Single-cell analysis reveals different age-related somatic mutation profiles between stem and differentiated cells in human liver.

Accumulating somatic mutations have been implicated in age-related cellular degeneration and death. Because of their random nature and low abundance, somatic mutations are difficult to detect except in single cells or clonal cell lineages. Here, we show that in single hepatocytes from human liver, an organ exposed to high levels of genotoxic stress, somatic mutation frequencies are high and increase substantially with age. Considerably lower mutation frequencies were observed in liver stem cells (LSCs) and organoids derived from them. Mutational spectra in hepatocytes showed signatures of oxidative stress that were different in old age and in LSCs. A considerable number of mutations were found in functional parts of the liver genome, suggesting that somatic mutagenesis could causally contribute to the age-related functional decline and increased incidence of disease of human liver. These results underscore the importance of stem cells in maintaining genome sequence integrity in aging somatic tissues.

The Pancreas Can Take the Cold: Lower Waitlist Times Through Importation.

BACKGROUND: Our center has used a strategy of pancreas importation owing to long regional waitlist times. Here we assess the clinical outcomes and financial considerations of this strategy. METHODS: This was a retrospective observational cohort study of patients who received a pancreas transplant at Montefiore Medical Center (MMC) from 2014 to 2017 (n = 28). Clinical parameters, including hemoglobin A1c and complications, were analyzed. The cohort was compared with United Network for Organ Sharing (UNOS) Region 9 with the use of the UNOS/Organ Procurement and Transplantation Network database. Cost analysis of length of stay (LOS), standard acquisition (SAC) fees, and transportation was performed with the use of internal financial data. RESULTS: Pancreas importation resulted in significantly shorter simultaneous pancreas kidney transplant waitlist times compared with Region 9: 518 days vs 1001 days (P = .038). In addition, postoperative complications and 1-year HbA1c did not differ between groups: local 6.30% vs import 6.17% (P = .87). Patients receiving local pancreata stayed an average of 9.2 days compared with 11 days for the import group (P = .36). As such, pancreas importation was associated with higher mean charges ($445,968) compared with local pancreas recipients ($325,470). CONCLUSIONS: Long waitlist times in Region 9 have encouraged our center's adoption of pancreas importation to address the needs of our patient population. This practice has resulted in a reduction of waitlist times by an average of 483 days. Understandably, centers have long been wary of importation owing to perceived risk in clinical outcomes. In our single-center experience, we have demonstrated equivalent postoperative glucose control and graft survival. Importantly, there does appear to be increased costs associated with importation, which are mainly driven by LOS. Curiously, importation from regions with lower SAC fees has the potential to offset costs related to transportation expenses. Notwithstanding these findings, pancreas importation does have the potential to lessen the financial societal burden through reduction in waitlist times.

Hypothermic machine preservation in human liver transplantation: the first clinical series.

Hypothermic machine perfusion (HMP) is widely used to preserve kidneys for transplantation with improved results over cold storage (CS). To date, successful transplantation of livers preserved with HMP has been reported only in animal models. In this, the first prospective liver HMP study, 20 adults received HMP-preserved livers and were compared to a matched group transplanted with CS livers. HMP was performed for 3-7 h using centrifugal perfusion with Vasosol solution at 4-6 degrees C. There were no cases of primary nonfunction in either group. Early allograft dysfunction rates were 5% in the HMP group versus 25% in controls (p = 0.08). At 12 months, there were two deaths in each group, all unrelated to preservation or graft function. There were no vascular complications in HMP livers. Two biliary complications were observed in HMP livers compared with four in the CS group. Serum injury markers were significantly lower in the HMP group. Mean hospital stay was shorter in the HMP group (10.9 +/- 4.7 days vs. 15.3 +/- 4.9 days in the CS group, p = 0.006). HMP of donor livers provided safe and reliable preservation in this pilot case-controlled series. Further multicenter HMP trials are now warranted.

Solving the organ shortage crisis: the 7th annual American Society of Transplant Surgeons' State-of-the-Art Winter Symposium.

The 2007 American Society of Transplant Surgeons' (ASTS) State-of-the-Art Winter Symposium entitled, 'Solving the Organ Shortage Crisis' explored ways to increase the supply of donor organs to meet the challenge of increasing waiting lists and deaths while awaiting transplantation. While the increasing use of organs previously considered marginal, such as those from expanded criteria donors (ECD) or donors after cardiac death (DCD) has increased the number of transplants from deceased donors, these transplants are often associated with inferior outcomes and higher costs. The need remains for innovative ways to increase both deceased and living donor transplants. In addition to increasing ECD and DCD utilization, increasing use of deceased donors with certain types of infections such as Hepatitis B and C, and increasing use of living donor liver, lung and intestinal transplants may also augment the organ supply. The extent by which donors may be offered incentives for donation, and the practical, ethical and legal implications of compensating organ donors were also debated. The expanded use of nonstandard organs raises potential ethical considerations about appropriate recipient selection, informed consent and concerns that the current regulatory environment discourages and penalizes these efforts.

Select utilization of obese donors in living donor liver transplantation: implications for the donor pool.

Living donor liver transplantation evolved in response to donor shortage. Current guidelines recommend potential living donors (LD) have a body mass index (BMI) <30. With the current obesity epidemic, locating nonobese LD is difficult. From September 1999 to August 2003, 68 LD with normal liver function test (LFTs) and without significant comorbidities underwent donor hepatectomy at our center. Post-operative complications were collected, including wound infection, pneumonia, hernia, fever, ileus, biliary leak, biliary stricture, thrombosis, bleeding, hepatic dysfunction, thrombocytopenia, deep venous thrombosis, pulmonary embolism, difficult to control pain, depression and anxiety. Complication rates for LD with BMI >30 (n = 16) and BMI <30 (n = 52) were compared. The incidence of wound infection increased with BMI, 4% for nonobese and 25% for obese LD (p = 0.024). There were no statistically significant differences for all other complications. No LD died. Recipient survival was 100% with obese LD and 80% with nonobese LD (p = 0.1). Select donors with a BMI >30 may undergo donor hepatectomy with acceptable morbidity and excellent recipient results. Updating current guidelines to include select LD with BMI >30 has the potential to safely increase the donor pool.

Pushing the envelope in renal preservation; improved results with novel perfusate modifications for pulsatile machine perfusion of cadaver kidneys.

INTRODUCTION: Novel preservation techniques may diminish ischemia/reperfusion (I/R) injury. Our preservation laboratory has modified Belzer MPS for machine perfusion (MP) with prostaglandin E1 (PGE 1), nitroglycerin (NTG), and polyethylene glycol-superoxide dismutase (PEG-SOD) to attenuate I/R injury. We reviewed our recent experience using this novel formulation (NF) compared with standard perfusates. RESULTS: Between January 1998 and March 2000, 1060 consecutive kidneys were preserved in our laboratory. One hundred forty-eight kidneys (14%) were discarded. Fifty-eight percent of kidneys during this time period underwent MP (n = 532). En bloc kidney pairs were randomly assigned to pulsatile MP using Waters RM3 or MOX-100 perfusion systems using 1 of 3 perfusates; NF (NF; n = 119), Belzer MPS (MPS; n = 201), or Belzer II albumin gluconate (ALB; n = 212) Significant improvements in delayed graft function (DGF) rate were seen with NF versus other perfusates (8% vs 14% vs 19%, respectively; P =.03). At 6 months, graft survival was significantly improved with NF compared with MPS and ALB (96% vs 90% vs 87%, respectively; P =.03). NF also produced a significantly higher percentage of recipients with a serum creatinine level < or = 1.5 mg/dL. CONCLUSIONS: Novel modifications of standard MP perfusate improved outcomes after renal transplantation. Preservation-based interventions targeted to ameliorate I/R injury can improve outcomes and may allow expansion of the donor pool.

The laparoscopic evaluation of ascites.

Laparoscopy is an invaluable technique for the evaluation of ascites in subgroups of patients with ascites. Indications for laparoscopic examination include determination of the causes of ascites when routine tests fail to disclose the source, evaluation for the presence of multiple causes of ascites formation, or histopathologic verification of malignancy within the peritoneal cavity. Several reported series have illustrated the efficacy of laparoscopy for the diagnosis of peritoneal carcinomatosis, tuberculous peritonitis, or unsuspected cirrhosis, securing its role in the management of selected patients with ascites.

Renin system activation and delayed function of the renal transplant.

Delayed graft function (DGF), defined as persistent renal failure that requires dialysis within the first week after kidney transplantation, occurs commonly after cadaveric renal transplantation (CRT). This has important implications for long-term outcome because the 1-year allograft survival rate is significantly reduced when DGF occurs. The mechanisms contributing to the development of DGF are not well established. However, several lines of evidence indicate that excess renin system activity, in both the cadaver kidney donor and recipient, contributes importantly to the pathogenesis of DGF. If this hypothesis can be verified in clinical studies, then pharmacologic agents that interrupt the renin-angiotensin system (eg, type 1 angiotensin II receptor blockade, angiotensin converting enzyme inhibition, and beta-adrenergic blockade) in the donor and recipient might significantly improve the outcome of cadaveric renal transplants.

Glutathione supplementation during cold ischemia does not confer early functional advantage in renal transplantation.

BACKGROUND: Reduced glutathione (GSH), a component of University of Wisconsin (UW) solution, is reported to oxidize during storage. Consequently the commercial manufacturer of UW recommends the supplemental addition of GSH to UW before utilization. We investigated the influence of supplemental GSH during cold ischemia on early renal allograft function. METHODS: One hundred kidneys were locally procured from heart-beating donors, preserved in our laboratory, and transplanted during an 18-month period. Selected donor, preservation, and outcome characteristics were collected and compared by presence of supplemental GSH and method of preservation. All kidneys were randomized to receive 3.0 mM supplemental GSH to perfusate or no supplementation (control) and were preserved by either cold storage (CS) in UW or machine perfused (MP) in UW-machine perfusate solution (MPS). During MP, perfusion characteristics (flow, resistance, perfusate electrolytes, and pH) were serially measured. RESULTS: There were no significant differences among the groups when the donor characteristics of age, serum creatinine, and intra-operative urine output were compared. Preservation characteristics were similar among the groups with the exception of cold ischemia time, which was longer in the MP group compared to CS (26.1 h vs. 21.9 h, P=0.03). When compared with CS, kidneys preserved by MP exhibited a 33.4% increase in immediate function (93% vs. 62%, P=0.01), a corresponding 29.4% decrease in the incidence of delayed graft function (10% vs. 34%, P=0.02), and a 10% improvement in short-term (6-month) graft survival (98% vs. 88%, P=0.02). The addition of GSH supplementation to perfusate resulted in no significant differences in graft outcomes. CONCLUSIONS: Despite recommendations by the manufacturer that UW solution be routinely supplemented with GSH, supplemental GSH does not influence early renal allograft function. Our data suggest that a far greater beneficial impact on early graft function is achieved by machine perfusion. We conclude that GSH supplementation of commercially available UW is not necessary.

The influence of pulsatile preservation on renal transplantation in the 1990s.

BACKGROUND: Unlike simple cold storage (CS), pulsatile machine preservation (MP) of kidneys for transplantation permits pharmacologic manipulation of the perfusate and aids in the pretransplant assessment of the kidney graft. These characteristics of MP may have importance in the era of increasing use of extended criteria donor kidneys. The overall aim of this article is to critically assess practices at our preservation unit with respect to graft function. Specific aims are to (1) compare the influence of MP versus CS on graft function, (2) determine which pretransplant variables have significance in pretransplant assessment, and (3) determine whether pharmacologic manipulation during MP is advantageous. METHODS: There were 650 consecutive kidneys preserved in our laboratory between January 1, 1993 and March 1, 999, by either MP or CS. All MP kidneys were preserved by continuous hypothermic pulsatile perfusion using Belzer-MPS or Belzer II solution. Perfusion parameters and electrolytes were measured serially during pulsatile perfusion. All CS kidneys were stored in University of Wisconsin solution. All kidneys obtained from donors exhibiting extended criteria features underwent pretransplant frozen section biopsies. Transmission electron microscopy (EM) was performed on a subset of kidneys undergoing pharmacologic manipulation. Four agents were assessed prospectively for their ability to influence MP characteristics when added to perfusate: PGE1, trifluoperazine, verapamil, and papaverine. RESULTS: MP was associated with improved immediate, 1-, and 2-year graft function and reduced length of initial hospital stay when compared with CS grafts. Changes in the machine perfusion variables flow and resistance, and the [Ca++] in perfusate, were significantly associated with delayed graft function (DGF) after the transplant. Biopsy information was not predictive of DGF. The addition of PGE1 to perfusate improved MP characteristics, reduced the release of [Ca++] into perfusate, and ameliorated mitochondrial ischemic injury in transmission EM images. Early graft function was improved in the presence of PGE1+MP, compared with function in the presence of other pharmacologic agents or CS alone. CONCLUSIONS: MP is associated with improved early and long term renal function. Moreover, PGE1 augments MP in improving graft function. The combination of MP+PGE1 may be important in optimizing the ability to use extended donor criteria kidneys and, thereby, improve the overall efficiency of cadaveric renal transplantation.

Donor treatment with phentolamine mesylate improves machine preservation dynamics and early renal allograft function.

BACKGROUND: It has been suggested that pharmacologic conditioning of the donor before organ procurement may protect the renal allograft from injuries associated with the cold ischemic period. We compared the administration of two vasoactive agents before organ procurement to: (1) determine their influence on machine perfusion characteristics and (2) determine their impact on delayed graft function (DGF) in transplanted renal allografts. METHODS: Between January 1997 and December 1998, 150 kidneys were procured from heart-beating donors and preserved in our laboratory by machine perfusion (MP) or cold storage (CS). The following vasoactive agents were randomly administered to the donor 5 min before aortic cross clamp: phentolamine mesylate (PM) or hydralazine (H). The control groups received no donor conditioning. Kidneys were grouped as follows: (1) MP+PM, (2) MP+H, (3) MP, (4) CS+PM, (5) CS+H, (6) CS. 10 mg PM/50 kg donor weight was administered to the PM groups and 20 mg H/50 kg donor weight was administered to the H groups. DGF was defined as the need for dialysis within the first 7 days after the transplant. RESULTS: MP+PM increased renal flow by 12% and decreased renal resistance by 18% compared with the MP+H group, and increased renal flow by 23% and decreased renal resistance by 30% compared with the MP group. Moreover, the MP+PM group was associated with improved early allograft function. CONCLUSIONS: Donor treatment with PM immediately before aortic cross-clamp is associated with improved machine perfusion dynamics (renal flow and renal resistance) and lower incidence of DGF compared with donor treatment with H or no treatment. Moreover, MP of renal allografts was associated with improved early function compared with CS grafts.

Calcium ion concentration of machine perfusate predicts early graft function in expanded criteria donor kidneys.

Pulsatile preservation offers the advantage of pretransplant assessment of donor kidneys. Selected electrolyte concentrations of machine perfusate were measured over time in order to: (1) describe electrolyte changes in perfusate during the pulsatile preservation of expanded-criteria donor (ECD) kidneys, and (2) to assess the prognostic significance of these characteristics to early graft function. One hundred and fifty ECD kidneys were preserved in our laboratory between 1 January 1995 and 11 January 1997. ECD kidneys were defined as those requiring pretransplant biopsy. Kidneys were grouped by the presence or absence of delayed graft function (DGF), and perfusion parameters were measured every hour during pulsatile perfusion. All kidneys were preserved by continuous hypothermic pulsatile perfusion using Belzer II solution. Renal flow is decreased and renal resistance is increased in the presence of DGF in machine-preserved ECD kidneys. In addition, ionized calcium concentration of the machine perfusate is significantly elevated in the DGF group compared with the No DGF group (0.091 vs 0.054, P = 0.0016). The incidence of DGF is significantly lower in the ECD kidney. Among the pretransplant variables of donor characteristics, perfusion parameters and histology, perfusion parameters are highly predictive of early graft function. In addition, we found that ionized calcium concentration in the perfusate is significantly elevated in kidneys exhibiting DGF, which may have implications for assessing the suitability of donor kidneys for transplantation.

Prostaglandin E1 influences pulsatile preservation characteristics and early graft function in expanded criteria donor kidneys.

INTRODUCTION: Unlike simple cold storage, machine preservation allows dynamic assessment and manipulation of the donor organ prior to transplantation. We prospectively compared the effects of five pharmacological agents added to the perfusate during machine preservation of expanded criteria donor (ECD) kidneys in order to (1) describe their influence on perfusion parameters and (2) determine their influence on early graft outcome. METHODS: Two hundred seventy-five consecutive ECD kidneys were preserved in our laboratory between 1/1/94 and 12/31/97 by either machine perfusion (MP) or cold storage (CS). ECD kidneys were defined as those requiring pretransplant biopsy. ECD kidneys were divided by method of preservation and MP kidneys were randomized to receive prostaglandin E1 (MP+PGE1), trifluoperazine (TFP), verapamil (VER), papaverine (PAP), mannitol (MAN), or no intervention during the period of machine perfusion. CS kidneys were randomized to receive PGE1 (CS+PGE1), TFP, VER, PAP, or no intervention. All MP kidneys were preserved by continuous hypothermic pulsatile perfusion using Belzer II solution and perfusion parameters were measured every hour during pulsatile perfusion. All CS kidneys were stored in 1.0 L of University of Wisconsin (UW) solution. RESULTS: The addition of PGE1 to machine perfusate increased renal flow and decreased renal resistance. Moreover, the MP+PGE1-treated group was associated with improved early graft function compared to all other groups. The addition of VER, TFP, PAP, or MAN influenced neither the perfusion characteristics nor the incidence of early graft function in MP kidneys. Similarly, the addition of VER, TFP, or PAP did not influence early graft function in the CS kidneys. The CS+PGE1 group exhibited a significantly lower incidence of early graft function than did the MP+PGE1 group. CONCLUSIONS: PGE1 treatment during machine preservation improves hydrostatic perfusion parameters and reduces the incidence of delayed graft function in ECD kidneys. Moreover, the addition of PGE1, TFP, VER, or PAP to UW does not influence early graft function in the CS kidney.

Resolution of recurrent hepatitis B in two liver transplant recipients treated with famciclovir.

Recurrent hepatitis B infection after orthotopic liver transplantation remains problematic despite prophylaxis with hepatitis B immune globulin (anti-HBs IgG). Recently, famciclovir (an oral nucleoside analog) has been shown to have potent antiviral activity against hepatitis B in vitro as well as in patients with chronic hepatitis B. We present two patients who developed recurrent hepatitis B after orthotopic liver transplantation and were treated with famciclovir, 500 mg t.i.d. Both patients subsequently responded with marked improvement in biochemical liver tests and histology, with subsequent loss of hepatitis B surface antigen. Famciclovir is a useful agent in the treatment of hepatitis B in the liver transplant recipient.

Prostaglandin E1 improves pulsatile preservation characteristics and early graft function in expanded criteria donor kidneys.

Unlike simple cold storage, machine preservation allows dynamic assessment and manipulation of the donor organ before transplantation. The effects of four pharmacologic agents added to the perfusate during machine preservation of expanded criteria donor (ECD) kidneys were prospectively compared to 1) describe their influence on perfusion parameters and 2) determine their influence on early graft outcome. Between 1 January 1995 and 1 October 1997, 125 consecutive ECD kidneys were preserved in the authors' laboratory. A definition of ECD was assigned to kidneys requiring pretransplant biopsy. The ECD kidneys were randomized to receive prostaglandin E1 (PGE1), trifluoperazine (TFP), verapamil (VER), mannitol (MAN), or no intervention (control) during machine preservation. All kidneys were preserved by continuous hypothermic pulsatile perfusion (CHPP) using Belzer II solution, and perfusion parameters were measured every 2 hours during pulsatile perfusion. The addition of PGE1 to the perfusate increased renal flow and decreased renal resistance. Moreover, the PGE1 treated group was associated with improved early graft function when compared with all other groups. The addition of VER, TFP, and MAN influenced neither the perfusion characteristics nor the incidence of early graft function. Treatment with PGE1 during machine preservation enhances hydrostatic perfusion parameters (renal flow and renal resistance) and reduces the incidence of delayed graft function in ECD kidneys.

Liver transplantation for adult polycystic liver disease.

Polycystic liver disease, commonly associated with polycystic kidney disease, can result in massive hepatomegaly and debilitating symptoms. Surgical intervention for symptomatic polycystic liver disease has been associated with significant morbidity and inconsistent long-term palliation; it is more appropriate in patients with a single dominant cyst or cysts which is/are confined to one lobe. At our institution, nine patients have undergone orthotopic liver transplantation for symptomatic hepatic cysts with excellent long-term results and minimal morbidity and mortality. Surgical candidates were selected based on severe limitations in daily activities and on sequelae of hepatic cystic involvement. Other factors considered were the extent and pattern of hepatic cystic disease, the degree of hepatic and renal dysfunction, and prior surgical intervention. Three patients (33%) required combined liver and kidney transplantation because of renal cystic involvement with renal insufficiency. The one-year survival rate was 89% with excellent symptomatic relief and improved quality of life in all the surviving patients. One death occurred in a significantly malnourished 62-year-old female. Complications included one case each of hepatic artery thrombosis requiring retransplantation, biliary leak necessitating biliary reconstruction, and postoperative bleeding requiring re-exploration. The mean hospital stay was 23 days and the mean intraoperative blood transfusion requirement was 18 units. Our experience demonstrates that appropriately selected patients with extensive hepatic involvement with adult polycystic liver disease can have an excellent outcome with transplantation, with morbidity comparable with other surgical options.

Significance of early aminotransferase elevation after liver transplantation.

BACKGROUND: We retrospectively reviewed 213 consecutive patients who received their first liver allograft between January 1 and December 31, 1993, in order to study the impact of ischemia/preservation/reperfusion injury (IPRI) on patient and graft outcome. METHODS: The extent of IPRI was assessed by the peak value of aspartate aminotransferase (ASTmax) observed within the first 72 hr after transplant. For the purpose of univariate analysis, categorical classification of recipients was done based upon ASTmax as follows: group 1, ASTmax<600 U/L (n=46); group 2, ASTmax=600-2000 U/L (n=97); group 3, ASTmax>2000-5000 U/L (n=50), and group 4, ASTmax>5000 U/L (n=17). For multivariate analysis, stepwise Cox regression was performed with age, ASTmax, and United Network for Organ Sharing (UNOS) status as covariates. RESULTS: Groups were comparable with respect to age, UNOS status at the time of transplantation, and diagnostic case mix. Median follow-up was 644 days. The overall incidence of primary graft nonfunction (PNF) was 7.6%. PNF incidence was significantly correlated with the severity of IPRI (0%, 4%, 10%, and 41% for groups 1 to 4, respectively, P < 0.0001), but this impact was confined to the respective rates of retransplantation as early patient survival was unaffected. The 1-year survival of patients whose initial grafts manifested extreme IPRI (group 4) was significantly inferior to recipients in the three other groups (77%, 71%, 73%, and 52% for groups 1 to 4, respectively, P=0.03). This increased mortality was confined to patients who never achieved discharge from their initial hospitalization, with no significant differences between groups being detected in the survival of those patients who were discharged (84%, 80%, 85%, and 81% for groups 1 to 4, respectively, P=NS). Although overall 1-year graft survival was strongly correlated with the extent of IPRI (77%, 67%, 62%, and 41% for groups 1 to 4, respectively, P=0.001), this correlation was abolished when survival of grafts not lost to PNF was examined at 1 and 2 years. Stepwise Cox regression analysis confirmed the independent association between ASTmax and patient and graft survival. The long-term quality of allograft function as well as the incidence of chronic rejection and biliary complications were unrelated to the extent of IPRI. CONCLUSIONS: We conclude that: (1) patient survival is influenced by IPRI only when it is extreme (ASTmax>5000 U/L), provided parameters of graft function are used in conjunction with aminotransferase values to assess the need for prompt retransplantation; (2) short-term graft survival is proportional to the extent of IPRI, but grafts that are not lost to PNF have equivalent 1- and 2-year survival irrespective of the magnitude of IPRI; (3) 40% of grafts with extreme IPRI are lost to PNF, but the same proportion also provide long-term function; and (4) for surviving grafts, long-term biochemical function as well as the incidence of biliary complications and of chronic rejection are unrelated to the extent of IPRI.