Nobuyuki Kikuchi

Technique for orthotopic reduced-size hepatic transplantation combined with ex vivo liver cut down in the rat

A technique is described for orthotopic reduced-size hepatic transplantation combined withex vivo liver cut down in the rat. Following perfusion of the donor liver with cold heparinized saline, the portal veins, bile ducts, and hepatic arteries to the median and left lobes together were dissectedin situ, encircled, and divided. After harvesting the donor liver, a hepatectomy was performed byex vivo liver cut down of the median and left lobes. The remnant amounted to 32% of the whole liver. As a result, the suprahepatic vena cava could be well visualized with adequate exposure for vascular anastomosis. Orthotopic reduced-size hepatic transplantation was performed using the right and caudate lobes of the liver. The suprahepatic vena cava was anastomosed with a 7-0 silk running suture. A simplified cuff without processes was made with an obliquely cut polyethylene tube and used for the portal and infrahepatic caval anastomoses. A Teflon tube stent was used for the biliary anastomosis. The newly devised angled clamp and flexible arm were used for the cuff attachment and operative procedure. Transplant survival followingex vivo liver cut down was as good as that with whole liver transplantation. Reestablishment of the hepatic artery restores liver function following transplantation. The maximum hepatocyte labeling index (LI) occurs 24 hr after a 68% hepatectomy, and at 36 hr following a reduced-size hepatic transplantation with or without hepatic arterialization. Possible explanations for the slight delay in achieving the maximal LI may include damage that is induced by the operation itself, pregraft preservation, and reperfusion injuries. In conclusion, the anatomical features of the hepatic lobes in rats are well suited to successful completion ofex vivo liver cut down.

Effect of cyclosporine on liver regeneration after orthotopic reduced-size hepatic transplantation in the rat

These experiments were undertaken to study the effects of cyclosporine A (CsA) on liver regeneration after an isogeneic orthotopic reduced-size hepatic transplantation (RSHT) in rats. Male Wistar rats were treated with or without a daily injection of CsA beginning 24 hr before surgery and were subjected to a 68% partial hepatectomy. A isogeneic orthotopic reduced-size hepatic transplantation was performed in recipient rats pretreated with or without CsA. A daily injection of CsA was continued until the recipient rats were sacrificed. Animals were sacrificed at various time points (12, 24, 36, 48, and 72 hr) postoperatively. The incorporation of bromodeoxyuridine (BrdU) into the DNA of the remnant hepatocytes was evaluated by immunohistochemical staining with a monoclonal antibody against BrdU. CsA (10 mg/kg/day) significantly augmented BrdU incorporation into hepatocytes after hepatectomy. The maximum labeling index (LI) was observed at 24 hr after hepatectomy. In contrast, the maximum LI in the recipient rats not receiving CsA was seen at 36 hr after RSHT, and 10 mg/kg/day of CsA decreased the LI at 36 hr after RSHT. A lower dose of CsA (3 mg/kg/day), however, significantly increased the LI in the recipient rats (P<0.01), and it reached a peak at 24 hr after RSHT when compared to the transplant recipients not receiving CsA. The time course of the increase in the LI in the transplant recipient rats receiving 3 mg/kg/day of CsA was similar to that observed in the rats after hepatectomy. This dosage improved the delay in the reduced-size hepatic transplant LI reaching its peak. These findings suggest that after RSHT the liver graft is more sensitive to both hepatotrophic and hepatotoxic effects of CsA.

PROLONGED SURVIVAL OF RAT HEPATIC ALLOGRAFTS AFTER TOTAL-BODY IRRADIATION OF THE DONORS

&NA; The effect of total-body irradiation of the donor on hepatic allograft survival was studied in the rat, with ACI(RT1a) as the donor and LEW(RT1) as the recipient. LEW recipients of ACI liver transplants experienced severe acute rejection, with a mean survival of only 10.2±0.3 days. The doses of irradiation were 450, 750, and 1000 rads administered 24 hr prior to harvesting or subsequent transplantation. TBI with a dose of 750 rads significantly prolonged the survival of the hepatic allograft to 30.3±1.7 days, without concomitant immunosuppression. However, neither 450 rads nor 1000 rads of TBI resulted in successful suppression of graft rejection. TBI appeared to have a beneficial effect on hepatic allograft survival and to have no deleterious effect on isograft survival, suggesting a possible modulation of the immunogenicity of the donor organ. Although the cause of this beneficial effect is not clear, TBI with a dose of 750 rads 24 hr prior to organ harvest seems to be optimal to eliminate-antigen presenting cells in the donor organs.

THE TIME COURSE OF CELL-MEDIATED LYMPHOLYSIS IN RAT HEPATIC ALLOGRAFT RECIPIENTS PRETREATED WITH A SINGLE DONOR-SPECIFIC BLOOD TRANSFUSION

A single intravenous injection of 1 ml freshly heparinized donor blood seven days before transplantation significantly prolonged the survival of subsequent donorspecific hepatic allografts in the fully allogeneic ACI(RT1a-to-LEW(RT11) rat combination. The time course of cell-mediated lympholysis was studied in this animal model. The activity of CML in lymphocytes infiltrating into the hepatic allograft (CML-G) and in the spleen (CML-S) was determined by measuring % lysis of donor Con A blast cervical lymph node cells. Preoperative DST resulted in an increased activity of CML-S with a peak (31.6%, E/T = 150) on day 7. This increased CML-S activity after DST rapidly declined during the first days following hepatic transplantation. The activities of both CML-S and CML-G then increased after transplantation and reached peaks on days 15 (48%, E/T = 150) and 20 (2.57%, E/T = 75), respectively. These were much higher than the peak values of CML-S (11.2%, E/T = 150) on day 7 and CML-G (19.5%, E/T = 75) on day 6 in untreated controls and were followed by a subsequent gradual decrease in those activities to pre-operative levels by day 113 posttransplant. Phenotypic analysis of lymphocytes infiltrating grafts in DST-treated hosts demonstrated that the CD4/CD8 ratio remained relatively constant (<1.0). While the ratio in control grafts increased and reached a peak (2.17) on day 9. Histological examination revealed that mononuclear cell infiltration of grafts reached a peak on day 9 in both DST-treated hosts and controls. This mononuclear cell accumulation gradually subsided in DST-enhanced grafts. The mitotic index of graft hepatocytes reached a peak on day 15 in DST-treated hosts and on day 7 in control. The evidence of prolonged survival of hepatic grafts in recipients pretreated with DST, despite the presence of cytotoxic T cells with increased CML activity in vitro, suggests that effector cytotoxic cell activity may not be necessary for rat liver allograft rejection and that there