Soluble Thrombomodulin Ameliorates Ischemia-Reperfusion Injury of Liver Grafts by Modulating the Proinflammatory Role of High-Mobility Group Box 1.

Transplantation using grafts obtained after cardiac death (CD) is considered a promising solution for graft shortages. However, no standard criteria for organ preservation have been established for CD donors. High-mobility group box 1 (HMGB1) is a DNA-binding protein that is released from dying hepatocytes as an early mediator of inflammation and organ tissue damage. HMGB1 stimulates immunocytes to produce inflammatory cytokines, thereby amplifying the inflammatory response. Thrombomodulin is an integral membrane protein that functions as an endothelial anticoagulant cofactor, and it binds HMGB1 through the extracellular domain. We investigated the effects of ART-123, recombinant human soluble thrombomodulin, on warm ischemia-reperfusion injury in liver grafts. Male Wistar rats were divided into four ex vivo groups: heart-beating (HB) group, in which livers were isolated from HB donors; CD group, in which livers were isolated from CD donors exposed to apnea-induced conditions and warm ischemic conditions for 30 min after cardiac arrest; and two CD groups pretreated with ART-123 (1 or 5 mg/kg). Each isolated liver was reperfused for 1 h after cold preservation for 6 h. The perfusate levels of HMGB1, LDH, TNF-α, and IL-6 were significantly lower in the CD group pretreated with ART-123 (5 mg/kg) than in the CD group. Bile production was significantly higher in the CD group pretreated with ART-123 (5 mg/kg) than in the CD group. The sinusoidal spaces were significantly narrower in the CD group than in the other groups. We propose that ART-123 maintains sinusoidal microcirculation by reducing endothelial cell damage during warm ischemia-reperfusion injury.

Hepatic venous reconstruction using the superficial femoral vein in a right-lobe living donor liver transplant patient with interrupted inferior vena cava.

Anatomical abnormalities in patients with BA often include polysplenia, preduodenal portal vein, interrupted retrohepatic IVC, cardiac abnormalities, and situs inversus. In LDLT patients who had congenital vascular anomalies, additional surgical modifications for the reconstruction of hepatic venous branches are sometimes necessary to prevent venous parenchymal congestion. We report a 12-yr-old female with post-Kasai BA with interrupted retrohepatic IVC who underwent right-lobe LDLT because the left liver graft volume was insufficient. The donor right liver graft had three major hepatic branches, including the RHV, IRHV, and MHV tributary (V8). We performed hepatic venous reconstruction by creating a large, wide triple orifice consisting of the RHV and two SFVs, which were anastomosed to the V8 and IRHV using the donor's SFV as an interposition graft. In conclusion, the reconstruction of venous orifices for right-lobe LDLT patients with the absent retrohepatic IVC is can be carried out using an SFV graft derived from the living donor or the recipient.

A new liver graft preparation method for uncontrolled non-heart-beating donors, combining short oxygenated warm perfusion and prostaglandin E1.

BACKGROUND: To resolve the shortage of donors associated with liver transplantation, the potential uncontrolled non-heart-beating donor (UNHBD) pool is expected to increase. However, warm ischemia-reperfusion injury leads to inferior survival in transplantation using the grafts from UNHBD compared with those from heart-beating donors. To overcome this problem, we developed a new method for preparation of liver grafts from UNHBDs consisting of a combination of short oxygenated warm perfusion (SOWP) and prostaglandin E1 (PGE1). METHODS: Using an ex vivo perfusion rat model, we examined the effectiveness of this new method. RESULTS: Using SOWP and PGE1 treatment, the total amount of bile production during reperfusion in UNHBD grafts was increased to the same level as that in the heart-beating donor grafts. The addition of PGE1 to SOWP buffer decreased aspartate aminotransferase/alanine aminotransferase and tumor necrosis factor α levels during 1 h of reperfusion. Necrosis and apoptosis were significantly decreased by SOWP + PGE1 treatment. SOWP + PGE1 ameliorated induction of mitochondrial permeability transition, and the total amount of mitochondrial cytochrome c in the SOWP + PGE1 group after reperfusion was kept significantly higher than that in the no treatment group. Cytosolic c-Jun N-terminal protein kinase activation was significantly suppressed by SOWP + PGE1. Decrease in mitochondrial Bcl-2 was suppressed by SOWP alone and SOWP + PGE1 treatment, and Bax in the mitochondria was significantly suppressed by SOWP + PGE1. CONCLUSION: SOWP and PGE1 prior to cold preservation significantly improved the function of liver grafts that underwent warm ischemia-reperfusion injury. Therefore, this method might be useful in liver transplantation using UNHBD grafts.

Contralateral glissonian pedicle occlusion in a case of portal vein tumor thrombosis.

To dissect portal vein branches directly and encircle them separately is a common procedure that is performed to control back flow bleeding during operations for hepatocellular carcinoma with portal vein tumor thrombosis. However, this technique has an increased risk of injuring contralateral portal branches and disseminating thrombosis fragments to the remnant liver. We present an alternative technique using right-sided glissonian pedicle occlusion for hepatocellular carcinoma with left portal vein tumor thrombosis due to complex anatomical vasculatures of the hepatic pedicle. This technique would be very useful for liver resection of hepatocellular carcinoma with the major type of portal vein tumor thrombosis.

Intestinal obstruction caused by colonic metastasis from intrahepatic cholangiocarcinoma 6 years after removal of the primary tumor: report of a case.

We report a case of intestinal obstruction caused by metastasis that manifested 6 years after surgery for intrahepatic cholangiocarcinoma (ICC). The patient, a 57-year-old man, had undergone resection of the hepatic left lobe, Spiegel lobe, and extrahepatic bile duct, following which histopathological examination had confirmed the diagnosis of ICC and that the resection margins were free from disease. There had been no signs of recurrence until an increase in the CA19-9 level was detected 6 years later. Colonoscopy revealed an ulcer-like lesion and stenosis at the level of the hepatic flexure. The patient was subsequently admitted to our hospital with abdominal pain and underwent right hemicolectomy with partial resection of hepatic segment V. Based on the immunohistological finding that the expression pattern of cytokeratins and mucins was consistent with ICC origin rather than colon cancer origin, we diagnosed colon metastasis from ICC.

Post-transplantation lymphoproliferative disorder in living-donor liver transplantation: a single-center experience.

BACKGROUND: Post-transplantation lymphoproliferative disorder (PTLD) is a group of life-threatening complications of organ transplantation, which occurs most frequently in pediatric patients. This retrospective study evaluates a single-institution experience of five cases of PTLD after living-donor liver transplantation (LDLT). PATIENTS AND METHOD: We reviewed the records of 78 pediatric patients (<18 years old) and 54 adult patients, who underwent LDLT between July 1991 and December 2009. RESULT: PTLD was diagnosed in five pediatric patients, yielding an overall incidence of 3.8%. There were no significant differences between the pediatric patients with and those without PTLD in terms of their age, sex, reason for transplantation, calcineurin inhibitor, Epstein-Barr virus (EBV) serostatus, ABO compatibility, lymphocyte cross-matching, or episodes of biopsy proven rejection. Two patients with abdominal lymphadenopathy and one with gastrointestinal PTLD responded to a reduction in immunosuppression. Treatment with rituximab was necessary for another gastrointestinal PTLD patient. Diffuse large-B-cell lymphoma was diagnosed in one patient with mediastinal and lung masses. This patient was treated with chemotherapy and rituximab, followed by surgical resection. All patients survived and no evidence of recurrence has been found since. CONCLUSION: Although PTLD is potentially life-threatening, it can be managed by appropriate and prompt treatment, with a good outcome.

Unique histopathological features of graft biopsies with liver function abnormalities in living donor liver transplant patients receiving basiliximab induction therapy.

Induction with basiliximab (BXM) has been confirmed as an effective treatment regimen for prophylaxis of acute cellular rejection (ACR). From 1991 to 2008, 116 living donor liver transplantations (LDLTs) were performed. Among these, 50 were included in this study. We compared calcineurin inhibitor plus steroid treatment without BXM (n = 14, control group) and with BXM (n = 36, BXM group). Although the rates of biopsied patients with abnormal serum biochemical tests (SBTs) were similar in the control (10/14, 71.4%) and BXM (21/36, 58.3%) groups, ACR was diagnosed in 9/10 (90.0%) patients in the control group compared with 4/21 (19.0%) patients in the BXM group. In accordance with the histopathological diagnosis, there was a significant difference in the ratios of peripheral CD4(+) CD25(+) T cells at five wk after LDLT between patients with and without ACR in the BXM group. Next, we divided the 32 patients without ACR in the BXM group into two groups: biopsied patients with abnormal SBTs and non-biopsied patients. The donor age of the biopsied patients was significantly higher than that of the non-biopsied patients. Induction with BXM reduced the incidence of ACR, and unique pathological phenomena responsible for graft dysfunction after LDLT with an increased incidence of abnormal SBTs were observed.

Carbon monoxide suppresses arteriosclerotic lesions associated with chronic graft rejection and with balloon injury.

Carbon monoxide (CO), one of the products of heme oxygenase action on heme, prevents arteriosclerotic lesions that occur following aorta transplantation; pre-exposure to 250 parts per million of CO for 1 hour before injury suppresses stenosis after carotid balloon injury in rats as well as in mice. The protective effect of CO is associated with a profound inhibition of graft leukocyte infiltration/activation as well as with inhibition of smooth muscle cell proliferation. The anti-proliferative effect of CO in vitro requires the activation of guanylate cyclase, the generation of cGMP, the activation of p38 mitogen-activated protein kinases and the expression of the cell cycle inhibitor p21Cip1. These findings demonstrate a protective role for CO in vascular injury and support its use as a therapeutic agent.

Nucleoside analogue therapy following one-year course of hepatitis B immunoglobulin in preventing hepatitis B virus reactivation after living donor liver transplantation.

The combination therapy with hepatitis B immunoglobulin (HBIG) and nucleoside analogue is well tolerated for the hepatitis B recipients after liver transplantation, but its cost is an important problem in these days. Here we report the efficacy of nucleoside analogue therapy following one-year course of HBIG plus nucleoside analogue after living donor liver transplantation (LDLT). Out of 103 LDLTs, we selected 14 recipients who received the post-transplant therapy against reactivation of hepatitis B virus for more than 30 months. Those were eight patients with chronic hepatitis B, three with fulminant hepatitis, and three whose donors were positive for antibody to HB core antigen (HBc). During two days after the operation, HBIG (40,000 units) was administered, and the serum level of antibody to HB surface antigen (HBs) was maintained at around 150 IU/L for one year by monthly administration of HBIG. After one year, HBIG was withdrawn. A nucleoside analogue was administered daily from just after LDLT, and it was continued up to the present. Among the 14 patients, two recipients had recurrence of hepatitis B. Three patients, including one patient with recurrence of hepatitis B, died due to hepatocellular carcinoma or its associated cirrhosis; namely, their deaths are unrelated to hepatitis B-related diseases. The remaining 11 patients are leading normal lives. In conclusion, nucleoside analogue therapy after one-year course of HBIG plus nucleoside analogue is feasible and cost-effective in preventing HBV reactivation. But the patients are still at risk of breakthrough and some patients may need continued prophylaxis with HBIG.

Long-term outcome of living related renal transplantation in a patient with short bowel syndrome.

Short-bowel syndrome (SBS) is defined as the malabsorptive state that occurs after extensive resection of the small intestine. In patients with SBS, oral administration of drugs usually becomes difficult because of the severity of intestinal failure. We describe a successful living related renal transplantation (LRRTx) in an 18-year-old male with SBS. Shortly after birth, the patient developed necrotizing enterocolitis requiring massive resection of the small intestine, which resulted in SBS. At seven years of age, the patient developed proteinuria and was diagnosed as focal segmental glomerulosclerosis (FSGS). His kidney function was gradually deteriorated toward the end-stage renal failure. The patient received LRRTx at age of 18 years. To evaluate the absorption capacity of the patient, we investigated pharmacokinetics of calcineurine inhibitors (tacrolimus and cyclosporine). The drug concentration, which is sufficient to provide effective immunosuppression, was achieved with cyclosporine, but not with tacrolimus. The patient therefore received a triple immunosuppressive therapy with oral cyclosporine, methyl-prednisolone and mycophenolate mofetil. To prevent both recurrent FSGS and rejection, we repeatedly analyzed the trough level and the pharmacokinetics of cyclosporine after LRRTx. The patient was successfully treated with oral immunosuppression for over 5 years, without hemodialysis. To our knowledge, this is the first report showing the long-term outcome of LRRTx treated with oral cyclosporine in a patient with SBS.

Long-term outcome of ABO-incompatible living-donor liver transplantation: a single-center experience.

PURPOSE: We report the long-term outcome of ABO-incompatible living donor liver transplantation (LDLT) performed in our hospital. METHODS: We started the LDLT program in 1991 and from that year up to now (2008) 11 patients have received an ABO-incompatible graft. RESULTS: Nine out of the 11 cases have survived from 3.7 years to 13.9 years (mean 7.3 years) and they are in good conditions at present. Seven patients were subjected to preoperative apheresis. Eight patients experienced acute rejection and of them, 6 experienced steroid-resistant rejection that was treated with deoxyspergualin and apheresis. One patient who suffered rapidly progressing rejection died due to liver failure. Three patients who were administered rituximab did not suffer severe rejection nor adverse effects. During the long-term follow up 5 recipients had major complications such as postoperative lymphoproliferative disease, post-transplantation diabetes mellitus, portal vein occlusion and biliary stenosis. But those complications were controlled under stable conditions. CONCLUSIONS: We concluded that long-term survival can be expected after ABO-incompatible LDLT provided perioperative complications such as humoral rejection are overcome.

The significance of preserving the energy status and microcirculation in liver grafts from non-heart-beating donor.

To complete a successful liver transplantation (LTx) from non-heart-beating donors (NHBD), it is necessary to both improve the energy status in liver grafts and to reduce the exposure to free radicals. This study investigated the effects of short perfusion with oxygenated buffer on the grafts prior to cold preservation. In addition, the effects of the antioxidant, biliverdin, for reduction of free radicals was investigated. Male Wistar rats were used. Livers were retrieved, preserved in UW solution, and perfused for 60 min with oxygenated Krebs-Henseleit solution. Rats were allocated to six groups as follows (n=5): (i) control group-no warm ischemia (WI) and cold preservation, (ii) HBD group--no WI with cold preservation for 6 h; (iii) NHBD group--with 30 min of WI and cold preservation, (iv) NM group--with WI including nafamostat mesilate infusion before cardiac arrest and cold preservation; (v) PRE group--with WI, 30-min pre-cold preservation perfusion with oxygenated buffer after cardiac arrest, and cold preservation, (vi) BV group-with the same treatment as the PRE group plus the addition of biliverdin to the pre-cold preservation perfusion. The portal flow volume, bile production, AST, and TNF-alpha in perfusate, energy charge (EC), and ATP level in the tissue, and histological findings were investigated. The portal flow volume in the NM, PRE, and BV groups were higher than in the NHBD group. The bile production in the PRE and BV groups were also higher than in the NHBD group. The EC and ATP level of the BV group after reperfusion were higher than those of the NHBD group. Pre-cold preservation perfusion and addition of biliverdin to perfusate improved viability of grafts from NHBD. The results indicate that the preservation of the energy status and microcirculation of the graft is important for successful LTx from NHBD.

Long-term outcome after living donor liver transplantation for two cases of homozygous familial hypercholesterolemia from a heterozygous donor.

AIM: We experienced two pediatric siblings with homozygous familial hypercholesterolemia (FH) who received living donor liver transplantation (LDLT) from their parents who were heterozygous for FH. METHODS: The elder brother presented orange cutaneous xanthomas and was diagnosed as homozygous FH at the age of one. The plasma lipid profile showed that his total cholesterol level was 898 mg/dL (23.2 mmol/L), LDL cholesterol level was 756 mg/dL (19.6 mmol/L) and triglyceride level was 60 mg/dL (0.7 mmol/L). There were no living donors with a normal LDL receptor in their family, and it was difficult to find a deceased donor in Japan; thus he underwent LDLT with his father as the donor. His sister was born 2 years after his LDLT. She underwent ABO-incompatible LDLT at the age of 2 with her mother as the donor. RESULTS: The boy's liver function tests normalized immediately after transplantation, and his cholesterol has remained controlled at around 280 mg/dL (7.2 mmol/L), with HMG-CoA reductase inhibitor for 6 years after LDLT. The girl's cholesterol remained stable at around 280 mg/dL (7.2 mmol/L) under treatment with HMG-CoA reductase inhibitor two years after LDLT. At present, the four patients, including the two donors, are leading normal daily lives. CONCLUSION: Living-donor liver transplantation from a donor with heterozygous FH is a feasible indication for the treatment of homozygous FH.

Management of anti-allogeneic antibody elimination by apheresis in living donor liver transplantation.

In this study, we report on the indications and efficacy of the elimination of antiallogeneic antibodies in living donor liver transplant recipients. Seven patients incompatible with the ABO-blood type were subjected to apheresis before transplantation. The procedure resulted in titers being decreased to less than a score of 8. After transplantation, apheresis was also performed in 6 cases and continuous hemodiafiltration in 1 case. In addition, three out of 11 ABO-blood type incompatible recipients were administered anti-CD20 antibody (rituximab). Two crossmatch positive patients were subjected to apheresis before transplantation, and in these cases the titers were reduced to less than a score of 2. Moreover, these two patients had no acute rejections after transplantation. We concluded that apheresis is effective for preventing acute rejection induced by pre-existing anti-A and/or anti-B antibodies, as well as antidonor specific antibodies, but is not effective in some patients who had accelerated humoral rejection.

Short Oxygenated Warm Perfusion With Prostaglandin E1 Administration Before Cold Preservation as a Novel Resuscitation Method for Liver Grafts From Donors After Cardiac Death in a Rat In Vivo Model.

BACKGROUND: We previously demonstrated that short oxygenated warm perfusion (SOWP) prevented warm ischemia-reperfusion injury in rat livers from donors after cardiac death (DCDs) in an ex vivo model. In the present study, we aimed to examine the in vivo effects of SOWP and SOWP with prostaglandin E1 (PGE1) in DCD rat liver transplants. METHODS: We performed liver transplantation after 6-hour cold preservation using grafts retrieved from DCD rats, divided into nontreatment (NT), SOWP, and SOWP with PGE1 (SOWP + PG) treatment groups. The SOWP grafts were perfused with oxygenated buffer at 37°C for 30 minutes before cold preservation. Prostaglandin E1 was added to the SOWP + PG group perfusate. Eleven liver transplants from each group were performed to evaluate graft function and survival; 5 rats were used for data collection after 1-hour reperfusion, and 6 rats were used for the survival study. As a positive control, the same experiment was performed in a heart-beating donor group. RESULTS: In both the SOWP and SOWP + PG groups, serum liver enzymes, intercellular adhesion molecule 1 levels, and cellular damage were significantly decreased compared with the NT group. In the SOWP + PG group, bile production and energy status were significantly improved compared with the NT group. The 4-week survival was 0% (0/6), 67% (4/6), 83% (5/6), and 100% (6/6) in the NT, SOWP, SOWP + PG, and heart-beating donor group, respectively. CONCLUSIONS: Short oxygenated warm perfusion before cold preservation and the addition of PGE1 to SOWP were thus beneficial in an in vivo rat model.

Successful use of anti-CD20 monoclonal antibody (rituximab) for ABO-incompatible living-related liver transplantation.

BACKGROUND: Humoral rejection after ABO-incompatible liver transplantation often causes graft loss and a life-threatening situation. We used rituximab, which can eliminate B cells highly selectively, as an additional therapy for ABO-incompatible living-related liver transplantation. CASES: Patient 1 was a 1-year-old girl with biliary atresia. Her blood type was O, and the donor's was A. She underwent two plasma exchanges before liver transplantation and had triple immunosuppressants (mycophenolate mofetil, tacrolimus, and methylprednisolone). She was diagnosed with humoral rejection by needle biopsy on postoperative day 6. Rituximab was used for 3 days at 375, 187, and 187 mg/m(2) and successfully reduced the antibody titer, transaminase, and CD19(+) cells count in peripheral blood lymphocytes. The patient has not had any severe rejection, infection, or serious complications 2 years posttransplantation. Patient 2 was a 42-year-old woman with primary biliary cirrhosis. The blood type was O, and the donor's was B. She received three plasma exchanges, triple immunosuppressants, splenectomy, intraarterial anticoagulant therapy, and rituximab (375 mg/m(2) immediately after transplantation). The titer and CD19(+) cells count remained persistently low throughout the recovery course. She did not develop humoral rejection 1 year after transplantation. CONCLUSIONS: Rituximab efficiently reduces anti-ABO antibody titer by selectively eliminating B cells and is safe and effective against humoral rejection after ABO-incompatible liver transplantation.

Heme oxygenase-1-derived carbon monoxide protects hearts from transplant associated ischemia reperfusion injury.

Heme oxygenase-1 (HO-1) degrades heme into iron, biliverdin, and carbon monoxide (CO). HO-1 expression can be used therapeutically to ameliorate undesirable consequences of ischemia reperfusion injury (IRI), but the mechanism by which this occurs, remains to be established. Rat hearts, exposed to a prolonged period (24 h) of cold (4 degrees C) ischemia, failed to function upon transplantation into syngeneic recipients. Induction of HO-1 expression by administration of cobalt protoporphyrin IX (CoPPIX) to the graft donor restored graft function. Inhibition of HO-1 enzymatic activity, by administration of zinc protoporphyrin (ZnPPIX) at the time of transplantation, reversed the protective effect of HO-1. Exposure of the graft donor as well as the graft (during ischemia) to exogenous CO mimicked the protective effect of HO-1. This was associated with a significant reduction in the number of cells undergoing apoptosis in the graft with no apparent decrease of intravascular fibrin polymerization, platelet aggregation, or P-selectin expression. In conclusion, HO-1-derived CO prevents IRI associated with cardiac transplantation based on its antiapoptotic action. The observation that exposure of the donor and the graft to CO is sufficient to afford this protective effect should have important clinical implications in terms of preventing IRI associated with heart transplantation in humans.

Effect of elimination of donor Kupffer cells and/or recipient macrophages on acute rejection in liver transplantation.

BACKGROUND/AIMS: Kupffer cells/macrophages have pivotal roles of antigen presentation and cytokine production in the acute rejection after liver transplantation. In this study, we attempted to eliminate donor Kupffer cells and/or recipient macrophages by administration of liposome encapsulated LE-Cl2MDP (dichloromethylene diphosphonate) for clarification of the effect of depletion of Kupffer cells and macrophages on acute rejection. METHODOLOGY: To compare the survival, five groups were prepared: i) non-treatment group, ii) Kupffer cells eliminated group: transplanted Kupffer cells eliminated graft, iii) administration after liver transplantation group: injected LE-Cl2MDP twice after liver transplantation of non-treated graft, iv) additional administration group: transplanted Kupffer cells eliminated graft and injected LE-Cl2MDP twice after liver transplantation, and v) frequent administrations after liver transplantation group: injected LE-Cl2MDP every day for one week after liver transplantation of non-treated. Liver function tests, histological examination, histochemical staining of Kupffer cells and macrophages were carried out. RESULTS: Survival days were not prolonged by Kupffer cells elimination of the graft or by depletion of migrated macrophages of recipient. In additional in the administration group, in which both Kupffer cells of the graft and migrated macrophages of recipient were eliminated, graft survival was prolonged significantly. But hyper bilirubinemia, hepatic fibrosis, and slight mononuclear cell infiltration were recognized. CONCLUSIONS: These results suggested that both donor Kupffer cells and recipient macrophages play important roles in acute rejection in liver transplantation.

Development of a novel ELISA for detection of anti-A and anti-B antibodies in recipients of ABO-incompatible living donor liver grafts.

The survival rate in ABO-incompatible (ABO-I) liver transplantation was much lower than that in ABO-compatible recipients for the early experiences. It is therefore essential to develop the precise and fast measurement of anti-A and anti-B antibodies (Abs) to prevent humoral rejection in ABO-I liver transplantation. Agglutination titer has been the standard method to measure these Abs, but the interpretation of the results is subject to bias. Here, we have developed an objective and quantitative enzyme-linked immunosorbent assay (ELISA) to measure anti-A and anti-B Abs. This test requires only a small amount (10 microl) of recipient's serum. We applied the newly developed ELISA to monitor living donor liver transplant recipients and investigated the correlation between ELISA and agglutination titer. The Spearman's correlation coefficient for Abs ranged from 0.461 to 0.812. Moreover, in one case of humoral rejection, the increase of Abs was detected by ELISA one day earlier than by the agglutination titer. In conclusion, our ELISA method proved useful to detect an increase of anti-A and anti-B Abs titers at an early stage, thereby contributing to a prompt treatment of humoral rejection due to ABO-I.