Runzhou Ni

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.

Expression of FOXJ1 in hepatocellular carcinoma: correlation with patients' prognosis and tumor cell proliferation.

FOXJ1 is a member of the forkhead box (FOX) family of transcription factors. Recent studies suggested that FOXJ1 may function as a tumor suppressor gene in breast cancer. To investigate the potential roles of FOXJ1 in hepatocellular carcinoma (HCC), expression of FOXJ1 was first examined in eight paired frozen HCC and adjacent noncancerous liver tissues by Western blot, and we found that FOXJ1 was upregulated in HCC specimens. In addition, immunohistochemistry was performed to confirm our results in 108 HCC samples. Moreover, we also evaluated its relation with clinicopathological variables and the prognostic significance. The data showed that high expression of FOXJ1 was associated with histological grade (P < 0.001), and FOXJ1 was positively correlated with proliferation marker Ki‐67 (P < 0.01). Univariate analysis suggested that FOXJ1 expression was associated with poor prognosis (P < 0.001). Multivariate analysis indicated that tumor grade (P < 0.0001), metastasis (P = 0.0451), tumor size (P = 0.0459), FOXJ1 (P = 0.0011), and Ki‐67 (P = 0.0006) were independent prognostic markers for HCC. Furthermore, we noted that there existed the change of the level of FOXJ1 subcellular localization during cell‐cycle transition in HepG2 cells by immunofluorescence and cell fractionation. Besides, we employed FOXJ1 overexpression/knockdown approaches to investigate the effects of FOXJ1 on HCC cell proliferation and cell‐cycle distribution and found that overexpression of FOXJ1 can promote tumor cell proliferation and cell‐cycle transition. Our results suggested that FOXJ1 was overexpressed in HCCs and associated with histological grade and poor prognosis. Overexpression of FOXJ1 was also involved in tumor cell proliferation and cell‐cycle progression in HCC cell lines.

S100 family signaling network and related proteins in pancreatic cancer (Review)

The occurrence and development of pancreatic cancer is a complex process convoluted by multi-pathogenies, multi-stages and multi-factors. S100 proteins are members of the S100 family that regulate multiple cellular pathways related to pancreatic cancer progression and metastasis. S100 proteins have a broad range of intracellular and extracellular functions, including the regulation of protein phosphorylation and enzyme activity, calcium homeostasis and the regulation of cytoskeletal components and transcriptional factors. S100 proteins interact with receptor for advanced glycation end-products (RAGE), p53 and p21, which play a role in the degradation of the extracellular matrix (ECM) and metastasis, and also interact with cytoskeletal proteins and the plasma membrane in pancreatic cancer progression and metastasis. S100A11 and S100P are significant tumor markers for pancreatic cancer and unfavorable predictors for the prognosis of patients who have undergone surgical resection. Recently, S100A2 has been suggested to be a negative prognostic biomarker in pancreatic cancer, and the expression of S100A6 may be an independent prognostic impact factor. The expression of S100A4 and S100P is associated with drug resistance, differentiation, metastasis and clinical outcome. This review summarizes the role and significance of the S100 family signaling network and related proteins in pancreatic cancer.

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.

Expression and clinical role of NF45 as a novel cell cycle protein in esophageal squamous cell carcinoma (ESCC)

NF45 (also known as ILF2), as one subunit of NF-AT (nuclear factor of activated T cells), repairs DNA breaks, inhibits viral replication, and also functions as a negative regulator in the microRNA processing pathway in combination with NF90. Recently, it was found that implicated in the mitotic control of HeLa cells and deletion of endogenous NF45 decreases growth of HeLa cells. While the role of NF45 in cancer biology remains under debate. In this study, we analyzed the expression and clinical significance of NF45 in esophageal squamous cell carcinoma ESCC. The expression of NF45 was evaluated by Western blot in 8 paired fresh ESCC tissues and immunohistochemistry on 105 paraffin-embedded slices. NF45 was highly expressed in ESCC and significantly associated with ESCC cells tumor stage and Ki-67. Besides, high NF45 expression was an independent prognostic factor for ESCC patients’ poor survival. To determine whether NF45 could regulate the proliferation of ESCC cells, we increased endogenous NF45 and analyzed the proliferation of TE1 ESCC cells using Western blot, CCK8, flow cytometry assays and colony formation analyses, which together indicated that overexpression of NF45 favors cell cycle progress of TE1 ESCC cells. While knockdown of NF45 resulted in cell cycle arrest at G0/G1-phase and thus abolished the cell growth. These findings suggested that NF45 might play an important role in promoting the tumorigenesis of ESCC, and thus be a promising therapeutic target to prevent ESCC progression.

Pyruvate Kinase M2 Regulates Apoptosis of Intestinal Epithelial Cells in Crohn’s Disease

BackgroundPyruvate kinase M2 (PKM2), a key glycolytic enzyme, is involved in multiple cellular processes including apoptosis. Recently increased fecal PKM2 has been found in Crohn’s disease (CD), but little is known regarding its function in the pathophysiology of the disease.AimThe intestinal expression of PKM2 and its involvement in CD was investigated.MethodsPyruvate kinase M2 expression in mucosal biopsies from patients with CD and normal controls was detected by immunohistochemistry. A murine model of colitis induced by trinitrobenzenesulphonic acid (TNBS) was established and expression of PKM2, B cell lymphoma-extra large (Bcl-xl), active caspase-3 as well as cleaved poly (ADP-ribose) polymerase (PARP) was examined for association of PKM2 with intestinal epithelial cell (IEC) apoptosis. Furthermore, we treated human IEC line HT-29 by tumor necrosis factor-α (TNF-α) and used RNA interference to analyze the role of PKM2 in IEC apoptosis.ResultsIntestinal expression of PKM2 was higher in patients with CD compared with normal controls mainly locating in IECs. In TNBS-induced colitis, up-regulation of PKM2 was accompanied by the elevated expression of Bcl-xl, active caspase-3, and cleaved PARP. PKM2 was co-localized with active caspase-3 in IECs marked by E-cadherin, suggesting its role in IEC apoptosis. Expression of PKM2 and Bcl-xl in TNF-α-induced HT-29 cells was increased, while TNF-α had no effect on cellular localization of PKM2. Furthermore, knockdown of PKM2 by siRNA could inhibit expression of Bcl-xl but enhance apoptosis in TNF-α-treated HT-29 cells.ConclusionThe up-regulation of PKM2 might protect IECs against apoptosis possibly through Bcl-xl in CD, indicating its important role in the pathophysiology of CD.

CtBP2 contributes to malignant development of human esophageal squamous cell carcinoma by regulation of p16INK4A

C‐terminal binding protein‐2 (CtBP2), as a transcriptional co‐repressor, has been shown to mediate the repression of p16INK4A, a tumor suppressor gene product, in primary human cells. Here we aimed to investigate how the correlation between CtBP2 and p16INK4A influenced the development of esophageal squamous cell carcinoma (ESCC). Immunohistochemistry of ESCC tissue sections indicated that the CtBP2 and p16INK4A expressions were inversely correlated to each other with a linear regression coefficient of −0.747 (P < 0.05), and Western blot analysis revealed that CtBP2 was higher expressed in tumorous tissues than in adjacent non‐tumorous tissues. Either CtBP2 or p16INK4A expression was significantly related to histological differentiation (P = 0.016 or 0.001) and to the expression of Ki‐67, a proliferating marker (P = 0.006 or 0.02), and patients with higher CtBP2 and lower p16INK4A expressions had shorter overall survival. We also observed that CtBP2 modulated the cell proliferation and cell cycle in ECA109 cells, an ESCC cell line, by inhibiting p16INK4A. Overexpression or knockdown of CtBP2 in ECA109 cells was found to inhibit or activate the mRNA or protein expression of p16INK4A, which in turn altered the cell proliferation and cell cycle in ECA109 cells, as measured by flow cytometry and cell count assay. Additionally, after ECA109 cells silenced for CtBP2 were treated with cisplatin (an anti‐ESCC agent), the p16INK4A expression was up‐regulated, and the cell apoptosis was promoted, thus confirming the repression of p16INK4A by CtBP2. Collectively, all results suggested that CtBP2 might contribute to the progression of ESCC through a negative transcriptional regulation of p16INK4A. J. Cell. Biochem. 114: 1343–1354, 2013.

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.