[Effect of Circular RNA hsa_circ_0067582 on the Proliferation and Invasion Ability of Gastric Cancer Cells].

Objective To investigate the effects on cell proliferation and invasion of the circular RNA hsa_circ_0067582 in gastric cancer(GC). Methods After hsa_circ_0067582 overexpression (Oe-circ_0067582) plasmid was transfected into AGS and SGC-7901 cells,the cell viability,proliferation,invasion ability,and apoptosis were detected by CCK-8,colony formation and EdU assays,Transwell assay,and flow cytometry,respectively.Western blotting was employed to detect the expression levels of proteins related to the cell apoptosis and epithelial-mesenchymal transition(EMT).The effect of Oe-circ_0067582 on the growth of SGC-7901 cells in nude mice was observed.Bioinformatics tools were used to predict the binding target miRNA of hsa_circ_0067582,and the competing endogenous RNA(ceRNA)regulatory network was established.Finally,functional enrichment was performed to analyze the biological functions of the target genes of the predicted miRNA. Results Compared with the pLO-ciR(empty plasmid)group,the Oe-circ_0067582 group in AGS and SGC-7901 cells attenuated the cell viability(t=7.883,P=0.001;t=5.679,P=0.005),proliferation(t=6.709,P=0.003;t=5.857,P=0.003),and invasion ability(t=7.782,P=0.002;t=6.342,P=0.003)and induced cell apoptosis(t=7.225,P=0.002;t=11.509,P=0.001).Western blotting showed that the Oe-circ_0067582 group in AGS and SGC-7901 cells up-regulated the protein levels of cysteinyl aspartate specific proteinase (Caspase) 3(t=6.863,P=0.002;t=7.024,P=0.001),Caspase 7(t=3.295,P=0.04;t=6.008,P=0.004),Caspase 9(t=4.408,P=0.012;t=6.278,P=0.004),and E-cadherin(t=12.453,P=0.002;t=10.867,P=0.001),while down-regulated those of Vimentin(t=7.242,P=0.002;t=5.694,P=0.004)and N-cadherin(t=6.480,P=0.003;t=7.446,P=0.001).Furthermore,Oe-circ_0067582 significantly inhibited the growth of tumor in the SGC-7901 tumor-bearing nude mice(t=3.526,P=0.017).The prediction based on TargetScan and miRnada suggested that hsa_circ_0067582 can competitively bind to hsa-miR-181b-3p,hsa-miR-337-3p,hsa-miR-421,and hsa-miR-548d-3p.The functional enrichment indicated that the target genes of miRNA were involved in multiple cancer-related biological processes including negative regulation of apoptotic process,gene expression,transcriptional misregulation in cancer,transforming growth factor-ß,and p53 signaling pathways. Conclusion Oe-circ_0067582 can inhibit the proliferation and attenuate EMT process to reduce the invasion ability of AGS and SGC-7901 cells,which provides a new target for the treatment of GC.

Activation of LXRß inhibits proliferation, promotes apoptosis, and increases chemosensitivity of gastric cancer cells by upregulating the expression of ATF4.

Recently, great advances have been achieved in both surgery and chemotherapy for the treatment of gastric cancer, but there is still poor prognosis for this disease. The aim of this study is to investigate the role of liver X receptor ß (LXRß) in chemosensitivity of gastric cancer SGC7901 cells. From 171 patients with gastric cancer, the gastric cancer and paracancerous tissues were selected to measure the expression of LXRß and ATF4. Gastric cancer cell lines were cultured and screened to figure out the proliferation and apoptosis of gastric cancer SGC7901 cells with the treatment of LXRß agonist (GW3965), ATF4 short hairpin RNA (shRNA), and chemotherapy drug paclitaxel. The expression of apoptosis-related gene cleaved caspase-3 was detected by Western blot analysis. First, we found that the expressions of LXRß and ATF4 in gastric cancer tissues and cells were significantly lower than those in their paracancerous tissues and gastric mucosal epithelial cells. In addition, activation of LXRß and paclitaxel treatment suppressed proliferation of SGC7901 cells, and the expression of ATF4 was upregulated in a concentration-dependent manner. Furthermore, shRNA significantly inhibited the expression of ATF4 and blocked the chemosensitivity of SGC7901 cells to LXRß activation. Our study demonstrates that the expression of LXRß was low in gastric cancer. In addition, activation of LXRß may inhibit the proliferation of gastric cancer cells, promote apoptosis, and increase chemosensitivity by upregulating the expression of ATF4.

Cannabidiol Induces Cell Cycle Arrest and Cell Apoptosis in Human Gastric Cancer SGC-7901 Cells.

The main chemical component of cannabis, cannabidiol (CBD), has been shown to have antitumor properties. The present study examined the in vitro effects of CBD on human gastric cancer SGC-7901 cells. We found that CBD significantly inhibited the proliferation and colony formation of SGC-7901 cells. Further investigation showed that CBD significantly upregulated ataxia telangiectasia-mutated gene (ATM) and p53 protein expression and downregulated p21 protein expression in SGC-7901 cells, which subsequently inhibited the levels of CDK2 and cyclin E, thereby resulting in cell cycle arrest at the G0-G1 phase. In addition, CBD significantly increased Bax expression levels, decreased Bcl-2 expression levels and mitochondrial membrane potential, and then upregulated the levels of cleaved caspase-3 and cleaved caspase-9, thereby inducing apoptosis in SGC-7901 cells. Finally, we found that intracellular reactive oxygen species (ROS) increased after CBD treatment. These results indicated that CBD could induce G0-G1 phase cell cycle arrest and apoptosis by increasing ROS production, leading to the inhibition of SGC-7901 cell proliferation, thereby suggesting that CBD may have therapeutic effects on gastric cancer.

The Growth of SGC-7901 Tumor Xenografts Was Suppressed by Chinese Bayberry Anthocyanin Extract through Upregulating KLF6 Gene Expression.

To investigate the antitumor effect of anthocyanins extracted from Chinese bayberry fruit (Myrica rubra Sieb. et Zucc.), a nude mouse tumor xenograft model was established. Treatments with C3G (cyanidin-3-glucoside, an anthocyanin) significantly suppressed the growth of SGC-7901 tumor xenografts in a dose-dependent manner. Immunohistochemical staining showed a significant increase in p21 expression, indicating that the cell cycle of tumor xenografts was inhibited. qPCR screening showed that C3G treatment up-regulated the expression of the KLF6 gene, which is an important tumor suppressor gene inactivated in many human cancers. Western blot showed that C3G treatments markedly increased KLF6 and p21 protein levels, inhibited CDK4 and Cyclin D1 expression, but did not notably change the expression of p53. These results indicated that KLF6 up-regulates p21 in a p53-independent manner and significantly reduces tumor proliferation. This study provides important information for the possible mechanism of C3G-induced antitumor activity against gastric adenocarcinoma in vivo.

MicroRNA-190 regulates FOXP2 genes in human gastric cancer.

OBJECTIVE: To investigate how microRNA-190 (miR-190) regulates FOXP2 genes in gastric cancer (GC) cell line SGC7901. METHODS: We identified that miR-190 could target FOXP2 genes by using dual luciferase enzyme assay. Precursor fragment transfection of miR-190 was performed with GC cell line SGC7901 and human gastric mucosal cell line GES-1. miR-190 expression was detected by reverse transcription-polymerase chain reaction (RT-PCR) and FOXP2 protein expression was measured by Western blotting. RESULTS: FOXP2-3'-untranslated region (UTR) in miR-190 transfection group was significantly decreased as compared with other groups. There were no significant differences in fluorescence signals of FOXP2mut-3'-UTR in each group. Therefore, it was assumed that miR-190 can target FOXP2 genes. Through RT-PCR verification, it was observed that the expression level of miR-190 was significantly higher in GC cell line SGC7901 than in human gastric mucosa cell line GES-1 after transfection with miR-190 mimics. The expression level of miR-190 was significantly higher in GES-1 cells than in SGC7901 cells after transfection with miR-190 inhibitors. Western blotting results showed the expression level of FOXP2 was significantly lower in GC cell line SGC7901 than in GES-1 cells. Compared with blank, mimics control, and inhibitors control groups, the miR-190 mimics group showed significantly enhanced proliferation, migration, and invasion abilities, while miR-190 inhibitors group showed decreased abilities toward proliferation, migration, and invasion (P<0.05). The transcription level of miR-190 and the expression level of FOXP2 in tumor tissues and adjacent normal tissues in GC patients were verified to be consistent with those of cell line experiments. CONCLUSION: Upregulation of miR-190 can lead to downregulation of FOXP2 protein expression. miR-190 may serve as a potential target for GC diagnosis.

Xiaotan Sanjie decoction inhibits interleukin-8-induced metastatic potency in gastric cancer.

AIM: To investigate the interaction between Xiaotan Sanjie (XTSJ) decoction and interleukin-8 (IL-8) and its effect on adhesion, migration and invasion of SGC-7901 gastric cancer cells. METHODS: SGC-7901 gastric cancer cells were exposed to serum containing XTSJ decoction and/or IL-8 (1 ng/mL). SGC-7901 cell adhesion to fibronectin, an extracellular matrix component, was detected using the Cell Counting Kit-8. Migration and invasion abilities of SGC-7901 cells were detected by scratch wound and Transwell chamber assays. Then, protein (immunofluorescence and Western blot) and mRNA levels (quantitative polymerase chain reaction) of cluster of differentiation 44 (CD44), a cell adhesion molecule, were measured in 72-h-cultured SGC-7901 cells. RESULTS: Cell adhesion was promoted by IL-8 (P = 0.001), but was inhibited by XTSJ decoction (P = 0.0001). Similarly, IL-8 promoted SGC-7901 cell invasion (P = 0.003), and XTSJ decoction inhibited cell invasion (P = 0.001). IL-8 induced SGC-7901 cell migration, but this was inhibited by XTSJ decoction. IL-8 up-regulated CD44 protein (P = 0.028) and mRNA expression (P = 0.002), whereas XTSJ decoction inhibited CD44 protein expression (P = 0.0001), but not mRNA expression (P = 0.275). An interaction between XTSJ decoction and IL-8 was confirmed in the invasion (P = 0.001) and CD44 mRNA expression of SGC-7901 cells (P = 0.010), but not in cell adhesion (P = 0.051). CONCLUSION: XTSJ decoction may inhibit adhesion, migration and invasion of gastric cancer cells, which is partly associated with down-regulation of IL-8.

Low-dose interleukin-8 induces the adhesion, migration and invasion of the gastric cancer SGC-7901 cell line.

Interleukin-8 (IL-8), an important inflammatory cytokine, is strongly associated with gastric cancer development and metastasis. High-dose (>1 ng/ml) IL-8 has been revealed to promote the adhesion, migration and invasion of human gastric cancer SGC-7901 cells in a dose-dependent manner. However, the IL-8 level produced by gastric cells is marginal, at even less than 1 ng/ml. It is unclear whether low-dose IL-8 also induces these capacities. In the present study, the effect of low-dose IL-8 on the adhesion, migration and invasion of the SGC-7901 cell line and the underlying molecular mechanism with regard to cluster of differentiation 44 (CD44) were investigated. The SGC-7901 cells were exposed to various concentrations of IL-8 (0, 0.2, 0.5, 0.8 and 1 ng/ml) in vitro. The adhesion of the SGC-7901 cells to fibronectin, an extracellular matrix component, was then detected by cell counting kit 8 assay. Migration and invasion abilities were evaluated by wound scratch and Transwell chamber assays. In addition, protein and mRNA levels of CD44 were measured using immunofluorescence and western blotting, and quantitative polymerase chain reaction, respectively, in cells cultured for 72 h. Following the exposure of the SGC-7901 cells to the various low doses of IL-8, the cell adhesion, migration and invasion capacities were promoted by IL-8, but not in a significant dose-dependent manner. Low-dose IL-8 upregulated the protein and mRNA expression of CD44. In conclusion, low-dose IL-8 potently induces the adhesion, migration and invasion of SGC-7901 cells, and the regulation of CD44 expression is one of the potential molecular mechanisms involved.

Studies on the identification of constituents in ethanol extract of Radix Glycyrrhizae and their anticancer activity.

BACKGROUND: The main functions of Radix Glycyrrhizae include regulating middle warmer, moistening lung, relieving toxicity, harmonizing property of drugs which is a traditional Chinese medicine widely used in clinical settings. The objective of the paper is to isolate and identify the constituents in ethanol extract of Radix Glycyrrhizae, and to study their anticancer activity. MATERIALS AND METHODS: Column chromatography, ODS column chromatography, preparative thin layer chromatography and NMR spectroscopy techniques were used to isolate compounds from ethanol extract of Radix Glycyrrhizae; optical microscopy and flow cytometry were used to determine the anticancer effect of Radix Glycyrrhizae extract. RESULTS: Four compounds were isolated from the ethanol extract of Radix Glycyrrhizae, namely oleanolic acid, isoliquiritin, glycyrrhetinic acid and licochalcone A. Optical microscopic observation showed that the growth of gastric cancer cell line SGC-7901 was inhibited in the experimental groups, and apoptotic morphological changes were observed in adherent cells; flow cytometry with PI staining showed that Radix Glycyrrhizae extract could induce SGC-7901 cell apoptosis within a concentration range of 0.5-1.5 mg/mL, compared with the control group, the apoptosis was positively correlated with the drug concentration, which exhibited an apparent dose-dependence. CONCLUSION: We conclude Ethanol extract of Radix Glycyrrhizae has an anti-proliferative activity on SGC-7901 cells.

A study on the extraction process of active ingredients from Akebia Stem and an analysis of their anti-gastric cancer activity.

The study investigated the extraction process of active ingredients from akebia stem and an analysis of their anti-gastric cancer activity. Three different extraction methods were used to obtain extracts, namely the decoction method (group A), reflux extraction method (group B), and maceration method (group C), of which reflux extraction method and maceration method used ethanol as the extraction solvent, while decoction method used distilled water for extraction. The differences in anti-gastric cancer activity of the three extracts were compared. MTT assay was used to test and compare the inhibitory effects of extracts obtained in A, B, and C groups on gastric cancer cells. The results showed that the dry extract obtained by heat reflux extraction with "water-ethanol" ratio of 1:2, extractant volume of 70 ml, with ethanol as extraction solvent presented the best inhibitory activity on gastric cancer SGC-7901 cells in this study. Its inhibitory effect did not change over time, and was directly proportional to the concentration.