Wang Hongyang

Independent Factors and Predictive Score for Extrahepatic Metastasis of Hepatocellular Carcinoma Following Curative Hepatectomy

BACKGROUND Postoperative extrahepatic metastasis (EHM) contributes to a poor prognosis in patients with hepatocellular carcinoma (HCC) after hepatectomy. This study was aimed to develop a practical method that can be used to predict postoperative EHM. METHODS In total, 578 patients were enrolled. We analyzed the clinicopathological features of the tumors and did a long-term follow-up to observe HCC recurrence. Postoperative EHM was detected in 136 patients, and multivariate analysis was used to confirm independent risk factors for postoperative EHM. After the factors were identified, a predictive scoring system was constructed as a weighted sum of these factors. The cutoff value that determines a high risk for EHM was defined by maximizing the Youdens index of the receiver operating characteristic curve. RESULTS Microvascular invasion, incomplete capsule, and larger tumor diameter were the three independent factors predictive for a high risk for EHM. The scoring system was derived with an area under the curve (AUC) of 0.81 for postoperative 10-year EHM prediction. A cutoff value of 43 was derived and validated with a sensitivity >90% and specificity >60% to predict the development of EHM. This system was further verified in a subgroup of Barcelona Clinic Liver Cancer stage 0-A patients with an AUC of 0.82. When the cutoff value was set at 43, the sensitivity and specificity were 90.38% and 64.88%, respectively. CONCLUSIONS Our predictive scoring system may be used to identify HCC patients who have a high risk for EHM following curative hepatectomy.

Knockdown of HBV surface antigen gene expression by a lentiviral microRNA-based system inhibits HBV replication and HCC growth.

Summary.  Current options for the treatment of hepatitis B virus (HBV) infections, a common liver cancer risk factor, are limited. While RNA interference (RNAi) technologies have been shown to inhibit HBV replication, the consequent effects on hepatocellular carcinoma (HCC) cell growth are not fully understood. The aim of this study was to evaluate the effect of RNAi‐mediated decrease in the HBV surface antigen (HBsAg) gene on HBV replication and HCC growth. A lentiviral microRNA‐based system expressing siRNAs targeting the HBsAg gene (LVshHBS) was developed and transfected into HepG2.2.15 cells (HBV stably expressing line). We found that LVshHBS significantly inhibited the HBsAg mRNA and protein levels in the HepG2.2.15 cells, while HBsAg secretion into the culture supernatant decreased by 70%. BALB/c (nu/nu) mice were injected with HepG2.2.15 cells transduced with LVshHBS or control vectors to investigate the effect of inhibiting the HBsAg on the development of tumour growth in a human HCC nude mice model. Compared with the control, the tumour growth in nude mice was significantly decreased after injection with LVshHBS. Microarray analysis of tumour‐related genes in LVshHBS‐transduced HepG2.2.15 cells showed that the expressions of genes involved in cell cycle, differentiation and oncogenesis such as ACP2, BHLHB2, CLK3, CTSC, FOS, NR1D1, PIM1 and SEPT6 genes were downregulated, while that of the E2F3 gene was upregulated. In conclusion, lentiviral microRNA‐based RNAi against the HBsAg gene not only inhibits HBV replication but also inhibits the growth of HCC. Downregulation of growth‐related genes is implicated in this mechanism of inhibition.

Quantitative Detection of Osteopontin High Expression in Human and Rat Hepatocellular Carcinoma

AbstractObjective: Osteopontin (OPN) is a secreted phosphorylated glycoprotein that is implicated in proliferation and migration of several malignancies including hepatocellular carcinoma (HCC). In present study, human HCC specimens were collected and rat HCC model was chemical-induced to elucidate the expression significance of OPN in HCC progression. Methods: OPN expression was detected quantitatively by real-time reverse transcription polymerase chain reaction (RT-PCR). Male Sprague-Dawley rats were administrated diethylnitrosamine (DENA) to induce HCC and OPN expression was dynamically assessed. Results: In 69 cases of 103 HCC patients (67%) OPN was highexpressed in HCC tissues than that in adjacent non-tumor liver tissues and in 58 cases of these 69 cases more than 2-fold. OPN expression was significantly different between HCC and adjacent liver tissues (0.53±0.91 vs 0.11±0.28, P<0.001). OPN expression was gradually elevated in occurrence and development of rat HCC. Conclusion: OPN was highexpressed in human HCC and gradually elevated in rat HCC progression.

Differential Expression of PKC Isoforms and Their Tumoricidal Activity in Two Macrophage Cell Lines: Involvement of Nitric Oxide-dependent Mechanisms

AbstractObjective: To investigate the role of PKC isoforms in the regulation of LPS-triggered tumoricidal activity in macrophages and further elucidate its signal mechanisms. Methods: Two macrophage cell lines (P388D1 and RAW264.7) were stimulated by LPS alone, or with long-term of PMA pretreatment. Then cytotoxicities to P815 cells (by MTT assay) and IL-1, TNF-α (by ELISA) and nitric oxide (NO) production (by Griess reagent) in supernatants were measured. Western blot for PKC isoforms after long-term PMA pretreatment was analyzed. Results: RAW264.7 cells were stimulated with LPS to kill target tumor cells P815, whereas P388D1 cells failed to develop such an ability. Down-regulation of PKC isoforms by chronic treatment with PMA significantly inhibited the LPS-induced cytotoxicity in RAW264.7 cells. In unstimulated state, Western blotting with rabbit antiserum specific for the PKCα, β1, β2, δ or ε showed all 5 isoforms were detected in P388D1 cells, while only PKCα, PKCβ1 and PKCδ were detected in RAW264.7 cells. Exposure of the cells to long-term of PMA treatment significantly down-regulated the expression of PKCα, PKCβ1 and PKCδ in RAW264.7 cells. But in P388D1 cells, although PKCα, PKCδ and PKCε were down-regulated, the expression of PKCβ1 and PKCβ2 could not be regulated. Comparing with LPS-induced IL-1, TNF- and NO production by the two macrophage cell lines, P388D1 failed to produce NO. In RAW264.7 cells, LPS-induced NO production and antitumor activity was attenuated by the addition of L-NAME, an iNOS inhibitor. Conclusion: The results indicated a critical role of PKCβ in LPS-induced antitumor activity and this cytotoxicity is mainly due to PKC-β mediated NO production by RAW264.7 cells, but not a direct cytotoxic activity.

The possible role of signal regulatory protein α1 in hepatocellular carcinoma

Objective: To study the relationship between signal regulatory protein α1 (SIRPα1) and hepatocellular carcinoma (HCC) and explore the possible molecular mechanisms.Methods: Total RNA was extracted from normal hepatocyte cell line Hs680, Hepatocellular carcinoma cell lines HepG2, H4–H7, Chang liver and SK-Hep1. The SIRPTα1 was detected in 42 pathologically confirmed hepatocellular carcinoma tissue samples and 6 normal liver tissues by Northern blot and immunohistochemical staining. The association of SIRPα1 and HGF was determined in HuH7.Results: SIRPα1 was highly expressed in Hs680, HepG2 and H4–H7, but weakly expressed in SK-Hep1 and Chang liver cell lines. Northern blot and Western blot both detected the reduced expression of SIRPα1 in hepatocellular carcinoma tissue as compare to paracancerous and normal tissues.In vitro experiments showed HGF stimulation elevated SIRPα1 phosphorylation. Over-expression of wild type SIRPα1 promoted cell proliferation in the presence of HGF.Conclusion: SIRPα1 might be involved in HCC development and metastasis. The underlying mechanisms may be related to regulatory tyrosine phosphorylation of SIRPα1 by HGF stimulation.