Toshiyasu Kawahara

Portal Vein Thrombosis Is a Potentially Preventable Complication in Clinical Islet Transplantation

Percutaneous transhepatic portal access avoids surgery but is rarely associated with bleeding or portal venous thrombosis (PVT). We herein report our large, single‐center experience of percutaneous islet implantation and evaluate risk factors of PVT and graft function. Prospective data were collected on 268 intraportal islet transplants (122 subjects). A portal venous Doppler ultrasound was obtained on Days 1 and 7 posttransplant. Therapeutic heparinization, complete ablation of the portal catheter tract with Avitene paste and limiting packed cell volume (PCV) to <5 mL completely prevented any portal thrombosis in the most recent 101 islet transplant procedures over the past 5 years. In the previous cumulative experience, partial thrombosis did not affect islet function. Standard liver volume correlated negatively (r =−0.257, p < 0.001) and PCV correlated positively with portal pressure rise (r = 0.463, p < 0.001). Overall, partial portal thrombosis occurred after 10 procedures (overall incidence 3.7%, most recent 101 patient incidence 0%). There were no cases of complete thrombosis and no patient developed sequelae of portal hypertension. In conclusion, portal thrombosis is a preventable complication in clinical islet transplantation, provided therapeutic anticoagulation is maintained and PCV is limited to <5 mL.

m-TOR inhibitors: what role in liver transplantation?

The development of calcineurin inhibitors (CNIs) led to marked improvements in patient and graft survival after liver transplantation (LTx). We have been left, however, with a dependence on immunosuppressive agents with nephrotoxicity, neurotoxicity, adverse impacts on cardiac risk profile, and risk for malignancy. These challenges need to be met against a dominance of hepatitis C virus (HCV) and hepatocellular carcinoma (HCC) as indications for liver transplant. Unmet needs for immunosuppression (IS) in LTx include: (1) Effective drugs that avoid CNIs toxicities. (2) Agents without adverse impact on HCV recurrence. (3) Compounds that minimize risk of HCC recurrence. New immunosuppressives will need to address the above needs while supporting patient and graft survival equivalent to those achievable with CNIs, ideally without important new toxicities. Two new classes of agents are currently in advanced clinical development: belatacept, and the mammalian target of rapamycin inhibitors (m-TORi). This manuscript will review evidence for a role for m-TORi in LTx in a range of clinical scenarios including patients with CNI nephrotoxicity or neurotoxicity, patients at risk of (or with) HCV recurrence, and patients at risk of HCC recurrence.

Factors affecting hepatocyte isolation, engraftment, and replication in an in vivo model

Human hepatocyte transplantation is an alternative treatment for acute liver failure and liver diseases involving enzyme deficiencies. Although it has been successfully applied in selected recipients, both isolation and transplantation outcomes have the potential to be improved by better donor selection. This study assessed the impact of various donor variables on isolation outcomes (yield and viability) and posttransplant engraftment, using the SCID/Alb‐uPA (severe combined immunodeficient/urokinase type plasminogen activator under the control of an albumin promoter) human liver chimeric mouse model. Human hepatocytes were obtained from 90 human liver donor specimens and were transplanted into 3942 mice. Multivariate analysis revealed improved viability with younger donors (P = 0.038) as well as with shorter warm ischemic time (P = 0.012). Hepatocyte engraftment, assessed by the posttransplant level of serum human α1‐antitrypsin, was improved with shorter warm ischemia time. Hepatocytes isolated from older donors (≥60 years) had lower viability and posttransplant engraftment (P ≤ 0.01). In conclusion, the selection of young donors (<60 years) and rapid liver specimen retrieval, allowing for shorter warm ischemia time, are key determinants for the success of both the isolation of high viability human hepatocytes and their subsequent posttransplantation capacity for engraftment and expansion. Liver Transpl 16:974‐982, 2010.

Late Cytomegalovirus Transmission and Impact of T-Depletion in Clinical Islet Transplantation

The epidemiology of cytomegalovirus infection (CMV) in islet transplantation (IT) is not well defined. This study defines incidence, transmission and clinical sequelae of CMV reactivation or disease in 121 patients receiving 266 islet infusions at a single institution. The donor (D)/recipient (R) serostatus was D+/R– 31.2%, D+/R+ 26.3%, D–/R+ 13.2% and D–/R– 29.3%. CMV prophylaxis with oral ganciclovir/valganciclovir was given in 68%. CMV infection occurred in 14/121 patients (11.6%); six had asymptomatic seroconversion and eight others had positive viremia (six asymptomatic and two with CMV febrile symptoms). Median peak viral loads were 1755 copies/mL (range 625–9 100 000). Risk factors for viremia included lymphocyte depletion (thymoglobulin or alemtuzumab, p < 0.001). Viremia was more common in D+/R+ versus D+/R– (p = 0.12), occurring mostly late after transplant (median 306 days). Presumed transmission from IT occurred in 8/83 of D+/R– procedures (9.6%). Of the two cases of CMV disease, one resulted from islet transmission from a CMV positive donor (D+/R–); the other was due to de novo exogenous infection (D–/R–). Therefore, CMV transmission presents rarely after IT and with low incidence compared to solid organ transplantation, but occurs late posttransplant. The use of lymphocyte depleting therapies is a primary risk factor.

A comparison of islet autotransplantation with allotransplantation and factors elevating acute portal pressure in clinical islet transplantation

BackgroundAcute portal pressure rise is occasionally observed during intraportal islet infusion, especially in islet autotransplantation (IAT) where tissue purification is rarely applied. In this paper we investigate factors associated with acute portal pressure rise, a known risk factor for portal vein thrombosis.MethodsRetrospective data was collected on 15 islet autotransplant and 122 allogeneic islet transplant subjects. Non-purified pancreatic cells were transplanted in islet autotransplants, and purified islet cells were transplanted in allogeneic transplants. Portal pressure was documented throughout the islet infusion.ResultsThe total numbers of transplanted islets were significantly smaller in autotransplants than allografts, although the packed cell volume in autotransplants was larger. Autoislet infusion, with a larger packed cell volume, caused higher transient portal venous pressures than allogeneic islet transplant. Univariate analysis and multivariate linear regression revealed that packed cell volume and the number of transplanted cells were significant risk factors for acute portal pressure rise in both autotransplants and allogeneic transplants.ConclusionsNon-purified IAT has a higher risk for acute portal pressure rise than allogeneic islet transplantation, and the rise is associated with the packed cell volume and the number of transplanted cells. Minimization of packed cell volume and cautious monitoring of portal pressure are important to avoid potential complications of portal hypertension.

Three-dimensional tumor volume and serum alpha-fetoprotein are predictors of hepatocellular carcinoma recurrence after liver transplantation: refined selection criteria

Total tumor volume (TTV), as a better predictor of hepatocellular carcinoma (HCC) recurrence after liver transplant, has been explored by our center. Some tumors are not typically spherical but rather ellipsoid or spheroid, and calculating their TTV based on one dimension only may overestimate their volume and exclude them from candidacy for transplantation. Our aim was to study the actual tumor volume (ATV) calculated using the ellipsoid formula and assess its impact on recurrence. HCC patients transplanted between 1990 and 2010 at University of Alberta Hospital were analyzed. Tumor volumes were calculated using both formulas: [(4/3) πr3] (r = max. radius) and [(4/3) πabc] (a, b, c = the 3 radiuses). A total of 115 patients were included with a mean follow‐up of 4.99 ± 4.23 yr. Five‐yr recurrence‐free survival was 79.8%. Univariate analysis for predictors of recurrence included: maximum tumor diameter, ATV, TTV, and alpha‐fetoprotein (AFP) ≥ 400 ng/mL. Multivariate analysis showed that ATV and AFP ≥ 400 ng/mL were the only predictors of recurrence. Combining both variables provides better predication of recurrence with accuracy that exceeds 80%. Three‐dimensional calculation of tumor volume is of critical importance for the group of patients with ellipsoid tumors where volumes are overestimated with the spherical formula and could lead to inappropriate exclusion from transplant.

Effect of olprinone, a phosphodiesterase III inhibitor, on hepatic ischemia-reperfusion injury in rats.

I/R injury is the main cause for hepatic dysfunction and failure after liver transplantation and liver resection. Therefore, reduction of I/R injury is the most important goal to improve the outcome of these procedures. Olprinone is a newly developed selective phosphodiesterase III inhibitor, which has been reported to ameliorate renal I/R injury in rats. However, no clear evidence for the actions of olprinone on inflammatory response after hepatic I/R injury has been disclosed thus far. Our study was designed to evaluate the action of olprinone on the hepatic I/R injury in rats. Olprinone increased the cyclic adenosine monophosphate level in injured liver tissue and ameliorated the liver injury after hepatic I/R. Moreover, olprinone suppressed the activation of p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, and nuclear factor-&kgr;B, cytokine production (TNF-&agr;, IL-6, and cytokine-induced neutrophil chemoattractant factor 1), and intercellular adhesion molecule 1 expression in liver after hepatic I/R. These observations suggest that olprinone protects liver against I/R injury via the elevation of cyclic adenosine monophosphate level and suppression of intercellular adhesion molecule 1 expression and cytokine production (TNF-&agr;, IL-6, and cytokine-induced neutrophil chemoattractant factor 1), possibly by interfering with the signaling pathways of p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, and nuclear factor-&kgr;B in rats.

Arylacetamide deacetylase: A novel host factor with important roles in the lipolysis of cellular triacylglycerol stores, VLDL assembly and HCV production

BACKGROUND & AIMS Very low density lipoproteins (VLDLs) are triacylglycerol (TG)-rich lipoproteins produced by the human liver. VLDLs derive the majority of their TG cargo from the lipolysis of TG stored in hepatocellular lipid droplets (LDs). Important roles for LDs and the VLDL secretory pathway in the cell culture production of infectious hepatitis C virus (HCV) have been established. We hypothesized that TG lipolysis and VLDL production are impaired during HCV infection so that these cellular processes can be diverted towards HCV production. METHODS We used an HCV permissive cell culture system (JFH-1/HuH7.5 cells) to examine the relationship between TG lipolysis, VLDL assembly, and the HCV lifecycle using standard biochemical approaches. RESULTS Lipolysis of cellular TG and VLDL production were impaired in HCV infected cells during the early peak of viral infection. This was partially explained by an apparent deficiency of a putative TG lipase, arylacetamide deacetylase (AADAC). The re-introduction of AADAC to infected cells restored cellular TG lipolysis, indicating a role for HCV-mediated downregulation of AADAC in this process. Defective lipolysis of cellular TG stores and VLDL production were also observed in HuH7.5 cells stably expressing a short hairpin RNA targeting AADAC expression, proving AADAC deficiency contributes to these defective pathways. Finally, impaired production of HCV was observed with AADAC knockdown cells, demonstrating a role for AADAC in the HCV lifecycle. CONCLUSIONS This insight into the biology of HCV infection and possibly pathogenesis identifies AADAC as a novel and translationally relevant therapeutic target.

Human cytomegalovirus infection in humanized liver chimeric mice

Cytomegalovirus is a common viral pathogen that influences the outcome of organ transplantation. To date, there is no established method to evaluate the effects of human CMV (HCMV) treatments in vivo except for human clinical trials. In the current study, we describe the development of a mouse model that supports the in vivo propagation of HCMV.

Critical role of natural killer cells in the rejection of human hepatocytes after xenotransplantation into immunodeficient mice

The severe combined immunodeficiency/albumin linked‐urokinase type plasminogen activator (SCID/Alb‐uPA) human liver chimeric mouse model has added a new dimension to studies of liver based human diseases and has important potential for study of human hepatic drug metabolism. However, it remains unclear if natural killer (NK) cell in SCID/Alb‐uPA mice has an important negative impact on engraftment and expansion of human hepatocytes after transplantation. Here, we explore the role of mouse NK cells in the rejection of transplanted human hepatocytes in SCID/Alb‐uPA mice. We assessed NK cell activity in vivo, using 125I‐iodo‐2′‐deoxyuridine incorporation assay. Low serum human alpha‐1 antitrypsin (hAAT, <10 μg/ml) recipients, representing graft failure, showed resistance to engraftment of MHC class I knockout marrow (indicating high NK cell activity), while NK cell‐depleted low hAAT recipients and high hAAT (>100 μg/ml) recipients accepted MHC class I knockout marrow, indicating a correlation between low NK cell activity, in vivo, and high level human hepatocyte engraftment. We also showed that higher level engraftment of human hepatocytes was achieved in both NK cell‐depleted SCID/Alb‐uPA mice and Rag2−/−γc−/−/Alb‐uPA (T,B and NK cell deficient) mice compared with untreated SCID/Alb‐uPA mice. These results support a critical role for mouse NK cells in the rejection of human hepatocytes xenotransplanted to immunodeficient mice.