Inhibited Gastric Cancer Cell Proliferation Research Articles

MicroRNA-128b suppresses tumor growth and promotes apoptosis by targeting A2bR in gastric cancer

MicroRNAs (miRNAs) play crucial roles in the development and progression of human cancers, including gastric cancer (GC). The discovery of miRNAs may provide a new and powerful tool for studying the mechanism, diagnosis, and treatment of GC. In this study, we aimed to investigate the role and mechanism of miR-128b in the development and progression of GC. Quantitative real-time PCR (qRT-PCR) was used to measure the expression level of miR-128b in GC tissues and cell lines. We found that miR-128b was significantly down-regulated in GC tissues and cell lines. In addition, over-expression of miR-128b inhibited GC cell proliferation, migration and invasion of GC cells in vitro. Gain-of-function in vitro experiments further showed that the miR-128b mimic significantly promoted GC cell apoptosis. Subsequent dual-luciferase reporter assay identified one of the proto-oncogene A2bR as direct target of miR-128b. Therefore, our results indicate that miR-128b is a proto-oncogene miRNA that can suppresses GC proliferation and migration through down-regulation of the oncogene gene A2bR. Taken together, our results indicate that miR-128b could serve as a potential diagnostic biomarker and therapeutic option for human GC in the near future.

Amplification of ACK1 promotes gastric tumorigenesis via ECD-dependent p53 ubiquitination degradation.

Amplification or over-expression of an activated Cdc42-associated kinase 1 (ACK1) gene is common in breast, lung and ovarian cancers. However, little is known about the role of ACK1 in gastric tumorigenesis. Here, we found that DNA copy numbers of the ACK1 gene and its mRNA expression levels were significantly increased in gastric cancer (GC) compared to normal gastric tissues. Additionally, silencing ACK1 inhibited GC cell proliferation and colony formation, induced G2/M arrest and cellular apoptosis in vitro, and suppressed tumor growth in vivo. Gene Ontology annotation revealed that 147 differential proteins regulated by ACK1 knockdown were closely related with cellular survival. A cell cycle regulator, ecdysoneless homolog (ECD), was found to be significantly down-regulated by ACK1 knockdown. Silencing of ECD inhibited colony formation and induced G2/M arrest and cell apoptosis, which is similar to the effects of ACK1 knockdown. Silencing of ECD did not further enhance the effects of ACK1 knockdown on G2/M arrest and apoptosis, while silencing of ECD blocked the enhancement of colony formation by ACK1 over-expression. Over-expression of ACK or ECD promoted the ubiquitination of tumor suppressor p53 protein and decreased p53 levels, while silencing of ACK1 or ECD decreased the p53 ubiquitination level and increased p53 levels. Silencing of ECD attenuated the ubiquitination enhancement of p53 induced by ACK1 over-expression. Collectively, we demonstrate that amplification of ACK1 promotes gastric tumorigenesis by inducing an ECD-dependent ubiquitination degradation of p53.

The inhibition of histone deacetylase 8 suppresses proliferation and inhibits apoptosis in gastric adenocarcinoma.

Histone deacetylase 8 (HDAC8), a unique member of class I HDACs, shows remarkable correlation with advanced disease stage. The depletion of HDAC8 leads to inhibition of proliferation, apoptosis and cell cycle arrest in multiple malignant tumors. However, little is known about the contribution of HDAC8 to the tumorigenesis of gastric cancer (GC). The present study investigated expression of HDAC8 in GC cell lines and tissues, and the roles of HDAC8 inhibition in the proliferation, cell cycle and apoptosis of gastric cancer cells and explored the potential mechanisms. In the present study, quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), western blotting, and immunohistochemistry were used to examine the mRNA and protein expression of HDAC8 in GC cell lines and tissues. Then, the correlation between the clinicopathological parameters and the expression of HDAC8 was assessed. Finally, siRNA transfection and HDAC8 plasmid was performed to explore the functions of HDAC8 in GC progression in vitro. We found that the expression of HDAC8 was significantly upregulated both in GC cell lines and tumor tissues compared to human normal gastric epithelial cell, GES-1 and matched non-tumor tissues. Furthermore, depletion of HDAC8 remarkably inhibited GC cell proliferation, increased the apoptosis rate and G0/G1 phase percentage in vitro. Western blotting showed that the expression of protein promoting apoptosis such as, Bmf, activated caspase-3, caspase-6 were elevated following HDAC8 depletion. Our data exhibited an important role of HDAC8 in promoting gastric cancer tumorigenesis and identify this HDAC8 as a potential therapeutic target for the treatment of gastric cancer.

Long noncoding RNA HOXA-AS2 promotes gastric cancer proliferation by epigenetically silencing P21/PLK3/DDIT3 expression.

Current evidence suggests that long noncoding RNAs (lncRNAs) may be an important class of functional regulators involved in human cancers development, including gastric cancer (GC). Here, we reported that HOXA cluster antisense RNA2 (HOXA-AS2), a 1048bp RNA, was upregulated in GC. Increased HOXA-AS2 expression in GC was associated with larger tumor size and higher clinical stage; patients with higher levels of HOXA-AS2 expression had a relatively poor prognosis. Further experiments revealed that HOXA-AS2 knockdown significantly inhibited GC cells proliferation by causing G1 arrest and promoting apoptosis, whereas HOXA-AS2 overexpression promoted cell growth. Furthermore, HOXA-AS2 could epigenetically repress the expression of P21, PLK3, and DDIT3 via binding with EZH2 (enhaner of zeste homolog 2), a key component of PRC2; ChIP assays demonstrated that EZH2 could directly bind to the promoter of P21, PLK3 and DDIT3, inducing H3K27 trimethylated. In conclusion, these data suggest that HOXA-AS2 could be an oncogene for GC partly through suppressing P21, PLK3, and DDIT3 expression; HOXA-AS2 may be served as a candidate prognostic biomarker and target for new therapies in human GC.

Myricetin inhibits proliferation and induces apoptosis and cell cycle arrest in gastric cancer cells.

Myricetin is a flavonoid that is abundant in fruits and vegetables and has protective effects against cancer and diabetes. However, the mechanism of action of myricetin against gastric cancer (GC) is not fully understood. We researched myricetin on the proliferation, apoptosis, and cell cycle in GC HGC-27 and SGC7901 cells, to explore the underlying mechanism of action. Cell Counting Kit (CCK)-8 assay, Western blotting, cell cycle analysis, and apoptosis assay were used to evaluate the effects of myricetin on cell proliferation, apoptosis, and the cell cycle. To analyze the binding properties of ribosomal S6 kinase 2 (RSK2) with myricetin, surface plasmon resonance (SPR) analysis was performed. CCK8 assay showed that myricetin inhibited GC cell proliferation. Flow cytometry analysis showed that myricetin induces apoptosis and cell cycle arrest in GC cells. Western blotting indicated that myricetin influenced apoptosis and cell cycle arrest of GC cells by regulating related proteins. SPR analysis showed strong binding affinity of RSK2 and myricetin. Myricetin bound to RSK2, leading to increased expression of Mad1, and contributed to inhibition of HGC-27 and SGC7901 cell proliferation. Our results suggest the therapeutic potential of myricetin in GC.

The MicroRNA-217 Functions as a Potential Tumor Suppressor in Gastric Cancer by Targeting GPC5.

Gastric cancer (GC) is one of the most common malignancies worldwide. Emerging evidence has shown that aberrant expression of microRNAs (miRNAs) plays important roles in cancer progression. However, little is known about the potential role of miR-217 in GC. In this study, we investigated the role of miR-217 on GC cell proliferation and invasion. The expression of miR-217 was down-regulated in GC cells and human GC tissues. Enforced expression of miR-217 inhibited GC cells proliferation and invasion. Moreover, Glypican-5 (GPC5), a new ocncogene, was identified as the potential target of miR-217. In addition, overexpression of miR-217 impaired GPC5-induced promotion of proliferation and invasion in GC cells. In conclusion, these findings revealed that miR-217 functioned as a tumor suppressor and inhibited the proliferation and invasion of GC cells by targeting GPC5, which might consequently serve as a therapeutic target for GC patients.

MicroRNA-425-5p is upregulated in human gastric cancer and contributes to invasion and metastasis in vitro and in vivo.

Emerging evidence indicates that microRNAs are involved in the development and progression of gastric cancer (GC), functioning as tumor suppressors or oncogenes. The aim of the current study was to explore the potential mechanism of microRNA-425-5p (miR-425-5p) in GC. Using reverse transcription quantitative polymerase chain reaction, miR-425-5p expression was detected in GC cell lines (SGC-7901, MKN-28, MKN-45, BGC-823 and AGS) and the GES-1 cell line. Cell proliferation, soft agar colony formation, flow cytometry, haptotactic migration and matrigel chemoinvasion assays were used to test the proliferation, apoptosis, invasion and migration of BGC-823 cells transfected with miR-425-5p mimics or an inhibitor. Female BALB/c mice were randomly divided into three groups. Each experimental group contained five mice. BGC 823 cells were stably transfected with miR-425-5p inhibitor, wild type or negative control and were injected into the mice through the tail vein. It was found that miR-425-5p expression was significantly upregulated in all the GC cell lines when compared with the GES-1 cell line. Downregulation of miR-425-5p expression inhibited GC cell proliferation, invasion and migration. In the in vivo studies, downregulation of miR-425-5p expression suppressed pulmonary metastasis in the nude mice. Thus, miR-425-5p was demonstrated to have the potential to become a novel metastasis-associated gene; however, the mechanisms associated with these effects require further study. In the future, the development of miR-425-5p as a novel therapeutic strategy for the treatment of GC may be possible.

MAWBP and MAWD inhibit proliferation and invasion in gastric cancer.

To investigate role of putative mitogen-activated protein kinase activator with WD40 repeats (MAWD)/MAWD binding protein (MAWBP) in gastric cancer (GC). MAWBP and MAWD mRNA expression level was examined by real-time reverse transcriptase-polymerase chain reaction and semi-quantitative polymerase chain reaction in six GC cell lines. Western blotting was used to examine the protein expression levels. We developed GC cells that stably overexpressed MAWBP and MAWD, and downregulated expression by RNA interference assay. Proliferation and migration of these GC cells were analyzed by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl tetrazolium bromide (MTT), soft agar, tumorigenicity, migration and transwell assays. The effect of expression of MAWBP and MAWD on transforming growth factor (TGF)-β1-induced epithelial-mesenchymal transition (EMT) was examined by transfection of MAWBP and MAWD into GC cells. We detected the levels of EMT markers E-cadherin, N-cadherin and Snail in GC cells overexpressing MAWBP and MAWD by Western blotting. The effect of MAWBP and MAWD on TGF-β signal was detected by analysis of phosphorylation level and nuclear translocation of Smad3 using Western blotting and immunofluorescence. Among the GC cell lines, expression of endogenous MAWBP and MAWD was lowest in SGC7901 cells and highest in BGC823 cells. MAWBP and MAWD were stably overexpressed in SGC7901 cells and knocked down in BGC823 cells. MAWBP and MAWD inhibited GC cell proliferation in vitro and in vivo. MTT assay showed that overexpression of MAWBP and MAWD suppressed growth of SGC7901 cells (P < 0.001), while knockdown of these genes promoted growth of BGC823 cells (P < 0.001). Soft agar colony formation experiments showed that overexpression of MAWBP and MAWD alone or together reduced colony formation compared with vector group in SGC7901 (86.25 ± 8.43, 12.75 ± 4.49, 30 ± 6.41 vs 336.75 ± 22.55, P < 0.001), and knocked-down MAWBP and MAWD demonstrated opposite effects (131.25 ± 16.54, 88.75 ± 11.12, 341.75 ± 22.23 vs 30.25 ± 8.07, P < 0.001). Tumorigenicity experiments revealed that overexpressed MAWBP and MAWD inhibited GC cell proliferation in vivo (P < 0.001). MAWBP and MAWD also inhibited GC cell invasion. Transwell assay showed that the number of traverse cells of MAWBP, MAWD and coexpression group were more than that in vector group (84 ± 16.57, 98.33 ± 9.8, 29 ± 16.39 vs 298 ± 11.86, P < 0.001). Coexpression of MAWBP and MAWD significantly decreased the cells traversing the matrix membrane. Conversely, knocked-down MAWBP and MAWD correspondingly promoted invasion of GC cells (100.67 ± 14.57, 72.66 ± 8.51, 330.67 ± 20.55 vs 27 ± 11.53, P < 0.001). More importantly, coexpression of MAWBP and MAWD promoted EMT. Cells that coexpressed MAWBP and MAWD displayed a pebble-like shape and tight cell-cell adhesion, while vector cells showed a classical mesenchymal phenotype. Western blotting showed that expression of E-cadherin was increased, and expression of N-cadherin and Snail was decreased when cells coexpressed MAWBP and MAWD and were treated with TGF-β1. Nuclear translocation of p-Smad3 was reduced by attenuating its phosphorylation. Coexpression of MAWBP and MAWD inhibited EMT, and EMT-aided malignant cell progression was suppressed.