Development of a Large Animal Model of Ischemia-free Liver Transplantation in Pigs.

Background: In organ transplantation, ischemia, and reperfusion injury (IRI) is considered as an inevitable event and the major contributor to graft failure. Ischemia-free liver transplantation (IFLT) is a novel transplant procedure that can prevent IRI and provide better transplant outcomes. However, a large animal model of IFLT has not been reported. Therefore, we develop a new, reproducible, and stable model of IFLT in pigs for investigating mechanisms of IFLT in IRI. Methods: Ten pigs were subjected to IFLT or conventional liver transplantation (CLT). Donor livers in IFLT underwent 6-h continuous normothermic machine perfusion (NMP) throughout graft procurement, preservation, and implantation, whereas livers in CLT were subjected to 6-h cold storage before implantation. The early reperfusion injury was compared between the 2 groups. Results: Continuous bile production, low lactate, and liver enzyme levels were observed during NMP in IFLT. All animals survived after liver transplantation. The posttransplant graft function was improved with IFLT when compared with CLT. Minimal histologic changes, fewer apoptotic hepatocytes, less sinusoidal endothelial cell injury, and proinflammatory cytokine (interleukin [IL]-1ß, IL-6, and tumor necrosis factor-α) release after graft revascularization were documented in the IFLT group versus the CLT group. Conclusions: We report that the concept of IFLT is achievable in pigs. This innovation provides a potential strategy to investigate the mechanisms of IRI and provide better transplant outcomes for clinical practice.

Metabolomics Differences of the Donor Livers Between In Situ and Ex Situ Conditions During Ischemia-free Liver Transplantation.

BACKGROUND: Ischemia-free liver transplantation (IFLT) has been innovated to avoid graft ischemia during organ procurement, preservation, and implantation. However, the metabolism activity of the donor livers between in the in situ and ex situ normothermic machine perfusion (NMP) conditions, and between standard criteria donor and extend criteria donor remains unknown. METHODS: During IFLT, plasma samples were collected both at the portal vein and hepatic vein of the donor livers in situ during procurement and ex situ during NMP. An ultra-high performance liquid chromatography-mass spectrometry was conducted to investigate the common and distinct intraliver metabolite exchange. RESULTS: Profound cysteine and methionine metabolism, and aminoacyl-tRNA biosynthesis were found in both in situ and ex situ conditions. However, obvious D-arginine and D-ornithine metabolism, arginine and proline metabolism were only found in the in situ condition. The suppressed activities of the urea cycle pathway during ex situ condition were confirmed in an RNA expression level. In addition, compared with extend criteria donor group, standard criteria donor group had more active intraliver metabolite exchange in metabonomics level. Furthermore, we found that the relative concentration of p-cresol, allocystathionine, L-prolyl-L-proline in the ex situ group was strongly correlated with peak alanine aminotransferase and aspartate aminotransferase at postoperative days 1-7. CONCLUSIONS: In the current study, we show the common and distinct metabolism activities during IFLT. These findings might provide insights on how to modify the design of NMP device, improve the perfusate components, and redefine the criteria of graft viability.

TUBB2B facilitates progression of hepatocellular carcinoma by regulating cholesterol metabolism through targeting HNF4A/CYP27A1.

Cholesterol metabolism plays a critical role in the progression of hepatocellular carcinoma (HCC), but it is not clear how cholesterol metabolism is regulated. The tubulin beta class I genes (TUBBs) are associated with the prognosis of many different cancers. To confirm the function of TUBBs in HCC, the Kaplan-Meier method and Cox analyses were performed using TCGA and GSE14520 datasets. A higher expression of TUBB2B is an independent prognostic factor for shorter over survival in HCC patients. Deletion of TUBB2B in hepatocytes inhibits proliferation and promotes tumor cell apoptosis, while over-expression of TUBB2B has the opposite function. This result was confirmed in a mouse xenograft tumor model. Mechanistically, TUBB2B induces the expression of CYP27A1, an enzyme responsible for the conversion of cholesterol to 27-hydroxycholesterol, which leads to the up-regulation of cholesterol and the progression of HCC. In addition, TUBB2B regulates CYP27A1 via human hepatocyte nuclear factor 4alpha (HNF4A). These findings indicated that TUBB2B functions as an oncogene in HCC, and plays a role in promoting cell proliferation and anti-apoptosis through targeting HNF4A/CYP27A1/cholesterol.

Neoadjuvant programmed cell death 1 inhibitor before liver transplantation for HCC is not associated with increased graft loss.

Immune checkpoint inhibitors (ICIs) may lead to rejection and even graft loss of solid organ transplant recipients, making them not widely used in transplant patients. There is insufficient clinical experience in using ICIs as a bridging or downstaging therapy before transplantation. We performed a retrospective review of patients receiving programmed cell death 1 inhibitor (PD1) before liver transplantation for HCC in our center and analyzed the data of these patients with the purpose of investigating the safety and feasibility of preoperative PD1 inhibitor among liver transplant recipients and exploring the preoperative correlation ICIs and the postoperative risk of rejection and immune-related graft loss. A total of 16 patients enrolled in this study. Acute rejection occurred in 9 patients, with an incidence of 56.3%. The median time of rejection was 7 days after surgery. The median FK506 concentration at the time of rejection was 7.1 µg/L. All rejection reactions were reversed after adjusting the immunosuppression regimen. The interval between the last PD1 inhibitor and transplantation in the rejection group was shorter than that in the nonrejection group, and there was a statistical difference [21.0 (15.5-27.5) days vs. 60.0 (34.0-167.0) days, p =0.01]. In conclusion, PD1 inhibitor is a safe and feasible method for bridging or downstaging treatment before liver transplantation. Although preoperative PD1 inhibitor may increase the incidence of postoperative rejection, it is not associated with increased immune-related graft loss and patient death.

Abrogation of graft ischemia-reperfusion injury in ischemia-free liver transplantation.

BACKGROUND: Ischemia-reperfusion injury (IRI) is considered an inherent component of organ transplantation that compromises transplant outcomes and organ availability. The ischemia-free liver transplantation (IFLT) procedure has been developed to avoid interruption of blood supply to liver grafts. It is unknown how IFLT might change the characteristics of graft IRI. METHODS: Serum and liver biopsy samples were collected from IFLT and conventional liver transplantation (CLT) recipients. Pathological, metabolomics, transcriptomics, and proteomics analyses were performed to identify the characteristic changes in graft IRI in IFLT. RESULTS: Peak aspartate aminotransferase (539.59 ± 661.76 U/L versus 2622.28 ± 3291.57 U/L) and alanine aminotransferase (297.64 ± 549.50 U/L versus 1184.16 ± 1502.76 U/L) levels within the first 7 days and total bilirubin levels by day 7 (3.27 ± 2.82 mg/dl versus 8.33 ± 8.76 mg/dl) were lower in the IFLT versus CLT group (all p values < 0.001). The pathological characteristics of IRI were more obvious in CLT grafts. The antioxidant pentose phosphate pathway remained active throughout the procedure in IFLT grafts and was suppressed during preservation and overactivated postrevascularization in CLT grafts. Gene transcriptional reprogramming was almost absent during IFLT but was profound during CLT. Proteomics analysis showed that "metabolism of RNA" was the major differentially expressed process between the two groups. Several proinflammatory pathways were not activated post-IFLT as they were post-CLT. The activities of natural killer cells, macrophages, and neutrophils were lower in IFLT grafts than in CLT grafts. The serum levels of 14 cytokines were increased in CLT versus IFLT recipients. CONCLUSIONS: IFLT can largely avoid the biological consequences of graft IRI, thus has the potential to improve transplant outcome while increasing organ utilization.

Single-Cell RNA Sequencing Identifies Intra-Graft Population Heterogeneity in Acute Heart Allograft Rejection in Mouse.

Transplant rejection remains a major barrier to graft survival and involves a diversity of cell types. However, the heterogeneity of each cell type in the allograft remains poorly defined. In the present study, we used single-cell RNA sequencing technology to analyze graft-infiltrating cells to describe cell types and states associated with acute rejection in a mouse heart transplant model. Unsupervised clustering analysis revealed 21 distinct cell populations. Macrophages formed five cell clusters: two resident macrophage groups, two infiltrating macrophage groups and one dendritic cell-like monocyte group. Infiltrating macrophages were predominantly from allogeneic grafts. Nevertheless, only one infiltrating macrophage cluster was in an active state with the upregulation of CD40, Fam26f and Pira2, while the other was metabolically silent. Re-clustering of endothelial cells identified five subclusters. Interestingly, one of the endothelial cell populations was almost exclusively from allogeneic grafts. Further analysis of this population showed activation of antigen processing and presentation pathway and upregulation of MHC class II molecules. In addition, Ubiquitin D was specifically expressed in such endothelial cell population. The upregulation of Ubiquitin D in rejection was validated by staining of mouse heart grafts and human kidney biopsy specimens. Our findings present a comprehensive analysis of intra-graft cell heterogeneity, describe specific macrophage and endothelial cell populations which mediate rejection, and provide a potential predictive biomarker for rejection in the clinic.

Ischemic-Free Liver Transplantation Reduces the Recurrence of Hepatocellular Carcinoma After Liver Transplantation.

Ischemia reperfusion injury (IRI) is an adverse factor for hepatocellular carcinoma (HCC) recurrence after liver transplantation. Ischemic-free liver transplantation (IFLT) is a novel transplant procedure that can largely reduce or even prevent IRI, but the clinical relevance of IFLT and the recurrence of HCC after liver transplantation are still unknown. This retrospective study compared survival outcomes, HCC recurrence, perioperative data and IRI severity following liver transplantation (LT). 30 patients received IFLT and 196 patients received conventional liver transplantation (CLT) were chosen for the entire cohort between June 2017 and August 2020. A 1:3 propensity score matching was performed, 30 IFLT recipients and 85 matched CLT patients were enrolled in propensity-matched cohorts. An univariate and multivariate Cox regression analysis was performed, and showed surgical procedure (CLT vs IFLT) was an independent prognostic factor (HR 3.728, 95% CI 1.172-11.861, P=0.026) for recurrence free survival (RFS) in HCC patients following liver transplantation. In the Kaplan-Meier analysis, the RFS rates at 1 and 3 years after LT in recipients with HCC in the IFLT group were significantly higher than those in the CLT group both in the entire cohort and propensity-matched cohort (P=0.006 and P=0.048, respectively). In addition, patients in the IFLT group had a lower serum lactate level, lower serum ALT level and serum AST level on postoperative Day 1. LT recipients with HCC in the IFLT group had a lower incidence of early allograft dysfunction than LT recipients with HCC in the CLT group. Histological analysis showed no obvious hepatocyte necrosis or apoptosis in IFLT group. In conclusion, IFLT can significantly reduce IRI damage and has the potential to be a useful strategy to reduce HCC recurrence after liver transplantation.

lncRNA NEAT1 regulates the proliferation and migration of hepatocellular carcinoma cells by acting as a miR­320a molecular sponge and targeting L antigen family member 3.

Long non­coding RNAs (lncRNAs) serve a pivotal role in hepatocellular carcinoma (HCC) progression and have been confirmed to participate in the carcinogenesis and development of HCC. Certain studies have focused on lncRNA nuclear enriched abundant transcript 1 (NEAT1) in HCC. However, the relationship between lncRNA NEAT1 and HCC remains unclear. The present study found that NEAT1 was significantly overexpressed in HCC cell lines compared with LX­2 hepatic stellate cells. NEAT1 expression in Huh7 and MHCC­97H cells was increased following transfection with lentivirus (LV)­NEAT1 but inhibited by LV­short hairpin NEAT1. Knockdown of NEAT1 significantly repressed HCC cell viability, increased cell apoptosis, and inhibited cell migration and invasion capacity. By contrast, upregulation of NEAT1 demonstrated the reverse effects. Furthermore, microRNA­320a (miR­230a) was predicted to be a direct target of NEAT1 and was significantly reduced in HCC cells. Additionally, a luciferase activity reporter assay and RNA immunoprecipitation assay were performed to confirm the interaction between miR­320a and NEAT1. Using a dual­luciferase activity assay, L antigen family member 3 (LAGE3) was found to be a target of miR­320a. Finally, in vivo nude mouse models were established, and the results indicated that NEAT1 suppressed HCC progression by targeting miR­320a. In conclusion, the present findings revealed that the NEAT1/miR­320a/LAGE3 axis participates in HCC development and that NEAT1 could be used as a biomarker for HCC.

Prospective, single-centre, randomised controlled trial to evaluate the efficacy and safety of ischaemia-free liver transplantation (IFLT) in the treatment of end-stage liver disease.

INTRODUCTION: During conventional liver transplantation (CLT), ischaemia-reperfusion injury (IRI) is inevitable and is associated with complications such as early allograft dysfunction (EAD), primary non-function and ischaemic-type biliary lesions. We have established a novel procedure called ischaemia-free liver transplantation (IFLT). The results from a pilot study suggest that IFLT might prevent IRI and yield better transplant outcomes than CLT. The purpose of this study was to further assess the efficacy and safety of IFLT versus CLT in patients with end-stage liver disease. METHODS AND ANALYSIS: This is an investigator-initiated, open-label, phase III, prospective, single-centre randomised controlled trial on the effects of IFLT in patients with end-stage liver disease. Adult patients (aged 18-75 years) eligible for liver transplantation will be screened for participation in this trial and will be randomised between the IFLT group (n=34) and the CLT group (n=34). In the IFLT group, the donor liver will be procured, preserved and implanted with continuous normothermic machine perfusion (NMP). In the CLT group, the donor liver will be procured after a fast cold flush, preserved in 0°C-4°C solution and implanted under hypothermic and hypoxic conditions. Patients in both groups will be managed according to the standard protocol of our centre. The primary end point is the incidence of EAD after liver transplantation. Intraoperative and postoperative parameters of donor livers and recipients will be observed and recorded, and postoperative liver graft function, complications and recipient and graft survival will be evaluated. After a 12-month follow-up of the last enrolled recipient, the outcomes will be analysed to evaluate the safety and efficacy of IFLT versus CLT in patients with end-stage liver disease. ETHICS AND DISSEMINATION: The protocol was reviewed and approved by the Ethics Committee of The First Affiliated Hospital of Sun Yat-sen University. The findings will be disseminated to the public through conference presentations and peer-reviewed scientific journals. TRIAL REGISTRATION NUMBER: ChiCTR1900021158.

Upregulated Seizure-Related 6 Homolog-Like 2 Is a Prognostic Predictor of Hepatocellular Carcinoma.

Seizure-related 6 homolog-like 2 (SEZ6L2), which is localized on the cell surface, has been found to be associated with tumor angiogenesis and lung cancer progression. However, the role of SEZ6L2 in hepatocellular carcinoma (HCC) is still unclear. We obtained data from The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) to investigate SEZ6L2 expression and regulation in HCC. Then, HCC tissue samples were collected to verify SEZ6L2 by quantitative real-time polymerase chain reaction (qRT-PCR) and immunohistochemical staining (IHC). Patient information was collected for survival and prognosis analysis. qRT-PCR, IHC, and bioinformatics analysis showed that the SEZ6L2 protein was highly expressed in HCC samples. Clinical data showed that high SEZ6L2 protein expression was correlated with tumor-node-metastasis (TNM) stages (P = 0.046), tumor number (P = 0.016), and tumor size (P = 0.029). Meanwhile, SEZ6L2 overexpression was closely associated with poor overall survival and disease-free survival in HCC patients. Moreover, SEZ6L2 is an independent prognostic predictor for the survival of HCC patients. This study suggests a significant correlation between SEZ6L2 and HCC, which means that SEZ6L2 may potentially serve as a useful prognostic biomarker for HCC patients.

lncRNA XIST regulates proliferation and migration of hepatocellular carcinoma cells by acting as miR-497-5p molecular sponge and targeting PDCD4.

BACKGROUND: MicroRNAs (miRNAs) play a pivotal role in hepatocellular carcinoma (HCC) progression and have been confirmed to participate in the carcinogenesis and development of HCC. However, the relationship between miR-497-5p and HCC remains unclear. METHODS: Kaplan-Meier curve analysis and the log-rank test were used to investigate the efficacy of miR-497-5p on overall survival (OS) and disease-free survival (DFS) in patients with HCC. According to in vitro experiments, programmed cell death 4 (PDCD4) was a target of miR-497-5p by the dual-luciferase activity assay. The efficacy of PDCD4 on cell proliferation and metastasis in HCC was examined by transwell assays, CCK-8 assays and reverse transcription quantitative PCR (RT-qPCR). Additionally, we conducted a luciferase activity reporter assay to confirm the interaction between lncRNA XIST and miR-49-5p. Then, to evaluate the relationship between lncRNA XIST and miR-497-5p, several mechanistic experiments, including qRT-PCR, Western blotting, transwell assays and tumor xenograft assays, were performed. RESULTS: miR-497-5p was upregulated in HCC tissues, and high expression of miR-497-5p resulted in increases in tumor size and tumor number and a higher tumor-node-metastasis (TNM) stage and Edmondson grade in patients with HCC. Silencing miR-497-5p inhibited the proliferation and migration of HCC cells. PDCD4, which was downregulated in HCC tissues, was shown to be a target of miR-497-5p and was negatively correlated with the expression of miR-497-5p. lncRNA XIST was found to act as a miR-497-5p sponge and to regulate the level of PDCD4, which is targeted by miR-497-5p. lncRNA XIST was observed to be downregulated in the HCC tissues and positively correlated with the expression of PDCD4. CONCLUSIONS: Our findings reveal that the XIST/miR-497-5p/PDCD4 axis participates in HCC development and that XIST could be used as a biomarker of HCC.

The long noncoding RNA AK002107 negatively modulates miR-140-5p and targets TGFBR1 to induce epithelial-mesenchymal transition in hepatocellular carcinoma.

The abnormal expression of long noncoding RNAs (lncRNAs) is associated with human carcinoma. The present study aimed to investigate the mechanisms underlying the function of lncRNA AK002107 in the progression of hepatocellular carcinoma (HCC). The differential expression of lncRNAs between HCC and paired nontumor tissues was identified using microarrays, and the correlation between the expression of lncRNA AK002107 and the clinical prognosis of HCC was analyzed. We investigated the role of lncRNA AK002107 in HCC tumor biology in vitro using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT), colony formation, and Matrigel invasion assays and in vivo by assessing the growth of xenografted HCC tumors. The potential microRNAs that interact with lncRNA AK002107 were identified using online tools and were verified using PCR and luciferase reporter assay. The levels of TGFBR1, E-cadherin, and vimentin were determined using western blot assays. We then further investigated the correlation between expression of lncRNA AK002107 with miR-140-5p and TGFBR1 expression in HCC tissues. The expression of lncRNA AK002107 is frequently upregulated in HCC samples and cell lines. Patients with HCC who have elevated lncRNA AK002107 expression exhibit poorer overall survival and disease-free survival. Silencing lncRNA AK002107 expression significantly inhibited HCC cell proliferation, colony formation, and invasion both in vitro and in vivo. Furthermore, lncRNA AK002107 directly binds to miR-140-5p and significantly inhibits miR-140-5p expression. The functions of lncRNA AK002107 in cell growth and tumor invasion are mediated via miR-140-5p. lncRNA AK002107 upregulated TGFBR1 expression and then induced epithelial-mesenchymal transition (EMT) by inhibiting miR-140-5p expression. The expression of lncRNA AK002107 inversely correlated with miR-140-5p expression and positively correlated with TGFBR1 expression in HCC tissues. In summary, lncRNA AK002107 functions as an oncogene in tumors by inhibiting miR-140-5p, targeting TGFBR1, and then inducing EMT. The lncRNA AK002107/miR-140-5p/TGFBR1/EMT regulatory network may be a valuable target for the development of novel diagnostic and treatment methods for HCC.

Overexpression of RALY promotes migration and predicts poor prognosis in hepatocellular carcinoma.

INTRODUCTION: RALY plays a critical role in promoting invasiveness and is associated with poor prognosis in different types of cancers. However, the prognostic value of RALY and its precise role in hepatocellular carcinoma (HCC) remain unknown. MATERIALS AND METHODS: We detected the expression of RALY in 127 clinical HCC tissue samples and seven HCC cell lines by immunohistochemical staining and Western blotting. The prognostic value of RALY expression was assessed using the Kaplan-Meier method. The expression and prognostic value of RALY were also studied by bioinformatics analysis of data from the Gene Expression Omnibus and The Cancer Genome Atlas. The biological influence of RALY on HCC cell lines was studied using proliferation, transwell migration, and invasion assays in vitro. RESULTS: The expression of RALY in HCC tissues was significantly higher than that in adjacent normal liver tissues. Abnormally high expression of RALY was associated with tumor size (P=0.031), TNM stage (P=0.026), presurgical serum AFP levels (P=0.025), and vascular invasion (P=0.001). Kaplan-Meier analysis demonstrated that higher expression of RALY correlated with poorer overall survival and disease-free survival in HCC patients. High RALY expression was an independent adverse prognostic factor for overall survival (HR =2.559, 95% CI: 1.710-3.827, P<0.001) and disease-free survival (HR =2.053, 95% CI: 1.384-3.047, P<0.001) in HCC. Moreover, knockdown of RALY expression using a specific shRNA suppressed the proliferation, migration, and invasion capabilities of HCC cells in vitro. Knockdown of RALY expression in HCC cell lines resulted in upregulation of E-cadherin and downregulation of N-cadherin, vimentin, and snail. CONCLUSION: Taken together, our results indicate that RALY represents a biomarker for the prognosis of patients with HCC and highlight the importance of RALY as an oncogene in HCC.

Outcomes of Organ Transplantation from Donors with a Cancer History.

BACKGROUND The inherent challenges of selecting an acceptable donor for the increasing number and acuity of recipients has forced programs to take increased risks, including accepting donors with a cancer history (DWCH). Outcomes of organ transplantation using organs from DWCH must be clarified. We assessed transplant outcomes of recipients of organs from DWCH. MATERIAL AND METHODS Retrospective analysis of the Scientific Registry of Transplant Recipients data from January 1, 2000 to December 31, 2014 identified 8385 cases of transplants from DWCH. A Cox-proportional hazard regression model and log-rank test were used to compare patient survival and hazard levels of various cancer types. RESULTS DWCH was an independent risk factor of 5-year patient survival (HR=1.089, 95% CI: 1.009-1.176, P=0.03) and graft survival (HR=1.129, 95% CI: 1.056-1.208, P<0.01) in liver and heart transplantation (patient survival: HR=1.112, 95% CI: 1.057-1.170, P<0.01; graft survival: HR=1.244, 95% CI: 1.052-1.472, P=0.01). There was no remarkable difference between the 2 groups in kidney and lung transplantation. Donors with genitourinary and gastrointestinal cancers were associated with inferior outcomes in kidney transplantation. Transplantation from donors with central nervous system cancer resulted in poorer survival in liver transplant recipients. Recipients of organs from donors with hematologic malignancy and otorhinolaryngologic cancer had poorer survival following heart transplantation. CONCLUSIONS Under the current donor selection criteria, recipients of organs from DWCH had inferior outcomes in liver and heart transplantation, whereas organs from DWCH were safely applied in kidney and lung transplantation. Specific cancer types should be cautiously evaluated before performing certain types of organ transplantation.

The first case of ischemia-free organ transplantation in humans: A proof of concept.

Ischemia and reperfusion injury (IRI) is an inevitable event in conventional organ transplant procedure and is associated with significant mortality and morbidity post-transplantation. We hypothesize that IRI is avoidable if the blood supply for the organ is not stopped, thus resulting in optimal transplant outcomes. Here we described the first case of a novel procedure called ischemia-free organ transplantation (IFOT) for patients with end-stage liver disease. The liver graft with severe macrovesicular steatosis was donated from a 25-year-old man. The recipient was a 51-year-old man with decompensated liver cirrhosis and hepatocellular carcinoma. The graft was procured, preserved, and implanted under continuous normothermic machine perfusion. The recipient did not suffer post-reperfusion syndrome or vasoplegia after revascularization of the allograft. The liver function test and histological study revealed minimal hepatocyte, biliary epithelium and vascular endothelium injury during preservation and post-transplantation. The inflammatory cytokine levels were much lower in IFOT than those in conventional procedure. Key pathways involved in IRI were not activated after allograft revascularization. No rejection, or vascular or biliary complications occurred. The patient was discharged on day 18 post-transplantation. This marks the first case of IFOT in humans, offering opportunities to optimize transplant outcomes and maximize donor organ utilization.

Combined liver-kidney perfusion enhances protective effects of normothermic perfusion on liver grafts from donation after cardiac death.

It has been shown that combined liver-kidney normothermic machine perfusion (NMP) is able to better maintain the circuit's biochemical milieu. Nevertheless, whether the combined perfusion is superior to liver perfusion alone in protecting livers from donation after circulatory death (DCD) is unclear. We aimed to test the hypothesis and explored the mechanisms. Livers from 15 DCD pig donors were subjected to either static cold storage (group A), liver-alone NMP (group B), or combined liver-kidney NMP (group C). Livers were preserved for 6 hours and reperfused ex vivo for 2 hours to simulate transplantation or were transplanted in situ. During perfusion, group C showed an improved acid-base and biochemical environment in the circuit over group B. After reperfusion, the architecture of the liver grafts was best preserved in group C, followed by group B, then group A, as shown by the histology and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining of both hepatocytes and biliary epithelium. Ki-67 staining showed substantial hepatocyte proliferation and biliary epithelial regeneration after perfusion in group B and group C. Group C produced more bile in the reperfusion phase than those in group A and group B, with more physiological bile composition and less severe biliary epithelium injury. Von Willebrand factor-positive endothelial cells and E-selectin expression decreased in both group B and group C. Combined liver-kidney NMP not only produced more adenosine triphosphate, protected the nitric oxide signaling pathway, but also diminished oxidative stress (high mobility group box-1 protein and 8-hydroxy-2-deoxy guanosine levels) and inflammatory cytokine (IL6 and IL8) release when compared with liver-alone NMP and CS. In addition, the 7-day survival rate of liver transplant recipients was higher in group C than that in groups A and B. In conclusion, combined liver-kidney NMP can better protect DCD livers from warm ischemia and reperfusion injury probably by maintaining the stability of the internal environment and by abolishing oxidative stress injury. Liver Transplantation 24 67-79 2018 AASLD.

Severe sclerosing cholangitis after Langerhans cell histiocytosis treated by liver transplantation: An adult case report.

BACKGROUND: Langerhans cell histiocytosis (LCH) is a rare hematopoietic disorder of unknown pathogenesis. LCH diseases may occur in a single organ or multisystem organ. The patients with multisystem involvement usually have a poor prognosis. Liver involvement in multisystem LCH results in severe complications, such as obvious sclerosing cholangitis (SC) with jaundice. METHODS: We reported a 31-year-old man developed severe SC due to multisystem LCH and was successfully treated by liver transplantation (LT). In addition, we firstly used tacrolimus and mycofenolate mofetil as immunosuppressants to treat LCH after LT. RESULTS: We performed the immunosuppressants to deal with the LCH after LT, now the patient is currently well with normal liver function and no evidence of recurrence of LCH for 4 and a half years follow-up. CONCLUSION: LT should be recommended as an effective treatment for these adults with severe SC due to multisystem LCH. Finally, using tacrolimus and mycofenolate mofetil as immunosuppressants to treat LCH might be favorable to prevent LCH recurrence.

Transperitoneal laparoscopic repair of vesicovaginal fistula for patients with supratrigonal fistula: comparison with open transperitoneal technique.

INTRODUCTION AND HYPOTHESIS: We investigated the treatment outcomes of laparoscopic vesicovaginal fistula repair (LVVFR) in patients with supratrigonal vesicovaginal fistula (VVF) in contrast with open transperitoneal vesicovaginal fistula repair (OVVFR). METHODS: We analyzed 58 VVF repairs from June 2005 to July 2014, with 22 patients in the LVVFR group and 36 in the OVVFR group. Demographic parameters, operative variables, and perioperative outcomes were retrospectively collected and analyzed. The chi-square test, Fisher's exact test, Student's t test and the Mann-Whitney U test were used for statistical analysis. RESULTS: Patients in both groups had comparable preoperative characteristics. Significantly shorter hospital stay (5.6 vs. 13.2 days, p < 0.05) and less blood loss (52 vs. 103 ml, p < 0.05) were observed in LVVFR group. Patients in the LVVFR group achieved a higher overall VVF success rate (95.5 % vs. 83.3 %, p > 0.05) and recurrent VVF success rate (90.0 % vs. 75.0 %, p > 0.05) than OVVFR group, but it was not statistically significant. Patients who underwent OVVFR experienced more postoperative symptomatic bladder spasms (8.3 % vs. 4.5 %, p > 0.05), urinary tract infections (UTIs) (5.6 % vs. 0.0 %, p > 0.05), and stress urinary incontinence (SUI) (5.6 % vs. 4.5 %, p > 0.05), but fewer incidents of postoperative ileus (0.0 % vs. 4.5 %, p > 0.05) than the LVVFR group; differences were not significant. CONCLUSIONS: Judging from this initial trial, LVVFR should be recommended as the primary intervention to treat supratrigonal VVF patients in view of its reduced blood loss and hospital stay.

CCR7 as a predictive biomarker associated with computed tomography for the diagnosis of lymph node metastasis in bladder carcinoma.

The aim of the present study was to investigate whether the expression levels of CC-chemokine receptor 7 (CCR7) combined with computed tomography (CT) was associated with lymph node metastasis in bladder transitional cell carcinoma (BTCC). For this purpose, 115 cases of BTCC were analyzed at the Department of Urology of Xiangya Hospital, Central South University (Changsha, China). Preoperative CT scans of abdomen and pelvis, immunohistochemistry of CCR7 expression in the tumor specimens and pathological findings for lymph node metastasis were assessed. In addition, the sensitivity, specificity and accuracy of CCR7 and CT for the diagnosis of lymph node metastasis in BTCC were evaluated separately and jointly. The expression levels of CCR7 were observed to be significantly higher in BTCC than in normal controls (P<0.01). Multivariate analysis indicated that the overexpression of CCR7 was an independent predictor for lymph node metastasis in BTCC (P<0.05). The sensitivity, specificity and accuracy of CCR7 combined with CT scan for the diagnosis of lymph node metastasis in BTCC were 92.3, 83.6 and 70.0%, respectively. By contrast, the sensitivity, specificity and accuracy of CCR7 alone were 88.1, 69.9 and 76.5%, respectively, while the sensitivity, specificity and accuracy of CT alone were 52.4, 79.5 and 69.6%, respectively. The results of the present study indicated that CCR7 is an independent predictor of lymph node metastasis in BTCC. Therefore, the use of CCR7 combined with CT may improve the accuracy of the diagnosis of lymph node metastasis in BTCC.

Maspin enhances cisplatin chemosensitivity in bladder cancer T24 and 5637 cells and correlates with prognosis of muscle-invasive bladder cancer patients receiving cisplatin based neoadjuvant chemotherapy.

BACKGROUND: Maspin, a non-inhibitory member of the serine protease inhibitor superfamily, has been characterized as a tumor suppressor gene in multiple cancer types. Chemotherapeutic insensitivity is one of major obstacles to effectively treating muscle invasive bladder cancer (MIBC). This study was conducted to investigate the role and probable mechanism of Maspin enhancing cisplatin chemosensitivity of bladder cancer in vitro and MIBC patients. METHODS: Maspin expression was quantified by qRT-PCR in two MIBC cell lines (T24 and 5637). After successful established Maspin overexpression model by lipidosome transfection, MTT and cell apoptosis assay were used to assess the MIBC's cisplatin sensitivity. Western blot method was used to test PI3K/ AKT/mTOR signal passway and apoptosis related molecules Caspase3 and Bcl-2. Additionally, we evaluated Maspin expression and prognosis in 62 MIBC cases who underwent cisplatin based neoadjuvant chemotherapy (NACT) using immunohistochemistry. RESULT: Upregulate Maspin expression could enhance the chemosensitivity induced by cisplatin in T24 and 5637 cell lines. The cell viability, cloning ability and IC50 were reduced while apoptosis rate was upregulated when cells were transfected Maspin. Phospho(p)-AKT, PI3K, mTOR, and Bcl-2 expression were significantly decreased, whereas Caspase3 was greatly increased in the Maspin group. In the clinic study, there was significant correlation between Maspin expression and overall survival (OS) and progression-free survival (PFS) rate in MIBC patients who received cisplatin based NACT. CONCLUSION: Maspin could enhance cisplatin chemosensitivity in T24 and 5637 cell lines. Its expression correlated with prognosis of MIBC patients who received cisplatin based neoadjuvant chemotherapy.