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Background: Early and late recurrence of hepatocellular carcinoma (HCC) have different clinical outcomes, especially for those accompanied by microvascular invasion (MVI), but the definition of early recurrence remains controversial. Therefore, a reasonable identification of the early recurrence time for HCC is urgently needed., Methods: Resected recurrence patients were enrolled and divided into two cohorts, one for identification of the early recurrence time and another for verification of the accuracy of the point. Univariable and multivariable Cox regression analyses were adopted to identify the prognostic factors of recurrence HCC (rHCC) and Kaplan-Meier method was applied to analyze the overall survival (OS). The appropriate cutoff value was determined by the exhaustive method using different recurrence intervals from 1 to 24 months in turn., Results: In total, 292 resected rHCC patients were analyzed to calculate the early recurrence interval, and another 421 resected rHCC patients with MVI were enrolled to verify the efficacy of adjuvant transarterial chemoembolization (TACE) in this recurrence interval. MVI was identified as an independent risk factor by multivariable analysis. The OS of rHCC patients without MVI is better than that of patients with MVI when the recurrence time was within 13 months, while not beyond 13 months. The verification cohort demonstrated that adjuvant TACE provided longer survival for rHCC with MVI when the recurrence time was within 13 months, while not beyond 13 months., Conclusion: For HCC patients with MVI who underwent R0 resection, 13 months may be a reasonable early recurrence time point, and within this interval, postoperative adjuvant TACE may result in longer survival compared with surgery alone., (© 2023 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.)

Purpose: This study aimed to assess the efficacy and safety of a triple therapy that comprises transarterial chemoembolization (TACE), antiangiogenic-targeted therapy, and programmed death-1 (PD-1) inhibitors in a real-world cohort of patients with unresectable hepatocellular carcinoma (HCC) with portal vein tumor thrombus (PVTT)., Methods: Consecutive patients treated with TACE combined with antiangiogenic therapy and PD-1 inhibitors at the Eastern Hepatobiliary Surgery Hospital between June 2019 and May 2021 were enrolled. The baseline characteristics and treatment course of the patients were recorded. The tumor response was evaluated based on the Response Evaluation Criteria in Solid Tumors (RECIST) 1.1 and HCC-specific modified RECIST (mRECIST). The overall survival (OS) and progression-free survival (PFS) of the patients were analyzed using the Kaplan-Meier method. Adverse events (AEs) were assessed according to the National Cancer Institute Common Terminology Criteria for Adverse Events version 5.0., Results: As of the data cutoff on 30 August 2021, the median follow-up time was 10.0 (3.9-28.4) months. A total of 39 eligible patients were included. The objective response rate (ORR) and the disease control rate (DCR) were 35.9% and 74.4% according to the RECIST 1.1, and 48.7% and 84.6% according to mRECIST criteria, respectively. The median OS and PFS were 14.0 and 9.2 months, respectively. Moreover, 34 (87.2%) patients experienced at least one treatment-related AE and 8 (20.5%) patients experienced grade 3/4 treatment-related AEs. The most common treatment- and laboratory-related AEs were hypertension (46.2%) and decreased albumin (53.8%), respectively. No treatment-related mortality occurred during the study period., Conclusions: TACE combined with antiangiogenic-targeted therapy and immune checkpoint inhibitors may have promising anticancer activity in unresectable HCC patients with PVTT. AEs were manageable, with no unexpected overlapping toxicities., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Feng, Liu, Fu, Chai, Sun, Wang, Cheng, Zhu, Xiang, Zhou, Shi, Guo, Zhai and Cheng.)

Background: Microvascular invasion (MVI) is a risk factor of post-hepatectomy tumor recurrence for hepatocellular carcinoma (HCC). The patterns, treatments, and prognosis have not been documented in HCC patients with MVI., Methods: A multicenter database of patients with HCC and MVI following resection was analyzed. The clinicopathological and initial operative data, timing and first sites of recurrence, recurrence management, and long-term survival outcomes were analyzed., Results: Of 1517 patients included, the median follow-up was 39.7 months. Tumor recurrence occurred in 928 patients, with 49% within 6 months of hepatectomy and 60% only in the liver. The incidence of intrahepatic only recurrence gradually increased with time after 6 months. Patients who developed recurrence within 6 months of hepatectomy had worse survival outcomes than those who developed recurrence later. Patients who developed intrahepatic only recurrence had better prognosis than those with either extrahepatic only recurrence or those with intra- and extrahepatic recurrence. Repeat resection of recurrence with curative intent resulted in better outcomes than other treatment modalities., Conclusion: Post-hepatectomy tumor recurrence in patients with HCC and MVI had unique characteristics and recurrence patterns. Early detection of tumor recurrence and repeat liver resection with curative intent resulted in improved long-term survival outcomes., (Copyright © 2021 International Hepato-Pancreato-Biliary Association Inc. Published by Elsevier Ltd. All rights reserved.)

Background: Hepatocellular carcinoma (HCC) with bile duct tumor thrombus (BDTT) is rare. The aim of this study is to evaluate the long-term prognosis of liver resection (LR) versus transcatheter arterial chemoembolization (TACE) in these patients., Methods: Data from HCC patients with BDTT who underwent liver resection and TACE were analyzed respectively. Propensity score matching (PSM) analysis was performed in these patients., Results: A total of 145 HCC patients with BDTT were divided into two groups: the LR group (n = 105) and the TACE group (n = 40). The median OS in the LR group was 8.0 months longer than that in the TACE group before PSM (21.0 vs. 13.0 months, P <0.001) and 9.0 months longer after PSM (20.0 vs. 11.0 months, P <0.001). The median DFS in the LR group was 3.5 months longer than that in the TACE group before PSM (7.0 vs. 3.5 months, P = 0.007) and 5 months longer after PSM (7.0 vs. 2.0 months, P = 0.007)., Conclusion: If surgery is technically feasible, liver resection provides better prognosis for HCC patients with BDTT compared with TACE., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Sun, Feng, Yang, Chen, Liu, Chai, Mao, Guo, Shi and Cheng.)

Background: Microvascular invasion (MVI) adversely affects long-term survival in patients with hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC). This study aimed to examine the association between preoperative type 2 diabetes mellitus (T2DM) with incidences of MVI and prognosis in HBV-related HCC after liver resection (LR)., Material and Methods: Data of HBV-related HCC patients who underwent LR as an initial therapy from four hospitals in China were retrospectively collected. Clinicopathological factors associated with the incidence of MVI were identified using univariate and multivariate logistic regression analysis. The recurrence-free survival (RFS) and overall survival (OS) curves between different cohorts of patients were generated using the Kaplan-Meier method and compared using the log-rank test., Results: Of 1473 patients who were included, 219 (14.9%) patients had T2DM. Preoperative T2DM, HBV DNA load, antiviral treatment, AFP level, varices, and tumor encapsulation were identified to be independent predictors of the incidence of MVI. Patients with HBV-related HCC and T2DM had a higher incidence of MVI (65.8%) than those without T2DM (55.4%) (P = 0.004). The RFS and OS were significantly worse in patients with T2DM than those without T2DM (median RFS: 11.1 vs 16.7 months; OS: 26.4 vs 42.6 months, both P < 0.001). Equivalent results were obtained in HCC patients with MVI who had or did not have T2DM (median RFS: 10.0 vs 15.9 months; OS: 24.5 vs 37.9 months, both P < 0.001)., Conclusions: Preoperative T2DM was an independent risk factor of incidence of MVI. Patients with HBV-related HCC and T2DM had worse prognosis than those without T2DM after LR., (Copyright © 2021. Published by Elsevier Ltd.)

Background: The presence of hepatic vein tumor thrombus (HVTT) is a major determinant of survival outcomes in hepatocellular carcinoma (HCC) patients. This study compared survival outcomes between liver resection (LR) and intensity-modulated radiation therapy (IMRT) in HCC patients with HVTT., Methods: Data from patients who underwent LR or IMRT for HCC with HVTT at the Eastern Hepatobiliary Surgery Hospital were retrospectively analyzed. Their survival outcomes were compared before and after propensity score matching (PSM)., Results: Three hundred and seven HCC patients with HVTT who underwent either LR (n=140) or IMRT (n=167) were enrolled. PSM matched 82 pairs of patients. The overall survival (OS) and recurrence-free survival (RFS) rates were significantly higher for patients in the LR group than those in the IMRT group. On subgroup analysis, significantly better survival outcomes were obtained after LR than IMRT in patients with peripheral type of HVTT (pHVTT) and major type of HVTT (mHVTT). However, similar survival outcomes were obtained after LR and IMRT when the HVTT had developed into inferior vena cava tumor thrombus (IVCTT)., Conclusions: LR resulted in significantly better survival outcomes in HCC patients with HVTT when compared to IMRT. Once the HVTT had developed IVCTT, LR and IMRT resulted in similarly bad survival outcomes., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://dx.doi.org/10.21037/hbsn.2020.03.20). The authors have no conflicts of interest to declare. Dr. WYL serves as the unpaid editorial board members of Hepatobiliary Surgery and Nutrition. The other authors have no conflicts of interest to declare., (2021 Hepatobiliary Surgery and Nutrition. All rights reserved.)

Background: Occurrence of portal vein tumor thrombus (PVTT) worsens the outcomes of hepatocellular carcinoma (HCC) and imparts high economic burden on society. Patients with high risks of having hypercoagulation are more likely to experience thrombosis. Herein, we examined how preoperative international normalized ratio (INR) was related to the incidence and extent of PVTT, and associated with survival outcomes in HCC patients following R0 liver resection (LR)., Methods: Patients with HCC and PVTT were enrolled from six major hospitals in China. The overall survival (OS) and recurrence-free survival (RFS) rates of individuals with different INR levels were assessed with Cox regression analysis as well as Kaplan-Meier method., Results: This study included 2207 HCC patients, among whom 1005 patients had concurrent PVTT. HCC patients in the Low INR group had a significantly higher incidence of PVTT and more extensive PVTT than the Normal and High INR groups (P<0.005). Of the 592 HCC subjects who had types I/II PVTT following R0 LR, there were 106 (17.9%), 342 (57.8%) and 144 (24.3%) patients in the High, Normal and Low INR groups, respectively. RFS and OS rates were markedly worse in patients in the Low INR group relative to those in the Normal and High INR groups (median RFS, 4.87 versus 10.77 versus 11.40 months, P<0.001; median OS, 6.30 versus 11.83 versus 12.67 months, P<0.001)., Conclusion: Preoperative INR influenced the incidence and extent of PVTT in HCC. Particularly, patients with HCC and PVTT in the Low INR group had worse postoperative prognosis relative to the High and Normal INR groups., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhang, Zhou, Feng, Sun, Zhen, Zhou, Zhang, Hu, Zhong, Chen, Chai, Wang, Shi, Guo, Wu, Lau and Cheng.)

Portal vein tumor thrombus (PVTT) is one of the most serious forms of hepatocellular carcinoma (HCC) vessel metastasis and has a poor survival rate. However, the molecular mechanism of PVTT has not yet been elucidated. In this study, the molecular mechanism of AXL expressed in tumor-derived endothelial cells (TECs) in vessel metastasis was investigated. High AXL expression was observed in TECs, but not in the tumor cells of HCC patients with PVTT and this was associated with poor overall survival (OS) and disease-free survival (DFS). AXL overexpression was positively associated with CD 31 expression both in vitro and in vivo . AXL promoted the cell proliferation, tube formation, and migration of both TECs and normal endothelial cells (NECs). High expression of AXL in TECs promoted the cell migration, but not the proliferation of HCC cells. Further studies demonstrated that AXL promoted cell migration and tube formation through activation of the PI3K/AKT/SOX2/DKK-1 axis. AXL overexpression in HUVECs promoted tumor growth and liver or vessel metastasis of HCC in xenograft nude mice, which could be counteracted by treatment with R428, an AXL inhibitor. R428 reduced tumor growth and CD 31 expression in HCC in PDX xenograft nude mice. Therefore, AXL over-expression in TECs promotes vessel metastasis of HCC, which indicates that AXL in TECs could be a potential therapeutic target in HCC patients with PVTT., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Chai, Zhang, Ao, Zhu, Wu, Lau, Sun and Cheng.)

Background: Hepatocellular carcinoma (HCC) commonly occurs in patients with splenomegaly. This study aimed to investigate the impact of splenomegaly with or without splenectomy on long-term survival of HCC patients with portal vein tumor thrombus (PVTT) treated with liver resection (LR)., Methods: HCC patients with PVTT who underwent LR from 2005 to 2012 from 6 hospitals were retrospectively studied. The long-term overall survival (OS) and recurrence-free survival (RFS) were compared between patients with or without splenomegaly, and between patients who did or did not undergo splenectomy for splenomegaly. Propensity score matching (PSM) analysis was performed to match patients in a 1:1 ratio., Results: Of 716 HCC patients with PVTT who underwent LR, 140 patients had splenomegaly (SM group) and 576 patients had no splenomegaly (non-SM group). The SM group was further subdivided into 49 patients who underwent splenectomy (SPT group), and 91 patients who did not received splenectomy (non-SPT group). PSM matched 140 patients in the SM group, and 49 patients in the SPT group. Splenomegaly was an independent risk factor of poor RFS and OS. The OS and RFS rates were significantly better for patients in the non-SM group than the SM group (OS: P<0.001; RFS: P<0.001), and for patients in the SPT group than the non-SPT group (OS: P<0.001; RFS: P<0.001)., Conclusions: Patients who had splenomegaly had significantly worse survival in HCC patients with PVTT. Splenectomy for splenomegaly significantly improved long-term survival in these patients., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-20-2229). The authors have no conflicts of interest to declare., (2021 Annals of Translational Medicine. All rights reserved.)

Background: The effect of bile duct tumor thrombus (BDTT) on the postoperative long-term prognosis of hepatocellular carcinoma (HCC) patients is still under debate., Methods: The PubMed, Embase, Cochrane Library, Web of Science databases were systematically searched to collect the clinicopathologic characteristics, perioperative indices, and postoperative survival outcomes in the BDTT and non-BDTT groups of HCC patients from inception to February 1, 2020. The study outcomes were extracted by two independent investigators., Results: A total of 15 studies involving 6,484 patients were included. The meta-analysis revealed that the levels of serum total bilirubin and alkaline phosphatase were notably higher in patients with HCC and BDTT than those without BDTT. Meanwhile, HCC patients with BDTT had more aggressive biological characteristics, such as poor tumor differentiation, macrovascular invasion, and lymph node metastasis, as compared to patients without BDTT. The 1-year [odds ratio (OR) 0.39, 95% confidence interval (CI): 0.31-0.48, P<0.01], 3-year (OR 0.33, 95% CI: 0.22-0.51, P<0.01) and 5-year overall survival (OS) rates (OR 0.31, 95% CI: 0.20-0.49, P<0.01) of the BDTT group were significantly worse than those of the non-BDTT group. The hazard ratio of HCC with BDTT was 4.27 (95% CI: 3.47-5.26, P<0.01) within 5 years after hepatectomy., Conclusions: HCC patients with BDTT had worse OS compared to patients free of BDTT after surgery. BDTT may be a potential prognostic factor for HCC patients., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/atm-20-4698). The authors have no conflicts of interest to declare., (2020 Annals of Translational Medicine. All rights reserved.)

Background: The optimal surgical modality for hepatocellular carcinoma (HCC) with bile duct tumor thrombus (BDTT) remains controversial, especially regarding deciding whether to perform concurrent bile duct resection (BDR)., Methods: PubMed, EMBASE, Cochrane Library, Web of Science and Scopus databases were systematically searched from inception to February 2020, in order to compare overall survival (OS) and recurrence-free survival (RFS) rates of HCC patients with BDTT who had either received hepatectomy with extrahepatic bile duct resection (BDR group) or hepatectomy without bile duct resection (NBDR group). Relevant outcomes were extracted by two investigators., Results: A total of 12 studies involving 355 patients was included. The 1-, 3- and 5-year OS rates were similar in the BDR and NBDR groups (OR =0.58, 95% CI: 0.31-1.09, P=0.09; OR =0.74, 95% CI: 0.43-1.28, P=0.28; OR =0.63, 95% CI: 0.36-1.11, P=0.11, respectively). However, the BDR group had better 1-, 3- and 5-year RFS rates than the NBDR group (OR =0.38, 95% CI: 0.22-0.65, P<0.01; OR =0.40, 95% CI: 0.22-0.72, P<0.01; OR =0.37, 95% CI: 0.19-0.71, P<0.01, respectively)., Conclusions: Concomitant bile duct resection results in decreased postoperative recurrence in HCC patients with BDTT. However, the OS rates were similar whether or not patients underwent bile duct resection., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at: http://dx.doi.org/10.21037/atm-20-3935). The authors have no conflicts of interest to declare., (2020 Annals of Translational Medicine. All rights reserved.)

Background: Hepatic vein tumor thrombus (HVTT) is a significant poor risk factor for survival outcomes in hepatocellular carcinoma (HCC) patients. Currently, the widely used international staging systems for HCC are not refined enough to evaluate prognosis for these patients. A new classification for macroscopic HVTT was established, aiming to better predict prognosis., Methods: This study included 437 consecutive HCC patients with HVTT who underwent different treatments. Overall survival (OS) and time-dependent receiver operating characteristic (ROC) curve area analysis were used to determine the prognostic capacities of the new classification when compared with the different currently used staging systems., Results: The new HVTT classification was defined as: type I, tumor thrombosis involving hepatic vein (HV), including microvascular invasion; type II, tumor thrombosis involving the retrohepatic segment of inferior vena cava; and type III, tumor thrombosis involving the supradiaphragmatic segment of inferior vena cava. The numbers (percentages) of patients with types I, II, and III HVTT in the new classification were 146 (33.4%), 143 (32.7%), and 148 (33.9%), respectively. The 1-, 2-, and 3-year OS rates for types I to III HVTT were 79.5%, 58.6%, and 29.1%; 54.8%, 23.3%, and 13.8%; and 24.0%, 10.0%, and 2.1%, respectively. The time-dependent-ROC curve area analysis demonstrated that the predicting capacity of the new HVTT classification was significantly better than any other staging systems., Conclusions: A new HVTT classification was established to predict prognosis of HCC patients with HVTT who underwent different treatments. This classification was superior to, and it may serve as a supplement to, the commonly used staging systems., Competing Interests: Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/hbsn.2019.10.07). Dr. WYL serves as an unpaid editorial board member of Hepatobiliary Surgery and Nutrition. The other authors have no conflicts of interest to declare., (2020 Hepatobiliary Surgery and Nutrition. All rights reserved.)

Background: The aim of this study was to evaluate the effect of portal vein tumor thrombus (PVTT) on the prognosis of patients undergoing liver resection (LR) for primary liver malignancies (PLC)., Methods: The recurrence-free survival (RFS) and overall survival (OS) for patients undergoing LR with and without PVTT for three primary liver malignancies, including hepatocellular carcinoma (HCC), intrahepatic cholangiocarcinoma (ICC) and hepato-cholangio carcinoma (CHC) were compared using the Kaplan-Meier method and Cox regression analysis., Results: In total, 3775 patients with PLC who underwent LR were included in this study. The incidence of PVTT in patients undergoing LR with HCC, IHC and CHC were 46%, 20%, and 17%, respectively. The median RFS and OS were significantly better for patients with HCC as compared to ICC or CHC (16 vs 11 vs 13 months; 21 vs 16 vs 18 months, respectively; P < 0.001). However, the presence of PVTT resulted in similarly poor RFS and OS in these 3 subgroups of patients (9 vs 8 vs 8 months, P = 0.062; 14 vs 13 vs 12 months, respectively, P = 0.052)., Conclusion: Although the prognosis of patients with PLC varied by histological subtype, once PVTT occurred, survival outcomes after LR were similarly poor across all three subgroups., (Copyright © 2019. Published by Elsevier Ltd.)

Background: Hepatic vein tumour thrombus (HVTT) is a major determinant of survival outcomes for patients with hepatocellular carcinoma (HCC). An Eastern Hepatobiliary Surgery Hospital (EHBH)-HVTT model was established to predict the prognosis of patients with HCC and HVTT after liver resection, in order to identify optimal candidates for liver resection., Methods: Patients with HCC and HVTT from 15 hospitals in China were included. The EHBH-HVTT model with contour plot was developed using a non-linear model in the training cohort, and subsequently validated in internal and external cohorts., Results: Of 850 patients who met the inclusion criteria, there were 292 patients who had liver resection and 198 who did not in the training cohort, and 124 and 236 in the internal and external validation cohorts respectively. Contour plots for the EHBH-HVTT model were established to predict overall survival (OS) rates of patients visually, based on tumour diameter, number of tumours and portal vein tumour thrombus. This differentiated patients into low- and high-risk groups with distinct long-term prognoses in the liver resection cohort (median OS 34·7 versus 12·0 months; P < 0·001), internal validation cohort (32·8 versus 10·4 months; P = 0·002) and external validation cohort (15·2 versus 6·5 months; P = 0·006). On subgroup analysis, the model showed the same efficacy in differentiating patients with HVTT in peripheral and major hepatic veins, the inferior vena cava, or in patients with coexisting portal vein tumour thrombus., Conclusion: The EHBH-HVTT model was accurate in predicting prognosis in patients with HCC and HVTT after liver resection. It identified optimal candidates for liver resection among patients with HCC and HVTT, including tumour thrombus in the inferior vena cava, or coexisting portal vein tumour thrombus., (© 2020 BJS Society Ltd Published by John Wiley & Sons Ltd.)

Study Design: To evaluate the effect of p38 pathway on spinal cord injury (SCI), a rat model of SCI was performed., Objective: We determined the effect of p38 on SCI and SCI related inflammation, apoptosis, and autophagy., Summary of Background Data: SCI is a severe clinical problem worldwide. It is difficult to prevent cell necroptosis and promote the survival of residual neurons after SCI. p38, a class of mitogen-activated protein kinases, its effect on SCI and SCI related inflammation, apoptosis, and autophagy have not been studied very well., Methods: The rats were randomly divided into the following four groups: the sham-operated (sham) group, the SCI group, the SCI + vehicle group, and the SCI + SB203580 (10 mg/kg) group. The p38 inhibitor SB203580 was administered by oral (10 mg/kg/d) gavage once per day for 14 days. Neurological recovery was assessed using the Basso, Beattie, and Bresnahan locomotion rating scale. Apoptosis, autophagy, and inflammation related proteins were measured by enzyme-linked immunosorbent assay kits or western blotting., Results: Our results showed that p38 was upregulated after SCI from day 3, which was paralleled with the levels of its proteins ATF-2, suggesting an increase in p38 activity. Our results showed administration of SB203580 attenuated histopathology and promoted locomotion recovery in rats after SCI. SB203580 administration significantly inhibited inflammatory cytokines levels as well as the inflammation signaling pathway. SB203580 administration also modulated the apoptosis and autophagy signaling pathway., Conclusion: Our findings suggest that p38 inhibitor SB203580 treatment alleviates secondary SCI by inhibiting inflammation and apoptosis, thereby promoting neurological and locomoter functional recovery, thus suggest the important role of p38 in neuronal protection after SCI., Level of Evidence: N/A.

Background: Microvascular invasion (MVI) is associated with poor postoperative survival outcomes in patients with hepatocellular carcinoma (HCC). An Eastern Hepatobiliary Surgery Hospital (EHBH) MVI scoring system was established to predict prognosis in patients with HCC with MVI after R0 liver resection (LR) and to supplement the most commonly used classification systems., Materials and Methods: Patients with HCC with MVI who underwent R0 LR as an initial therapy were included. The EHBH-MVI score was developed from a retrospective cohort from 2003 to 2009 to form the training cohort. The variables associated with overall survival (OS) on univariate analysis were subsequently investigated using the log-rank test, and the EHBH-MVI score was developed using the Cox regression model. It was validated using an internal prospective cohort from 2011 to 2013 as well as three independent external validation cohorts., Results: There were 1,033 patients in the training cohort; 322 patients in the prospective internal validation cohort; and 493, 282, and 149 patients in the three external validation cohorts, respectively. The score was developed using the following factors: α-fetoprotein level, tumor encapsulation, tumor diameter, hepatitis B e antigen positivity, hepatitis B virus DNA load, tumor number, and gastric fundal/esophageal varicosity. The score differentiated two groups of patients (≤4, >4 points) with distinct long-term prognoses outcomes (median OS, 55.8 vs. 19.6 months; p < .001). The predictive accuracy of the score was greater than the other four commonly used staging systems for HCC., Conclusion: The EHBH-MVI scoring system was more accurate in predicting prognosis in patients with HCC with MVI after R0 LR than the other four commonly used staging systems. The score can be used to supplement these systems., Implications for Practice: Microvascular invasion (MVI) is a major determinant of survival outcomes after curative liver resection for patients with hepatocellular carcinoma (HCC). Currently, there is no scoring system aiming to predict prognosis of patients with HCC and MVI after R0 liver resection (LR). Most of the widely used staging systems for HCC do not use MVI as an independent risk factor, and they cannot be used to predict the prognosis of patients with HCC and MVI after surgery. In this study, a new Eastern Hepatobiliary Surgery Hospital (EHBH) MVI scoring system was established to predict prognosis of patients with HCC and MVI after R0 LR. Based on the results of this study, postoperative adjuvant therapy may be recommended for patients with HCC and MVI with an EHBH-MVI score >4. This score can be used to supplement the currently used HCC classifications to predict postoperative survival outcomes in patients with HCC and MVI., Competing Interests: Disclosures of potential conflicts of interest may be found at the end of this article., (© AlphaMed Press 2019.)

Background: Microvascular invasion (MVI) is a major determinant of survival outcome for hepatocellular carcinoma (HCC). This study aimed to investigate the efficacy of postoperative adjuvant Sorafenib (PA-Sorafenib) in HCC patients with MVI after R0 liver resection (LR)., Methods: The data of patients who underwent R0 LR for HCC with histologically confirmed MVI at the Eastern Hepatobiliary Surgery Hospital were retrospectively analyzed. The survival outcomes for patients who underwent PA-Sorafenib were compared with those who underwent R0 LR alone. Propensity score matching (PSM) analysis was performed., Results: 728 HCC patients had MVI in the resected specimens after R0 resection, with 581 who underwent LR alone and 147 patients who received in additional adjuvant sorafenib. PSM matched 113 patients in each of these two groups. The overall survival (OS) and recurrence free survival (RFS) were significantly better for patients in the PA-sorafenib group (for OS: before PSM, P = 0.003; after PSM, P = 0.007), (for RFS: before PSM, P = 0.029; after PSM, P = 0.001), respectively. Similar results were obtained in patients with BCLC 0-A, BCLC B and Child-Pugh A stages of disease., Conclusions: PA-Sorafenib was associated with significantly better survival outcomes than LR alone for HCC patients with MVI., (Copyright © 2019 International Hepato-Pancreato-Biliary Association Inc. Published by Elsevier Ltd. All rights reserved.)

Background: Microvascular invasion (MVI) predicts poor prognosis in patients with hepatocellular carcinoma (HCC). HCC patients with hypercoagulability are prone to develop thrombosis; however, the relationship between preoperative coagulability state, as reflected by the international normalized ratio (INR) level, and MVI remains unclear., Methods: From January 2009 to December 2012, HCC patients who underwent R0 liver resection (LR) from four cancer centers entered into this study. The overall survival (OS) and recurrence-free survival (RFS) rates were compared using the Kaplan-Meier method and Cox regression analysis., Results: Of the 2509 HCC patients who were included into this study, 1104 were found to have MVI in the resected specimens. These patients were divided into the low (n = 151), normal (n = 796), and high (n = 157) INR subgroups based on the preoperative INR levels. The low INR subgroup had a significantly higher incidence of MVI than the normal or high INR subgroups (61.6% vs. 41.6% vs. 44.6%; p < 0.001). HCC patients with MVI were significantly more likely to have a low preoperative INR level (p < 0.001); the INR level (p < 0.001) was an independent risk factor of OS and RFS. HCC patients with MVI in the low INR subgroup had significantly worse RFS and OS than the normal or high INR subgroups (median RFS 13.5 vs. 20.2 vs. 21.6 months, p < 0.001; median OS 35.5 vs. 59.5 vs. 57.0 months, p < 0.001)., Conclusions: Preoperative hypercoagulability was associated with poor long-term prognosis in HCC patients with MVI after R0 LR.

Background: The benefits of adjuvant transarterial chemoembolization (TACE) in hepatocellular carcinoma (HCC) patients with microvascular invasion (MVI) remain controversial. We compared the efficacy and safety of adjuvant TACE and hepatic resection (HR) alone for HCC patients with MVI., Methods: The PubMed, EMBASE, Cochrane Library, VIP, Wan Fang, and Sino Med databases were systematically searched to compare adjuvant TACE and HR alone for the treatment of HCC with MVI from inception to January 1, 2019. The study outcomes, including overall survival (OS) and disease-free survival (DFS), were extracted independently by two authors., Results: 12 trials involving 2190 patients were evaluated. A meta-analysis of 11 studies suggested that the 1-, 3-, and 5-year overall survival (OS) rates (OR = 0.33, P < 0.001; OR = 0.49, P < 0.001; and OR = 0.59, P < 0.01; respectively), favored adjuvant TACE over HR alone. 11 studies were included in the meta-analysis of DFS, and adjuvant TACE showed better 1-, 3-, and 5-DFS (OR = 0.45, P < 0.001; OR = 0.50, P < 0.001; and OR = 0.58, P < 0.001; respectively) compared to HR alone. Subgroup analysis demonstrated that adjuvant TACE could benefit HCC patients with MVI with tumor diameter >5 cm or multinodular tumors., Conclusion: Adjuvant TACE may improve OS and DFS for HCC patients with MVI compared to HR alone and should be recommended for selected HCC patients with MVI. However, these results need to be validated through further high-quality clinical studies., Lay Summary: The benefits of adjuvant TACE in HCC patients with microvascular invasion remain controversial. Twelve studies involving 2190 patients were include in our meta-analysis. Adjuvant TACE may improve OS and DFS for HCC patients with MVI compared to HR alone and should be recommended for selected HCC patients with MVI., (Copyright © 2019 Elsevier Ltd, BASO ~ The Association for Cancer Surgery, and the European Society of Surgical Oncology. All rights reserved.)

Background: Sepsis is the main cause of death in Stevens-Johnson syndrome (SJS) and toxic epidermal necrolysis (TEN)., Objectives: Our aim was to identify admission risk factors predictive of bacteremia and the accompanying clinical or biochemical markers associated with positive blood cultures., Methods: A retrospective cohort study over a 14-year period (2003-2016) was performed., Results: The study included 176 patients with SJS (n = 59), SJS-TEN overlap (n = 51), and TEN (n = 66). During hospitalization, bacteremia developed in 52 patients (29.5%), who experienced poorer outcomes, including higher intensive care unit admission (P < .0005), longer length of stay (P < .0005), and higher mortality (P < .0005). There were 112 episodes of bacteremia, and isolates included Acinetobacter baumannii (27.7%, n = 31) and Staphylococcus aureus (21.4%, n = 24). On multivariate analysis, clinical factors present at admission that were predictive of bacteremia included hemoglobin ≤10 g/dL (odds ratio [OR] 2.4, confidence interval [CI] 2.2-2.6), existing cardiovascular disease (OR 2.10, CI 2.0-2.3), and body surface area involvement ≥10% (OR 14.3, CI 13.4-15.2). The Bacteremia Risk Score was constructed with good calibration. Hypothermia (P = .03) and procalcitonin ≥1 μg/L (P = .02) concurrent with blood culture sampling were predictive of blood culture positivity., Limitations: This is a retrospective study performed in a reference center., Conclusion: Hemoglobin ≤10 g/dL, cardiovascular disease, and body surface area involvement ≥10% on admission were risk factors for bacteremia. Hypothermia and elevated procalcitonin are useful markers for the timely detection of bacteremia., (Copyright © 2019 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.)

Background: The effect of microvascular invasion (MVI) on the postoperative long-term prognosis of solitary small hepatocellular carcinoma remains controversial. We compared the long-term outcomes of MVI-positive and MVI-negative groups of patients with solitary small hepatocellular carcinoma., Methods: The PubMed, EMBASE, Cochrane Library, VIP, Wan Fang, and Sino Med databases were systematically searched to compare the long-term outcomes of MVI-positive and MVI-negative groups of patients with solitary small hepatocellular carcinoma from inception to November 1, 2018. The study outcomes, including overall survival (OS) and disease-free survival (DFS), were extracted independently by two authors., Results: Fourteen studies involving 3033 patients were evaluated. A meta-analysis of all 14 studies suggested that the OS of the MVI-positive group was significantly worse than that of the MVI-negative group (HR = 2.39, 95% CI = 2.02-2.84, I 2  = 22.8%; P < 0.001). Twelve studies were included in the meta-analysis of DFS, and MVI showed a worse prognosis (HR = 1.79, 95% CI = 1.59-2.02, I 2  = 25.3%; P < 0.001). Subgroup analysis demonstrated that MVI still showed a negative effect on the long-term OS and DFS of patients with solitary small HCC measuring up to 2 cm, 3 cm, or 5 cm., Conclusion: Microvascular invasion was a risk factor for poorer prognosis for solitary small hepatocellular carcinoma., (Copyright © 2019 International Hepato-Pancreato-Biliary Association Inc. Published by Elsevier Ltd. All rights reserved.)

Portal vein tumor thrombus (PVTT) is a significant poor prognostic factor for hepatocellular carcinoma (HCC). Patients with PVTT limited to a first-order branch of the main portal vein (MPV) or above could benefit from negative margin (R0) liver resection (LR). An Eastern Hepatobiliary Surgery Hospital (EHBH)/PVTT scoring system was established to predict the prognosis of HCC patients with PVTT after R0 LR and guide selection of subgroups of patients that could benefit from LR. HCC patients with PVTT limited to a first-order branch of the MPV or above who underwent R0 LR as an initial therapy were included. The EHBH-PVTT score was developed from a retrospective cohort in the training cohort using a Cox regression model and validated in a prospective internal validation cohort and three external validation cohorts. There were 432 patients in the training cohort, 285 in the prospective internal validation cohort, and 286, 189, and 135 in three external validation cohorts, respectively. The score was calculated using total bilirubin, α-fetoprotein (AFP), tumor diameter, and satellite lesions. The EHBH-PVTT score differentiated two groups of patients (≤/>3 points) with distinct long-term prognoses (median overall survival [OS], 17.0 vs. 7.9 months; P < 0.001). Predictive accuracy, as determined by the area under the time-dependent receiver operating characteristic curves (AUCs; 0.680-0.721), was greater than that of the other commonly used staging systems for HCC and PVTT. Conclusion: The EHBH-PVTT scoring system was more accurate in predicting the prognosis of HCC patients with PVTT than other staging systems after LR. It selected appropriate HCC patients with PVTT limited to a first-order branch of the MPV or above for LR. It can be used to supplement the other HCC staging systems., (© 2018 by the American Association for the Study of Liver Diseases.)

Background: Vascular invasion is a major determinant of survival outcomes after curative resection for hepatocellular carcinoma (HCC) patients. This study was designed to investigate the efficacy of postoperative adjuvant transarterial chemoembolization (PA-TACE) in patients with HCC with hepatic vein tumor thrombus (HVTT)., Methods: Data from patients who underwent LR for HCC with HVTT at the Eastern Hepatobiliary Surgery Hospital were retrospectively analyzed. The survival outcomes for patients who underwent PA-TACE after LR were compared with those who underwent LR alone. Propensity score matching (PSM) analysis was performed to match patients in a ratio of 1:1., Results: All included 319 patients who underwent LR for HCC with HVTT, 134 underwent LR alone (the LR group), and 185 patients underwent in adjuvant TACE (the PA-TACE group). PSM matched 107 patients in two groups. The overall survival (OS) and recurrence-free survival (RFS) were significantly better for patients in the PA-TACE group than the LR group (for OS: before PSM, P < 0.001; after PSM, P = 0.004; for RFS: before PSM, P < 0.001; after PSM, P = 0.013), respectively. On subgroup analysis, equivalent acceptable results were obtained in patients with peripheral HVTT (pHVTT) and major HVTT (mHVTT). However, PA-TACE resulted in no survival benefits for patients when the HVTT had extended to the inferior vena cava (IVCTT)., Conclusions: PA-TACE was associated with significantly better survival outcomes than LR alone for patients with HCC and HVTT (pHVTT and mHVTT). There was no survival benefits in patients whose HVTT had extended to form IVCTT.

Aim: Because of the rarity of hepatic vein tumor thrombus (HVTT) in patients with hepatocellular carcinoma (HCC), little is known about HVTT. Thus, the survival benefit of liver resection (LR) versus transcatheter arterial chemoembolization (TACE) for HCC patients with HVTT or inferior vena cava tumor thrombus (IVCTT) remains controversial. We aimed to explore the survival benefits of LR versus TACE for the treatment of these patients., Methods: From 2012 to 2016, a total of 276 patients with HVTT or IVCTT who underwent liver resection or TACE were enrolled in this study. Patients in the LR group were matched at a 1:1 ratio with patients treated with TACE as an initial treatment (TACE group). Clinical characteristics, overall survival, and disease-free survival were analyzed., Results: The median survival time in the LR group was 4.7 months longer than that in the TACE group before PSM (19.4 vs. 14.7 months, P = 0.006) and 6.9 months longer than that in the TACE group after PSM (20.9 vs. 14.0 months, P = 0.019). The median disease-free survival time in the LR group was 3.2 months longer than that in the TACE group before PSM (12.3 vs. 9.1 months, P = 0.038) and 5.8 months longer than that in the TACE group after PSM (13.0 vs. 7.2 months, P = 0.011)., Conclusion: Liver resection provides a good prognosis for HCC patients with HVTT or IVCTT compared with patients undergoing TACE, and coexistence with portal vein tumor thrombus is the most important factor related to survival., (© 2018 The Japan Society of Hepatology.)

The S100 protein family is widely involved in the pathological process of various types of cancer. However, the prognostic value of the S100 protein family member S100A12 in hepatocellular carcinoma (HCC) remains unknown. A total of 139 patients undergoing curative surgical resection for HCC from December 2005 to June 2006 were investigated. Immunohistochemistry of S100A12 tissue was performed and expression was classified according to the total positive staining area. Co-expression of S100A12 with cluster of differentiation (CD)11B, CD15 and CD68 was evaluated using immunofluorescence. Associations between S100A12 expression and preoperative clinicopathological parameters were assessed using a χ 2 test or independent sample Student's t-test. Kaplan-Meier estimator survival analysis and multivariate Cox's proportional hazard regression model were used to evaluate the prognostic value of S100A12 expression. The expression of S100A12 was restricted exclusively to stroma cells, primarily to myeloid-derived immune cells, CD15-positive neutrophils and CD68-positive macrophages in particular. A total positive staining area of 1,600 µm 2 was selected as the threshold between high and low S100A12 expression. There was a statistically significant association between intratumoral S100A12 expression and tumor differentiation (P=0.010). High expression of S100A12 on intratumoral stroma cells was an independent prognostic factor for the overall (P=0.002) and disease-free survival (P=0.007) rates of HCC following curative surgical resection. No significant association was identified between peritumoral S100A12 expression and HCC prognosis. The results of the present study demonstrated that high expression of S100A12 on intratumoral stroma cells is associated with poor HCC prognosis following curative resection, which may serve as a potential target for an adjuvant therapy.

Platelet endothelial cell adhesion molecule-1 (PECAM-1 or CD31) is a well-known marker of endothelial cells and a key factor for adhesion and accumulation of platelets. CD31 plays roles in cell proliferation, apoptosis, migration, and cellular immunity. CD31 is also expressed on tumor cells, such as breast cancer cells and non-Hodgkin's lymphomas, and contributes to tumor cell invasion. Here, our experiments show that CD31 promotes metastasis by inducing the epithelial-mesenchymal transition in hepatocellular carcinoma by up-regulating integrin β1 via the FAK/Akt signaling pathway., (Copyright © 2018 Elsevier B.V. All rights reserved.)

IL-8 over-expression could enhance cancer metastasis. In present study, berberine hydrochloride (BER) triggered proliferative inhibition and G2/M arrest in AGS cells, down-regulated protein expression of cyclin B1, Bcl-2, up-regulated expression of p21, p53 and cleaved caspase 3, but showed no effect on protein expression of CHOP, Bip, and caspase 4. BER could down-regulate the enhanced IL-8 expression through down-regulating ERK1/2 and p38 MAPK over-activation induced by SN 38. The increased IL-8 mediated adhesive ability of AGS cells to HUVECs induced by SN 38, could be reduced by BER. Thus, BER could reduce the side-effect of SN 38 in clinic.

Rnd1, a member of Rho GTPases, was found to be downregulated in human malignancies and downregulation of Rnd1 promotes tumor invasion via various mechanisms. However, the role of Rnd1 in hepatocellular carcinoma (HCC) progression remains unclear. In this study, our results demonstrated that Rnd1 was downregulated in HCC cells and in human HCC tissues. Low expression of Rnd1 was associated with aggressive clinic-pathologic characteristics, such as vascular invasion, and poor prognosis in patients who underwent curative surgery for HCC. Overexpression of Rnd1-suppressed cell growth, migration, invasion, and EMT processes in vitro and in vivo. Furthermore, Rnd1 blocked HCC progression by restricting EMT process through inhibition of the Raf/MEK/ERK cascade, and this was correlated with a reduction in RhoA activity. Combination of Rnd1 overexpression with sorafenib, a Raf signaling pathway inhibitor, showed a more potent inhibition on HCC metastasis. Moreover, epigenetic inhibitors (5-Aza and SAHA) increased the expression of Rnd1, and potentiated sorafenib-induced toxicity in HCC cells. In a conclusion, Rnd1-suppressed EMT-mediated metastasis of HCC by reducing the activity of the RhoA/Raf/MEK/ERK signaling pathway, functioning as a favorable anti-metastasis target for HCC patients. Rnd1 overexpression in combination with sorafenib may result in enhanced anti-metastasis efficacy in HCC.

Colony-stimulating factor-1 (CSF-1) and its receptor, CSF-1R, regulate the differentiation and function of macrophages and play an important role in macrophage infiltration in the context of hepatocellular carcinoma. The therapeutic effects of CSF-1R blockade in hepatocellular carcinoma remain unclear. In this study, we found that CSF-1R blockade by PLX3397, a competitive inhibitor with high specificity for CSF-1R tyrosine kinase, significantly delayed tumor growth in mouse models. PLX3397 inhibited the proliferation of macrophages in vitro , but intratumoral macrophage infiltration was not decreased by PLX3397 in vivo Gene expression profiling of tumor-associated macrophages (TAM) showed that TAMs from the PLX3397-treated tumors were polarized toward an M1-like phenotype compared with those from vehicle-treated tumors. In addition, PLX3397 treatment increased CD8 + T-cell infiltration, whereas CD4 + T-cell infiltration was decreased. Further study revealed that tumor cell-derived CSF-2 protected TAMs from being depleted by PLX3397. In conclusion, CSF-1R blockade delayed tumor growth by shifting the polarization rather than the depletion of TAMs. CSF-1R blockade warrants further investigation in the treatment of hepatocellular carcinoma. Mol Cancer Ther; 16(8); 1544-54. ©2017 AACR ., (©2017 American Association for Cancer Research.)

M2-polarized (alternatively activated) macrophages play an important role in the progression of hepatocellular carcinoma (HCC). Allograft inflammatory factor 1 (AIF1) is overexpressed in M2-polarized macrophages. This study explored the role of AIF1 in tumor-associated macrophages in HCC. Macrophages were stimulated with colony-stimulating factor 1 (CSF1) to characterize the regulatory pathway of AIF1 in macrophages. The chromatin immunoprecipitation and luciferase reporter gene assay were conducted to examine transcription factors associated with AIF1 expression. AIF1 was down or upregulated, and the effects on tumor progression were evaluated by using in vitro and in vivo co-culture systems. A cytokine array was performed to screen the downstream functional components of AIF1. Tumor tissue from 206 patients with HCC were used to explore the clinical significance of AIF1. AIF1 induced a M2-like phenotype of macrophages. By facilitating the binding of c-Jun to the promoter of AIF1, CSF1 secreted from hepatoma cells increased AIF1 expression through the CSF1R-MEK1/2-Erk1/2-c-Jun axis. AIF1 expressed in macrophages promoted the migration of hepatoma cells in co-culture system of RAW264.7 and Hepa1-6 and tumor growth in an animal model. The cytokine array showed that CXCL16 was increased in RAW264.7 cells with overexpressed AIF1, leading to enhanced tumor cell migration. In human HCC tissue, AIF1-positive macrophages in the adjacent microenvironment was associated with microvascular invasion and advanced TNM stages and with patients' overall and disease-free survival ( p = 0.002 for both). AIF1 expression in macrophages plays a pivotal role in the interaction between macrophages and hepatoma cells.

Tumor-associated endothelial cells (TEC) directly facilitate tumor progression, but little is known about the mechanisms. We investigated the function of CD109 in TEC and its clinical significance in hepatocellular carcinoma (HCC). The correlation between CD109 expressed on tumor vessels and the prognosis after surgical resection of HCC was studied. The effect of human umbilical vein endothelial cells (HUVEC) with different CD109 expression on hepatoma cell proliferation, migration, and invasion was compared in co-culture assay. Associated key factors were screened by human cytokine antibody array and validated thereafter. HUVEC with different CD109 expression were co-implanted with HCCLM3 or HepG2 cells in nude mice to investigate the effect of CD109 expression on tumor growth and metastasis. Reduced expression of CD109 on tumor vessels was associated with large tumor size, microvascular invasion, and advanced tumor stage. CD109 was an independent risk factor for disease-free survival (P = 0.001) after curative resection of HCC. CD109 knockdown in HUVEC promoted hepatoma cell proliferation, migration, and invasion. Interleukin-8 (IL-8) was a key tumor-promoting factor secreted from CD109 knockdown HUVEC. CD109 knockdown upregulated IL-8 expression through activation of TGF-β/Akt/NF-κB pathway in HUVEC. Co-implantation with CD109 knockdown HUVEC accelerated tumor growth and metastasis in mice models. In conclusion, CD109 expression on tumor vessels is a potential prognostic marker for HCC, and its reduced expression on TEC promoted tumor progression by paracrine IL-8., Competing Interests: Conflicts of Interest: None declared.

Background: Our previous study reported that microRNA-26a (miR-26a) inhibited tumor progression by inhibiting tumor angiogenesis and intratumoral macrophage infiltration in hepatocellular carcinoma (HCC). The direct roles of miR-26a on tumor cell invasion remain poorly understood. In this study, we aim to explore the mechanism of miR-26a in modulating epithelial-mesenchymal transition (EMT) in HCC., Methods: In vitro cell morphology and cell migration were compared between the hepatoma cell lines HCCLM3 and HepG2, which were established in the previous study. Overexpression and down-regulation of miR-26a were induced in these cell lines, and Western blot and immunofluorescence assays were used to detect the expression of EMT markers. Xenograft nude mouse models were used to observe tumor growth and pulmonary metastasis. Immunohistochemical assays were conducted to study the relationships between miR-26a expression and enhancer of zeste homolog 2 (EZH2) and E-cadherin expression in human HCC samples., Results: Down-regulation of miR-26a in HCCLM3 and HepG2 cells resulted in an EMT-like cell morphology and high motility in vitro and increased in tumor growth and pulmonary metastasis in vivo. Through down-regulation of EZH2 expression and up-regulation of E-cadherin expression, miR-26a inhibited the EMT process in vitro and in vivo. Luciferase reporter assay showed that miR-26a directly interacted with EZH2 messenger RNA (mRNA). Furthermore, the expression of miR-26a was positively correlated with E-cadherin expression and inversely correlated with EZH2 expression in human HCC tissue., Conclusions: miR-26a inhibited the EMT process in HCC by down-regulating EZH2 expression.

Robo1 is a member of the Robo immunoglobulin superfamily of proteins, and it plays an important role in angiogenesis and cancer. In this study, we investigate the role of roundabout 1 (Robo1) in tumor angiogenesis in hepatocellular carcinoma (HCC). Firstly, the relationship between Robo1 expression on tumors and patient's survival and endothelial cells in tumor blood vessels and patient's survival was studied. Secondly, Robo1 was overexpressed or knocked down in human umbilical vein endothelial cells (HUVECs). Cell proliferation, motility, and tube formation were compared in HUVEC with different Robo1 expression. Also, HUVECs with different Robo1 expression were mixed with HCCLM3 and HepG2 hepatoma cells and then implanted in a nude mouse model to examine the effects of Robo1 in endothelial cells on tumor growth and angiogenesis. Cell motility-related molecules were studied to investigate the potential mechanism how Robo1 promoted tumor angiogenesis in HCC. The disease-free survival of the patients with high Robo1 expression in tumoral endothelial cells was significantly shorter than that of those with low expression (P = 0.021). Overexpression of Robo1 in HUVECs resulted in increased proliferation, motility, and tube formation in vitro. In the implanted mixture of tumor cells and HUVECs with an increased Robo1 expression, tumor growth and microvessel density were enhanced compared with controls. Robo1 promoted cell division cycle 42 (Cdc42) expression in HUVECs, and a distorted actin cytoskeleton in HUVECs was observed when Robo1 expression was suppressed. In conclusion, Robo1 promoted angiogenesis in HCC mediated by Cdc42.

Background: microRNAs (miRNAs) have been reported to modulate macrophage colony-stimulating factor (M-CSF) and macrophages. The aim of this study was to find whether miR-26a can suppress M-CSF expression and the recruitment of macrophages., Methods: Hepatocellular carcinoma (HCC) cell lines with decreased or increased expression of miR-26a were established in a previous study. M-CSF expression by tumor cells was measured by enzyme-linked immunosorbent assay, and cell migration assays were used to explore the effect of HCC cell lines on macrophage recruitment in vitro. Real-time PCR measured a panel of mRNAs expressed by macrophages. Xenograft models were used to observe tumor growth. Immunohistochemistry was conducted to study the relation between miR-26a expression and M-CSF expression and macrophage recruitment in patients with HCC., Results: Ectopic expression of miR-26a reduced expression of M-CSF. The conditioned medium (CM) from HepG2 cells that overexpressed miR-26a reduced the migration ability of THP-1 cells stimulated by phorbol myristate acetate (PMA) increased expression of interleukin (IL)-12b or IL-23 mRNA and decreased expression of chemokine (C-C motif) ligand (CCL)22, CCL17, and IL-10 mRNA, in comparison to the medium from the parental HepG2 cells. These effects could be interrupted by the PI3K/Akt pathway inhibitor LY294002. Ectopic expression of miR-26a in HCC cells suppressed tumor growth, M-CSF expression, and infiltration of macrophages in tumors. Similar results were also found when using HCCLM3 cells. Furthermore, the expression of miR-26a was inversely correlated with M-CSF expression and macrophage infiltration in tumor tissues from patients with HCC., Conclusions: miR-26a expression reduced M-CSF expression and recruitment of macrophages in HCC.

Background: Radiofrequency ablation (RFA) is one of the curative therapies for hepatocellular carcinoma (HCC), however, accelerated progression of residual HCC after incomplete RFA has been reported more frequently. The underlying molecular mechanism of this phenomenon remains to be elucidated. In this study, we used an incomplete RFA orthotopic HCC nude mouse model to study the invasive and metastatic potential of residual cancer as well as the correlated mechanism., Methods: The incomplete RFA orthotopic nude mouse models were established using high metastatic potential HCC cell line HCCLM3 and low metastatic potential HCC cell line HepG2, respectively. The changes in cellular morphology, motility, metastasis and epithelial-mesenchymal transition (EMT), and HCC cell molecular markers after in vitro and in vivo incomplete RFA intervention were observed., Results: Pulmonary and intraperitoneal metastasis were observed in an in vivo study. The underlying pro-invasive mechanism of incomplete RFA appeared to be associated with promoting EMT, including down-regulation of E-cadherin and up-regulation of N-cadherin and vimentin. These results were in accordance with the in vitro response of HCC cells to heat intervention. Further studies demonstrated that β-catenin was a pivotal factor during this course and blocking β-catenin reduced metastasis and EMT phenotype changes in heat-treated HCCLM3 cells in vitro., Conclusion: Incomplete RFA enhanced the invasive and metastatic potential of residual cancer, accompanying with EMT-like phenotype changes by activating β-catenin signaling in HCCLM3 cells.

Background: Cancer stem cells (CSCs) play a key role in the posthepatectomy recurrence of hepatocellular carcinoma (HCC). CD133+ HCC cells exhibit liver CSC-like properties, and CSC differentiation-inducing therapy may lead these cells to lose their self-renewal ability and may induce terminal differentiation, which may in turn allow their malignant potential to be controlled. Because arsenic trioxide (As₂O₃) increases remission rates and prolongs survival among patients with acute promyelocytic leukemia by inducing differentiation and apoptosis of leukemic cells, we hypothesized that As₂O₃ might also inhibit HCC recurrence and prolong survival time after hepatectomy by inducing differentiation of HCC CSCs., Methods: We evaluated the As₂O₃ induced differentiation of human HCC CSCs and its mechanism in vitro, and we investigated the effects of treatment with As₂O₃ on recurrence rates and median survival in a mouse xenograft model., Results: We found that As₂O₃ induced HCC CSC differentiation by down-regulating the expression of CD133 and some stemness genes, thus inhibiting the cells' self-renewal ability and tumorigenic capacity without inhibiting their proliferation in vitro. In vivo experiments indicated that As₂O₃ decreased recurrence rates after radical resection and prolonged survival in a mouse model. As₂O₃, which shows no apparent toxicity, may induce HCC CSC differentiation by down-regulating the expression of GLI1., Conclusions: We found that As₂O₃ induced HCC CSC differentiation, inhibited recurrence, and prolonged survival after hepatectomy by targeting GLI1expression. Our results suggest that the clinical safety and utility of As₂O₃ should be further evaluated.

Background & Aims: microRNAs (miRNAs) have been reported to regulate angiogenesis by down-regulating the expression of pro-angiogenic or anti-angiogenic factors. The aims of this study were to investigate whether miR-26a inhibited angiogenesis by down-regulating vascular endothelial growth factor A (VEGFA) and its clinical relevance in hepatocellular carcinoma (HCC)., Methods: The expression of miR-26a was modified in HepG2 and HCCLM3 cell lines respectively, and a panel of angiogenic factors was measured by real-time PCR in the cells. A luciferase reporter assay was used to validate the target gene of miR-26a. Specific inhibitors of signal transduction pathway and siRNA approaches were used to explore the regulatory mechanism of miR-26a. Migration and tube forming assays were conducted to show the changes of angiogenesis induced by miR-26a and its target genes. Finally animal studies were used to further validate those findings., Results: Ectopic expression of miR-26a exhibited decreased levels of VEGFA in HepG2 cells. Migration and tube forming of human umbilical vein endothelial cells (HUVECs) were decreased in the conditioned medium from ectopic expression of miR-26a in HepG2 cells compared to control HepG2 cells. The pro-angiogenic effects of the conditioned medium of HepG2 cells on HUVECs were specifically decreased by LY294002, YC-1, and bevacizumab. Integrated analysis disclosed PIK3C2α as a downstream target gene of miR-26a. Ectopic expression of miR-26a suppressed ectopic and orthotopic tumor growth and vascularity in nude mice. The results in HCCLM3 were consistent with those in HepG2. miR-26a expression was inversely correlated with VEGFA expression in HCC patients., Conclusions: miR-26a modulated angiogenesis of HCC through the PIK3C2α/Akt/HIF-1α/VEGFA pathway. The expression of VEGFA was inversely correlated with miR-26a expression in HCC tumors.

Antiangiogenic therapy, specially sorafenib, has become the standard of care for patients with advanced hepatocellular carcinoma (HCC), however, the improvement in survival time is not satisfactory. Previous studies have found that, in some circumstances, antiangiogenic therapy promoted tumor metastasis and the mechanistic studies were mainly focus on cancer-cell-autonomous manners. In two experimental metastasis models with tail-vein injection with hepatoma cells and an orthotopic HCC mouse model, we found that pretreatment with two vascular endothelial growth factor receptor (VEGFR) inhibitors, sunitinib and sorafenib, facilitated tumor cell survival in blood stream and promoted lung metastasis from tumors that were subsequently incubated after drug discontinuation, indicating that host response joined into the pro-metastatic effects. An antibody microarray identified that interleukin (IL)-12b was decreased in the peripheral blood of the mice treated with the two VEGFR inhibitors. IL-12b suppression in macrophages and dendritic cells from host organs was found to play a crucial role in treatment-induced metastasis. Supplement with recombinant mouse IL-12b or restoration of IL-12b expression in the host by zoledronic acid, which was previously reported to enhance IL-12 expression in vitro and in vivo, alleviated the metastasis-promoting effects of sunitinib and sorafenib. These studies suggest that host response to VEGFR inhibitors facilitates HCC metastasis and restoration of IL-12b expression could translate into clinical benefits.

Endothelial cells (ECs) are critical for angiogenesis, and microRNAs play important roles in this process. We investigated the regulatory role of microRNAs in ECs of hepatocellular carcinoma (HCC) by examining the microRNA expression profile of human umbilical vein endothelial cells (HUVECs) in the absence or presence of human HCC cells, and identified miR-146a as the most highly upregulated microRNA. Furthermore, we revealed that miR-146a promoted the expression of platelet-derived growth factor receptor α (PDGFRA) in HUVECs, and this process was mediated by BRCA1. Overexpression of PDGFRA in the ECs of HCC tissues was associated with microvascular invasion and predicted a poorer prognosis. These results suggest that miR-146a plays a key role in regulating the angiogenic activity of ECs in HCC through miR-146a-BRCA1-PDGFRA pathway. MiR-146a and PDGFRA may emerge as potential anti-angiogenic targets on ECs for HCC therapy.

Background Aims: We previously demonstrated the pro-metastasis effect of sorafenib in hepatocellular carcinoma (HCC), which is mediated by down-regulation of tumor suppressor HTATIP2. The aim of the present study was to determine whether aspirin minimizes this effect and improves survival., Methods: The effects of sorafenib, aspirin, and combined sorafenib and aspirin were observed in HCCLM3 and HepG2 xenograft nude mice. Tumor growth, intrahepatic metastasis (IHM), lung metastasis, and survival were assessed. Polymerase chain reaction (PCR) array, real-time (RT)-PCR, and Western blotting were used to examine gene expression. The anti-invasion and anti-metastasis effects of aspirin were studied in HTATIP2-knockdown and HTATIP2-overexpressing HCC cell lines. The molecular mechanism of HTATIP2 regulation by aspirin was explored., Results: Aspirin suppressed the pro-invasion and pro-metastasis effects of sorafenib in HCC and up-regulated HTATIP2 expression. Aspirin did not inhibit the proliferation of HCC cells, but it decreased the invasiveness of HCC with lower expression of HTATIP2 and increased expression of a set of markers, indicating a mesenchymal-to-epithelial transition in tumor cells. The up-regulation of HTATPI2 expression by aspirin is most likely mediated through inhibition of cyclooxygenase (COX) 2 expression., Conclusions: Aspirin minimized the pro-metastasis effect of sorafenib by up-regulating the tumor suppressor HTATIP2; this mechanism is mediated through inhibition of COX2.

Sorafenib, a multi-tyrosine kinase inhibitor, is a standard treatment for advanced hepatocellular carcinoma (HCC). The present study was undertaken to determine whether the growth and metastasis of HCC were influenced in mice receiving sorafenib prior to implantation with tumors, and to investigate the in-vivo and in-vitro effect of sorafenib on natural killer (NK) cells. In sorafenib-pretreated BALB/c nu/nu mice and C57BL/6 mice, tumor growth was accelerated, mouse survival was decreased, and lung metastasis was increased. However, the depletion of NK1.1(+) cells in C57BL/6 mice eliminated sorafenib-mediated pro-metastatic effects. Sorafenib significantly reduced the number of NK cells and inhibited reactivity of NK cells against tumor cells, in both tumor-bearing and tumor-free C57BL/6 mice. Sorafenib down-regulated the stimulatory receptor CD69 in NK cells of tumor-bearing mice, but not in tumor-free mice, and inhibited proliferation of NK92-MI cells, which is associated with the blocking of the PI3K/AKT pathway, and inhibited cytotoxicity of NK cells in response to tumor targets, which was due to impaired ERK phosphorylation. These results suggest immunotherapeutic approaches activating NK cells may enhance the therapeutic efficacy of sorafenib in HCC patients.

Background: Promotion of endothelial normalization restores tumor oxygenation and obstructs tumor cells invasion, intravasation, and metastasis. We therefore investigated whether a vasoactive drug, tanshinone IIA, could inhibit metastasis by inducing vascular normalization after palliative resection (PR) of hepatocellular carcinoma (HCC)., Methods: A liver orthotopic double-tumor xenograft model in nude mouse was established by implantation of HCCLM3 (high metastatic potential) and HepG2 tumor cells. After removal of one tumor by PR, the effects of tanshinone IIA administration on metastasis, tumor vascularization, and survival were evaluated. Tube formation was examined in mouse tumor-derived endothelial cells (TECs) treated with tanshinone IIA., Results: PR significantly accelerated residual hepatoma metastases. Tanshinone IIA did not inhibit growth of single-xenotransplanted tumors, but it did reduce the occurrence of metastases. Moreover, it inhibited PR-enhanced metastases and, more importantly, prolonged host survival. Tanshinone IIA alleviated residual tumor hypoxia and suppressed epithelial-mesenchymal transition (EMT) in vivo; however, it did not downregulate hypoxia-inducible factor 1α (HIF-1α) or reverse EMT of tumor cells under hypoxic conditions in vitro. Tanshinone IIA directly strengthened tube formation of TECs, associated with vascular endothelial cell growth factor receptor 1/platelet derived growth factor receptor (VEGFR1/PDGFR) upregulation. Although the microvessel density (MVD) of residual tumor tissue increased after PR, the microvessel integrity (MVI) was still low. While tanshinone IIA did not inhibit MVD, it did dramatically increase MVI, leading to vascular normalization., Conclusions: Our results demonstrate that tanshinone IIA can inhibit the enhanced HCC metastasis associated with PR. Inhibition results from promoting VEGFR1/PDGFR-related vascular normalization. This application demonstrates the potential clinical benefit of preventing postsurgical recurrence.