Xiaowu Li

A randomized controlled trial of radiofrequency ablation and surgical resection in the treatment of small hepatocellular carcinoma

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 4 cm 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 15 min (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.

Pim kinase-dependent inhibition of c-Myc degradation.

Pim kinases are found to be highly expressed in leukemia, lymphoma, prostate and pancreatic cancer. Bitransgenic mice overexpressing either Pim-1 or Pim-2 and c-Myc succumb to pre-B-cell lymphoma at a strikingly accelerated speed. Despite that Pim-1/Pim-2 has long been recognized as a strong synergistic partner with c-Myc in tumorigenesis, the mechanism underlying the synergism is still not well understood. Overexpression of Pim-1/Pim-2 kinase dramatically stabilizes c-Myc in vivo, and the stabilization is partially mediated by phosphorylation of c-Myc by Pim kinase on a novel site, Ser329. We provide evidence that Pim-2 is more efficient in directly phosphorylating c-Myc Ser329 to stabilize c-Myc. In contrast, we find that Pim-1 is more effective in mediating a decrease in c-Myc Thr58 phosphorylation and an increase in c-Myc Ser62 phosphorylation than in phosphorylating Ser329. In either case, through stabilizing c-Myc, Pim-1/Pim-2 kinases enhance the transcriptional activity of c-Myc. Also knocking down either Pim-1 or Pim-2 dramatically decreases the endogenous levels of c-Myc and thus, its transcriptional activity. Finally, coexpression of the Pim kinases and c-Myc enhances the transforming activity of c-Myc as does the phosphomimic mutant of c-Myc on Ser329. We conclude that these findings appear to explain at least in part the mechanism underlying the synergism between the Pim kinases and c-Myc in tumorigenesis.

Hypoxia induces epithelial-mesenchymal transition via activation of SNAI1 by hypoxia-inducible factor -1α in hepatocellular carcinoma

BackgroundHigh invasion and metastasis are the primary factors causing poor prognosis of patients with hepatocellular carcinoma (HCC). However, the molecular mechanisms underlying these biological behaviors have not been completely elucidated. In this study, we investigate the molecular mechanism by which hypoxia promotes HCC invasion and metastasis through inducing epithelial-mesenchymal transition (EMT).MethodsThe expression of EMT markers was analyzed by immunohistochemistry. Effect of hypoxia on induction of EMT and ability of cell migration and invasion were performed. Luciferase reporter system was used for evaluation of Snail regulation by hypoxia-inducible factor -1α (HIF-1α).ResultsWe found that overexpression of HIF-1α was observed in HCC liver tissues and was related to poor prognosis of HCC patients. HIF-1α expression profile was correlated with the expression levels of SNAI1, E-cadherin, N-cadherin and Vimentin. Hypoxia was able to induce EMT and enhance ability of invasion and migration in HCC cells. The same phenomena were also observed in CoCl2-treated cells. The shRNA-mediated HIF-1α suppression abrogated CoCl2-induced EMT and reduced ability of migration and invasion in HCC cells. Luciferase assay showed that HIF-1α transcriptional regulated the expression of SNAI1 based on two hypoxia response elements (HREs) in SNAI1 promoter.ConclusionsWe demonstrated that hypoxia-stabilized HIF1α promoted EMT through increasing SNAI1 transcription in HCC cells. This data provided a potential therapeutic target for HCC treatment.

Interaction between Cancer Cells and Stromal Fibroblasts Is Required for Activation of the uPAR-uPA-MMP-2 Cascade in Pancreatic Cancer Metastasis

Purpose: Interaction between tumor cells and surrounding stromal fibroblast (SF) plays a critical role in tumor growth and invasion. The aim of the study is to determine the role of SF in regulating the invasive behaviors of pancreatic cancer by evaluating the mode of SF activating the urokinase plasminogen activator (uPA)-plasmin-matrix metalloproteinase (MMP)-2 cascade. Experimental Design: The expression patterns of uPA, MMP-2, and uPA receptor (uPAR) in human metastatic pancreatic cancer were analyzed by immunohistochemistry and the roles of SF in activation of the uPA-plasmin-MMP-2 cascade were evaluated by coculturing pancreatic cancer cell lines with SF. Results: uPA expression and fibroblastic uPAR expression were correlated with liver metastasis of human pancreatic cancer. MMP-2 rather than MMP-9 was activated in the metastatic pancreatic cancer. In the in vitro culture system, the coculture of peritumor fibroblasts with metastatic pancreatic cancer BxPc3 cells resulted in activation of MMP-2 and up-regulation of uPAR expression. In this coculture system, the uPA-plasminogen cascade was involved in MMP-2 activation. This activation required a direct interaction between SF and cancer cells. In the coculture system, intergrin α6β1 expression was increased in BxPc3 cells, and blocking the function of integrin α6β1 decreased the activation of uPA and MMP-2. This suggests that interaction between integrins of cancer cells and the uPARs of the SF might be involved in the activation of the uPAR-uPA-MMP-2 cascade. Conclusion: Our results suggest that SF plays a role in promoting pancreatic cancer metastasis via activation of the uPA-plasminogen-MMP-2 cascade.

Fyn requires HnRNPA2B1 and Sam68 to synergistically regulate apoptosis in pancreatic cancer

PURPOSE The Src family kinase Fyn, heterogenous nuclear ribonucleoprotein (HnRNP) A2B1 and Sam68 are thought to be associated with the metastasis of tumors, but their roles in the regulation of apoptosis remain unclear. This study investigated the role of Fyn and its potential relationship with HnRNPA2B1 and Sam68 in the regulation of apoptosis in pancreatic cancer. Experimental design. We examined both the activity of Fyn and the expression of HnRNPA2B1 in human pancreatic cancer tissues and systematically investigated the apoptotic mechanisms induced by Fyn activity using multiple experimental approaches. RESULTS We found that Fyn activity was increased in metastatic pancreatic cancer tissues. In the pancreatic cancer BxPc3 cell line, the inhibition of Fyn activity by kinase-dead Fyn downregulated HnRNPA2B1 expression. Further analysis showed that HnRNPA2B1 expression was associated with pancreatic cancer progression. In BxPc3 cells, HnRNPA2B1 bound to Bcl-x messenger RNA (mRNA), which affected splicing and therefore, the formation of Bcl-x(s). Downregulation of HnRNPA2B1 by RNA interference (RNAi) resulted in the increased formation of the pro-apoptotic Bcl-x(s) and promoted apoptosis of BxPc3 cells. In addition, deactivation of Fyn in BxPc3 cells reduced Sam68 phosphorylation. This resulted in increased binding between Sam68 and Bcl-x mRNA, promoting the formation of the anti-apoptotic Bcl-x(L). The knockdown of Sam68 by RNAi also increased the formation of Bcl-x(L). Finally, HnRNPA2B1 overexpression or Sam68 knockdown could rescue pancreatic cancer cells from apoptosis. CONCLUSION Our results suggest a mechanism by which Fyn requires HnRNPA2B1 and Sam68 to coordinate and regulate apoptosis, thus promoting the proliferation and metastasis of pancreatic cancer.

Ars2 is overexpressed in human cholangiocarcinomas and its depletion increases PTEN and PDCD4 by decreasing microRNA-21†

Due to the lack of effective diagnostic tools, most patients with cholangiocarcinoma (CCA) have no chance of surgical resection. Ars2 is a protein that was reported to be important for microRNA (miR) biogenesis, and its depletion can reduce the levels of several miRs, including miR‐21, which is overexpressed in CCAs. We hypothesized that Ars2 was also present in CCAs and could be an early diagnostic marker. In our experiments, Ars2, PTEN, PDCD4, and miR‐21 were evaluated in 18 CCAs and paired normal tissues. ShArs2, miR‐21 mimics, and Ars2 were transfected into CCA and bile duct epithelial cells either alone or together. Cell proliferation, tumorigenicity analysis and expression changes of Ars2, PTEN, PDCD4, and miR‐21 were evaluated. We found that both Ars2 and miR‐21 were overexpressed, with 95% sensitivity and 100% specificity, and an ROC of 0.995 in distinguishing between CCAs and paired normal tissues by qRT‐PCR. PTEN and PDCD4 were reversed in immunohistochemistry, but no difference was observed using qRT‐PCR. The knockdown of Ars2 in CCA cells decreased the level of miR‐21, inhibited cell proliferation and prevented tumor formation in nude mice. Ars2 knockdown also led to an increase in both PTEN and PDCD4 protein levels. Both proteins decreased when the miR‐21 mimic was con‐transfected. However, the overexpression of Ars2 alone could not get the opposite results. Based on our data, we conclude that Ars2 is overexpressed in human CCA and may be a diagnostic marker. Ars2 depletion increases PTEN and PDCD4 protein levels via the reduction of miR‐21.

Increased liver-infiltrating CD8+FoxP3+ regulatory T cells are associated with tumor stage in hepatocellular carcinoma patients.

Hepatocellular carcinoma (HCC) is the most common primary malignant tumor of the liver, and patients who are diagnosed with this tumor typically have a poor prognosis. The suppressive effects of CD4(+)FoxP3(+) regulatory T cells on antitumor immune response in HCC have been studied in great detail. CD8(+)FoxP3(+) regulatory T cells have recently been detected in tumors; however, the role of CD8(+)FoxP3(+) regulatory T cells in HCC is still unknown. Here, the frequency and phenotype of CD8(+)FoxP3(+) regulatory T cells were analyzed by multicolor flow cytometry in liver of HCC patients and healthy donors. We observed that the percentage of these cells in HCC patients was significantly higher than that observed in healthy control donors (p = 0.0155); their phenotype was close to that of CD4(+) regulatory T cells. Furthermore, we show that CD8(+)FoxP3(+) regulatory T cells are activated and act as effector memory cells (EM, CD45RA(-)CCR7(-)CD27(+/-)CD28(+)). Most importantly, a higher percentage of intrahepatic CD8(+)FoxP3(+) regulatory T cells was found in patients with advanced HCC than in those with early HCC in terms of tumor-node-metastasis (TNM) stage (stage I vs III, p = 0.0007). These data suggest that CD8(+)FoxP3(+) regulatory T cells may contribute to HCC immune escape and disease progression.

Autologous bone marrow-derived mesenchymal stem cell transplantation promotes liver regeneration after portal vein embolization in cirrhotic rats

BACKGROUND Preexisting cirrhosis usually leads to an inadequate and delayed regeneration of the future liver remnant (FLR) after portal vein embolization (PVE). Bone marrow-derived mesenchymal stem cells (BMSC) are promising candidates for therapeutic applications in liver diseases. In this study, the efficacy of autologous BMSCs transplantation to promote FLR regeneration was investigated in a rat cirrhotic model. METHODS Autologous BMSCs were expanded and labeled with PKH26, and then were injected immediately into nonembolized lobes after PVE through portal vein in cirrhotic rat. At 7, 14, and 28 d after this, liver weight and Ki-67 labeling index were measured, and blood analysis was performed. Cirrhotic degree of FLR was assessed by hydroxyproline content assay and histopathology. Gene expressions of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), interleukin-10 (IL-10), and matrix metalloproteinase-9 (MMP-9) were detected with real-time reverse transcriptase-polymerase chain reaction. Distribution and hepatocyte differentiation of BMSCs in FLR were determined by confocal microscopy. RESULTS Autologous BMSCs significantly increased the FLR weight ratio to the total liver and the Ki-67 labeling index, and serum albumin levels were significantly higher and total bilirubin levels were significantly lower in the BMSCs group compared with the controls without BMSCs transplantation 14 and 28 d post-PVE. BMSCs significantly decreased the hydroxyproline content and collagen accumulation, up-regulated the expressions of HGF, IL-10, VEGF, and MMP-9 28 d post-PVE, and expressed hepatocyte-specific markers, such as α-fetoprotein, cytokeratin 18, and albumin in a time-dependent manner in FLR. CONCLUSIONS Autologous BMSCs can differentiate into hepatocyte and promote FLR regeneration after PVE in cirrhotic liver, which may be through improving local microenvironment by decreasing cirrhosis, up-regulating the gene expressions of VEGF, HGF, IL-10, and MMP-9.

Differences in Yes-associated protein and mRNA levels in regenerating liver and hepatocellular carcinoma

Yes-associated protein (YAP) has been identified as an oncoprotein that regulates organ size by modulating proliferative and apoptotic activities. The aim of this study was to measure YAP gene expression and protein levels during rat liver regeneration and in liver cancer, and to explore possible correlations between YAP levels and cancer cell proliferation following partial hepatectomy. We examined YAP expression in rat regenerating liver using real-time PCR, Western blotting and immunohistochemical methods. YAP mRNA and protein levels were measured in human hepatocellular carcinoma. The results demonstrated that YAP protein levels were markedly increased in the regenerating liver (2.5-3 times), while YAP mRNA levels decreased slightly (1.47-2.17 times) (P=0.017). YAP mRNA and protein levels were both higher in hepatocellular carcinoma than in para-cancerous tissue. In conclusion, liver regeneration alone elevated YAP protein levels without elevating YAP mRNA levels. YAP mRNA and protein levels were both elevated in liver cancer cells.

Detailed analysis of temporal features on contrast enhanced ultrasound may help differentiate intrahepatic cholangiocarcinoma from hepatocellular carcinoma in cirrhosis.

Aim To verify if detailed analysis of temporal enhancement patterns on contrast enhanced ultrasound (CEUS) may help differentiate intrahepatic cholangiocarcinoma (ICC) from hepatocellular carcinoma (HCC) in cirrhosis. Methods Thirty three ICC and fifty HCC in cirrhosis were enrolled in this study. The contrast kinetics of ICC and HCC was analyzed and compared. Results Statistical analysis did not reveal significant difference between ICC and HCC in the time of contrast first appearance and arterial peak maximum time. ICC displayed much earlier washout than that of HCC (47.93±26.45 seconds vs 90.86±31.26 seconds) in the portal phase, and most ICC (87.9%) showed washout before 60 seconds than HCC (16.0%). Much more ICC (78.8%) revealed marked washout than HCC (12.0%) while most HCC (88.0%) showed mild washout or no washout in late part of the portal phase (90–120 seconds). Twenty six out of thirty three ICC (78.8%) demonstrated both early washout(<60seconds) and marked washout in late part of the portal phase, whereas, only six of fifty HCC (12.0%)showed these temporal enhancement features (p = 0.000).When both early washout and marked washout in the portal phase are taken as diagnostic criterion for ICC, the diagnostic sensitivity, specificity, positive predictive value, negative predictive value and accuracy were 78.8%,88.0%,81.3%,86.3%,and 84.3% respectively by CEUS. Conclusions Analysis of detailed temporal enhancement features on CEUS is helpful differentiate ICC from HCC in cirrhosis.If a nodule in cirrhotic liver displays hyper-enhancement in the arterial phase followed by early and marked washout in the portal phase, the nodule is highly suspicious of ICC rather than HCC.