James F. Markmann

Therapeutic siRNA silencing in inflammatory monocytes in mice

Excessive and prolonged activity of inflammatory monocytes is a hallmark of many diseases with an inflammatory component. In such conditions, precise targeting of these cells could be therapeutically beneficial while sparing many essential functions of the innate immune system, thus limiting unwanted effects. Inflammatory monocytes—but not the noninflammatory subset—depend on the chemokine receptor CCR2 for localization to injured tissue. Here we present an optimized lipid nanoparticle and a CCR2-silencing short interfering RNA that, when administered systemically in mice, show rapid blood clearance, accumulate in spleen and bone marrow, and localize to monocytes. Efficient degradation of CCR2 mRNA in monocytes prevents their accumulation in sites of inflammation. Specifically, the treatment attenuates their number in atherosclerotic plaques, reduces infarct size after coronary artery occlusion, prolongs normoglycemia in diabetic mice after pancreatic islet transplantation, and results in reduced tumor volumes and lower numbers of tumor-associated macrophages.

Homeostatic proliferation is a barrier to transplantation tolerance

Despite the ease of inhibiting immune responses by blockade of T-cell costimulation in naive rodent models, it is difficult to suppress those responses in animals with memory cells. Studies demonstrating the importance of alloreactive T-cell deletion during tolerance induction have promoted use of peritransplant T-cell-depleting therapies in clinical trials. But potentially complicating wide-scale, nonspecific T-cell depletion is the finding that extensive T-cell proliferation can occur under conditions of lymphopenia. This process, termed homeostatic proliferation, may induce acquisition of functional memory T cells. Here, using clinically relevant mouse models of peripheral T-cell depletion, we show that residual nondepleted T cells undergo substantial homeostatic expansion. In this setting, costimulatory blockade neither significantly suppresses homeostatic proliferation nor prevents allograft rejection. In addition, T cells that have completed homeostatic proliferation show dominant resistance to tolerance when adoptively transferred into wild-type recipients, consistent with known properties of memory cells in vivo. These findings establish the importance of homeostatic proliferation in clinically relevant settings, demonstrate the barrier that homeostatic proliferation can present to the induction of transplantation tolerance, and have important implications for transplantation protocols that use partial or complete peripheral T-cell depletion.

Survival following liver transplantation from non-heart-beating donors

Objective:To determine whether patient and graft survival following transplantation with non-heart-beating donor (NHBD) hepatic allografts is equivalent to heart-beating-donor (HBD) allografts. Summary Background Data:With the growing disparity between the number of patients awaiting liver transplantation and a limited supply of cadaveric organs, there is renewed interest in the use of hepatic allografts from NHBDs. Limited outcome data addressing this issue exist. Methods:Retrospective evaluation of graft and patient survival among adult recipients of NHBD hepatic allografts compared with recipients of HBD livers between 1993 and 2001 using the United Network of Organ Sharing database. Results:NHBD (N = 144) graft survival was significantly shorter than HBD grafts (N = 26,856). One- and 3-year graft survival was 70.2% and 63.3% for NHBD recipients versus 80.4% and 72.1% (P = 0.003 and P = 0.012) for HBD recipients. Recipients of an NHBD graft had a greater incidence of primary nonfunction (11.8 vs. 6.4%, P = 0.008) and retransplantation (13.9% vs. 8.3%, P = 0.04) compared with HBD recipients. Prolonged cold ischemic time and recipient life support were predictors of early graft failure among recipients of NHBD livers. Although differences in patient survival following NHBD versus HBD transplant did not meet statistical significance, a strong trend was evident that likely has relevant clinical implications. Conclusions:Graft and patient survival is inferior among recipients of NHBD livers. NHBD donors remain an important source of hepatic grafts; however, judicious use is warranted, including minimization of cold ischemia and use in stable recipients.

Liver transplantation from controlled non-heart-beating donors: an increased incidence of biliary complications.

Background. Hepatic allografts from non–heart-beating donors (NHBD) have been cited as a means to expand the supply of donor livers. Concern exists that donor warm ischemic time in addition to subsequent cold ischemia-reperfusion injury may result in damage to sensitive cell populations within the liver. Because the biliary epithelium is sensitive to ischemia-reperfusion injury, the authors surmised that an increased incidence of biliary complications might occur among recipients of an NHBD allograft. Methods. This study was a retrospective evaluation of NHBD recipients compared to a group of heart-beating donor (HBD) recipients from a single institution. Results. Fifteen patients received a hepatic allograft from a controlled NHBD donor. NHBD and HBD (n=221) graft survival did not differ at 1 (71.8% vs. 85.4%, P =0.23) or 3 years (71.8% vs. 73.9%, P =0.68). Patient survival at 1 (79% vs. 90.9%, P =0.16) and 3 years (79.0% vs. 77.7%, P =0.8) was also similar. Major biliary complications occurred in five (33.3%) NHBD recipients; 66.6% of the NHBD biliary complications consisted of intrahepatic strictures versus 19.2% among HBD recipients (P <0.01). Major biliary complications in the NHBD recipients resulted in multiple interventional procedures, retransplantation, and death. Conclusions. Donor warm ischemic time may predispose hepatic allografts to an increased incidence of ischemic type strictures. Although graft and patient survival was similar to a cohort of HBD recipients, caution is urged with the use of these organs.

ASTS recommended practice guidelines for controlled donation after cardiac death organ procurement and transplantation

The American Society of Transplant Surgeons (ASTS) champions efforts to increase organ donation. Controlled donation after cardiac death (DCD) offers the family and the patient with a hopeless prognosis the option to donate when brain death criteria will not be met. Although DCD is increasing, this endeavor is still in the midst of development. DCD protocols, recovery techniques and organ acceptance criteria vary among organ procurement organizations and transplant centers. Growing enthusiasm for DCD has been tempered by the decreased yield of transplantable organs and less favorable posttransplant outcomes compared with donation after brain death. Logistics and ethics relevant to DCD engender discussion and debate among lay and medical communities. Regulatory oversight of the mandate to increase DCD and a recent lawsuit involving professional behavior during an attempted DCD have fueled scrutiny of this activity. Within this setting, the ASTS Council sought best‐practice guidelines for controlled DCD organ donation and transplantation. The proposed guidelines are evidence based when possible. They cover many aspects of DCD kidney, liver and pancreas transplantation, including donor characteristics, consent, withdrawal of ventilatory support, operative technique, ischemia times, machine perfusion, recipient considerations and biliary issues. DCD organ transplantation involves unique challenges that these recommendations seek to address.

Predicting outcome after liver transplantation: utility of the model for end-stage liver disease and a newly derived discrimination function1

Background. The Model for End-Stage Liver Disease (MELD) has been found to accurately predict pretransplant mortality and is a valuable system for ranking patients in greatest need of liver transplantation. It is unknown whether a higher MELD score also predicts decreased posttransplant survival. Methods. We examined a cohort of patients from the United Network for Organ Sharing (UNOS) database for whom the critical pretransplant recipient values needed to calculate the MELD score were available (international normalized ratio of prothrombin time, total bilirubin, and creatinine). In these 2,565 patients, we analyzed whether the MELD score predicted graft and patient survival and length of posttransplant hospitalization. Results. In contrast with its ability to predict survival in patients with chronic liver disease awaiting liver transplant, the MELD score was found to be poor at predicting posttransplant outcome except for patients with the highest 20% of MELD scores. We developed a model with four variables not included in MELD that had greater ability to predict 3-month posttransplant patient survival, with a c-statistic of 0.65, compared with 0.54 for the pretransplant MELD score. These pretransplant variables were recipient age, mechanical ventilation, dialysis, and retransplantation. Recipients with any two of the three latter variables showed a markedly diminished posttransplant survival rate. Conclusions. The MELD score is a relatively poor predictor of posttransplant outcome. In contrast, a model based on four pretransplant variables (recipient age, mechanical ventilation, dialysis, and retransplantation) had a better ability to predict outcome. Our results support the use of MELD for liver allocation and indicate that statistical modeling, such as reported in this article, can be used to identify futile cases in which expected outcome is too poor to justify transplantation.

In situ splitting of the cadaveric liver for transplantation

BACKGROUND The shortage of cadaveric donor livers is the rate-limiting step in clinical liver transplantation. Split liver transplantation provides a means to expand the cadaveric donor pool. However, this concept has not reached its full potential because of inferior patient and graft survival and high complication rates when traditional ex vivo split techniques are used. Therefore we sought to evaluate the safety, applicability, and effectiveness of a new technique for split liver transplantation. METHODS This study consists of 15 in situ split liver procurements, which resulted in 28 liver transplants. In situ splitting of selected livers from hemodynamically stable cadaveric donors was performed at the donor hospital without any additional work-up or equipment being needed. In situ liver splitting is accomplished in a manner identical to the living-donor procurement. This technique for liver splitting results in a left lateral segment graft (segments 2 and 3) and a right trisegmental graft (segments 1 and 4-8). This procedure required the use of the donor hospital operating room for an additional 1.5-2.5 hr and did not interfere with the procurement of 30 kidneys, 12 hearts, 7 lungs, and 9 pancreata from these same donors. RESULTS The 6-month and 1-year actuarial patient survival rates were 92% and 92%, respectively, while the 6-month and 1-year actuarial graft survival rates were 86% and 86%, respectively. The 6-month and 1-year actuarial patient survival rate of patients who received a left lateral segment graft was 100% and 100%, respectively, while those who received a right trisegmental graft had 6-month and 1-year rates of 86% and 86%, respectively. The actuarial death-censored graft survival rates at 6 months and 1 year were 80% and 80%, respectively, for the left lateral segment grafts, and 93% and 93%, respectively, for the right trisegmental grafts. Alograft and patient survival was independent of United Network for Organ Sharing status at the time of liver transplantation. No patient developed a biliary stricture, required re-exploration for intra-abdominal hemorrhage, or suffered from portal vein, hepatic vein, or hepatic artery thrombosis CONCLUSIONS In situ split liver transplantation can be accomplished without complications and provides results that are superior to those obtained previously with ex vivo methods. It abolishes ex vivo benching and prolonged ischemia times and provides two optimal grafts with hemostasis accomplished. This technique decreases pediatric waiting time and allows adult recipients to receive right-sided grafts safely. In situ splitting is the method of choice for expanding the cadaveric liver donor pool.

Preoperative factors associated with outcome and their impact on resource use in 1148 consecutive primary liver transplants.

BACKGROUND Hepatic transplantation is a highly effective but costly treatment for end-stage hepatic dysfunction. One approach to improve efficiency in the use of scarce organs for transplantation is to identify preoperative factors that are associated with poor outcome posttransplantation. This may assist both in selecting patients optimal for transplantation and in identifying strategies to improve survival. METHODS In the present work, we retrospectively reviewed consecutive liver transplants performed at the University of California at Los Angeles during a 6-year period and determined preoperative variables that were associated with outcome in primary grafts. In addition, we used the hospitals cost accounting database to determine the impact of these variables on the degree of resource use by high-risk patients. RESULTS We found five variables to have independent prognostic value in predicting graft survival after primary liver transplantation: (1) donor age, (2) recipient age, (3) donor sodium, (4) recipient creatinine, and (5) recipient ventilator requirement pretransplant. Recipient ventilator requirement and elevated creatinine were associated with significant increases in resource use during the transplant admission. CONCLUSIONS Patients at high risk for graft failure and costly transplants can be identified preoperatively by a set of parameters that are readily available, noninvasive, and inexpensive. Selection of recipients on the basis of these data would improve the efficiency of liver transplantation and reduce its cost.

Orthotopic liver transplantation for hepatitis C : Outcome, effect of immunosuppression, and causes of retransplantation during an 8-year single-center experience

OBJECTIVE To determine the outcome of orthotopic liver transplantation (OLT) for end-stage liver disease caused by hepatitis C virus (HCV). SUMMARY BACKGROUND DATA HCV has become the leading cause of cirrhosis and hepatic failure leading to OLT. Recurrent HCV after OLT is associated with significant complications and may lead to graft loss that requires retransplantation (re-OLT). The authors studied the outcome of transplantation for HCV, the effect of primary immunotherapy, and causes of retransplantation. METHODS The authors conducted a retrospective review of their experience during an 8-year period (1990-1997), during which 374 patients underwent transplants for HCV (298 [79.6%] received one OLT; 76 [20.4%] required re-OLT). Median follow-up was 2 years (range 0 to 8.3). Immunosuppression was based on cyclosporine in 190 patients and tacrolimus in 132 patients. In a third group of patients, therapy was switched from cyclosporine to tacrolimus or from tacrolimus to cyclosporine (cyclosporine/tacrolimus group). RESULTS Overall, 1-, 2-, and 5-year actuarial patient survival rates were 86%, 82%, and 76%, respectively. The 2-year patient survival rate was 81 % in the cyclosporine group, 85% in the tacrolimus group, and 82% in the cyclosporine/tacrolimus group. In patients receiving one OLT, overall 1-, 2-, and 5-year patient survival rates were 85%, 81%, and 75%, respectively. The 2-year patient survival rate was 79% in the cyclosporine group, 84% in the tacrolimus group, and 80% in the cyclosporine/tacrolimus group. The overall graft survival rates were 70%, 65%, and 60% at 1, 2, and 5 years, respectively. The graft survival rate at 2 years was similar under cyclosporine (68.5%), tacrolimus (64%), or cyclosporine/tacrolimus (60%) therapy. Re-OLT was required in 42 (11.2%) patients for graft dysfunction in the initial 30 days after OLT. Other causes for re-OLT included hepatic artery thrombosis in 10 (2.6%), chronic rejection in 8 (2.1%), and recurrent HCV in 13 (3.4%) patients. The overall survival rates after re-OLT were 63% and 58% at 1 and 2 years. The 1-year survival rate after re-OLT was 61 % for graft dysfunction, 50% for chronic rejection, 60% for hepatic artery thrombosis, and 60% for recurrent HCV. At re-OLT, 85.3% of the patients were critically ill (United Network for Organ Sharing [UNOS] status 1); only 14.7% of the patients were UNOS status 2 and 3. In re-OLT for chronic rejection and recurrent HCV, the 1-year survival rate of UNOS 1 patients was 38.4%, compared with 87.5% for UNOS 2 and 3 patients. In patients requiring re-OLT, there was no difference in the 1-year patient survival rate after re-OLT when cyclosporine (60%), tacrolimus (63%), or cyclosporine/tacrolimus (56%) was used for primary therapy. With cyclosporine, three patients (1.5%) required re-OLT for chronic rejection versus one patient (0.7%) with tacrolimus. Re-OLT for recurrent HCV was required in four (3%) and seven (3.6%) patients with tacrolimus and cyclosporine therapy, respectively. CONCLUSIONS Orthotopic liver transplantation for HCV is performed with excellent results. There are no distinct advantages to the use of cyclosporine versus tacrolimus immunosuppression when patient and graft survival are considered. Re-OLT is an important option in the treatment of recurrent HCV and should be performed early in the course of recurrent disease. Survival after re-OLT is not distinctively affected by cyclosporine or tacrolimus primary immunotherapy. The incidence of re-OLT for recurrent HCV or chronic rejection is low after either tacrolimus or cyclosporine therapy.

Insulin independence following isolated islet transplantation and single islet infusions

ObjectiveTo restore islet function in patients whose labile diabetes subjected them to frequent dangerous episodes of hypoglycemic unawareness, and to determine whether multiple transplants are always required to achieve insulin independence. Summary Background DataThe recent report by the Edmonton group documenting restoration of insulin independence by islet transplantation in seven consecutive patients with type 1 diabetes differed from previous worldwide experience of only sporadic success. In the Edmonton patients, the transplanted islet mass critical for success was approximately more than 9,000 IEq/kg of recipient body weight and required two or three separate transplants of islets isolated from two to four cadaveric donors. Whether the success of the Edmonton group can be recapitulated by others, and whether repeated transplants using multiple donors will be a universal requirement for success have not been reported. MethodsThe authors report their treatment with islet transplantation of nine patients whose labile type 1 diabetes was characterized by frequent episodes of dangerous hypoglycemia. ResultsIn each of the seven patients who have completed the treatment protocol (i.e., one or if necessary a second islet transplant), insulin independence has been achieved. In five of the seven patients only a single infusion of islets was required. To date, only one recipient has subsequently lost graft function, after an initially successful transplant. This patient suffered recurrent hyperglycemia 9 months after the transplant. ConclusionsThis report confirms the efficacy of the Edmonton immunosuppressive regimen and indicates that insulin independence can often be achieved by a single transplant of sufficient islet mass.