Chengqi Guan

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.

Interaction with CCNH/CDK7 facilitates CtBP2 promoting esophageal squamous cell carcinoma (ESCC) metastasis via upregulating epithelial-mesenchymal transition (EMT) progression

CtBP2, as a transcriptional corepressor of epithelial-specific genes, has been reported to promote tumor due to upregulating epithelial-mesenchymal transition (EMT) in cancer cells. CtBP2 was also demonstrated to contribute to the proliferation of esophageal squamous cell carcinoma (ESCC) cells through a negative transcriptional regulation of p16INK4A. In this study, for the first time, we reported that CtBP2 expression, along with CCNH/CDK7, was higher in ESCC tissues with lymph node metastases than in those without lymph node metastases. Moreover, both CtBP2 and CCNH/CDK7 were positively correlated with E-cadherin, tumor grade, and tumor metastasis. However, the concrete mechanism of CtBP2’s role in enhancing ESCC migration remains incompletely understood. We confirmed that CCNH/CDK7 could directly interact with CtBP2 in ESCC cells in vivo and in vitro. Furthermore, our data demonstrate for the first time that CtBP2 enhanced the migration of ESCC cells in a CCNH/CDK7-dependent manner. Our results indicated that CCNH/CDK7-CtBP2 axis may augment ESCC cell migration, and targeting the interaction of both may provide a novel therapeutic target of ESCC.

Far upstream element-binding protein 1 (FUBP1) is a potential c-Myc regulator in esophageal squamous cell carcinoma (ESCC) and its expression promotes ESCC progression.

The human far upstream element (FUSE) binding protein 1 (FUBP1) belongs to an ancient family which is required for proper regulation of the c-Myc proto-oncogene. Although c-Myc plays an important role in development of various carcinomas, the relevance of FUBP1 and their contribution to esophageal squamous cell carcinoma (ESCC) development remain unclear. In this study, we aimed to investigate the relationship between FUBP1 and c-Myc as well as their contribution to ESCC development. Western blot and immunohistochemical analyses were performed to evaluate FUBP1 expression. Coimmunoprecipitation analysis was performed to explore the correlation between FUBP1 and c-Myc in ESCC. In addition, the role of FUBP1 in ESCC proliferation was studied in ESCC cells through knocking FUBP1 down. The regulation of FUBP1 on proliferation was confirmed by Cell Counting Kit-8 (CCK-8) assay, flow cytometric assays, and clone formation assays. The expressions of FUBP1 and c-Myc were both upregulated in ESCC tissues. In addition to correlation between expression of FUBP1 and tumor grade, we also confirmed the correlation of FUBP1, c-Myc, and Ki-67 expression by twos. Moreover, upregulation of FUBP1 and c-Myc in ESCC was associated with poor survival. FUBP1 was confirmed to activate c-Myc in ESCC tissues and cells. FUBP1 was demonstrated to promote proliferation of ESCC cells. Moreover, downregulation of both FUBP1 and c-Myc was confirmed to inhibit proliferation of ESCC cells. Our results indicated that FUBP1 may potentially stimulate c-Myc expression in ESCC and its expression may promote ESCC progression.

Upregulation of SYF2 in esophageal squamous cell carcinoma promotes tumor cell proliferation and predicts poor prognosis

SYF2, also known as CCNDBP1-interactor or p29, is reported in pre-mRNA splicing and cell cycle progression. However, the role of SYF2 in esophageal squamous cell carcinoma (ESCC) development remains elusive. In the present study, Western blot and immunohistochemistry assays demonstrated that SYF2 was overexpressed in ESCC tumor tissues and cell lines. In addition, immunohistochemistry analysis revealed that SYF2 expression was positively correlated with tumor grade and predicted poor prognosis of ESCC. In vitro studies using serum starvation-refeeding experiment and SYF2-siRNA transfection assay demonstrated that SYF2 expression promoted proliferation of ESCC cells, while SYF2 knockdown led to decreased cell growth rate and colony formation resulted from growth arrest of cell cycle at G0/G1 phase. Furthermore, our results indicated that SYF2 can down-regulate the sensitivity of ESCC cells for cisplatin. Our findings for the first time supported that SYF2 might play an important role in the regulation of ESCC proliferation and would provide a novel therapeutic strategy against human ESCC.

Enhanced expression of early mitotic inhibitor-1 predicts a poor prognosis in esophageal squamous cell carcinoma patients

Early mitotic inhibitor-1 (Emi1), as a key cell cycle regulatory gene, induces S phase and mitotic entry by controlling anaphase-promoting complex substrates. Emi1 overexpression may be a prognostic factor for patients with invasive breast cancer. However, its expression and clinical significance in esophageal squamous cell carcinoma (ESCC) remain unknown. In the present study, Emi1 was overexpressed in ESCC samples, contrarily to their neighboring normal tissues. The expression of Emi1 was correlated with histological differentiation (P=0.032), lymphatic metastasis (P=0.006) and Ki-67 expression (P=0.028). Multivariate analysis indicated that the presence of lymphatic metastasis and the protein expression levels of Emi1 and Ki-67 were all independent prognostic factors for ESCC patients (P=0.042, 0.018 and 0.001, respectively). In vitro, however, the expression of Emi1 was upregulated in the ECA109 cell line following release from serum starvation. In addition, depletion of endogenous Emi1 by small interfering RNA could effectively reduce cell proliferation. Thus, the present data indicated that Emi1 expression was upregulated in ESCC tissues and correlated with poor survival in ESCC patients, and suggested that Emi1 may be an independent prognostic factor for ESCC patients.

High NUSAP1 expression predicts poor prognosis in colon cancer

BACKGROUND AND AIM Nucleolar and spindle-associated protein 1 (NUSAP1) is an indispensable mitotic regulator. Aberrant NUSAP1 expression is associated with perturbed mitosis and tumorigenesis. In this study, we investigated the clinical significance of NUSAP1 expression in colon cancer. METHODS AND MATERIALS Immunohistochemical staining was performed to determine NUSAP1 protein levels in paraffin colon tumor specimens. Real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) was conducted to detect NUSAP1 mRNA levels in colon tumor samples. The association between NUSAP1 protein expression and clinicopathological characteristics of patients with colon cancer was assessed. A Kaplan-Meier analysis was performed to determine the prognostic significance of NUSAP1 in colon cancer. A Cox proportional hazards model was used to calculate univariate and multivariate hazard ratios for the NUSAP1 and other clinicopathological variables. RESULT NUSAP1 protein and mRNA levels were significantly higher in colon tumor tissues than in paired non-cancerous adjacent tissues (P < 0.001, respectively). NUSAP1 protein expression was significantly correlated with histopathological grading (P < 0.001), depth of invasion (P = 0.001), lymph node metastasis (P < 0.001) and TNM stage (P < 0.001). The overall survival rate of patients with high NUSAP1 expression was significantly lower than for patients with low NUSAP1 expression (log-rank test, P < 0.001). A multivariate Cox model demonstrated that NUSAP1 is an independent risk factor for overall survival (P = 0.025). CONCLUSION NUSAP1 is overexpressed in colon cancer and high expression of NUSAP1 acts as an independent predictive factor for poor prognosis in colon cancer.

Correlation between S100A11 and the TGF-β 1 /SMAD4 pathway and its effects on the proliferation and apoptosis of pancreatic cancer cell line PANC-1

S100A11 as a S100 protein family member has been documented to play dual-direction regulation over cancer cell proliferation. We explored the role of S100A11 in the proliferation and apoptosis of pancreatic cancer cell line PANC-1 and the potential mechanisms involving the TGF-β1/SMAD4/p21 pathway. S100A11 and TGF-β1 protein expressions in 30 paraffin-embedded specimens were evaluated by immunohistochemistry. S100A11 and TGF-β1 expression in PANC-1 cell line was suppressed using small interfering RNA (siRNA), respectively. Subsequently, pancreatic cancer cell apoptosis was measured by Cell Counting Kit-8 and flow cytometry, and S100A11 and TGF-β1/SMAD4/p21 pathway proteins and genes were detected with Western blotting and quantitative polymerase chain reaction (qPCR). S100A11 cytoplasmic/nuclear protein translocation was examined using NE-PER® cytoplasm/nuclear protein extraction in cells interfered with TGF-β1 siRNA. Our results showed that S100A11 expression was positively correlated with TGF-β1 expression in pancreatic cancerous tissue. Silencing TGF-β1 down-regulated intracellular P21WAF1 expression by 90%, blocked S100A11 from cytoplasm entering nucleus, and enhanced cell proliferation. Silencing S100A11 down-regulated intracellular P21 expression and promoted cell apoptosis without significantly changing TGF-β1 and SMAD4 expression. Our findings revealed that S100A11 and TGF-β1/SMAD4 signaling pathway were related but mutually independent in regulating PANC-1 cells proliferation and apoptosis. Other independent mechanisms might be involved in S100A11’s regulation of pancreatic cell growth. S100A11 could be a potential gene therapy target for pancreatic cancer.