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Mass spectrometry imaging (MSI) of lipids in biological tissues is useful for correlating molecular distribution with pathological results, which could provide useful information for both biological research and disease diagnosis. It is well understood that the lipidome could not be clearly deciphered without tandem mass spectrometry analysis, but this is challenging to achieve in MSI due to the limitation in sample amount at each image spot. Here we develop a multiplexed MS2 imaging (MS2I) method that can provide MS2 images for 10 lipid species or more for each sampling spot, providing spatial structural lipidomic information. Coupling with on‐tissue photochemical derivatization, imaging of 20 phospholipid C=C location isomers is also realized, showing enhanced molecular images with high definition in structure for mouse brain and human liver cancer tissue sections. Spatially mapped t‐distributed stochastic neighbor embedding has also been adopted to visualize the tumor margin with enhancement by structural lipidomic information. [ABSTRACT FROM AUTHOR]

A majority of hepatocellular carcinomas (HCCs) combine with liver cirrhosis. The cirrhotic liver has been implicated in interfering with the effects of HCC‐targeted drugs, including sorafenib. Alterations in the tumor microenvironment of the cirrhotic liver include both biochemical and biomechanical factors. In this study, we induced sorafenib resistance in HCC cells. We observed changes in cell morphology, cytoskeletal architecture, and cellular stiffness in these sorafenib‐resistant cells, resembling those adapted to stiffer substrates. To examine the contribution of mechanical factors in HCC cell growth and drug resistance, we used an in vitro cell culture system with adjustable stiffness mimicking the normal or cirrhotic liver tissues. We identified that mechanical adaptation conferred HCC cells with increased motility and sorafenib resistance. We further reported the mechanism underlying the involvement of the transcription coactivator YAP. Our results underline the important role of mechanical factors in the interaction between tumor cells and their microenvironment. Our work described an unexpected and very surprising connection between the drug resistance of hepatocellular carcinoma (HCC) and cellular mechanotransduction. We pointed out the critical role of environmental mechanics in targeted therapy for HCC. [ABSTRACT FROM AUTHOR]

14‐3‐3ζ, a phosphopeptide‐binding molecule, is reportedly overexpressed in the cancerous tissues of patients with hepatocellular carcinoma (HCC). Hepatitis B virus (HBV) protein X (HBx) draws intensive attention in HBV‐related HCC because it not only regulates HBV replication, but also promotes carcinogenesis by interacting with various tumor or antitumor molecules. This study is performed to investigate whether and how 14‐3‐3ζ interacts with HBx. The coimmunoprecipitation (Co‐IP) results showed that 14‐3‐3ζ bond to HBx in HBV‐infected Hep3B HCC cells and CSQT‐2 portal vein tumor thrombosis (PVTT) cells. By performing Co‐IP assay in HBV‐free Huh7 cells expressing wild‐type HBx, mutant HBx‐S31A, or HBx‐S31D (serine31 was mutated into alanine31 or aspartic acid31), we found that the phosphorylated serine31 with its near amino acid residues constituted a RPLphosphoS31GP (R, arginine; P, proline; L, leucine; S, serine; G, glycine) motif in HBx for 14‐3‐3ζ docking. This 14‐3‐3ζ‐HBx interaction was partly impaired when Akt signaling transduction was blocked by LY294002. Furthermore, 14‐3‐3ζ silencing augmented HBx ubiquitination and decreased its expression in cancer cells and xenograft tumor. The migratory and invasive abilities of CSQT‐2 cells were inhibited upon 14‐3‐3ζ silencing, whereas partly restored by HBx overexpression. Additionally, 14‐3‐3ζ positively correlated with HBx to be overexpressed in the primary HCC tissues (r = 0.344) and metastatic PVTT (r = 0.348). In summary, findings of this study reveal a novel 14‐3‐3ζ‐HBx interaction in HCC cells and suggest 14‐3‐3ζ as a candidate target for treating HBV‐related HCC. 14‐3‐3ζ binds to HBx by docking to the RPLphosphoS31GP motif of HBx. The knockdown of 14‐3‐3ζ downregulates HBx expression in HBV‐infected HCC cells via inducing HBx ubiquitylation and degradation. [ABSTRACT FROM AUTHOR]

Background/Aims: Hepatocellular carcinoma (HCC) is a lethal disease with nearly equal morbidity and mortality. Thus, the discovery and application of more useful predictive biomarkers for improving therapeutic effects and prediction of clinical outcomes is of crucial significance. Methods: A total of 475 HBV-related HCC patients were enrolled. Ataxin 7 (ATXN7) single nucleotide polymorphisms (SNPs) were genotyped by Sanger DNA sequencing after PCR amplification. The associations between ATXN7 SNPs and mRNA expression with the prognosis of HBV-related HCC were analyzed. Results: In all, rs3774729 was significantly associated with overall survival (OS) of HBV-related HCC (P = 0.013, HR = 0.66, 95% CI: 0.48- 0.94). And patients with the AA genotype and a high level of serum alpha fetoprotein (AFP) had significantly worse OS when compared to patients with AG/GG genotypes and a low level of AFP (adjusted P = 0.007, adjusted HR = 1.83, 95% CI = 1.18-2.82). Furthermore, low expression of ATXN7 was significantly associated with poor recurrence-free survival (RFS) and OS (P = 0.007, HR = 2.38, 95% CI = 1.27-4.45 and P = 0.025, HR = 1.75, 95% CI = 1.18-2.62). Conclusion: ATXN7 may be a potential predictor of post-operative prognosis of HBV-related HCC. [ABSTRACT FROM AUTHOR]

Background: To understand the molecular mechanisms of caveolin-1 down regulation by hepatitis B virus X protein (HBx). Methods: The DNA methylation status of the caveolin-1 promoter was examined by nested methylation-specific PCR of 33 hepatitis B virus (HBV)-infected hepatocellular carcinoma (HCC) samples. The SMMC-7721 hepatoma cell line was transfected with a recombinant HBx adenoviral vector, and the effects of HBx protein on caveolin-1expression and promoter methylation were examined and confirmed by sequencing. A reporter gene containing the caveolin-1 promoter region was constructed, and the effects of HBx on the transcriptional activity of the promoter were also studied. Results: Methylation of the caveolin-1 promoter was detected in 84.8% (28/33) of HBV-infected HCC samples. Expression of caveolin-1 was significantly downregulated (P = 0.022), and multiple CpG sites in the promoter region of caveolin-1 were methylated in SMMC-7721 cells after HBx transfection. Transfected HBx significantly suppressed caveolin-1 promoter activity (P = 0.001). Conclusions: HBx protein induces methylation of the caveolin-1 promoter region and suppresses its expression. [ABSTRACT FROM AUTHOR]

Abstract: Viral cirrhotic primary liver cancer (VCPLC) is characterized by intra-hepatic metastasis (IHM) at early stage. By contrast, simple primary liver cancer without cirrhosis (SPLC) has high extra-hepatic metastasis (EHM) rate. Historically, studies on mechanism of metastasis of PLC mainly focused on biological behaviors of cancer cells, however tumor microenvironment is always neglected, especially blood flow dynamics and Kupffer cells. In SPLC, few damages of hepatic lobules occur, to the contrary, VCPLC is characterized by obvious hepatic fibrosis which results in varieties of blood flow disturbances, such as central lobular vein obstruction, hepatic artery-portal vein fistula (HAPVF), and portal vein countercurrent (PVC). Traditionally, Kupffer cells in liver are regarded as powerful defenses against PLC and metastasis lesions in liver, however, recent studies show under long term ischemic–hypoxic stress, Kupffer cells can secrete varieties of pro-inflammatory mediators to promote fibrosis or even cancerogenesis. Tumor associated macrophages (TAMs) are a group of macrophages living in ischemic tumor lesions which can promote tumor progression and metastasis. Based on the totally different metastasis routes and significantly different tumor microenvironment of VCPLC and SPLC, we hypothesize: (1) unlike the normal Kupffer cells in non-cirrhosis liver, Kupffer cells of VCPLC, a group of dominant hepatic resident macrophages living in extremely cirrhotic ischemic–hypoxic sinusoids, might not act as fighters against cancer cells, what is worse, ischemic–hypoxic situation might incite Kupffer cells to secrete cytokines and chemokines, just like TAMs, to promote tumor growth and metastasis; (2) the blood flow disturbances, not the behaviors of cancer cells, are the main force to determine the different metastasis preferences. In VCPLC, central lobular vein is mechanically compressed by cirrhosis nodes, so cancer cells cannot go extra-hepatically, instead, the hepatic artery-portal vein fistula (HAPVF) and portal vein countercurrent (PVC) load cancer cells intra-hepatically. In SPLC, cancer cells can run directly extra-hepatically through central lobular vein fluently. In short, the different blood flow dynamics between these two PLC is the predominant factor to determine the preferences of metastasis route. In VCPLC, the Kupffer cells should not be the powerful defenses against tumor and metastasis lesions, instead, they should be tamed to be some TAMs by long term ischemic–hypoxic stress and tumor derived factors to play important roles in tumorigenesis or even metastasis, so Kupffer cells can be a new target for preventing hepatocancerogenesis and metastasis. [Copyright &y& Elsevier]

The purpose of the study is to evaluate the protective effects of Verapamil on hepatic ischemia–reperfusion injury in cirrhotic patients with HCC. A controlled study was performed on 86 consecutive patients with cirrhosis who underwent liver resection for suffering HCC. Thirty-nine patients were injected with Verapamil through the mesenteric vein before Pringle's maneuver to alleviate hepatic ischemia–reperfusion injury, while 47 patients were not. The serum AST, ALT, and total bilirubin on days 1 and 3 were markedly lower in the Verapamil group than that in the control group postoperatively (AST, P < 0.001 and P < 0.001; ALT, P < 0.001 and P = 0.042; TB, P = 0.037 and 0.049; respectively). The serum hyluronic acid on day 1 was also significantly lower in Verapamil group than that in the control group ( P < 0.01), but no marked difference on days 3 and 7. Meanwhile, the levels of IL6 on days 1 and 3 were markedly lower in the Verapamil group than that in control group ( P < 0.05 and P < 0.01, respectively. The new Clavien's classification of surgical complications showed marked severity in the control group than that in the Verapamil group ( P = 0.021), especially in the grade I level ( P = 0.045). Verapamil may serve as protective agent for hepatic ischemia–reperfusion injury in cirrhotic patients with HCC. Therefore, Verapamil can be used for treatment of patients who are to undergo hepatectomy with occlusion of hepatic blood flow. [ABSTRACT FROM AUTHOR]

Detection of hepatocellular carcinoma circulating tumor cells performed with conventional strategies, is significantly limited due to inherently heterogeneous and dynamic expression of EpCAM, as well as degradation of cytokeratins during epithelial-to-mesenchymal transition, which inevitably lead to non-negligible false negative detection of such "uncapturable and invisible" CTCs. A novel SE-iFISH strategy, improved for detection of HCC CTCs in this study, was applied to comprehensively detect, in situ phenotypically and karyotypically characterize hepatocellular and cholangiocarcinoma CTCs (CD45-/CD31-) in patients subjected to surgical resection. Clinical significance of diverse subtypes of CTC was systematically investigated. Existence of small cell size CTCs (≤5 μm of WBCs) with cytogenetic abnormality of aneuploid chromosome 8, which constituted majority of the detected CTCs in HCC patients, was demonstrated for the first time. The stemness marker EpCAM+ aneuploid circulating tumor stem cells (CTSCs), and EpCAM- small CTCs with trisomy 8, promote tumor growth. Postsurgical quantity of small triploid CTCs (≥5 cells/6 ml blood), multiploid (≥pentasomy 8) CTSCs or CTM (either one ≥ 1) significantly correlated to HCC patients' poor prognosis, indicating that detection of those specific subtypes of CTCs and CTSCs in post-operative patients help predict neoplasm recurrence. [ABSTRACT FROM AUTHOR]

Background The aim of this study was to determine the effect of anatomic resection (AR) versus non-anatomic resection (NAR) on recurrence rates in patients with hepatocellular carcinoma (HCC). Methods Eligible patients were randomized to AR or NAR from January 2006 to July 2007 at a single center. The primary outcome was the 2-year recurrence-free survival (RFS). Secondary outcomes were postoperative complications, time to first recurrence, 1-, 3-, and 5-year RFS, and overall survival (OS). Results Fifty-three (51%) and 52 (50%) patients underwent NAR and AR, respectively. A larger proportion of patients achieved margins ≥20 mm in the AR group (52% vs. 30%; P = 0.023). Complications (blood loss, transfusion requirement, and hospital stay) were similar between the two groups. Median follow-up was 33 (range, 2–77) months. Incidence of local recurrence at 2 years was 30% and 59% in the AR and NAR groups, respectively. Median time to first local recurrence in the AR group was significantly longer than in the NAR group (53 vs. 10 months, P = 0.010). There was no difference in overall RFS between the two groups (P = 0.290). Discussion AR decreased the 2-year local recurrence rate and increased the time to first local recurrence compared to NAR in patients with HCC. [ABSTRACT FROM AUTHOR]

Long non-coding RNAs (lncRNAs) had been proved to be pivotal regulators in carcinogenesis. On the basis of competitive endogenous RNAs (ceRNAs) system, lncRNAs significantly expanded their regulating networks. In our research, we aimed to figure out the exact role of lncRNA HNF1A-AS1 in the pathogenesis of hepatocellular carcinoma (HCC), in a ceRNA-dependent way. First, we revealed: HNF1A-AS1 was frequently overexpressed in HCC tissues and cell lines and its relative high expression was closely related to larger tumor size, multiple tumor lesions, poor differentiation and advanced TNM stage. Then we found: HNF1A-AS1 functioned as an oncogene in tumor growth and apoptosis through sponging tumor-suppressive hsa-miR-30b-5p (miR-30b) and de-repressing Bcl-2. Further experiments identified: HNF1A-AS1-miR-30b axis significantly promoted autophagy under starvation and ATG5 was first proved to be a target of miR-30b. In summary, we identified HNF1A-AS1-miR-30b axis as a key regulator in hepatocarcinogenesis, which may be promising biomarkers and therapeutic targets in the future. [ABSTRACT FROM AUTHOR]

Background & Aims: The aim of this study was to compare the efficacy of radiofrequency ablation (RFA) with surgical resection (RES) in the treatment of small hepatocellular carcinoma (HCC). Methods: A total of 168 patients with small HCC with nodular diameters of less than 4cm and up to two nodules were randomly divided into RES (n=84) and RFA groups (n=84). Outcomes were carefully monitored and evaluated during the 3-year follow-up period. Results: The 1-, 2-, and 3-year survival rates for the RES and RFA groups were 96.0%, 87.6%, 74.8% and 93.1%, 83.1%, 67.2%, respectively. The corresponding recurrence-free survival rates for the two groups were 90.6%, 76.7%, 61.1% and 86.2%, 66.6%, 49.6%, respectively. There were no statistically significant differences between the two groups in overall survival rate (p =0.342) or recurrence-free survival rate (p =0.122). Multivariate analysis demonstrated that the independent risk factors associated with survival were multiple occurrences of tumors at different hepatic locations (relative risk of 2.696; 95% CI: 1.189–6.117; p =0.018) and preoperative indocyanine green retention rate at 15min (ICG-15) (relative risk of 3.853; 95% CI: 1.647–9.015; p =0.002). Conclusions: In patients with small hepatocellular carcinomas, percutaneous RFA may provide therapeutic effects similar to those of RES. However, percutaneous RFA is more likely to be incomplete for the treatment of small HCCs located at specific sites of the liver, and open or laparoscopic surgery may be the better choice. [Copyright &y& Elsevier]

Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer-related deaths around the world; Asian countries account for nearly 78% of the roughly 600,000 cases of HCC reported globally each year. Europe, the US, Asian- Pacific nations, South Korea, and Japan have published evidence-based guidelines for the management of HCC. The management of HCC in Japan has achieved remarkable results, which are attributed to a combination of quantitative and qualitative evaluation incorporated in the Japanese guidelines. However, many of the control methods and interventions in current HCC guidelines cannot be implemented in some Asian countries. The majority of HCC patients in Asia still present with advanced HCC and long-term outcomes following treatment are unsatisfactory because of a lack of effective adjuvant and systemic therapies. Asian countries should formulate evidence-based clinical practice guidelines and pay particularly close attention to combining quantitative and qualitative evaluation when drafting and implementing HCC guidelines. The guidelines should also be updated by incorporating new evidence. [ABSTRACT FROM AUTHOR]

Background: Cell migration is a basis for invasion and metastasis of malignant tumors. Receptor tyrosine kinases are recognized as important therapeutic targets in antineoplastic strategies. C-met is a receptor tyrosine kinase highly expressed in human hepatocellular carcinoma (HCC) cell line MHCC97-H. Higher expression of c-met in tumor tissue can lead to scattering, angiogenesis, proliferation, enhanced cell motility, invasion, and eventually, metastasis. To explore the roles of c-met in modulating the motility of cell, we silenced c-met expression in the HCC line MHCC97-H by RNA interference (RNAi). Materials and Methods: For transient expression, c-met-siRNA 1,2 recombinant plasmids were transfected into phoenix A cells. The MHCC97-H cells were cultured in Dulbecco''s modified Eagles''s medium (DMEM) with 10% fetal bovine serum (FBS) to establish MHCC97-H HCC cells stably expressing c-met-siRNA. MHCC97-H cells were treated with the recombinant virus for assay of c-met mRNA and protein, evaluation of growth and invasion of MHCC97-H cells, and identification of hepatitis B virus X (HBX) protein correlation with c-met. Results: After transfection of c-met-siRNA for 48 h, the expression of c-met decreased markedly in MHCC97-H cells; the most effective site of the siRNA target sequence is at the 537 upstream, far from the transcription start. In addition, the proliferation, motility, and invasive ability of MHCC97-H cells were significantly inhibited. Furthermore, we showed that hepatitis B virus (HBV) X protein (HBX) potentiated the activities of the extracellular signal-regulated kinase 1/2 (ERK1/2) in MHCC97-H cells. Treatment with extracellular signal-regulated kinase (ERK) inhibitor (U0126), but not P38 MAPK inhibitor (SB203580) or phosphatidylinositol 3-kinase (PI3K) inhibitor (wortmannin), markedly suppressed the expression of c-met protein in MHCC97-H cells. Conclusion: These results indicate that the over-expression of c-met protein plays an important role in the cell invasion of MHCC97-H, and HBX protein may promote the expression of c-met by ERKs pathway. [Copyright &y& Elsevier]