Qiang Tong

MG-132 Inhibits Telomerase Activity, Induces Apoptosis and G 1 Arrest Associated with Upregulated p27kip1 Expression and Downregulated Survivin Expression in Gastric Carcinoma Cells

Ubiquitin-proteasome pathway (UPP) is the major system for the selective degradation of cellular proteins that play key roles in cellular processes. Previous study indicated that ubiquitin-proteasome inhibitor MG-132 could inhibit growth of some carcinoma. However, anti-carcinoma mechanism of MG-132 is unclear. Our objective was to investigate mechanisms of growth inhibitory effect of MG-132 on gastric carcinoma cells. Gastric carcinoma cell SGC-7901 was treated with ubiquitin-proteasome inhibitor MG-132. Cell growth suppression was evaluated with 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. DNA synthesis was evaluated by 3H-thymidine (3H-TdR) incorporation. Activity of telomerase was examined by telomeric repeat amplification protocol (TRAP) PCR-ELISA. Cell cycle and apoptosis were detected by flow cytometry (FCM). DNA fragment analysis was used to confirm the presence of apoptosis. Expression of p27kip1 and survivin was detected using the western blot method. After exposed to MG-132, the growth and value of 3H-TdR incorporation of gastric carcinoma cells were obviously inhibited. TRAP PCR-ELISA showed that light absorption of cells gradually decreased after exposed to 5 μ M of MG-132 for 24, 48, 72 and 96 h (P < 0.01). The percentage of cells at G0/G1 phase was increased and that at S and G2/M phase was decreased (P < 0.01). The ratio of apoptotic cells treated with 5 μ M MG-132 for 96 h was 53.7 ± 6.4%. Agarose electrophoresis showed marked ladders. Moreover, expression of p27kip1 of cells was increased and expression of survivin was decreased. Our results suggest that MG-132 inhibits telomerase activity, induces apoptosis and G1 arrest which is associated with upregulated p27kip1 expression and downregulated survivin expression in gastric carcinoma cells.

Up-regulation of P-glycoprotein is involved in the increased paclitaxel resistance in human esophageal cancer radioresistant cells

Abstract Objective. Development of drug and radiation resistance is one of the major causes of cancer treatment failure with chemoradiotherapy. Whether radiotherapy affects drugs resistance in esophageal cancer cells remain to be determined. The purpose of the study was to investigate the change of drug-sensitivity and P-glycoprotein (P-gp) expression in ionization radiation-induced human esophageal cancer radioresistant cells. Materials and methods. Radioresistant cells were established by means of continuous fractionated gamma-ray irradiation on human esophageal squamous cancer cell line EC9706. The radiosensitivity and drug-sensitivity between established radioresistant cells and parental cells were detected by a colony-forming assay and 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide (MTT) assay, respectively. The expressions of multidrug resistance type 1 gene (MDR1) mRNA and protein for P-gp were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot methods. The roles of P-gp activity in irradiation-induced drugs resistance were studied by using verapamil, an inhibitor of P-gp activity. Results. The esophageal cancer radioresistant cells showed an increased cisplatin or paclitaxel resistance. Compared with their parental cells, the expressions of MDR1 mRNA and protein for P-gp were increased significantly in radioresistant cells. Verapamil reduced paclitaxel resistance but had no effect on cisplatin resistance in human esophageal cancer radioresistant cells. Conclusions. These results suggested that up-regulation of P-gp is involved in the increased paclitaxel resistance but not cisplatin resistance in ionization radiation-induced human esophageal squamous cancer radioresistant cells.

Mechanism of Growth Inhibitory Effects of Cyclooxygenase-2 Inhibitor–NS398 on Cancer Cells

Cyclooxygenase (COX)-2 appears to play an important role in gastrointestinal carcinogenesis, and COX-2 overexpression has been demonstrated both in esophageal adenocarcinomas and lymph nodes metastasis. The aim of our study was to investigate the mechanism of growth inhibitory effect of selective inhibition of COX-2 by NS-398 on human cancer cells. The esophageal cancer cell lines (EC9706) that express COX-2 permanently and hepatocellular carcinoma cell lines (SMMC7721) while no expression of COX-2 were studied. Two kinds of cell lines were treated with various concentrations of NS-398 (selective for COX-2 inhibition) at 0.01–0.1 mM for 24 h, 48 h and 72 h. Antiproliferation effect was measured by 3H-TdR incorporation. The cell apoptosis were determined by flow cytometry (FCM) and DNA fragmentation analysis. Survivin was detected by immunocytochemical technique. The growth inhibition could be induced by NS398 in a dose- and time-dependent manner in two kinds of cell lines. FCM analysis revealed a high sub-G1 cell peak in EC9706 group. Agarose electrophroesis showed marked apoptosis ladder pattern, but no apoptosis by NS-398 in SMMC7721. The difference of apoptosis percentage in EC9706 and SMMC7721 was (45.23 ± 1.08)% and (3.05 ± 0.15)% (p < 0.001). After 24 h incubation with NS-398 at concentration of 0.1 dmM, the expression of survivin was markedly reduced in EC9706, but not in SMMC7721. We conclude that the administration of a selective inhibitor of COX-2 significantly decreases cell growth in cancer cell lines by different mechanism. NS-398 could inhibit cell proliferation in cancer cells whether or no COX-2 expression. Nevertheless, apoptosis in the cancer cells expressing COX-2 protein increase more than those lacking COX-2.

The relationship between p27kip1 expression and the change of radiosensitivity of esophageal carcinoma cells

Abstract Objective. Radioresistance is considered the main reason for therapeutic failure in radiotherapy of esophageal carcinoma. However, the underlying mechanisms of radioresistance remain elusive. The purpose of this study was to investigate the relationship between p27kip1 expression and the change of radiosensitivity of esophageal carcinoma cells. Material and methods. Radioresistant cells were gradually isolated by means of repeated gamma-ray irradiation upon esophageal carcinoma cells. The radiosensitivity of established radioresistant cells and parental cells was measured by standard colony-forming assay. Cell cycle was detected by flow cytometry. Western blot method was performed to identify the expression of p27kip1. Results. Colony-forming assay showed that the radioresistant cells had obvious radioresistance. Percentage of the radioresistant cells at G0/G1 and G2/M phase was significantly decreased, and the percentage of S phase cells was significantly increased compared with the parent cells (p < 0.05). Western blotting revealed that p27kip1 expression of the radioresistant cells was lower than that of parent cells. Conclusions. Our results suggest that cell phase change due to the decrease of p27kip1 expression is one of the mechanisms of radioresistance in esophageal carcinoma cells.

Upregulated p27kip1 can Downregulate Survivin Expression and Inhibit Telomerase Activity in Gastric Carcinoma Cells

ABSTRACT Previous study indicated that p27kip1 gene transfer can obviously inhibit the growth of some carcinoma cells. The purpose of this study was to investigate anticarcinoma mechanism of p27kip1. Gastric carcinoma cell strain SGC-7901 was transfected with recombinant adenovirus vector carrying human p27kip1 gene (Ad-p27kip1). Expression of survivin was detected using the western blot method and activity of telomerase was examined by telomeric repeat amplification protocol (TRAP) PCR-ELISA in gastric carcinoma cells. Our results suggest that upregulated p27kip1 can downregulate survivin expression and inhibit telomerase activity in gastric carcinoma cells.

A Cyclooxygase-2 Inhibitor NS-398-Enhanced Apoptosis of Esophageal Carcinoma Cell EC9706 by Adjusting Expression of Survivin and Caspase-3

The aim of this study is to examine the effect and mechanism of a selective cyclooxygenase-2 (COX-2) inhibitor NS-398 on inducing apoptosis of esophageal cancer cells. After the treatment with NS-398 on esophageal carcinoma EC9706 cell, MTT assay was used to observe the inhibition of EC9706 cell growth and apoptosis was determined by electronic microscopy and flow cytometry. The expression of survivin and caspase-3 was examined using immunocytochemical technique. The dose of 0.01–0.1 mM NS398 showed the inhibitory effect on growth of EC9706 cell lines and induce apoptosis in a dose- and time-effective manner; moreover, NS-398 also downregulated the level of expression of survivin and elevated the expression of capase-3. NS-398 can induce apoptosis of the esophageal carcinoma EC9706 cells by means of adjusting expression of survivin and caspase-3.

The usefulness of endoscopic ultrasound in the differential diagnosis between benign and malignant gastric ulcer

Abstract Objective. Gastric cancer can present as an exophytic lesion, diffuse infiltration of the gastric mucosa or even as a gastric ulcer, which can mimic a benign gastric ulcer. The purpose of this study was to evaluate the value of endoscopic ultrasound (EUS) in the differential diagnosis between benign and malignant gastric ulcer. Material and methods. 176 patients with gastric ulcer were divided into two groups on the basis of the cause of ulcer. Benign gastric ulcer group consisted of 102 patients and malignant gastric ulcer group consisted of 74 patients. All patients were examined by radial scanning echoendoscope (Olympus GF-UM 2000). Results. For diagnosis of malignant gastric ulcer, the sensitivity of EUS is 83.8%, the specificity is 62.7% and the accuracy is 71.6%. Conclusions. The results demonstrate that EUS is a useful examination in differential diagnosis between benign and malignant gastric ulcer. However, it also has certain limitation which may be solved with more newer EUS applications and development.

T lymphocyte Subsets Determination and DNA Ploidy Analysis in the Differential Diagnosis between Benign and Malignant Ascites

Differentiating malignant from benign ascites often leads to confusion and an inability to exclude its multitude of causes in many patients. In the present study, T lymphocyte subsets and DNA ploidy in ascitic fluid were detected by flow cytometer. There were significant differences in T lymphocyte subsets between benign and malignant ascites. For malignant ascites, the sensitivity of DNA aneuploid is 75.6%, the specificity is 79.0%, and the accuracy is 77.6%. The results demonstrate that T lymphocyte subsets determination and DNA ploidy analysis can be used in the differential diagnosis between benign and malignant ascites.

Combined detection of IL-6 and IL-8 is beneficial to the diagnosis of early stage esophageal squamous cell cancer: a preliminary study based on the screening of serum markers using protein chips

Background The diagnosis rate of early stage esophageal squamous cell carcinoma (ESCC) is low due to the lack of specific tumor markers. Seeking for these markers is beneficial to improve the early diagnosis rate and the prognosis of patients. This study profiles the differentially expressed proteins of early stage ESCC patients via the AAH-BLG-507 protein chip, which further consolidates the clinical evidence of ESCC diagnosis. Materials and methods In this study, 20 serum samples were collected from Taihe Hospital between August 2016 and June 2017. Ten of them carried ESCC, while the rest were healthy controls. To profile the proteins’ expression level, the AAH-BLG-507 protein chip was used, and both highly expressed and lowly expressed proteins were fished out. Meanwhile, their biological roles were examined by using Gene Ontology (GO) database and String database, and they were further verified by ELISA. Results Results showed that the expression levels of AXL, ARTN, Ang2, BDNF, BMP7, cripto-1, CCL28, E-selectin, IL-6, IL-8 and SHH in the serum of early ESCC were significantly upregulated (P<0.05), particularly IL-6 and IL-8. The expression levels of TSP1 and MMP-8 were markedly downregulated (P<0.05). Analysis showed that these proteins were mainly involved in angiogenesis, signal transduction, cell proliferation and migration, indicating the close relationship with the development of ESCC. Conclusion It suggested that IL-6 and IL-8 proteins could be considered as the markers for ESCC diagnosis.