METTL3/16-mediated m6A modification of ZNNT1 promotes hepatocellular carcinoma progression by activating ZNNT1/osteopontin/S100A9 positive feedback loop-mediated crosstalk between macrophages and tumour cells.

Macrophages are the major components of tumour microenvironment, which play critical roles in tumour development. N6-methyladenosine (m6A) also contributes to tumour progression. However, the potential roles of m6A in modulating macrophages in hepatocellular carcinoma (HCC) are poorly understood. Here, we identified ZNNT1 as an HCC-related m6A modification target, which was upregulated and associated with poor prognosis of HCC. METTL3 and METTL16-mediated m6A modification contributed to ZNNT1 upregulation through stabilizing ZNNT1 transcript. ZNNT1 exerted oncogenic roles in HCC. Furthermore, ZNNT1 recruited and induced M2 polarization of macrophages via up-regulating osteopontin (OPN) expression and secretion. M2 Macrophages-recruited by ZNNT1-overexpressed HCC cells secreted S100A9, which further upregulated ZNNT1 expression in HCC cells via AGER/NF-κB signaling. Thus, this study demonstrates that m6A modification activated the ZNNT1/OPN/S100A9 positive feedback loop, which promoted macrophages recruitment and M2 polarization, and enhanced malignant features of HCC cells. m6A modification-triggered ZNNT1/OPN/S100A9 feedback loop represents potential therapeutic target for HCC.

A randomized-controlled trial of ischemia-free liver transplantation for end-stage liver disease.

BACKGROUND & AIMS: Ischemia-reperfusion injury (IRI) has thus far been considered as an inevitable component of organ transplantation, compromising outcomes, and limiting organ availability. Ischemia-free organ transplantation is a novel approach designed to avoid IRI, with the potential to improve outcomes. METHODS: In this randomized-controlled clinical trial, recipients of livers from donors after brain death were randomly assigned to receive either an ischemia-free or a 'conventional' transplant. The primary endpoint was the incidence of early allograft dysfunction. Secondary endpoints included complications related to graft IRI. RESULTS: Out of 68 randomized patients, 65 underwent transplants and were included in the analysis. 32 patients received ischemia-free liver transplantation (IFLT), and 33 received conventional liver transplantation (CLT). Early allograft dysfunction occurred in two recipients (6%) randomized to IFLT and in eight (24%) randomized to CLT (difference -18%; 95% CI -35% to -1%; p = 0.044). Post-reperfusion syndrome occurred in three recipients (9%) randomized to IFLT and in 21 (64%) randomized to CLT (difference -54%; 95% CI -74% to -35%; p <0.001). Non-anastomotic biliary strictures diagnosed with protocol magnetic resonance cholangiopancreatography at 12 months were observed in two recipients (8%) randomized to IFLT and in nine (36%) randomized to CLT (difference, -28%; 95% CI -50% to -7%; p = 0.014). The comprehensive complication index at 1 year after transplantation was 30.48 (95% CI 23.25-37.71) in the IFLT group vs. 42.14 (95% CI 35.01-49.26) in the CLT group (difference -11.66; 95% CI -21.81 to -1.51; p = 0.025). CONCLUSIONS: Among patients with end-stage liver disease, IFLT significantly reduced complications related to IRI compared to a conventional approach. CLINICAL TRIAL REGISTRATION: chictr.org. ChiCTR1900021158. IMPACT AND IMPLICATIONS: Ischemia-reperfusion injury has thus far been considered as an inevitable event in organ transplantation, compromising outcomes and limiting organ availability. Ischemia-free liver transplantation is a novel approach of transplanting donor livers without interruption of blood supply. We showed that in patients with end-stage liver disease, ischemia-free liver transplantation, compared with a conventional approach, led to reduced complications related to ischemia-reperfusion injury in this randomized trial. This new approach is expected to change the current practice in organ transplantation, improving transplant outcomes, increasing organ utilization, while providing a clinical model to delineate the impact of organ injury on alloimmunity.

Association of Perfusion Characteristics and Posttransplant Liver Function in Ischemia-Free Liver Transplantation.

It has been shown that normothermic machine perfusion (NMP), a novel preservation method, is able to assess and resuscitate liver grafts with risk factors. However, there is no consistent criteria for the assessment of liver grafts with NMP. Ischemia-free liver transplantation (IFLT) includes innovative surgical techniques and NMP, which can protect liver grafts from ischemia throughout organ procurement, preservation, and implantation. In our center, 28 human livers from donation after brain death donors were subjected to IFLT between July 2017 and October 2018. The correlation between posttransplant liver function tests with the perfusion parameters, blood gas analysis of perfusate, and bile biochemistry were analyzed. During the preservation phase, the vascular flow was stable, and the lactate level decreased rapidly. The transaminase release in the perfusate was low but stable, whereas the glucose level remained high. The perfusate lactate and aspartate aminotransferase (AST) levels at 1 hour of perfusion were correlated with the posttransplant peak AST level. There were negative correlations between the portal vein and hepatic artery flows at the end of perfusion and the peak transaminase levels within 7 days after transplantation. In conclusion, during IFLT, NMP is able to bridge the liver grafts from donors to recipients and can allow the assessment of liver function by perfusion characteristics.

Systematic Evaluation of Research Progress on Natural Language Processing in Medicine Over the Past 20 Years: Bibliometric Study on PubMed.

BACKGROUND: Natural language processing (NLP) is an important traditional field in computer science, but its application in medical research has faced many challenges. With the extensive digitalization of medical information globally and increasing importance of understanding and mining big data in the medical field, NLP is becoming more crucial. OBJECTIVE: The goal of the research was to perform a systematic review on the use of NLP in medical research with the aim of understanding the global progress on NLP research outcomes, content, methods, and study groups involved. METHODS: A systematic review was conducted using the PubMed database as a search platform. All published studies on the application of NLP in medicine (except biomedicine) during the 20 years between 1999 and 2018 were retrieved. The data obtained from these published studies were cleaned and structured. Excel (Microsoft Corp) and VOSviewer (Nees Jan van Eck and Ludo Waltman) were used to perform bibliometric analysis of publication trends, author orders, countries, institutions, collaboration relationships, research hot spots, diseases studied, and research methods. RESULTS: A total of 3498 articles were obtained during initial screening, and 2336 articles were found to meet the study criteria after manual screening. The number of publications increased every year, with a significant growth after 2012 (number of publications ranged from 148 to a maximum of 302 annually). The United States has occupied the leading position since the inception of the field, with the largest number of articles published. The United States contributed to 63.01% (1472/2336) of all publications, followed by France (5.44%, 127/2336) and the United Kingdom (3.51%, 82/2336). The author with the largest number of articles published was Hongfang Liu (70), while Stéphane Meystre (17) and Hua Xu (33) published the largest number of articles as the first and corresponding authors. Among the first author's affiliation institution, Columbia University published the largest number of articles, accounting for 4.54% (106/2336) of the total. Specifically, approximately one-fifth (17.68%, 413/2336) of the articles involved research on specific diseases, and the subject areas primarily focused on mental illness (16.46%, 68/413), breast cancer (5.81%, 24/413), and pneumonia (4.12%, 17/413). CONCLUSIONS: NLP is in a period of robust development in the medical field, with an average of approximately 100 publications annually. Electronic medical records were the most used research materials, but social media such as Twitter have become important research materials since 2015. Cancer (24.94%, 103/413) was the most common subject area in NLP-assisted medical research on diseases, with breast cancers (23.30%, 24/103) and lung cancers (14.56%, 15/103) accounting for the highest proportions of studies. Columbia University and the talents trained therein were the most active and prolific research forces on NLP in the medical field.

A simplified multivisceral transplantation procedure for patients with combined end-stage liver disease and type 2 diabetes mellitus.

In liver transplant patients with type 2 diabetes mellitus (DM), the disease worsens after transplantation because of longterm use of diabetogenic immunosuppressive drugs, making management of those patients a great challenge. The objective of our study was to evaluate the safety and efficacy of a simplified multivisceral transplantation (SMT) procedure for the treatment of patients with end-stage liver disease and concurrent type 2 DM. Forty-four patients who had pretransplant type 2 DM were included. A total of 23 patients received SMT, and 21 patients received orthotopic liver transplantation (OLT). Patient and graft survivals, complications, diabetic control, and quality of life (QOL) were retrospectively analyzed in both groups. The 1-, 3-, and 5-year cumulative patient and graft survival rates were 91.5%, 75.4%, and 75.4% in the SMT group and were 94.4%, 64.4%, and 64.4% in the OLT group, respectively (P = 0.70). Interestingly, 95.7% (22/23) of patients achieved complete remission from DM after SMT compared with 16.7% (3/18) of patients after OLT. The occurrence of biliary complication was significantly higher in the OLT group than that in the SMT group (23.8% versus 0.0%; P = 0.01). Moreover, better QOL was observed in the SMT group than that in the OLT group. In conclusion, the SMT procedure we described here is a safe and viable option for patients with end-stage live disease and concurrent type 2 DM. This SMT procedure offers excellent transplant outcomes and QOL. Liver Transplantation 23 1161-1170 2017 AASLD.

Immune roles of dendritic cells in stem cell transplantation.

Dendritic cells (DCs) are professional antigen-presenting cells and initial stimulators for immune response. DCs can shape their functions based on their immune states, which are crucial for the balance of immunity and tolerance to preserve homeostasis. In the immune response involved in stem cell transplantation, DCs also play important roles in inducing immune tolerance and antitumor immunity. After the rapid development of stem cell transplantation technology in recent years, the risks of graft rejection, tumor recurrence, and tumorigenicity are still present after stem cell transplantation. It is important to understand the mechanisms of DC-mediated immune tolerance and stimulation during stem cell transplantation. In this review, we will summarize and analyze the regulatory mechanisms of DCs in stem cell transplantation and their application in clinical settings. It may help to promote the innovation in basic theories and therapeutic approaches of stem cell transplantation.

[Survival outcome comparative analysis of Liver transplantation and radical resection surgery for Klatskin tumor].

OBJECTIVE: To compare the survival results of liver transplantation with radical resection for klatskin tumor based on the published literatures. METHODS: We retrieved Medline for articles on treatment of klatskin tumor published from January 1996 to December 2014, and choose the literatures including liver transplantation and radical resection. Review Manager 5. 0 software was used to perform the Meta-analysis of the extracted data. RESULTS: After performing meta-analysis of 11 studies including 676 patients, we found comparable patient survival rate between liver transplantation group and radical resection group (1-year RR:1. 00, 95% CI:0. 82 - 1. 23, P = 0. 97; 3-year RR:1. 15, 95% CI:0. 75 - 1. 77, P 0. 51, 5-year RR: 1. 37, 95% CI: 0. 74 - 2. 53, P = 0. 32, respectively). No statistical significance of overall survival rate were shown between this 2 groups whether in short-term or long-time, but the total survival trends show the long-time survival of LT group is better than that of RR group which has the potential longer survival time. CONCLUSIONS: Liver transplantation was equivalent to radical resection for klatskin tumor treatment in terms of patient survival, with potential higher long-term patient survival rate and lower recurrence. especially for the patients with unrespectable KT or insufficient liver function, LT is superior for clinical outcome.

Antiapoptotic gene BAG-1 vector structure of RNA interference and endogenous targeted screening in colon cancer cell lines.

The purposes of the present work were to construct the shRNA plasmids for BAG-1 gene of human and test the expression of mRNA and protein of BAG-1. Recombinant plasmids containing green fluorescent protein reporter genes are constructed using gene cloning methods. The shRNA plasmids for the BAG-1 gene are constructed by RNA interference technology. We applied fluorescent plasmid-transfected target cells in the cell transfection experiments and monitored the transfection rate of plasmids by observing the fluorescence amount. We transfected three synthesized shRNA in target screening cell and adopted RT-PCR and Western test to identify the difference of target gene transfection and translation level in cells. The specific shRNA plasmid for the BAG-1 gene was successfully recombined, and stably transfected colon cancer Lo Vo cell lines were obtained. The results present that the constructed shRNA plasmids significantly inhibited the expression of mRNA and protein of Lo Vo cell BAG-1, and can maintain the effect for a long term. pGPH1/GFP/Neo-BAG-1-homo-825 was screened as the optimum sequence of interference so as to lay a solid foundation to explore into the research on the BAG-1 gene and the biological behavior of colon cancer cells. It showed the remarkable advantage of RNAi in the generation of posttranscriptional gene silencing.

Radiofrequency-assisted versus clamp-crush liver resection: a systematic review and meta-analysis.

BACKGROUND: Conflicting results were found between radiofrequency-assisted liver resection (RF-LR) and clamp-crush liver resection (CC-LR) during liver surgery. We conducted a systematic review and meta-analysis that included randomized controlled trials (RCTs) and non-RCTs to compare the effectiveness and safety of RF-LR versus CC-LR during liver surgery. METHODS: Articles comparing RF-LR and CC-LR that were published before December 2012 were retrieved and subjected to a systematic review and meta-analysis. Data synthesis and statistical analysis were carried out by Review Manager Version 5.2 software. RESULTS: In all, four RCTs and five nonrandomized studies evaluating 728 patients were included. Compared with CC-LR, the RF-LR group had significantly reduced total intraoperative blood loss (weighted mean difference [WMD] = -187 mL; 95% confidence interval [CI] = -312, -62; data on 628 patients), and blood loss during liver transection (WMD = -143.7 mL; 95% CI = -200, -87; data on 190 patients). However, RF-LR is associated with a higher rate of intra-abdominal abscess than the clamp-crushing method (odds ratio = 3.61; 95% CI = 1.26, 10.32; data on 366 patients). No significant difference was observed between both the groups for the incidence of both blood transfusion and bile leak. CONCLUSIONS: There is currently not sufficient evidence to support or refute the use of RF-LR in liver surgery. RF-LR has advantages in terms of reducing blood loss. However, RF-LR may increase the rates of both bile leak and abdominal abscess. So, the safety of RF-LR has not been established. Future well-designed RCTs are awaited to further investigate the efficacy and safety of RF devices in liver resection.

Liver transplantation using organs from deceased organ donors: a single organ transplant center experience.

BACKGROUND: In 2011, a pilot program for deceased organ donation was initiated in China. We describe the first successful series of liver transplants in the pilot program. METHODS: From July 2011 to August 2012, our center performed 26 liver transplants from a pool of 29 deceased donors. All organ donation and allograft procurement were conducted according to the national protocol. The clinical data of donors and recipients were collected and summarized retrospectively. RESULTS: Among the 29 donors, 24 were China Category II donors (organ donation after cardiac death), and five were China Category III donors (organ donation after brain death followed by cardiac death). The recipients were mainly the patients with hepatocellular carcinoma. The one-year patient survival rate was 80.8% with a median follow-up of 422 (2-696) days. Among the five mortalities during the follow-up, three died of tumor recurrence. In terms of post-transplant complications, 9 recipients (34.6%) experienced early allograft dysfunction, 1 (3.8%) had non-anastomotic biliary stricture, and 1 (3.8%) was complicated with hepatic arterial thrombosis. None of these complications resulted in patient death. Notably, primary non-function was not observed in any of the grafts. CONCLUSION: With careful donor selection, liver transplant from deceased donors can be performed safely and plays a critical role in overcoming the extreme organ shortage in China.

Atomic force microscopy of chronic lymphatic leukaemia cells activation induced by Staphylococcus aureus.

Activation of lymphatic cells is associated with changes in morphology, ultrastructure and adhesion force. We have investigated the activation efficiency of Staphylococcus aureus (SAC) on B-cell chronic lymphatic leukaemia (B-CLL) cells using atomic force microscopy (AFM), and found changes in the above properties. Cell viability and proliferation were measured using Cell Counting Kit-8 (CCK-8) and enzyme-linked immunosorbent assay (ELISA). AFM clearly showed that the volume and nuclear-cytoplasm ratio of cells increased significantly with activated time. It also showed that pseudopodia and immunological synapses began to appear at 24 h. In the activation process, nano-structures of the cell surface became aggregated, and adhesion increased. In conclusion, the results indicate a close relationship between membrane reconstruction and multiplication process of B-CLL cells.

Ultrasound-Guided Human Islet Transplantation: Safety, Feasibility, and Efficacy Analysis.

RATIONALE AND OBJECTIVES: We aimed to analyze the safety, feasibility, and efficacy of human islet transplantation (IT) using ultrasound (US) throughout the entire procedure. MATERIALS AND METHODS: A total of 22 recipients (18 males; mean age 42.6 ± 17.5years) with 35 procedures were retrospective included. Under US guidance, percutaneous transhepatic portal catheterization was successfully performed through a right-sided transhepatic approach, and islets were infused into the main portal vein. Color Doppler and contrast-enhanced ultrasound were used to guide the procedure and monitor the complications. After infusion of the islet mass, the access track was embolized by embolic material. If hemorrhage persisted, US-guided radiofrequency ablation (RFA) was performed to stop bleeding. Factors that could affect the complication were analyzed. After transplantation, primary graft function was evaluated with a ß-score 1month after the last islet infusion. RESULTS: The technical success rates were 100% with a single puncture attempt. Six (17.1%) abdominal bleeding episodes were immediately stopped by US-guided RFA. No portal vein thrombosis were encountered. Dialysis (OR (Odd Ratio): 32.0; 95% CI: 1.561-656.054; and P = .025) was identified as a significant factor associated with bleeding. Primary graft function was optimal in eight patients (36.4%), suboptimal in 13 patients (59.1%), and poor in one patient (4.5%). CONCLUSION: In conclusion, whole-procedure US-guided IT is a safe, feasible, and effective method for diabetes. Complications are either self-limiting or manageable with noninvasive treatment.

Living donor vs. deceased donor liver transplantation for patients with hepatitis C virus-related diseases.

BACKGROUND & AIMS: Living donor liver transplantation (LDLT) provides a timely alternative to deceased donor liver transplantation (DDLT) for patients with hepatitis C virus-related (HCV-related) diseases in the circumstances of severe organ dearth. However, the patient and graft outcomes, and recurrence of HCV after LDLT remain controversial. Here we sought to compare the post-transplant outcomes after LDLT and DDLT. METHODS: A systematic review and meta-analysis were performed. PubMed/MEDLINE, EMBASE, and the Cochrane database were searched for eligible literatures. The major end points were patient survival, graft survival, recurrence rate, and acute rejection. The pooled odds ratio (OR) was calculated using random-effects model to synthesize the results. Heterogeneity and publication bias were quantitatively evaluated. RESULTS: Fourteen studies with a total of 2024 participants were included in this analysis. We found comparable patient survival between groups (1-year: OR, 0.78, 95% CI, 0.48-1.26, p=0.31; 2-year: OR, 0.71, 95% CI, 0.41-1.23, p=0.23; 3-year: OR, 0.79, 95% CI, 0.5-1.12, p=0.18; 4-year: OR, 0.92, 95% CI, 0.43-1.95, p=0.83; 5-year: OR, 1.06, 95% CI, 0.53-2.14, p=0.86, respectively). Although 1- and 3-year graft survivals were inferior in LDLT, 2-, 4- and 5-year graft survivals were similar. HCV recurrence rates and acute rejection rates were equivalent. CONCLUSIONS: LDLT was equivalent to DDLT in terms of patient survival, long-term graft survival, HCV recurrence, and acute rejection rates, with potentially lower short-term patient and graft survival.

Steroid elimination within 24 hours after orthotopic liver transplantation: effectiveness and tolerability.

BACKGROUND: Steroids have been the mainstay of immunosuppressive regimen in liver transplantation. However, the use of steroids is associated with various post-transplant complications. This study evaluated the efficacy and safety of reduced immunosuppressive regimen with steroids (steroid elimination within 24 hours post-transplant) in a cohort of Chinese liver transplant recipients. METHODS: Seventy-six patients in line with the selection criteria were enrolled in this prospective study. All patients received anti-IL-2 receptor antibody induction and tacrolimus-based maintenance therapy. The recipients were divided into two groups according to the duration of steroid use: 40 transplant in a 3-month withdrawal group and the remaining 36 in a 24-hour elimination group. Recipient survival, post-operative infections, biopsy-proven acute rejection and steroid-resistant acute rejection, non-healing wound, recurrence of hepatitis B virus (HBV) and hepatocellular carcinoma (HCC), de novo diabetes, hyperlipidemia and hypertension were assessed in the two groups. RESULTS: There was no significant difference in patient survival, incidence of acute rejection episodes and hyperlipidemia, and recurrence of HBV and HCC between the two groups. However, the incidence rates of post-transplant infection, non-healing wound, de novo diabetes and hypertension were significantly lower in the 24-hour elimination group than in the 3-month withdrawal group (all P values <0.05). CONCLUSION: Under anti-IL-2 receptor antibody induction and tacrolimus-based maintainance, steroid elimination within 24 hours post-transplant is associated with reduced steroid-related complications without increasing the risk of rejection.

[Roles of E-cadherin in enhancing the in vitro differentiation of hepatocytes from murine embryonic stem cells].

OBJECTIVE: To investigate the roles of E-cadherin (E-cad) in enhancing the in vitro differentiation of hepatocytes from murine embryonic stem cells (ESCs). METHODS: Exogenous E-cad was transfected into BALB/c murine ESCs to enable its stable expression. Then hepatic differentiation from E-cad-ESCs was induced by such growth factors as hepatocyte growth factor (HGF), fibroblast growth factor (FGF) and transforming growth factor (TGF). And the expressions of hepatic markers ALB, TAT, Cyp7a1 and urea were detected. The morphology of hepatic differentiation was observed under microscopy. RESULTS: E-cad expression gradually decreased in normal ESC differentiation, but was stably expressed in E-cad-ESCs. In E-cad-ESC group, hepatic markers ALB, TAT and CYP7a1 were expressed earlier or higher than that in normal ESC group, and the concentrations of ALB and urea were significantly higher than that in normal ESC group. The adhesion of the differentiated E-cad-ESCs was significantly enhanced compared with the normal ESCs. They maintained close connections and multidimensional growth. Cell number of hepatocytes from ESC increased significantly in E-cad-ESC group. CONCLUSION: E-cad enhances the hepatic differentiation of ESC by increasing the number of differentiated cells and increasing the synthetic capacity of ALB and urea.

[Efficacy and safety of sorafenib in the prevention and treatment of hepatocellular carcinoma recurrences after liver transplantation].

OBJECTIVE: To evaluate the efficacy and safety of sorafenib in the prevention and treatment of hepatocellular carcinoma (HCC) relapse after liver transplantation. METHODS: A retrospective cohort study was performed to assess the efficacy and safety of sorafenib for HCC. Forty-four patients who underwent liver transplant for HCC beyond Milan criteria form July 2007 to May 2010 were included study group (sorafenib, n = 22) and control group (without sorafenib, n = 22). The primary endpoints of the study were disease-free survival (DFS), overall survival (OS). Secondary outcomes included the rates of acute rejection and graft survival. RESULTS: The clinical data of 44 patients were completely collected. There were significantly differences between sorafenib group and control group in 1-year DFS (81.8% (n = 18) vs 63.6% (n = 14), P < 0.05) and OS (90.9% (n = 20) vs 72.7% (n = 16), P < 0.05) respectively. The acute rejection rates in Sorafenib were 13.6% (3/22), compared with 18.2% (4/22) in control group (P = 0.524) and 1-year graft survival in Sorafenib group were 86.4% (19/22), compared with 72.7% (16/22) in control group (P = 0.086). The overall incidence of treatment-related adverse events was 68.1% (n = 15) in sorafenib group and 31.8% (n = 7) in the control group (P < 0.01). Adverse events that were reported for patients receiving sorafenib were predominantly grade 1 or 2 in severity including diarrhea (45.5%, n = 10), liver dysfunction (40.9%, n = 9), hand-foot skin reaction (31.8%, n = 7) and pains of head and four limbs (22.7%, n = 5). Two patients with grade 3 adverse events in study group were stopped continuing to use the sorafenib. Three patients with the dose of 400 mg twice daily and 17 patients with the dose reduction of sorafenib continued to the study endpoint. CONCLUSION: Patients with HCC undergoing liver transplantation could get the benefits of Sorafenib in reducing the incidence of tumor recurrence and extending disease-free and overall survival time.

[Clinical characteristics, diagnosis and treatment of digestive tract leakage after orthotopic liver transplantation].

OBJECTIVE: To investigate the clinical characteristics, diagnosis and treatment of digestive tract leakage after orthotopic liver transplantation (OLT). METHODS: Sixty-one recipients had digestive tract leakage in early stage after OLT among 1173 cases from January 2000 to December 2010. There were 55 male and 6 female patients, aging from 36 to 61 years, with a median of 45 years. Digestive tract leakage included bile leakage (46 cases), gastric leakage (5 cases), duodenal leakage (1 case), jejunal leakage (4 cases), ileal leakage (1 case) and colon transversum leakage (4 cases). Ten of recipients with gastrointestinal leakage had 1 to 3 times of abdominal surgery before OLT. Abdominal drainage was used in 28 cases with bile leakage, and additionally, endoscopic retrograde cholangiopancreatography, endoscopic nasobiliary drainage and stenting were performed for 8 of them, and surgical neoplasty for another 18 patients with bile leakage. Simple surgical neoplasty of perforation was performed for 13 patients with gastrointestinal leakage, and diverticulectomy and neoplasty for 1 case with duodenal leakage, and partial jejunostomy for one severe jejunal leakage. Nutritional support was administered for all of cases. RESULTS: The incidence rate of digestive tract leakage in early stage after OLT was 5.20% (61/1173). Intra-operative iatrogenic injury of gastrointestinal tract was occurred in 6 cases with gastrointestinal leakage. After treatment, 11 cases died of multiple organ failure resulted from severe infection, with mortality of 18.0% (11/61), including 4 cases with bile leakage, with the mortality of 8.6% (4/46), and 7 cases with gastrointestinal tract leakage, with the mortality of 46.6% (7/15). The remanent 50 cases through comprehensive treatment with a span of 1 to 3 months recovered and discharged healthily. No digestive tract leakage reoccurred in the follow-up of 6 to 84 months. CONCLUSIONS: The morbidity of digestive tract leakage in early stage after OLT is low, but its mortality is high, especially for gastrointestinal tract leakage. High dose corticosteroids therapy, history of abdominal operation and intra-operative iatrogenic injury may be high risk factor. Comprehensive treatment is crucial for improving prognosis.

Generation of a human induced pluripotent stem cell line (SYSUTFi001-A) from infiltrating cytotoxic T cells in hepatocellular carcinoma (HCC).

In this study, we report a novel induced pluripotent stem cell (iPSC) line SYSUTFi001-A derived from cytotoxic T cells (CTLs) infiltrating in hepatocellular carcinoma (HCC), using an integrative Sendai virus vector. This pluripotent cell line shows a normal karyotype and can be redifferentiated to the rejuvenated CTLs targeted to HCC. The cell line SYSUTFi001-A can be further used to perform vitro and vivo anti-tumor assays and design future cell replacement therapies.

Integrative network analysis of circular RNAs reveals regulatory mechanisms for hepatic specification of human iPSC-derived endoderm.

BACKGROUND: Human-induced pluripotent stem cell (hiPSC)-derived functional hepatic endoderm (HE) is supposed to be an alternative option for replacement therapy for end-stage liver disease. However, the high heterogeneity of HE cell populations is still challenging. Hepatic specification of definitive endoderm (DE) is an essential stage for HE induction in vitro. Recent studies have suggested that circular RNAs (circRNAs) determine the fate of stem cells by acting as competing endogenous RNAs (ceRNAs). To date, the relationships between endogenous circRNAs and hepatic specification remain elusive. METHODS: The identities of DE and HE derived from hiPSCs were determined by qPCR, cell immunofluorescence, and ELISA. Differentially expressed circRNAs (DEcircRNAs) were analysed using the Arraystar Human circRNA Array. qPCR was performed to validate the candidate DEcircRNAs. Intersecting differentially expressed genes (DEGs) of the GSE128060 and GSE66282 data sets and the DEcircRNA-predicted mRNAs were imported into Cytoscape for ceRNA networks. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were involved in the enrichment analysis. Hepatic markers and Wnt/ß-catenin were detected in hsa_circ_004658-overexpressing cells by western blotting. Dual-luciferase reporter assays were used to evaluate the direct binding among hsa_circ_004658, miRNA-1200 and CDX2. DE cells were transfected with miR-1200 mimics, adenovirus containing CDX2, and Wnt/ß-catenin was detected by western blotting. RESULTS: hiPSC-derived DE and HE were obtained at 4 and 9 days after differentiation, as determined by hepatic markers. During hepatic specification, 626 upregulated and 208 downregulated DEcircRNAs were identified. Nine candidate DEcircRNAs were validated by qPCR. In the ceRNA networks, 111 circRNA-miRNA-mRNA pairs were involved, including 90 pairs associated with hsa_circ_004658. In addition, 53 DEGs were identified among the intersecting mRNAs of the GSE128060 and GSE66282 data sets and the hsa_circ_004658-targeted mRNAs. KEGG and GO analyses showed that the DEGs associated with hsa_circ_004658 were mainly enriched in the WNT signalling pathway. Furthermore, hsa_circ_004658 was preliminarily verified to promote hepatic specification, as determined by hepatic markers (AFP, ALB, HNF4A, and CK19) (p < 0.05). This promotive effect may be related to the inhibition of the Wnt/ß-catenin signalling pathway (detected by ß-catenin, p-ß-catenin, and TCF4) when hsa_circ_004658 was overexpressed (p < 0.05). Dual-luciferase reporter assays showed that there were binding sites for miR-1200 in the hsa_circ_004658 sequence, and confirmed the candidate DEG (CDX2) as a miR-1200 target. The level of miR-1200 decreased and the level of CDX2 protein expression increased when hsa_circ_004658 was overexpressed (p < 0.05). In addition, the results showed that CDX2 may suppress the Wnt/ß-catenin signalling during hepatic specification (p < 0.05). CONCLUSIONS: This study analysed the profiles of circRNAs during hepatic specification. We identified the hsa_circ_004658/miR-1200/CDX2 axis and preliminarily verified its effect on the Wnt/ß-catenin signalling pathway during hepatic specification. These results provide novel insight into the molecular mechanisms involved in hepatic specification and could improve liver development in the future.