Impact of Portable Normothermic Machine Perfusion for Liver Transplantation From Adult Deceased Donors.

OBJECTIVE: To assess how liver allografts preserved using portable normothermic machine perfusion (NMP) compare against those that underwent ischemic cold storage (ICS) in the setting of donation after brain death (DBD) and donation after circulatory death (DCD) liver transplantation (LT). BACKGROUND: Compared with conventional ICS, NMP may offer more homeostatic preservation, permit physiological assessment of organ function, and provide opportunities for graft improvement/modification. We report a single-center US experience of liver NMP. METHODS: A single-center, retrospective analysis of collected data on 541 adult whole LTs from 469 DBD donors [NMP (n = 58) vs ICS (n = 411)] and 72 DCD donors [NMP (n = 52) vs ICS (n = 20)] between January 2016 and December 2022. RESULTS: In DBD LT, male sex [odds ratio (95% CI): 1.83 (1.08-3.09)] and >10% macrosteatosis of the donor liver [1.85 (1.10-3.10)] were statistically significant independent risk factors of early allograft dysfunction (EAD). Donor age >40 years and cold ischemia time >7 hours were independent risk factors of reperfusion syndrome (RPS). One-year, 3-year, and 5-year incidences of ischemic cholangiopathy (IC) did not differ significantly in DBD cases between the NMP and ICS cohorts. In DCD LT, NMP was an independent protective factor against EAD [0.11 (0.03-0.46)] and RPS [0.04 (0.01-0.25)]. The incidence of IC in the DCD cases at 1-year and 3-year time points was significantly lower in the NMP cohort (1.9% compared with 20% in the ICS group). CONCLUSIONS: Compared with conventional ICS, NMP can significantly reduce the incidence of EAD, RPS, and IC after DCD LT.

Impact of donor sex on hepatocellular carcinoma recurrence in liver transplantation after brain death.

BACKGROUND: Hepatocellular carcinoma (HCC) is predominantly seen in males but has a better prognosis in females. No prior studies have investigated HCC recurrence based on sex combination following liver transplant donated after brain death (DBDLT). This study sought to elucidate the effects of donor and recipient sex on HCC recurrence rates. METHODS: 9232 adult recipients from the United Network for Organ Sharing (UNOS) database who underwent DBDLT for HCC from 2012 to 2018 were included. Donor-recipient pairs were divided into (1) female donor/female recipient (F-F) (n = 1089); (2) male donor/female recipient (M-F) (n = 975); (3) female donor/male recipient (F-M) (n = 2691); (4) male donor/male recipient (M-M) (n = 4477). The primary prognostic outcome was HCC recurrence. A multivariable competing risk regression analysis was used to assess prognostic influences. RESULTS: The median recipient age and model for end-stage liver disease (MELD) scores were similar among the four groups. Livers of male recipients demonstrated greater in size and number of HCC (both p-values were <.0001). There was also a higher rate of vascular invasion in male recipients compared to female (p < .0001). Competing risk analyses showed that the cumulative HCC recurrence rate was significantly lower in the M-F group (p = .013). After adjusting for tumor characteristics, liver grafts from male donors were associated with a lower HCC recurrence rate in female recipients (HR: .62 95%CI: .42-.93) (p = .021). CONCLUSION: In DBDLT, male donor to female recipient pairing exhibited lower HCC recurrence rates. SUMMARY: Lowest rates of HCC recurrence were confirmed among the female recipients of male donor grafts group in the deceased donor LT cohort. A competing risk multivariable regression analysis demonstrated that male donor sex was significantly associated with low HCC recurrence in female but not male recipients.

Resection of NAFLD/NASH-related Hepatocellular Carcinoma (HCC): Clinical Features and Outcomes Compared with HCC Due to Other Etiologies.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) are the leading causes of hepatocellular carcinoma (HCC) worldwide. Limited data exist on surgical outcomes for NAFLD/NASH-related HCC compared with other HCC etiologies. We evaluated differences in clinicopathological characteristics and outcomes of patients undergoing surgical resection for NAFLD/NASH-associated HCC compared with other HCC etiologies. METHODS: Demographic, clinicopathological features, and survival outcomes of patients with surgically resected HCC were collected. NAFLD activity score (NAS) and fibrosis score were assessed by focused pathologic review in a subset of patients. RESULTS: Among 492 patients screened, 260 met eligibility (NAFLD/NASH [n = 110], and other etiologies [n = 150]). Median age at diagnosis was higher in the NAFLD/NASH HCC cohort compared with the other etiologies cohort (66.7 vs. 63.4 years, respectively, P = .005), with an increased percentage of female patients (36% vs. 18%, P = .001). NAFLD/NASH-related tumors were more commonly >5 cm (66.0% vs. 45%, P = .001). There were no significant differences in rates of lymphovascular or perineural invasion, histologic grade, or serum AFP levels. The NAFLD/NASH cohort had lower rates of background liver fibrosis, lower AST and ALT levels, and higher platelet counts (P < .01 for all). Median overall survival (OS) was numerically shorter in NAFLD/NASH vs other etiology groups, however, not statistically significant. CONCLUSIONS: Patients with NAFLD/NASH-related HCC more commonly lacked liver fibrosis and presented with larger HCCs compared with patients with HCC from other etiologies. No differences were seen in rates of other high-risk features or survival. With the caveat of sample size and retrospective analysis, this supports a similar decision-making approach regarding surgical resection for NAFLD/NASH and other etiology-related HCCs.

Revisiting the "Weekend Effect" on Adult and Pediatric Liver and Kidney Offer Acceptance.

BACKGROUND Weekends can impose resource and manpower constraints on hospitals. Studies using data from prior allocation schemas showed increased adult organ discards on weekends. We examined the impact of day of the week on adult and pediatric organ acceptance using contemporary data. MATERIAL AND METHODS Retrospective analysis of UNOS-PTR match-run data of all offers for potential kidney and liver transplant from 1/1/2016 to 7/1/2021 were examined to study the rate at which initial offers were declined depending on day of the week. Risk factors for decline were also evaluated. RESULTS Of the total initial adult/pediatric liver and kidney offers, the fewest offers occurred on Mondays and Sundays. The decline rate for adult/pediatric kidneys was highest on Saturdays and lowest on Tuesdays. The decline rate for adult livers was highest on Saturday and lowest on Wednesday. In contrast, the decline rate for pediatric livers was highest on Tuesdays and lowest on Wednesdays. Independent risk factors from multivariate analysis of the adult/pediatric kidney and liver decline rate were analyzed. The weekend offer remains an independent risk factor for adult kidney and liver offer declines, but for pediatric offers, these were not significant independent risk factors. CONCLUSIONS Although allocation systems have changed, and the availability of kidneys and livers have increased in the USA over the past 5 years, the weekend effect remains significant for adult liver and kidney offers for declines. Interestingly, the weekend effect was not seen for pediatric liver and kidney offers.

Suppression of allograft rejection by regulatory B cells induced via TLR signaling.

B lymphocytes have long been recognized for their critical contributions to adaptive immunity, providing defense against pathogens through cognate antigen presentation to T cells and Ab production. More recently appreciated is that B cells are also integral in securing self-tolerance; this has led to interest in their therapeutic application to downregulate unwanted immune responses, such as transplant rejection. In this study, we found that PMA- and ionomycin-activated mouse B cells acquire regulatory properties following stimulation through TLR4/TLR9 receptors (Bregs-TLR). Bregs-TLR efficiently inhibited T cell proliferation in vitro and prevented allograft rejection. Unlike most reported Breg activities, the inhibition of alloimmune responses by Bregs-TLR relied on the expression of TGF-ß and not IL-10. In vivo, Bregs-TLR interrupted donor-specific T cell expansion and induced Tregs in a TGF-ß-dependent manner. RNA-Seq analyses corroborated the involvement of TGF-ß pathways in Breg-TLR function, identified potential gene pathways implicated in preventing graft rejection, and suggested targets to foster Breg regulation.

Stent Diameter, Not Cephalic Arch Anatomy, Predicts Stent Graft Patency in Cephalic Arch Stenosis.

PURPOSE: To investigate the relationship between anatomic factors and primary patency of brachiocephalic arteriovenous fistulae (AVFs) after stent graft (SG) placement for cephalic arch stenosis (CAS). MATERIALS AND METHODS: This retrospective study reviewed all cephalic arch SGs placed in brachiocephalic AVFs in a tertiary academic medical center between 2014 and 2017. Sixty-three patients were included in the study. The mean patient age at the time of SG placement was 62.6 years ± 19, and the mean patient follow-up was 1,994 days ± 353. A cohort of patients (n = 31) who underwent brachiocephalic fistulograms for CAS but only received percutaneous transluminal angioplasty (PTA) was the control group. Patient demographic characteristics, AVF anatomy, SG type, and clinical outcomes were reviewed. The duration of primary cephalic arch patency after SG placement was compared with that after previous PTA. RESULTS: The median AVF age at the time of data retrieval was 345 days. The primary patency of CAS after SG placement at 6 months, 12 months, and 3 years was 64%, 49.9%, and 23.5%, respectively. Primary cephalic arch patency was significantly associated with the SG diameter (P = .007) but not with cephalic vein-axillary vein junction anatomy, size of feeding artery, or SG length (P > .05). The primary patency of CAS in patients treated with PTA only (n = 31) at 6 months, 12 months, and 3 years was 61%, 35%, and 0%, respectively, which was significantly lower than that in patients treated with SG placement (P = .01). CONCLUSIONS: This study showed that the primary patency of CAS after SG placement was significantly higher than that of PTA-only treatment. Moreover, primary cephalic arch patency after SG placement was significantly associated with the SG diameter.

Impact of Portable Normothermic Blood-Based Machine Perfusion on Outcomes of Liver Transplant: The OCS Liver PROTECT Randomized Clinical Trial.

IMPORTANCE: Ischemic cold storage (ICS) of livers for transplant is associated with serious posttransplant complications and underuse of liver allografts. OBJECTIVE: To determine whether portable normothermic machine perfusion preservation of livers obtained from deceased donors using the Organ Care System (OCS) Liver ameliorates early allograft dysfunction (EAD) and ischemic biliary complications (IBCs). DESIGN, SETTING, AND PARTICIPANTS: This multicenter randomized clinical trial (International Randomized Trial to Evaluate the Effectiveness of the Portable Organ Care System Liver for Preserving and Assessing Donor Livers for Transplantation) was conducted between November 2016 and October 2019 at 20 US liver transplant programs. The trial compared outcomes for 300 recipients of livers preserved using either OCS (n = 153) or ICS (n = 147). Participants were actively listed for liver transplant on the United Network of Organ Sharing national waiting list. INTERVENTIONS: Transplants were performed for recipients randomly assigned to receive donor livers preserved by either conventional ICS or the OCS Liver initiated at the donor hospital. MAIN OUTCOMES AND MEASURES: The primary effectiveness end point was incidence of EAD. Secondary end points included OCS Liver ex vivo assessment capability of donor allografts, extent of reperfusion syndrome, incidence of IBC at 6 and 12 months, and overall recipient survival after transplant. The primary safety end point was the number of liver graft-related severe adverse events within 30 days after transplant. RESULTS: Of 293 patients in the per-protocol population, the primary analysis population for effectiveness, 151 were in the OCS Liver group (mean [SD] age, 57.1 [10.3] years; 102 [67%] men), and 142 were in the ICS group (mean SD age, 58.6 [10.0] years; 100 [68%] men). The primary effectiveness end point was met by a significant decrease in EAD (27 of 150 [18%] vs 44 of 141 [31%]; P = .01). The OCS Liver preserved livers had significant reduction in histopathologic evidence of ischemia-reperfusion injury after reperfusion (eg, less moderate to severe lobular inflammation: 9 of 150 [6%] for OCS Liver vs 18 of 141 [13%] for ICS; P = .004). The OCS Liver resulted in significantly higher use of livers from donors after cardiac death (28 of 55 [51%] for the OCS Liver vs 13 of 51 [26%] for ICS; P = .007). The OCS Liver was also associated with significant reduction in incidence of IBC 6 months (1.3% vs 8.5%; P = .02) and 12 months (2.6% vs 9.9%; P = .02) after transplant. CONCLUSIONS AND RELEVANCE: This multicenter randomized clinical trial provides the first indication, to our knowledge, that normothermic machine perfusion preservation of deceased donor livers reduces both posttransplant EAD and IBC. Use of the OCS Liver also resulted in increased use of livers from donors after cardiac death. Together these findings indicate that OCS Liver preservation is associated with superior posttransplant outcomes and increased donor liver use. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT02522871.

Hepatectomy for Solitary Hepatocellular Carcinoma: Resection Margin Width Does Not Predict Survival.

BACKGROUND: The survival benefit of negative margins for hepatocellular carcinoma (HCC) has been demonstrated. However, there is no consensus regarding the optimal resection margin width. We assessed the impact of hepatic resection margin width for solitary HCC on overall (OS), recurrence-free (RFS), and liver-specific recurrence-free survival (LSRFS). METHODS: Clinicopathologic data were retrospectively collected for solitary HCC patients who underwent a negative margin hepatectomy (1992-2015). Margin width was categorized in tertiles as "narrow" (≤ 0.3 cm), "intermediate" (0.31-1.0 cm), or "wide" (> 1.0 cm). Survival was compared among groups. RESULTS: Of the 178 included patients, most were male (76%); median age, MELD score, and tumor size were 63 years, 8, and 5.2 cm, respectively; 93% were Child-Pugh class A. Median margin width was 0.5 cm. Median follow-up and OS were 47.8 months and 76.7 months, respectively. There was no significant survival difference among narrow, intermediate, and wide margin groups with a median OS of 53 months (IQR 21-not reached [NR]), 74 months (IQR 14-138), and 97 months (IQR 37-142) (p = 0.87), respectively. Median RFS was 33.0 months; again, there was no difference among narrow, intermediate, and wide margin groups with a median of 31 months (IQR 18-NR), 45 months (IQR 14-NR), and 27 months (IQR 11-NR), respectively (p = 0.66). Median LSRFS was 63.0 months (IQR 14-NR) with no difference among groups (p = 0.87). In multivariate analyses, margin width was not associated with OS (p = 0.77), RFS (p = 0.74), or LSRFS (p = 0.92). Findings were similar in all subgroups analyzed (≤ 5 cm, > 5 cm, microvascular invasion, T1, T2/T3, anatomical or non-anatomical resection, major or minor hepatectomy). CONCLUSIONS: Narrow margins appear to be oncologically safe and the feasibility of achieving wide margins should not determine resectability.

Multi-Center Analysis of Liver Transplantation for Combined Hepatocellular Carcinoma-Cholangiocarcinoma Liver Tumors.

BACKGROUND: Combined hepatocellular-cholangiocarcinoma liver tumors (cHCC-CCA) with pathologic differentiation of both hepatocellular carcinoma (HCC) and intrahepatic cholangiocarcinoma within the same tumor are not traditionally considered for liver transplantation due to perceived poor outcomes. Published results are from small cohorts and single centers. Through a multicenter collaboration, we performed the largest analysis to date of the utility of liver transplantation for cHCC-CCA. STUDY DESIGN: Liver transplant and resection outcomes for HCC (n = 2,998) and cHCC-CCA (n = 208) were compared in a 12-center retrospective review (2009 to 2017). Pathology defined tumor type. Tumor burden was based on radiologic Milan criteria at time of diagnosis and applied to cHCC-CCA for uniform analysis. Kaplan-Meier survival curves and log-rank test were used to determine overall survival and disease-free survival. Cox regression was used for multivariate survival analysis. RESULTS: Liver transplantation for cHCC-CCA (n = 67) and HCC (n = 1,814) within Milan had no significant difference in overall survival (5-year cHCC-CCA 70.1%, HCC 73.4%, p = 0.806), despite higher cHCC-CCA recurrence rates (23.1% vs 11.5% 5 years, p < 0.001). Irrespective of tumor burden, cHCC-CCA tumor patient undergoing liver transplant had significantly superior overall survival (p = 0.047) and disease-free survival (p < 0.001) than those having resection. For cHCC-CCA within Milan, liver transplant was associated with improved disease-free survival over resection (70.3% vs 33.6% 5 years, p < 0.001). CONCLUSIONS: Regardless of tumor burden, outcomes after liver transplantation are superior to resection for patients with cHCC-CCA. Within Milan criteria, liver transplant for cHCC-CCA and HCC result in similar overall survival, justifying consideration of transplantation due to the higher chance of cure with liver transplantation in this traditionally excluded population.

Donor plasmacytoid dendritic cells modulate effector and regulatory T cell responses in mouse spontaneous liver transplant tolerance.

We assessed the role of donor liver non-conventional plasmacytoid dendritic cells (pDCs) in spontaneous liver transplant tolerance in a fully MHC-mismatched (C57BL/6 (H2b ) to C3H (H2k )) mouse model. Compared with spleen pDCs, liver pDCs expressed higher levels of DNAX-activating protein of 12 kDa and its co-receptor, triggering receptor expressed by myeloid cells 2, and higher ratios of programed death ligand-1 (PD-L1):costimulatory CD80/CD86 in the steady state and after Toll-like receptor 9 ligation. Moreover, liver pDCs potently suppressed allogeneic CD4+ and CD8+ T cell proliferative responses. Survival of pDC-depleted livers was much poorer (median survival time: 25 days) than that of either untreated donor livers or pDC-depleted syngeneic donor livers that survived indefinitely. Numbers of forkhead box p3 (FoxP3)+ regulatory T cells in grafts and mesenteric lymph nodes of mice given pDC-depleted allogeneic livers were reduced significantly compared with those in recipients of untreated livers. Graft-infiltrating CD8+ T cells with an exhausted phenotype (programed cell death protein 1+ , T cell immunoglobulin and mucin domain-containing protein 3+ ) were also reduced in recipients of pDC-depleted livers. PD1-PD-L1 pathway blockade reversed the reduction in exhausted T cells. These novel observations link immunoregulatory functions of liver interstitial pDCs, alloreactive T cell exhaustion, and spontaneous liver transplant tolerance.

Liver transplantation with suprahepatic caval anastomosis including inferior vena cava stent.

Budd Chiari syndrome (BCS) results from hepatic outflow obstruction. Endovascular management to restore venous patency, including inferior vena cava (IVC) angioplasty with stenting, and transjugular intrahepatic shunt (TIPS) placement to decompress liver congestion, have become standard of care. Herein, we describe a patient with BCS requiring liver transplantation and the surgical technique of suprahepatic IVC anastomosis including thoracic extension of an IVC stent with a review of the relevant literature. A 29-year-old female with BCS due to polycythemia vera, who had been previously managed with TIPS and IVC stent placement, was taken for liver transplantation. Preoperative imaging confirmed stent extension into the thoracic IVC and the stent was unable to be removed intraoperatively. The thoracic IVC was clamped through the diaphragm at the level of the right atrium and the stent was left in place and incorporated within the suprahepatic anastomosis with good vascular outcome at one year follow up. Diligent preoperative preparation is essential with adequate imaging and cardiac surgical consultation in patients with malpositioned stents. Review of the literature shows four cases in which performing the suprahepatic anastomosis including an embedded stent is a viable alternative that allows for avoidance of a thoracotomy.

Immediate administration of antiviral therapy after transplantation of hepatitis C-infected livers into uninfected recipients: Implications for therapeutic planning.

The practice of transplanting hepatitis C (HCV)-infected livers into HCV-uninfected recipients has not previously been recommended in transplant guidelines, in part because of concerns over uncontrolled HCV infection of the allograft. Direct-acting antivirals (DAAs) provide an opportunity to treat donor-derived HCV-infection and should be administered early in the posttransplant period. However, evidence on the safety and efficacy of an immediate DAA treatment approach, including how to manage logistical barriers surrounding timely DAA procurement, are required prior to broader use of HCV-positive donor organs. We report the results of a trial in which 14 HCV-negative patients underwent successful liver transplantation from HCV-positive donors. Nine patients received viremic (nucleic acid testing [NAT]-positive) livers and started a 12-week course of oral glecaprevir-pibrentasvir within 5 days of transplant. Five patients received livers from HCV antibody-positive nonviremic donors and were followed using a reactive approach. Survival in NAT-positive recipients is 100% at a median follow-up of 46 weeks. An immediate treatment approach for HCV NAT-positive liver transplantation into uninfected recipients is safe and efficacious. Securing payer approval for DAAs early in the posttransplant course could enable need-based allocation of HCV-positive donor organs irrespective of candidate HCV status, while averting chronic HCV allograft infection.

Regulatory B cells require antigen recognition for effective allograft tolerance induction.

Through multiple mechanisms, regulatory B cells (Breg) have been shown to play an important role in the development of allograft tolerance. However, a careful understanding of the role of antigen-specificity in Breg-mediated allograft tolerance has remained elusive. In experimental models of islet and cardiac transplantation, it has been established that Bregs can be induced in vivo by anti-CD45RB ± anti-TIM1antibody treatment, resulting in prolonged, Breg-dependent allograft tolerance. The importance of Breg antigen recognition has been suggested but not confirmed through adoptive transfer experiments, using tolerant WT C57BL/6 animals challenged with either BALB/c or C3H grafts. However, the importance of receptor-specificity has not been formally tested. Here, we utilize the novel ovalbumin-specific B cell receptor transnuclear (OBI) mice in multiple primary tolerance and adoptive transfer experiments to establish that Breg-dependent allograft tolerance relies on antigen recognition by B cells. Additionally, we identify that this Breg-dependent tolerance relies on the function of transforming growth factor-ß. Together, these experiments mark important progress toward understanding how best to improve Breg-mediated allograft tolerance.

Charting the Path Forward for Risk Prediction in Liver Transplant for Hepatocellular Carcinoma: International Validation of HALTHCC Among 4,089 Patients.

Prognosticating outcomes in liver transplant (LT) for hepatocellular carcinoma (HCC) continues to challenge the field. Although Milan Criteria (MC) generalized the practice of LT for HCC and improved outcomes, its predictive character has degraded with increasing candidate and oncological heterogeneity. We sought to validate and recalibrate a previously developed, preoperatively calculated, continuous risk score, the Hazard Associated with Liver Transplantation for Hepatocellular Carcinoma (HALTHCC), in an international cohort. From 2002 to 2014, 4,089 patients (both MC in and out [25.2%]) across 16 centers in North America, Europe, and Asia were included. A continuous risk score using pre-LT levels of alpha-fetoprotein, Model for End-Stage Liver Disease Sodium score, and tumor burden score was recalibrated among a randomly selected cohort (n = 1,021) and validated in the remainder (n = 3,068). This study demonstrated significant heterogeneity by site and year, reflecting practice trends over the last decade. On explant pathology, both vascular invasion (VI) and poorly differentiated component (PDC) increased with increasing HALTHCC score. The lowest-risk patients (HALTHCC 0-5) had lower rates of VI and PDC than the highest-risk patients (HALTHCC > 35) (VI, 7.7%[ 1.2-14.2] vs. 70.6% [48.3-92.9] and PDC:4.6% [0.1%-9.8%] vs. 47.1% [22.6-71.5]; P < 0.0001 for both). This trend was robust to MC status. This international study was used to adjust the coefficients in the HALTHCC score. Before recalibration, HALTHCC had the greatest discriminatory ability for overall survival (OS; C-index = 0.61) compared to all previously reported scores. Following recalibration, the prognostic utility increased for both recurrence (C-index = 0.71) and OS (C-index = 0.63). Conclusion: This large international trial validated and refined the role for the continuous risk metric, HALTHCC, in establishing pre-LT risk among candidates with HCC worldwide. Prospective trials introducing HALTHCC into clinical practice are warranted.

Reframing the approach to patients with hepatocellular carcinoma: Longitudinal assessment with hazard associated with liver transplantation for HCC (HALTHCC) improves ablate and wait strategy.

Patients with hepatocellular carcinoma (HCC) are screened at presentation for appropriateness of liver transplantation (LT) using morphometric criteria, which poorly specifies risk. Morphology is the crux of measuring tumor response to locoregional therapy (LRT) using modified Response Evaluation Criteria in Solid Tumors (mRECIST). This study investigated the utility of following a continuous risk score (hazard associated with liver transplantation in hepatocellular carcinoma; HALTHCC) to longitudinally assess risk. This multicenter, retrospective study from 2002 to 2014 enrolled 419 patients listed for LT for HCC. One cohort had LRT while waiting (n = 351), compared to the control group (n = 68) without LRT. Imaging studies (n = 2,085) were collated to laboratory data to calculate HALTHCC, MORAL, Metroticket 2.0, and alpha fetoprotein (AFP) score longitudinally. Cox proportional hazards evaluated associations of HALTHCC and peri-LRT changes with intention-to-treat (ITT) survival (considering dropout or post-LT mortality), and utility was assessed with Harrell's C-index. HALTHCC better predicted ITT outcome (LT = 309; dropout = 110) when assessed closer to delisting (P < 0.0001), maximally just before delisting (C-index, 0.742 [0.643-0.790]). Delta-HALTHCC post-LRT was more sensitive to changes in risk than mRECIST. HALTHCC score and peri-LRT percentage change were independently associated with ITT mortality (hazard ratio = 1.105 [1.045-1.169] per point and 1.014 [1.004-1.024] per percent, respectively). CONCLUSIONS: HALTHCC is superior in assessing tumor risk in candidates awaiting LT, and its utility increases over time. Peri-LRT relative change in HALTHCC outperforms mRECIST in stratifying risk of dropout, mortality, and recurrence post-LT. With improving estimates of post-LT outcomes, it is reasonable to consider allocation using HALTHCC and not just waiting time. Furthermore, this study supports a shift in perspective, from listing to allocation, to better utilize precious donor organs. (Hepatology 2018).

Perturbed Calcineurin-NFAT Signaling Is Associated with the Development of Alzheimer's Disease.

Down syndrome (DS), the most common genetic disorder, is caused by trisomy 21. DS is accompanied by heart defects, hearing and vision problems, obesity, leukemia, and other conditions, including Alzheimer's disease (AD). In comparison, most cancers are rare in people with DS. Overexpression of dual specificity tyrosine-phosphorylation-regulated kinase 1A and a regulator of calcineurin 1 located on chromosome 21 leads to excessive suppression of the calcineurin-nuclear factor of activated T cells (NFAT) signaling pathway, resulting in reduced expression of a critical angiogenic factor. However, it is unclear whether the calcineurin-NFAT signaling pathway is involved in AD pathology in DS patients. Here, we investigated the association between the calcineurin-NFAT signaling pathway and AD using neuronal cells. Short-term pharmacological stimulation decreased gene expression of tau and neprilysin, and long-term inhibition of the signaling pathway decreased that of amyloid precursor protein. Moreover, a calcineurin inhibitor, cyclosporine A, also decreased neprilysin activity, leading to increases in amyloid-ß peptide levels. Taken together, our results suggest that a dysregulation in calcineurin-NFAT signaling may contribute to the early onset of AD in people with DS.

Orthotopic mouse liver transplantation to study liver biology and allograft tolerance.

Orthotopic liver transplantation in the mouse is a powerful research tool that has led to important mechanistic insights into the regulation of hepatic injury, liver immunopathology, and transplant tolerance. However, it is a technically demanding surgical procedure. Setup of the orthotopic liver transplantation model comprises three main stages: surgery on the donor mouse; back-table preparation of the liver graft; and transplant of the liver into the recipient mouse. In this protocol, we describe our procedure in stepwise detail to allow efficient completion of both the donor and recipient operations. The protocol can result in consistently high technical success rates when performed by personnel experienced in the protocol. The technique can be completed in ∼2-3 h when performed by an individual who is well practiced in performing mouse transplantation in accordance with this protocol. We have achieved a perioperative survival rate close to 100%.

Template-dependent nucleotide addition in the reverse (3'-5') direction by Thg1-like protein.

Thg1-like protein (TLP) catalyzes the addition of a nucleotide to the 5'-end of truncated transfer RNA (tRNA) species in a Watson-Crick template-dependent manner. The reaction proceeds in two steps: the activation of the 5'-end by adenosine 5'-triphosphate (ATP)/guanosine 5'-triphosphate (GTP), followed by nucleotide addition. Structural analyses of the TLP and its reaction intermediates have revealed the atomic detail of the template-dependent elongation reaction in the 3'-5' direction. The enzyme creates two substrate binding sites for the first- and second-step reactions in the vicinity of one reaction center consisting of two Mg(2+) ions, and the two reactions are executed at the same reaction center in a stepwise fashion. When the incoming nucleotide is bound to the second binding site with Watson-Crick hydrogen bonds, the 3'-OH of the incoming nucleotide and the 5'-triphosphate of the tRNA are moved to the reaction center where the first reaction has occurred. That the 3'-5' elongation enzyme performs this elaborate two-step reaction in one catalytic center suggests that these two reactions have been inseparable throughout the process of protein evolution. Although TLP and Thg1 have similar tetrameric organization, the tRNA binding mode of TLP is different from that of Thg1, a tRNA(His)-specific G-1 addition enzyme. Each tRNA(His) binds to three of the four Thg1 tetramer subunits, whereas in TLP, tRNA only binds to a dimer interface and the elongation reaction is terminated by measuring the accepter stem length through the flexible ß-hairpin. Furthermore, mutational analyses show that tRNA(His) is bound to TLP in a similar manner as Thg1, thus indicating that TLP has a dual binding mode.

Contribution of alloantigens to hepatic ischemia/reperfusion injury: Roles of natural killer cells and innate immune recognition of nonself.

Hepatic ischemia/reperfusion injury (IRI) remains a major clinical problem and involves the innate immune system's recognition of "nonself." Considering the efficient nonself recognition by natural killer (NK) cells, we hypothesize in this study that hepatic IRI associated with liver transplantation (LT) could be augmented in allogeneic rather than in syngeneic (Syn) grafts due to alloantigen recognition by innate immune cells, especially by NK cells. Using green fluorescent protein (GFP)/Sprague-Dawley rats, we tested our hypothesis in a rat LT model with 18 hours of cold storage in University of Wisconsin solution. Hepatic IRI was significantly augmented in allografts with higher alanine transaminase levels, increased necrosis, and vigorous proinflammatory mediator up-regulation compared to Syn grafts. Injury increased in allografts associated with augmented GFP+ host leukocyte infiltration due to significantly increased host CD11b/c+ and RP-1(+) neutrophil recruitment. A large number of liver-resident (donor) mature CD11b/c+ NK cells quickly diminished from allografts, but not from Syn grafts. Depletion of mature NK cells from liver grafts with anti-asialo monosialotetrahexosylganglioside significantly improved hepatic IRI and reduced neutrophil infiltration and proinflammatory mediators. In conclusion, early innate immune responses were more significantly enhanced in allografts than in Syn grafts during hepatic IRI, in part through NK cell recognition of "missing self."