolation, including those in which hepatocytes are isolated for transplantation. The use of collagenase, however, has several drawbacks. We describe the use of a highly concentrated ethylenediaminetetraacetate solution for isolation of hepatocytes followed by purification using a Percoll gradient in the rabbit model. Isolated hepatocytes were then preserved at 4 degrees C for up to three days in either University of Wisconsin solution or Dulbeccos modified eagles medium. Morphological studies of hepatocytes at 24 hr and 72 hr in either medium were performed using Papanicolaou and PAS stains of cytopreparations for the presence of glycogen. Similarly, functional studies of the preserved hepatocytes included LDH release, total tissue water content, and amount of 99m-technetium mebrofenin uptake. The use of EDTA perfusion for hepatocyte isolation was highly reproducible in this model. The cell yield was comparable to that achieved previously using collagenase. The morphologic studies demonstrated pure hepatocytes with well-preserved architecture when preserved for up to three days in UW solution. Functional studies showed a statistically significant lower LDH release and total tissue water content and a higher technetium-99m mebrofenin uptake for hepatocytes preserved in UW solution. We conclude that a pure and viable hepatocyte suspension can be obtained from rabbit livers using concentrated EDTA solutions. These hepatocytes can be well preserved at 4 degrees C for up to 72 hr in UW solution, based on morphologic and functional criteria.

Pancreas-Kidney Transplantation for Intensivists: Perioperative Care and Complications

Simultaneous pancreas-kidney transplantation is a therapeutic option for type I diabetics with end-stage renal disease. It aims to correct the uremic state, to normalize glucose hemeostasis, and to ameliorate diabetic complications. Careful donor-recipient selection and meticulous intra-operative and postoperative care will substantially impact recipient morbidity. An understanding of the technical aspects of the surgical procedure and its metabolic and immunological consequences is necessary to successfully manage a pancreas-kidney transplant recipient, many of whom are nursed in intensive care units. A successful outcome is predicted in early recognition of technical complications and aggressive management of rejection to achieve the current 1-year graft survival rates of 75% for pancreas transplants and 84% for kidney transplants.