Indications of pro-inflammatory cytokines in laparoscopic and open liver resection for early-stage hepatocellular carcinoma.

BACKGROUND: Our clinical practice of laparoscopic liver resection (LLR) had achieved better short-term and long-term benefits for patients with hepatocellular carcinoma (HCC) over open liver resection (OLR), but the underlying mechanisms are not clear. This study was to find out whether systemic inflammation plays an important role. METHODS: A total of 103 patients with early-stage HCC under liver resection were enrolled (LLR group, n = 53; OLR group, n = 50). The expression of 9 inflammatory cytokines in patients at preoperation, postoperative day 1 (POD1) and POD7 was quantified by Luminex Multiplex assay. The relationships of the cytokines and the postoperative outcomes were compared between LLR and OLR. RESULTS: Seven of the circulating cytokines were found to be significantly upregulated on POD1 after LLR or OLR compared to their preoperative levels. Compared to OLR, the POD1 levels of granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin-6 (IL-6), IL-8, and monocyte chemoattractant protein-1 (MCP-1) in the LLR group were significantly lower. Higher POD1 levels of these cytokines were significantly correlated with longer operative time and higher volume of blood loss during operation. The levels of these cytokines were positively associated with postoperative liver injury, and the length of hospital stay. Importantly, a high level of IL-6 at POD1 was a risk factor for HCC recurrence and poor disease-free survival after liver resection. CONCLUSIONS: Significantly lower level of GM-CSF, IL-6, IL-8, and MCP-1 after liver resection represented a milder systemic inflammation which might be an important mechanism to offer better short-term and long-term outcomes in LLR over OLR.

The Landscape of Aberrant Alternative Splicing Events in Steatotic Liver Graft Post Transplantation via Transcriptome-Wide Analysis.

The application of steatotic liver graft has been increased significantly due to the severe donor shortage and prevalence of non-alcoholic fatty liver disease. However, steatotic donor livers are vulnerable to acute phase inflammatory injury, which may result in cancer recurrence. Alternative splicing events (ASEs) are critical for diverse transcriptional variants in hepatocellular carcinoma (HCC). Here, we aimed to depict the landscape of ASEs, as well as to identify the differential ASEs in steatotic liver graft and their association with tumor recurrence after transplantation. The overall portrait of intragraft transcripts and ASEs were elucidated through RNA sequencing with the liver graft biopsies from patients and rat transplant models. Various differential ASEs were identified in steatotic liver grafts. CYP2E1, ADH1A, CYP2C8, ADH1C, and HGD, as corresponding genes to the common pathways involved differential ASEs in human and rats, were significantly associated with HCC patients' survival. The differential ASEs related RNA-binding proteins (RBPs) were enriched in metabolic pathways. The altered immune cell distribution, particularly macrophages and neutrophils, were perturbated by differential ASEs. The cancer hallmarks were enriched in steatotic liver grafts and closely associated with differential ASEs. Our work identified the differential ASE network with metabolic RBPs, immune cell distribution, and cancer hallmarks in steatotic liver grafts. We verified the link between steatotic liver graft injury and tumor recurrence at post-transcriptional level, offered new evidence to explore metabolism and immune responses, and provided the potential prognostic and therapeutic markers for tumor recurrence.

Mutational Signature Analysis Reveals Widespread Contribution of Pyrrolizidine Alkaloid Exposure to Human Liver Cancer.

BACKGROUND AND AIMS: Mutational signature analyses are an effective tool in identifying cancer etiology. Humans are frequently exposed to pyrrolizidine alkaloids (PAs), the most common carcinogenic phytotoxins widely distributed in herbal remedies and foods. However, due to the lack of human epidemiological data, PAs are classified as group II hepatocarcinogens by the World Health Organization. This study identified a PA mutational signature as the biomarker to investigate the association of PA exposure with human liver cancer. APPROACH AND RESULTS: Pyrrole-protein adducts (PPAs), the PA exposure biomarker, were measured and found in 32% of surgically resected specimens from 34 patients with liver cancer in Hong Kong. Next, we delineated the mode of mutagenic and tumorigenic actions of retrorsine, a representative PA, in mice and human hepatocytes (HepaRG). Retrorsine induced DNA adduction, DNA damage, and activation of tumorigenic hepatic progenitor cells, which initiated hepatocarcinogenesis. PA mutational signature, as the unique molecular fingerprint of PA-induced mutation, was derived from exome mutations in retrorsine-exposed mice and HepaRG cells. Notably, PA mutational signature was validated in genomes of patients with PPA-positive liver cancer but not patients with PPA-negative liver cancer, confirming the specificity of this biomarker in revealing PA-associated liver cancers. Furthermore, we examined the established PA mutational signature in 1,513 liver cancer genomes and found that PA-associated liver cancers were potentially prevalent in Asia (Mainland China [48%], Hong Kong [44%], Japan [22%], South Korea [6%], Southeast Asia [25%]) but minor in Western countries (North America [3%] and Europe [5%]). CONCLUSIONS: This study provides a clinical indication of PA-associated liver cancer. We discovered an unexpectedly extensive implication of PA exposure in patients with liver cancer, laying the scientific basis for precautionary approaches and prevention of PA-associated human liver cancers.

Long-term outcomes of entecavir monotherapy for chronic hepatitis B after liver transplantation: Results up to 8 years.

Long-term antiviral prophylaxis is required to prevent hepatitis B recurrence for patients with chronic hepatitis B after liver transplantation. We determined the long-term outcome of 265 consecutive chronic hepatitis B liver transplant recipients treated with entecavir monotherapy without hepatitis B immune globulin. Viral serology, viral load, and liver biochemistry were performed at regular intervals during follow-up. The median duration of follow-up was 59 months. The cumulative rates of hepatitis B surface antigen (HBsAg) seroclearance were 90% and 95% at 1 and 5 years, respectively. At 1, 3, 5, and 8 years, 85%, 88%, 87.0%, and 92% were negative for HBsAg, respectively, and 95%, 99%, 100%, and 100% had undetectable hepatitis B virus (HBV) DNA, respectively. Fourteen patients remained persistently positive for HBsAg, all of whom had undetectable HBV DNA. There was no significant difference in liver stiffness for those who remained HBsAg-positive compared to those who achieved HBsAg seroclearance (5.5 versus 5.2 kPa, respectively; P = 0.52). The overall 9-year survival was 85%. There were 37 deaths during the follow-up period, of which none were due to hepatitis B recurrence. CONCLUSION: Long-term entecavir monotherapy is highly effective at preventing HBV reactivation after liver transplantation for chronic hepatitis B, with a durable HBsAg seroclearance rate of 92%, an undetectable HBV DNA rate of 100% at 8 years, and excellent long-term survival of 85% at 9 years. (Hepatology 2017;66:1036-1044).

NLRP3 inflammasome induced liver graft injury through activation of telomere-independent RAP1/KC axis.

Acute-phase inflammation plays a critical role in liver graft injury. Inflammasomes, multi-molecular complexes in the cytoplasm, are responsible for initiating inflammation. Here, we aimed to explore the role of inflammasomes in liver graft injury and further to investigate the regulatory mechanism. In a clinical liver transplant cohort, we found that intragraft expression of nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3) inflammasomes was significantly up-regulated post-transplantation. Importantly, overexpression of NLRP3 was strongly associated with poor liver function characterized by high levels of ALT, AST, and urea, as well as neutrophil infiltration after transplantation. The significant correlation between NLRP3 and IL-1ß mRNA levels led us to focus on one of the associated upstream regulators, telomere-independent repressor activator protein 1 (RAP1), which was further proved to be co-localized with NLRP3 in neutrophils. In the liver of a mouse model (hepatic ischaemia/reperfusion and hepatectomy model) and isolated neutrophils from RAP1-/- mice, the expression levels of NLRP3 and keratinocyte chemoattractant (KC) were significantly down-regulated in contrast to those in wild types. The levels of ALT and AST, as well as the neutrophil infiltration, were also decreased by RAP1 deficiency. In our clinical validation, intragraft KC expression was associated with NLRP3 and co-localized with RAP1 in neutrophils. Furthermore, NLRP3 inflammasomes were up-regulated by recombinant KC in the isolated neutrophils and liver of the mouse model. Our data demonstrated that NLRP3 inflammasomes, activated by the RAP1/KC axis, played a critical role in initiating inflammation during the early stage of liver graft injury. Targeting RAP1/KC/NLRP3 inflammasomes may offer a new therapeutic strategy against liver graft injury. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

CXCL10/CXCR3 signaling mobilized-regulatory T cells promote liver tumor recurrence after transplantation.

BACKGROUND & AIMS: Liver graft injury and tumor recurrence are the major challenges of liver transplantation for the patients with hepatocellular carcinoma (HCC). Here, we aimed to explore the role and mechanism of liver graft injury mobilizing regulatory T cells (Tregs), which lead to late phase tumor recurrence after liver transplantation. METHODS: The correlation among tumor recurrence, liver graft injury and Tregs mobilization were studied in 257 liver transplant recipients with HCC and orthotopic rat liver transplantation models. The direct roles of CXCL10/CXCR3 signaling on Tregs mobilization and tumor recurrence were investigated in CXCL10-/- and CXCR3-/- mice models with hepatic IR injury. RESULTS: Clinically, patients received the graft with graft weight ratio (GWR) <60% had higher HCC recurrence after liver transplantation than the recipients with GWR ⩾60% graft. More circulating Tregs and higher intragraft TLR4/CXCL10/CXCR3 levels were detected in recipients with GWR <60% graft. These results were further validated in rat transplantation model. Foxp3+ cells and expressions of TLR4, CXCL10, TGFß, CTLA-4 and CD274 were increased in rat liver tumor tissues from small-for-size graft group. In mouse model, the mobilization and recruitment of Tregs were decreased in TLR4-/-, CXCL10-/- and CXCR3-/- mice compared to wild-type mice. Moreover, less CXCR3+ Tregs were recruited into liver in CXCL10-/- mice after hepatic IR injury. The knockout of CXCL10 and depletion of Tregs inhibited tumor recurrence after hepatic IR injury. CONCLUSION: CXCL10/CXCR3 signaling upregulated at liver graft injury directly induced the mobilization and intragraft recruitment of Tregs, which further promoted HCC recurrence after transplantation. LAY SUMMARY: There were positive correlation among tumor recurrence, circulating Tregs and liver graft injury after human transplantation for HCC patients. The knockout of CXCL10 decreased hepatic recruitment of CXCR3+ Tregs and late phase tumor recurrence after hepatic IR injury.

Fusion with stem cell makes the hepatocellular carcinoma cells similar to liver tumor-initiating cells.

BACKGROUND: Cell fusion is a fast and highly efficient technique for cells to acquire new properties. The fusion of somatic cells with stem cells can reprogram somatic cells to a pluripotent state. Our research on the fusion of stem cells and cancer cells demonstrates that the fused cells can exhibit stemness and cancer cell-like characteristics. Thus, tumor-initiating cell-like cells are generated. METHODS: We employed laser-induced single-cell fusion technique to fuse the hepatocellular carcinoma cells and human embryonic stem cells (hESC). Real-time RT-PCR, flow cytometry and in vivo tumorigenicity assay were adopted to identify the gene expression difference. RESULTS: We successfully produced a fused cell line that coalesces the gene expression information of hepatocellular carcinoma cells and stem cells. Experimental results showed that the fused cells expressed cancer and stemness markers as well as exhibited increased resistance to drug treatment and enhanced tumorigenesis. CONCLUSIONS: Fusion with stem cells transforms liver cancer cells into tumor initiating-like cells. Results indicate that fusion between cancer cell and stem cell may generate tumor initiating-like cells.

Clinical relevance and therapeutic potential of angiopoietin-like protein 4 in hepatocellular carcinoma.

BACKGROUND: Development of novel adjuvant therapy to eradicate tumor angiogenesis and metastasis is a pressing need for patients with advanced hepatocellular carcinoma (HCC). We aimed to investigate the clinical relevance and therapeutic potential of angiopoietin-like 4 (ANGPTL4) in HCC. METHODS: ANGPTL4 mRNA levels in tumor and non-tumor liver tissues of HCC patients were analyzed to investigate its clinical relevance. The mechanisms of deregulation of ANGPTL4 in HCC were studied by copy number variation (CNV) and CpG methylation analyses. The orthotopic liver tumor nude mice model was applied using a human metastatic cell line. ANGPTL4-overexpressing adenovirus (Ad-ANGPTL4) was injected via portal vein to investigate its anti-tumorigenic and anti-metastatic potentials. RESULTS: HCC tissues expressed significantly lower levels of ANGPTL4 mRNA than non-tumor tissues. The copy number of ANGPTL4 gene in tumor tissues was significantly lower than in non-tumor tissues of HCC patients. Higher frequency of methylation of CpG sites of ANGPTL4 promoter was detected in tumor tissues compared to non-tumor tissues. Downregulation of ANGPTL4 mRNA in HCC was significantly associated with advanced tumor stage, presence of venous infiltration, poor differentiation, higher AFP level, appearance of tumor recurrence, and poor postoperative overall and disease-free survivals of HCC patients. Treatment with Ad-ANGPTL4 significantly inhibited the in vivo tumor growth, invasiveness and metastasis by promoting tumoral apoptosis, inhibiting tumoral angiogenesis and motility, and suppressing tumor-favorable microenvironment. Moreover, administration of recombinant ANGPTL4 protein suppressed the motility of HCC cells and altered the secretion profile of cytokines from macrophages. CONCLUSION: ANGPTL4 is a diagnostic and prognostic biomarker for HCC patients and a potential therapeutic agent to suppress HCC growth, angiogenesis and metastasis.

The inhibition of aldose reductase attenuates hepatic ischemia-reperfusion injury through reducing inflammatory response.

OBJECTIVE: We aim to investigate the role of aldose reductase (AR) in hepatic ischemia-reperfusion injury (IRI) of normal and fatty livers and to explore the underlying mechanisms. BACKGROUND: Hepatic IRI is a typical inflammatory response during liver surgery. It contributes to liver graft failure or nonfunction after transplantation. Increasing evidence implicates that AR plays a key role in a number of inflammatory diseases. However, the role of AR in hepatic IRI is still unknown. METHODS: Intragraft AR expression profile and the association with liver graft injury were investigated in both human and rat liver transplantation using normal or fatty graft. The direct role of AR in hepatic IRI was studied in the AR knockout mice IRI model with or without fatty liver. They were further validated by the simulated IRI in vitro model using fatty LO2 cells with or without AR inhibitor zopolrestat and primary peritoneal macrophages isolated from AR knockout and wild-type mice. Gene expression of inflammatory cytokines/chemokines, the infiltration of macrophages/neutrophils, and NF-κB pathway activation were compared among different groups. RESULTS: AR was overexpressed in liver graft after human and rat liver transplantation and correlated with consequent liver injuries. The knockout of AR significantly attenuated hepatic sinusoidal damage and apoptosis in both normal and fatty livers after IRI. The expression of proinflammatory cytokines/chemokines and neutrophil chemoattractants, infiltration of macrophage and neutrophil, and activation of inflammation-associated NF-κB and JNK pathway were downregulated in AR knockout mice. Furthermore, the inhibition of AR effectively suppressed macrophage migration and decreased lipopolysaccharide (LPS)-induced production of proinflammatory cytokines/chemokines in isolated macrophages. CONCLUSIONS: The deficiency of AR attenuated hepatic IRI in both normal and fatty livers by reducing liver inflammatory responses.

Prognostic MicroRNA Fingerprints Predict Recurrence of Early-Stage Hepatocellular Carcinoma Following Hepatectomy.

Purpose: This study aims to develop liquid biopsy assays for early HCC diagnosis and prognosis. Methods: Twenty-three microRNAs were first consolidated as a panel (HCCseek-23 panel) based on their reported functions in HCC development. Serum samples were collected from 103 early-stage HCC patients before and after hepatectomy. Quantitative PCR and machine learning random forest models were applied to develop diagnostic and prognostic models. Results: For HCC diagnosis, HCCseek-23 panel demonstrated 81% sensitivity and 83% specificity for identifying HCC in the early-stage; it showed 93% sensitivity for identifying alpha-fetoprotein (AFP)-negative HCC. For HCC prognosis, the differential expressions of 8 microRNAs (HCCseek-8 panel: miR-145, miR-148a, miR-150, miR-221, miR-223, miR-23a, miR-374a, and miR-424) were significantly associated with disease-free survival (DFS) (Log-rank test p-value = 0.001). Further model improvement using these HCCseek-8 panel in combination with serum biomarkers (i.e. AFP, ALT, and AST) demonstrated a significant association with DFS (Log-rank p-value = 0.011 and Cox proportional hazards analyses p-value = 0.002). Conclusion: To the best of our knowledge, this is the first report to integrate circulating miRNAs, AST, ALT, AFP, and machine learning for predicting DFS in early HCC patients undergoing hepatectomy. In this setting, HCCSeek-23 panel is a promising circulating microRNA assay for diagnosis, while HCCSeek-8 panel is promising for prognosis to identify early HCC recurrence.

Post-transplant inflammatory cytokine signature adds value for predicting tumor recurrence after liver transplantation for hepatocellular carcinoma.

BACKGROUND: Cytokines are key regulators of post-transplant inflammation responses which reconstitute post-transplant hepatic and systemic environments to influence the likelihood of tumor relapse. This study investigated the prognostic value of post-transplant cytokines on tumor recurrence after liver transplantation (LT) for hepatocellular carcinoma (HCC). METHODS: A retrospective analysis was conducted in prospectively collected 150 adult HCC patients who received liver transplantation from 1997 to 2015. The post-transplant 41 inflammatory cytokines were quantified by multiplexing analysis and determined their prognostic value for predicting post-LT tumor recurrence by receiver operative characteristic analysis. A prediction model for post-LT tumor recurrence was generated by the logistic regression and internally validated Bootstrapping and compared with external prediction models. RESULTS: Post-transplant circulating CCL11, IFNα2, and IL17A cytokines were identified to be significant predictors of post-LT tumor recurrence and survival. A prediction score composed of the post-transplant 3-cytokine (P3C) signature, UCSF criteria, and pre-LT AFP was established. The P3C-UCSF-AFP score significantly predicted post-LT tumor recurrence and poor survival both in deceased donor liver transplantation (DDLT) and living donor liver transplantation (LDLT). The P3C-UCSF-AFP score was validated to significantly predict post-LT 2-year and 5-year tumor recurrence, outperforming the RETREAT score, French AFP model, up-to-seven, UCSF criteria, and Milan criteria. Importantly, the P3C-UCSF-AFP score could cost-effectively stratify high-risk recipients subjected to a refinement of post-recurrence survival. CONCLUSION: The integrated P3C-UCSF-AFP score not only compensated for the pre-LT unpredictability and predicted post-LT tumor recurrence accurately, but also guided the clinical refinements of post-LT surveillance and therapeutic strategies in transplant oncology.

Type III TGF-ß Receptor Down-Regulation Promoted Tumor Progression via Complement Component C5a Induction in Hepatocellular Carcinoma.

Background and Aims-Transforming growth factor-beta (TGF-ß) signaling orchestrates tumorigenesis and one of the family members, TGF-ß receptor type III (TGFßR3), are distinctively under-expressed in numerous malignancies. Currently, the clinical impact of TGFßR3 down-regulation and the underlying mechanism remains unclear in hepatocellular carcinoma (HCC). Here, we aimed to identify the tumor-promoting roles of decreased TGFßR3 expression in HCC progression. Materials and Methods-For clinical analysis, plasma and liver specimens were collected from 100 HCC patients who underwent curative resection for the quantification of TGFßR3 by q-PCR and ELISA. To study the tumor-promoting mechanism of TGFßR3 downregulation, HCC mouse models and TGFßR3 knockout cell lines were applied. Results-Significant downregulation of TGFßR3 and its soluble form (sTGFßR3) were found in HCC tissues and plasma compared to healthy individuals (p < 0.01). Patients with <9.4 ng/mL sTGFßR3 exhibited advanced tumor stage, higher recurrence rate and shorter disease-free survival (p < 0.05). The tumor-suppressive function of sTGFßR3 was further revealed in an orthotopic mouse HCC model, resulting in 2-fold tumor volume reduction. In TGFßR3 knockout hepatocyte and HCC cells, increased complement component C5a was observed and strongly correlated with shorter survival and advanced tumor stage (p < 0.01). Interestingly, C5a activated the tumor-promoting Th-17 response in tumor associated macrophages. Conclusion-TGFßR3 suppressed tumor progression, and decreased expression resulted in poor prognosis in HCC patients through upregulation of tumor-promoting complement C5a.

Plasmacytoid dendritic cells recruited by HIF-1α/eADO/ADORA1 signaling induce immunosuppression in hepatocellular carcinoma.

Plasmacytoid dendritic cells (pDCs) play immunosuppressive roles in the tumor microenvironment (TME). However, the molecular mechanisms underlying the recruitment and dysfunction of pDCs in the TME remain largely elusive, especially in hepatocellular carcinoma (HCC). In this study, we observed the accumulation of pDCs in the blood, tumor tissue, and ascitic fluid of HCC patients. A high density of tumor-infiltrating pDCs was correlated with poor prognosis in patients with HCC. Hypoxia-induced extracellular adenosine (eADO) significantly enhanced pDC recruitment into tumors via the adenosine A1 receptor (ADORA1). Mechanistically, hypoxia-inducible factor 1-alpha (HIF-1α) transcriptionally upregulated the expression of the ectonucleotidases CD39 and CD73 in HCC cells, both of which are essential for the generation of eADO. Moreover, eADO-stimulated pDCs promoted the induction of regulatory T cells and suppressed proliferation and cytotoxicity of CD8+ T cells. Depletion of pDCs using a monoclonal antibody or an ADORA1 antagonist significantly improved antitumor immunity and suppressed HCC growth in the immunocompetent HCC mouse model. Thus, targeting pDC recruitment may serve as a potential adjuvant strategy for immunotherapies in HCC.

Glutathione S-transferase A2 promotes hepatocellular carcinoma recurrence after liver transplantation through modulating reactive oxygen species metabolism.

Hepatocellular carcinoma (HCC) recurrence after liver transplantation remains a significant clinical problem. Ischemia-reperfusion injury (IRI) occurred inevitably at the early phase after liver transplantation (LT) spawns a significant risk of HCC recurrence. However, their linkage and IRI-derived risk factors for HCC recurrence remain exclusive. Understanding the mechanism of post-transplantation hepatic injury could provide new strategies to prevent the later event of HCC recurrence. We demonstrated that glutathione S-transferase A2 (GSTA2) expression was significantly associated with early phase hepatic and systemic injury and ROS level after liver transplantation. Early phase circulating GSTA2 (EPCGSTA2) protein was a significant predictor of HCC recurrence and survival. Heterogeneous single nucleotide polymorphism at G335C of GSTA2 was significantly associated with poor survival of HCC recipients. Enhancement of GSTA2 could protect HCC cells against H2O2-induced cell death by compensating for the elevated ROS stress. We also demonstrated that GSTA2 played crucial roles in regulating the ROS-associated JNK and AKT signaling pathways and ROS metabolism in HCCs in responding to a dynamic ROS environment. Functionally, endogenous or exogenous upregulation of GSTA2 could promote HCC growth and invasion through activating the epithelial-mesenchymal-transition process. Targeted inhibition of GSTA2 could suppress HCC growth and metastasis. In conclusion, GSTA2 could be a novel prognostic and therapeutic target to combat HCC recurrence after liver transplantation.

FTY720 Suppresses Liver Tumor Growth and Metastasis by Reducing Circulating Regulating T Cells and Enhancing the Anti-Tumor Effect of Rapamycin.

BACKGROUND: In this study, we aimed to study the effect of FTY720 treatment in reducing circulating Tregs level and then suppressing liver tumor metastasis after hepatectomy and I/R injury in animal models. Furthermore, we also investigated the synergistic anti-tumor effect of FTY720 combined with rapamycin on hepatocellular carcinoma. METHODS: The effect of FTY720 on suppressing Tregs mobilization and tumor metastasis after hepatectomy was investigated in an orthotopic liver tumor rat model with hepatectomy and hepatic ischemia/reperfusion (I/R) injury. The synergistic anti-tumor effect of FTY720 combined with rapamycin was further explored both in in vitro functional study and in orthotopic liver tumor mouse model. RESULTS: In rat model, hepatic I/R promoted tumor metastasis and increased circulating Tregs after hepatectomy. The treatment of FTY720 reduced liver tumor metastasis and the number of circulating Tregs. Furthermore, FTY720 enhanced the anti-tumor capacity of rapamycin by inhibiting tumor cell proliferation and migration in vitro and reducing tumor growth in vivo through suppressing hepatic stellate cell activation and tumor angiogenesis. CONCLUSION: FTY720 suppressed liver tumor growth and metastasis by reducing the population of circulating Tregs and enhancing the anti-tumor effect of rapamycin. It was suggested that FTY720 single or combined with rapamycin might provide novel insight for suppressing tumor growth and metastasis for HCC patients.

IL-17a exacerbates hepatic ischemia-reperfusion injury in fatty liver by promoting neutrophil infiltration and mitochondria-driven apoptosis.

Hepatic ischemia-reperfusion (IR) injury is a critical issue during liver transplantation (LT). Recent studies have demonstrated that IL-17a contributes to IR injury and steatohepatitis. However, the underlying mechanism is not understood. This study aimed to examine the role of IL-17a on hepatic IR injury in fatty liver and to investigate the underlying mechanisms. The correlation between serum IL-17a levels and liver function was analyzed in LT patients receiving fatty (n = 42) and normal grafts (n = 44). Rat LT model was applied to validate the clinical findings. IL-17a knockout (KO) and wild-type mice were fed with high-fat diets to induce fatty liver and subjected to hepatic IR injury with major hepatectomy. Frequency of circulating neutrophils and IL-17a expression on PBMCs were analyzed by flow cytometry. Mitochondrial outer membrane permeabilization (MOMP) was examined by a living intravital image system. Serum IL-17a was elevated after human LT, especially with fatty grafts. The aspartate aminotransferase and alanine transaminase levels were increased in recipients with fatty grafts compared with normal grafts. In rat LT model, the intragraft IL-17a expression was significantly higher in fatty grafts than normal ones post-LT. KO of IL-17a in mice notably attenuated liver damage after IR injury in fatty liver, characterized by better-preserved liver architecture, improved liver function, and reduced neutrophil infiltration. MOMP triggered cell death after hepatic IR injury in a caspase-independent way via IL-17a/NF-κB signaling pathway. KO of IL-17a protected the fatty liver against IR injury through the suppression of neutrophil infiltration and mitochondria-driven apoptosis.

Clinical significance and functional role of transmembrane protein 47 (TMEM47) in chemoresistance of hepatocellular carcinoma.

Chemoresistance is the main cause of chemotherapy failure in patients with hepatocellular carcinoma (HCC). The gene encoding transmembrane protein 47 (TMEM47) was previously identified to be significantly upregulated in HCC cell lines with acquired chemoresistance. The aim of the present study was to characterize the clinical significance and function of TMEM47 in HCC chemoresistance. The results demonstrated that the TMEM47 expression levels in the tumors of patients not responding to cisplatin­based transarterial chemoembolization (TACE) treatment was significantly higher compared with those in patients who responded to TACE treatment. Moreover, analyses from clinical samples and HCC cell lines indicated that TMEM47 expression may be upregulated in HCC in response to cisplatin treatment. Furthermore, the TMEM47 mRNA expression levels were positively correlated with the degree of cisplatin resistance of HCC cells. Overexpression of TMEM47 in HCC cells significantly promoted cisplatin resistance. The present study also demonstrated that targeted inhibition of TMEM47 could significantly reduce cisplatin resistance of cisplatin­resistant HCC cells via enhancing caspase­mediated apoptosis. In addition, targeted inhibition of TMEM47 enhanced the sensitivity of cisplatin­resistant cells to cisplatin via suppressing cisplatin­induced activation of the genes involved in drug efflux and metabolism. The present study also validated that TMEM47 expression was significantly correlated with multidrug resistance­associated protein 1 in patients with HCC who received TACE treatment. In conclusion, the findings of the present study demonstrated that TMEM47 may be a useful biomarker for predicting the response to chemotherapy and a potential therapeutic target for overcoming HCC chemoresistance.

Neonatal exposure of ketamine inhibited the induction of hippocampal long-term potentiation without impairing the spatial memory of adult rats.

Ketamine is one of general anesthetics and has been commonly used in obstetric and pediatric anesthesia. However, effects of exposure to ketamine on neonatal brain are largely unknown. In this study, we aim to investigate the effect of neonatal exposure of ketamine on spatial memory and long-term potentiation (LTP) in the hippocampus of adult rats. One-week-old neonatal rats were separated into ketamine group and control group. Neonatal rats in ketamine group were received intraperitoneal injection of 25 mg/kg (low-dose group, N = 8) or 50 mg/kg ketamine (high-dose group, N = 8). Neonatal Rats in control group received saline injection (N = 8). After 10 weeks, the spatial memory of adult rats was examined by using Morris Water Maze, and LTP in the hippocampus of adult rats was assessed by electrophysiological experiment. We found that exposure of ketamine to neonatal rats, either low-dose or high-dose, had not induced alteration on their adulthood's escape latency, swimming speed and the percentage of time spent in original quadrant compared with the control. The electrophysiological examination showed that the induction of LTP in hippocampus was significantly reduced in adult rats of ketamine group (either low-dose or high-dose). Our study showed that neonatal exposure of ketamine inhibited the induction of hippocampal LTP without impairing the spatial memory of adult rats.

ApoA-1 accelerates regeneration of small-for-size fatty liver graft after transplantation.

OBJECTIVES: Apolipoprotein A-1 (ApoA-1) is involved in regulating both lipid and energy metabolism, which may play important roles in liver regeneration, especially for the liver with steatosis. We here intended to investigate the role of ApoA-1 in regeneration of small-for-size fatty liver graft and to explore the underlying mechanism. METHODS: The association of ApoA-1 expression with liver regeneration was studied in rat liver transplantation models using small-for-size normal graft or small-for-size fatty graft. The direct role of ApoA-1 in liver regeneration was studied in mouse hepatectomy model in vivo and hepatocytes in vitro. RESULTS: Compared to small-for-size normal graft, decreased expression of ApoA-1 associated with delayed regeneration were detected in small-for-size fatty liver graft after transplantation. In functional study, the expression of ApoA-1 was decreased in hepatocytes with steatosis and was inversely associated with the concentration of oleic acid. The ApoA-1 administration effectively attenuated hepatocytes steatosis and accelerated hepatocytes proliferation. In mouse model, ApoA-1 treatment promoted liver regeneration at day 2 after major hepatectomy. In addition, the treatment of ApoA-1 increased the expressions of PGC-1α and its target genes Tfam, Ucp2 and SDHB. CONCLUSIONS: ApoA-1 may accelerate regeneration of small-for-size fatty liver graft at day 2 after transplantation through regulating mitochondrial function. ApoA-1 may be the potential new therapy of promoting liver regeneration.

Glutathione Peroxidase 3 Delivered by hiPSC-MSCs Ameliorated Hepatic IR Injury via Inhibition of Hepatic Senescence.

Background and Aims: Down-regulation of GPx3 accelerated hepatic senescence, which further caused overwhelming inflammation and severe liver graft injury. MSCs derived from human induced pluripotent stem cells (hiPSC-MSCs) have been developed as more efficient delivery vehicle with the property of injury tropism. Here, we aimed to explore the suppressive role of GPx3 in hepatic IR injury using novel delivery system of hiPSC-MSCs. Methods: The mice IR injury model with partial hepatectomy was established. The engineered hiPSC-MSCs delivering GPx3 was constructed. All the mice were segregated into three groups. hiPSC-MSC-GPx3, hiPSC-MSC-pCDH (vector control) or PBS were injected via portal vein after reperfusion. Liver injury was evaluated by histological and serological test. Hepatic apoptosis was detected by Tunel staining and remnant liver regeneration was assessed by Ki67 staining. The role of hepatic senescence in liver graft injury was evaluated in rat orthotopic liver transplantation model. The suppressive effect of GPx3 on hepatic senescence was examined in mice IR injury model and confirmed in vitro. Hepatic senescence was detected by SA-ß-Gal and P16/ink4a staining. Results: GPx3 can be successfully delivered by hiPSC-MSCs into liver tissues. Histological examination showed that hiPSC-MSC-GPx3 treatment significantly ameliorated hepatic IR injury post-operation. Significantly lower LDH (891.43±98.45 mU/mL, P<0.05) and AST (305.77±36.22 IU/L, P<0.01) were observed in hiPSC-MSC-GPx3 group compared with control groups. Less apoptotic hepatocytes were observed and the remnant liver regeneration was more active in hiPSC-MSC-GPx3 group. In rat orthotopic liver transplantation model, more senescent hepatocytes were observed in small-for-size liver graft, in which GPx3 expression was significantly compromised. In mice IR injury model, hiPSC-MSC-GPx3 significantly suppressed hepatic senescence. In addition, rGPx3 inhibited cellular senescence of liver cells in a dose dependent manner. Four candidate genes (CD44, Nox4, IFNG, SERPERINB2) were identified to be responsible for suppressive effect of GPx3 on hepatic senescence. Conclusion: Engineered hiPSC-MSCs delivering GPx3 ameliorated hepatic IR injury via inhibition of hepatic senescence.