Cuihua Lu

The expressions and clinical significances of tissue and serum galectin-3 in pancreatic carcinoma.

PurposeGalectin-3, a member of the beta-galactoside-binding protein family, is involved in many biological processes, including cell proliferation, regulating cell cycle, angiogenesis, tumorigenesis, metastasis, etc. The aim of this study is to elucidate the relationship between galectin-3 and clinicopathological variables and to evaluate the clinical significance of serum galectin-3 in the diagnosis of pancreas carcinoma.MethodsGalectin-3 expression in 78 pairs of pancreatic carcinoma tissues and the adjacent nontumorous tissues was tested by immunohistochemistry. The relationship between galectin-3 expression and clinical variables was analyzed. A sensitive method of time-resolved fluorescence immunological assay (TRFIA) for the detection of galectin-3 was established, and serum galectin-3 in cases with different pancreatic diseases was measured by TRFIA and ELISA. Further we compared the sensitivity and specificity of determining galectin-3, carcinoembryonic antigen (CEA) and carbohydrate antigen199 (CA199) for diagnosis of pancreatic carcinoma and assessed the complementary diagnostic value of galectin-3, CEA and CA199 for pancreatic carcinoma.ResultsImmunohistochemistry showed that galectin-3 expression was significantly higher in the human pancreatic carcinoma tissues than in the adjacent nontumorous tissues. The expression levels were correlated with the differentiation degree with the higher expression in poor differentiation tissues. Serum galectin-3 detected by both TRFIA and ELISA was much higher in patients with pancreatic carcinoma than in other groups. Serum galectin-3 was not correlated with CEA and CA199. Combined determination of these three markers has the complementary diagnostic value for human pancreatic carcinoma and may increase the diagnostic sensitivity to 97.5%.ConclusionsGalectin-3 is overexpressed in pancreatic carcinoma tissues, and it is correlated with the tumor differentiation. Serum galectin-3 is higher in cases with pancreatic carcinoma than in benign pancreatic diseases and healthy persons. Combined determination of serum galectin-3, CEA and CA199 may improve the diagnostic power for pancreatic carcinoma.

Association of hepatitis B virus pre-S deletions with the development of hepatocellular carcinoma in Qidong, China.

Background/Aim To investigate the roles of mutations in pre-S and S regions of hepatitis B virus (HBV) on the progression of hepatocellular carcinoma (HCC) in Qidong, China. Methods We conducted an age matched case-control study within a cohort of 2387 male HBV carriers who were recruited from August, 1996. The HBV DNA sequence in pre-S/S regions was successfully determined in 96 HCC cases and 97 control subjects. In addition, a consecutive series of samples from 11 HCC cases were employed to evaluate the pre-S deletion patterns before and after the occurrence of HCC. Results After adjustment for age, history of cigarette smoking and alcohol consumption, HBeAg positivity, pre-S deletions, pre-S2 start codon mutations, and T53C mutation were significantly associated with HCC, showing adjusted odds ratios (ORs) from 1.914 to 3.199. HCC patients also had a lower frequency of T31C mutation in pre-S2 gene, compared with control subjects (0.524; 95% CI 0.280-0.982). HBV pre-S deletions were clustered mainly in the 5′ end of pre-S2 region. Multivariate analysis showed that pre-S deletions and pre-S2 start codon mutations were independent risk factors for HCC. The OR (95% CI) were 2.434 (1.063–5.573) and 3.065 (1.099–8.547), respectively. The longitudinal observation indicated that the pre-S deletion mutations were not acquired at the beginning of HBV infection, but that the mutations occurred during the long course of liver disease. Conclusion Pre-S deletions and pre-S2 start codon mutations were independently associated with the development of HCC. The results also provided direct evidence that pre-S deletion mutations were not acquired from the beginning of infection but arose de novo during the progression of liver disease.

Long non-coding RNA SNHG20 predicts a poor prognosis for HCC and promotes cell invasion by regulating the epithelial-to-mesenchymal transition

BACKGROUND Recently, Accumulating evidence indicates that long noncoding RNAs (lncRNAs) have been shown to have critical regulatory roles in human tumor biology and development. However, the expression pattern and biological function of lncRNA small nucleolar RNA host gene 20 (SNHG20) in hepatocellular carcinoma (HCC) remains largely unknown. METHODS The expression of SNHG20 in 96 paired HCC tissues and cell lines were detected by quantitative real-time PCR (qRT-PCR) analysis. Kaplan-Meier survival analysis and log-rank test was used to reveal the association between SNHG20 expression and the overall survival time in HCC patients. CCK8 cell proliferation and transwell invasion assays were performed to analyze the cell proliferation and cell invasion ability. QRT-PCR and western-blotting analysis were performed to demonstrate the mRNA levels and protein expression of ZEB1, ZEB2 and relative epithelial-to-mesenchymal transition (EMT) markers (E-cadherin, N-cadherin and Vimentin). RESULTS We showed that the expression level of SNHG20 was significantly up-regulated in 96 pairs of HCC tissues and adjacent normal tissues. Higher SNHG20 expression was positively correlated with larger tumor size and advanced TNM stage, and negatively correlated with the over survival (OS) time for HCC patients. In vitro, loss-function assays revealed that knockdown of SNHG20 inhibited cell proliferation and invasion, whereas, gain-of-function promoted cell proliferation and invasion. Furthermore, knockdown of SNHG20 inhibited ZEB1, ZEB2, N-cadherin and Vimentin expression and up-regulated the E-cadherin expression in HCC cells. Mechanistic investigation revealed that SNHG20 could bind to enhancer of zeste homolog 2 (EZH2) and regulated E-cadherin expression. CONCLUSION Our results showed that the SNHG20/EZH2/E-cadhein regulator pathway might contribute to the development of novel therapeutic strategies for HCC patients.

Early mitotic inhibitor-1, an anaphase-promoting complex/cyclosome inhibitor, can control tumor cell proliferation in hepatocellular carcinoma: correlation with Skp2 stability and degradation of p27Kip1

Early mitotic inhibitor-1 (Emi1) is a key cell-cycle regulator that promotes S-phase and M-phase entry by inhibiting anaphase-promoting complex/cyclosome (APC/C) activity. Immunohistochemical analysis was performed in 114 human hepatocellular carcinoma (HCC) samples, and the data were correlated with clinicopathologic features. Univariate and multivariate survival analyses were performed to determine the prognostic significance of the proteins. Expression of Emi1 correlated directly with the stage of HCC. More importantly, high expression of Emi1 was associated with a poor outcome. Western blot analysis showed that Emi1 was highly expressed in HCC compared with the adjacent noncancerous tissue. In vitro, after the release of HCC cell lines from serum starvation, the expression of Emi1 APC/C substrates (cyclins A, B) and Skp2 was up-regulated, whereas p27(Kip1) was down-regulated. In addition, we used small interfering RNA to knock out Emi1 expression and observed its effects on HCC growth in vitro to determine whether loss of Emi1 could inhibit cell proliferation by blocking S-phase and mitotic entry. Western blot analyses indicated that deletion of Emi1 was positively correlated with APC/C substrates (cyclins A, B) and Skp2 but was negatively correlated with p27(Kip1). Emi1 inhibits APC/C activity, whereas Skp2 degradation is mediated by APC/C, and degradation of Skp2 can stabilize p27(kip1). These results suggested that Emi1 participates in HCC cell proliferation and that progression is controlled by APC/C inhibition, which stabilized Skp2 and enabled p27(kip1) degradation. These findings provide a potential therapeutic strategy for HCC.

Overexpressed nuclear BAG‐1 in human hepatocellular carcinoma is associated with poor prognosis and resistance to doxorubicin

Bcl‐2‐associated athanogene‐1 (BAG‐1) is a multifunctional anti‐apoptotic protein which regulates an array of cellular processes, including apoptosis, signaling, proliferation, transcription, and cell motility and has been reported to be over‐expressed in a number of human malignancies. To investigate the possible involvement of BAG‐1 in tumorigenesis of hepatocellular carcinoma (HCC), we performed Western blot analysis in eight paired samples of HCC and adjacent peritumoral tissues and immunohistochemistry in 65 paraffin sections of HCC, which both showed an enhanced expression of nuclear BAG‐1 isoform in HCC tissues. Statistical analysis confirmed that overexpression of nuclear BAG‐1 in HCC tissues was significantly associated with histological grading (P < 0.001), poor prognosis (P = 0.004), and was found to be an independent prognostic indicator for HCC (P = 0.023). We also noted that BAG‐1 was overexpressed in four HCC cell lines compared with a normal hepatocyte cell line, and BAG‐1 overexpression increased resistance of HCC cells to doxorubicin, a common chemotherapeutic agent for HCC. Furthermore, we observed that knock down of BAG‐1 with siRNA in HepG2 cells increased the chemosensitivity of cells, a process mediated through inhibition of doxorubicin‐triggered NF‐κB activation; and knock down of BAG‐1 suppressed proliferation and cell cycle transition of HepG2 cells. In consequence, our results for the first time indicated that BAG‐1 was dysregulated in HCC and suppression of BAG‐1 expression which resulted in inhibiting of NF‐κB signaling might be developed into a new strategy in HCC therapy. J. Cell. Biochem. 114: 2120–2130, 2013.

Expression of SGTA correlates with prognosis and tumor cell proliferation in human hepatocellular carcinoma.

To investigate the potential role of small glutamine-rich TPR-containing protein A (SGTA) in hepatocarcinogenesis, immunohistochemistry and Western blot were performed to detect the expression of SGTA in clinical Hepatocellular carcinoma (HCC) samples, adjacent nontumorous liver tissues and HCC cell lines. In addition, expression of SGTA was correlated with clinicopathological variables and univariate and multivariate survival analyses were performed to determine the prognostic significance. Moreover, the biological significance of the aberrant expression of SGTA was investigated in vitro. Both immunohistochemistry evaluation and Western blot analyses demonstrated that SGTA was overexpressed in HCC tissues compared with adjacent nontumorous liver tissues. Expression of SGTA directly correlated with the histological grades of HCC and high expression of SGTA was associated with a poor prognosis. SGTA depletion by siRNA inhibited cell proliferation, blocked S-phase and mitotic entry in Huh7 cells. Western blot analyses showed that SGTA depletion decreased cyclin A and cyclin B levels. Taken together, owing to overexpression of SGTA in HCC and its important role in predicting poor prognosis and the development of HCC, SGTA could be a potential prognostic marker and therapeutic target of HCC.

Effect of serum hepatitis B surface antigen levels on predicting the clinical outcomes of chronic hepatitis B infection: A meta-analysis

The impact of serum hepatitis B surface antigen (HBsAg) levels on the prognosis of chronic hepatitis B virus (HBV) infection remains unclear. This meta‐analysis aimed to determine whether serum HBsAg levels influenced the risk of cirrhosis and hepatocellular carcinoma (HCC) development. Furthermore, we explored the role played by serum HBsAg levels in prediction of spontaneous HBsAg seroclearance.

Combined analysis of serum γ-glutamyl transferase isoenzyme II, α-L-fucosidase and α-fetoprotein detected using a commercial kit in the diagnosis of hepatocellular carcinoma.

γ-glutamyl transferase isoenzyme II (GGT-II) is a sensitive biomarker of hepatocellular carcinoma (HCC). However, numerous disadvantages of the traditional manual method affected its application. The commercial kit provided a convenient and fast method for the determination of GGT-II levels. The purposes of the present study were to compare the reproducibility and sensitivity between the manual and commercial kit methods and to evaluate the diagnostic efficiency for HCC with the combined analysis of GGT-II, α-L-fucosidase (AFU) and α-fetoprotein (AFP). In patients with various liver diseases (HCC, liver cirrhosis and chronic hepatitis) and normal subjects, GGT-II was detected by manual and commercial polyacrylamide gel electrophoresis (PAGE). The levels of AFU and AFP were assayed by colorimetry and a chemiluminescence immunoassay, respectively. The commercial PAGE had equal diagnostic efficiency with traditional manual PAGE and no significant differences were observed in intra- and average-gel reproducibility and GGT-II sensitivities between the manual and commercial PAGE (P>0.05). The incidence of GGT-II detected by commercial PAGE in HCC patients was 84.1% and <8% in benign liver disease. The levels of AFU and AFP in the benign liver diseases and normal subjects were lower than those in HCC. According to the cut-off value obtained by receiver operating characteristic curves, a total of 56.6 and 59.3% of HCC patients (64 out of 113 and 67 out of 113) had AFU >636.5 μmol/l h and AFP >44.0 μg/l, respectively. There were no significant correlations between GGT-II and AFU or AFP. Combined detection of GGT-II with AFU or AFP increased the diagnostic sensitivity to 92.9 and 93.8%, respectively. These results suggest that commercial PAGE provides a simple and reproducible method for GGT-II detection. Combined determination of GGT-II with AFU or AFP exhibited superior sensitivity and specificity for the diagnosis of HCC.

The presence of IGHG1 in human pancreatic carcinomas is associated with immune evasion mechanisms.

Objectives: To investigate the expression of Ig&ggr;-1 chain C region (IGHG1) in human pancreatic carcinomas and determine the biological function of IGHG1 expression in immune evasion mechanisms. Methods: Comparative proteomic analysis was used to detect the differential expression of IGHG1 in human pancreatic cancer tissues versus adjacent noncancerous tissues, followed by confirmatory tests including quantitative real-time reverse transcription-polymerase chain reaction, Western blot analysis, immunohistochemistry, and immunofluorescence. A murine pancreatic tumor model was established by transplantation of IGHG1-overexpressing Panc02 cells. The cytotoxic responses of natural killer (NK) cells were assessed with a lactate dehydrogenase release assay. Results: Ig&ggr;-1 chain C region was found to be present in human pancreatic cancer tissues but nearly absent or expressed lower in adjacent noncancerous tissues. In the murine pancreatic tumor model, the tumor growth was significantly accelerated from day 12 to 20 after tumor injection, and the survival time of animals was decreased. Blockage of IGHG1 led to retarded tumor growth and improved survival. The cytotoxicity assay revealed that IGHG1 downregulated the cytotoxic activity of NK cells through inhibition of antibody-dependent cellular cytotoxicity function. Conclusions: The presence of IGHG1 in pancreatic cancer cells might constitute an important element responsible for tumor cell proliferation and immune evasion mechanisms.Abbreviations: IGHG1 - Ig&ggr;-1 chain C region, ADCC - antibody-dependent cellular cytotoxicity, NK cells - natural killer cells, PCR - polymerase chain reaction, LDH - lactate dehydrogenase, CH - constant region of heavy chain

Upregulated expression of ubiquitin-conjugating enzyme E2Q1 (UBE2Q1) is associated with enhanced cell proliferation and poor prognosis in human hapatocellular carcinoma.

Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world. Ubiquitin-proteasome system has been shown to play a pivotal role in the pathophysiology of HCC and other malignancies. UBE2Q1 is a putative E2 ubiquitin conjugating enzyme, and may be involved in the regulation of cancer-related proteins. In this study, we investigated the expression pattern of UBE2Q1 in HCC cell lines and human HCC specimens, and its potential clinical and biological significance in HCC. Western blot and immunohistochemical analyses revealed that UBE2Q1 was significantly upregulated in HCC tumorous tissues compared with the adjacent noncancerous ones. Next, univariate and multivariate survival analyses were performed to determine the prognostic significance of UBE2Q1 in HCC. The results showed that upregulated expression of UBE2Q1 was positively correlated with high histological grades of HCC and predicted poor prognosis. In addition, the expression of UBE2Q1 was progressively increased in serum-refed HCC cells. UBE2Q1 depletion by small interfering RNA inhibited cell proliferation and led to G1 phase arrest in HepG2 and BEL-7404 cells. Furthermore, we showed that cells transfected with UBE2Q1-targeting siRNA resulted in significant increase in the levels of p53, p21 in HepG2 and BEL-7404 cells. These data imply that UBE2Q1 is upregulated in liver cancer cell lines and tumorous samples and may play a role in the development of HCC.