Human Gastric Carcinoma Cell Research Articles

RETRACTED ARTICLE: Characterization of cancer stem-like cells in the side population cells of human gastric cancer cell line MKN-45

Side population (SP) cells may play a crucial role in tumorigenesis and the recurrence of cancer. Many kinds of cell lines and tissues have demonstrated the presence of SP cells, including several gastric cancer cell lines. This study is aimed to identify the cancer stem-like cells in the SP of gastric cancer cell line MKN-45. We used fluorescence activated cell sorting (FACS) to sort SP cells in the human gastric carcinoma cell line MKN-45 (cells labeled with Hoechst 33342) and then characterized the cancer stem-like properties of SP cells. This study found that the SP cells had higher clone formation efficiency than major population (MP) cells. Five stemness-related gene expression profiles, including OCT-4, SOX-2, NANOG, CD44, and adenosine triphosphate (ATP)-binding cassette transporters gene ABCG2, were tested in SP and MP cells using quantitative real-time reverse transcription polymerase chain reaction (RT-PCR). Western blot was used to show the difference of protein expression between SP and MP cells. Both results show that there was significantly higher protein expression in SP cells than in MP cells. When inoculated into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, SP cells show higher tumorigenesis tendency than MP cells. These results indicate that SP cells possess cancer stem cell properties and prove that SP cells from MKN-45 are gastric cancer stem-like cells.

Integrin-Linked Kinase Silencing Induces a S/G2/M Phases Cell Cycle slowing and Modulates Metastasis-Related Genes in SGC7901 Humancc Gastric Carcinoma Cells

Integrin-linked kinase has been implicated in metastasis of human tumors. Recent studies have also shown that the down-regulation of integrin-linked kinase has anti-tumor potential by inhibiting the metastatic potential of several types of cultured human cancer cells. However, the mechanism by which integrin-linked kinase regulates metastasis in human gastric carcinoma is not fully clear. We investigated the effect of integrin-linked kinase deletion on metastasis-associated markers in human gastric carcinoma SGC-7901 cell lines. We generated cell lines depleted for integrin-linked kinase. Cell adhesion and invasion were measured by the MTS assay and transwell assay. The cell cycle was measured by flow cytometry. Expression of metastasis-related genes was assessed by reporter assay, quantitative RT-PCR and western blotting. Our data showed an inhibitory effect on cell adhesion and invasion after silencing of integrin-linked kinase. The cell cycle was slowed in S/G2/M phases. Metastasis-related genes E-cadherin, MMP-2/9 and cystatin B, as well as the signaling molecules p-Akt, NF-κB, and AP-1 activation, were also modulated. Our results indicate that integrin-linked kinase plays an important role in metastasis of human gastric carcinoma cells. Down-regulation of integrin-linked kinase resulted in the impairment of the metastatic potential of gastric tumor cells by regulating metastasis-related gene expression, by inhibiting the Akt pathway as well as the activity of transcription factors. Integrin-linked kinase could be used as a potential therapeutic target.

Aquaporin 5 promotes the proliferation and migration of human gastric carcinoma cells

Aquaporin 5 (AQP5) promotes the progression and invasion of several cancers, but its role in the tumorigenesis of human gastric carcinoma (GC) has not been clearly defined. Here, we investigated the potential functions of AQP5 in the proliferation and migration of human GC. RT-PCR and western blotting were used to detect the expression of AQP5 in human GC cell lines. Immunohistochemistry was applied to evaluate the expression of AQP5 in human GC tissues and corresponding normal tissues. Following ectopic overexpression of AQP5 or inhibition of AQP5 by its inhibitor, acetazolamide (AZA), cell proliferation and migration of AGS cells were analyzed by MTT assay, colony formation assay, and wound healing assay. Heterogeneous expression of AQP5 mRNA and protein was observed in human GC cell lines MKN45, MKN28, AGS, and SGC7901. AQP5 was up-regulated in GC tissues in comparison to corresponding normal tissues. AQP5 protein was mainly localized in the cell membrane. Overexpression of AQP5 was correlated with enhanced lymph node metastasis. In vitro, overexpression of AQP5 notably enhanced, while inhibition of AQP5 by AZA significantly attenuated the proliferation and migration of AGS cells. Our data indicate that AQP5 may play an important role in the tumorigenesis and progression of human GC and suggest that AQP5 is a potential therapeutic target against GC.

Novel Liposomes Composed of Dimyristoylphosphatidylcholine and Trehalose Surfactants Inhibit the Growth of Tumor Cells along with Apoptosis

Novel liposomes composed of L-α-dimyristoylphosphatidylcholine (DMPC) and trehalose surfactant (DMTre) were produced and inhibitory effects of DMTre on the growth of human colon carcinoma (HCT116) and gastric carcinoma (MKN45) in vitro were examined. The remarkably high inhibitory effects of DMTre on the growth of HCT116 and MKN45 cells were obtained without affecting the growth of normal cells. The thickness of fixed aqueous layer of DMTre was larger than that of DMPC liposomes and increased in a dose-dependent manner. The induction of apoptosis by DMTre was revealed on the basis of flow cytometric analysis. DMTre induced apoptosis through the activation of caspases and mitochondria via Bax. It is noteworthy that remarkable inhibitory effects of DMTre on the growth of human colon and gastric carcinoma cells leading to apoptosis were obtained for the first time.

Amorphous calcium silicate hydrate/block copolymer hybrid nanoparticles: synthesis and application as drug carriers

Amorphous calcium silicate hydrate (CSH)/block copolymer monomethoxy(polyethyleneglycol)-block-poly(lactide-co-glycolide) (mPEG-PLGA) (CSHP) hybrid nanoparticles have been prepared in aqueous solution via a facile coprecipitation route at room temperature. The block copolymer mPEG-PLGA micelles in aqueous solution serve as the templates, and the CSH links to the micelles by hydrogen bonding between the silanol groups and the hydrophilic mPEG segments. The obtained core/shell hybrid nanoparticles can be transformed to solid hybrid nanoparticles by washing with water. The drug ibuprofen (IBU) loading capacity of the CSHP hybrid nanoparticles is ultrahigh (~1.9 g drug per g carrier), and the IBU loading efficiency can reach as high as ~100%. The drug release of loaded IBU in the CSHP hybrid nanoparticles has been investigated in simulated body fluid (SBF). The loaded IBU drug in CSHP hybrid nanoparticles can release in SBF for a long period of time (about 300 h), during which the CSHP nanocarrier is completely transformed to hydroxyapatite, exhibiting a good bioactivity. The IBU drug release profile can be interpreted by the Higuchi model. The loading capacity for the anticancer drug docetaxel (DTX) in the CSHP hybrid nanoparticles is about 82 mg g(-1). More importantly, the release of the loaded DTX in phosphate buffer saline (PBS) at pH 5.5 is obviously faster than that at pH 7.4, which is promising for the application in cancer therapy. Human gastric carcinoma (SGC-7901) cells can be effectively killed in the presence of the DTX-loaded CSHP hybrid nanoparticles at appropriate concentrations. Thus, the CSHP hybrid nanoparticles are promising nanocarriers in drug delivery. Furthermore, the hemoglobin (Hb) loading capacity of CSHP hybrid nanoparticles can achieve an utrahigh value of 995 mg g(-1), suggesting that the as-prepared CSHP hybrid nanoparticles are excellent protein adsorbents.

Identification of 5-hydroxy-3,6,7,8,3´,4´-hexamethoxyflavone from Hizikia fusiforme involved in the induction of the apoptosis mediators in human AGS carcinoma cells.

An 80% ethanol extract of Hizikia fusiforme was obtained and followed by successive fractionation using the organic solvents n-hexane, ethyl acetate, and n-butanol to identify the antioxidative substance. The aqueous part of the nbutanol fractionation step, showing high antioxidative activity, was subjected to reverse-phase liquid chromatography. As a result, a substance purified from a BB-2 fraction showed high antioxidative activity. The m/z 419 [M+H] molecular ion peak in the fraction was observed by the analysis of the ESI-LC/MS spectrum. By the analysis of 1H NMR (500 MHz, DMSO-d6) and 13C NMR (125 MHz, DMSO-d6) spectra, a unique compound of the fraction was biochemically identified as a 5-hydroxy-3,6,7,8,3´,4´- hexamethoxyflavone (5HHMF). We also investigated the effect of 5HHMF on human gastric AGS carcinoma cells. Western blot analysis suggested that the flavone substantially increased the levels of the death receptor-associated apoptosis mediators Fas, Fas L, FADD, TRADD, and DR4 in a concentration-dependent manner. The levels of Fas, Fas L, TRADD, and DR4 in the cells treated with 5HHMF (5 microgram/ml) were approximately 26.4-, 12.8-, 6.7-, and 9.8- times higher than those of non-treated cells, respectively. Of note, the level of FADD protein in the cells exposed to 5HHMF (1 microgram/ml) increased approximately 9.6-times. In addition, the cleavage of caspase-3, -8, and -9 in cultured AGS cells treated with 5HHMF was significantly confirmed. Therefore, our results suggest that 5HHMF from H. fusiforme is involved in the induction of death receptor-associated apoptosis mediators in human gastric AGS carcinoma cells.

Expression and regulatory function of miRNA-34a in targeting survivin in gastric cancer cells

We aimed to investigate the expression of microRNA-34a (miR-34a) in human gastric cancer cells and to evaluate the effects of miR-34a, acting via its gene survivin, on gastric cancer cell HGC-27 to provide potential new strategies for treating gastric cancer. In vitro cultures of the human gastric cancer cell lines MGC80-3, HGC-27, NCI-N87, and SGC-7901 and the normal human gastric epithelial cell line GES-1 were established. The expression of miR-34a in each gastric cancer cell line and GES-1 normal human gastric epithelial cell line was detected using quantitative real-time polymerase chain reaction (qRT-PCR). After the HGC-27 cells were transfected with a miR-34a mimic for 48 h, the changes in the expression levels of miR-34a were detected using qRT-PCR. The effect of miR-34a on HGC-27 cell viability was measured using a tetrazolium-based colorimetric [-(4,5)-dimethylthiahiazo-(-z-y1)-3,5-di-phenytetrazoliumromide (MTT)] assay. Flow cytometry was used to analyze the effects of miR-34a on HGC-27 cell proliferation. Annexin V/propidium iodide double staining and flow cytometry were used to analyze the effects of miR-34a on HGC-27 cell apoptosis. A Transwell invasion chamber was used to detect the effects of miR-34a on HGC-27 cell invasion. Finally, western blotting was used to analyze the effects of miR-34a on survivin protein expression. The qRT-PCR test determined that miR-34a expression in gastric cancer cells was significantly reduced compared to the normal gastric epithelial cell line GES-1 (p < 0.01). Compared to the control group, cellular miR-34a expression levels were significantly increased in HGC-27 human gastric carcinoma cells after transfection with a miR-34a mimic for 48 h (p < 0.01). The MTT assay demonstrated that after overexpressing miR-34a in HGC-27 cells, cellular viability was significantly reduced (p < 0.05). Flow cytometry analysis determined that upon miR-34a overexpression, the proliferation index decreased significantly (p < 0.05), and cellular apoptosis was significantly increased (p < 0.01). The Transwell invasion chamber assay illustrated that after increasing the expression of miR-34a, the number of cells passing through the Transwell chamber was significantly reduced (p < 0.01). Based on western blotting, compared with the control group, survivin protein expression levels were significantly decreased in the HGC-27 cells transfected with the miR-34a mimic for 48 h (p < 0.01). In conclusion, the expression level of miR-34a was downregulated in human gastric cancer cell lines. miR-34a can negatively regulate survivin protein expression and inhibit gastric cancer cell proliferation and invasion. Therapeutically enhancing miR-34a expression or silencing the survivin gene may benefit patients with gastric cancer.

MicroRNA-372 maintains oncogene characteristics by targeting TNFAIP1 and affects NFκB signaling in human gastric carcinoma cells

Aberrant microRNA (miRNA) expression has been investigated in gastric cancer, which is one of the most common malignancies. However, the roles of miRNAs in gastric cancer remain largely unknown. In the present study, we found that microRNA-372 (miR-372) directly targets tumor necrosis factor, α-induced protein 1 (TNFAIP1), and is involved in the regulation of the NFκB signaling pathway. Furthermore, overexpression of TNFAIP1 induced changes in AGS cells similar to those in AGS cells treated with miR-372-ASO. Collectively, these findings demonstrate an oncogenic role for miR-372 in controlling cell growth and apoptosis through downregulation of TNFAIP1. This novel molecular basis provides new insights into the etiology of gastric cancer.

Synthesis, topoisomerase-targeting activity and growth inhibition of lycobetaine analogs

The plant alkaloid lycobetaine has potent topoisomerase-targeting properties and shows anticancer activity. Based on these findings, several lycobetaine analogs were synthesized mainly differing in their substituents at 2, 8 and 9 position and their biological activities were evaluated. The topoisomerase-targeting properties and cytotoxicity of these structural analogs were assessed in the human gastric carcinoma cell line GXF251L. Performing a plasmid relaxation assay, an increased inhibition of topoisomerase I was found with N-methylphenanthridinium chlorides bearing a 8,9-methylenedioxy moiety or a methoxy group in 2-position. Furthermore, quaternized phenanthridinium derivatives bearing either a 2-methoxy or a 8,9-methylenedioxy moiety in conjunction with a 2-hydroxy or 2-methoxy group display potent topoisomerase II inhibition as shown by decatenation of kinetoplast DNA. In general, the N-methylphenanthridinium chlorides possess more potency in inhibiting topoisomerase I than topoisomerase II. All quaternized derivatives also exhibited potent inhibition of tumor cell growth in the low micromolar concentration range. Hence, N-methylphenanthridinium compounds were found to represent a promising class of compounds, potently inhibiting both, topoisomerases I and II, and may be further developed into clinically useful topoisomerase inhibitors.

Involvement of Aquaporin 3 in Helicobacter pylori-Related Gastric Diseases

BackgroundGastric cancer is one of the most common and lethal malignant cancers worldwide, and numerous epidemiological studies have demonstrated that Helicobacter pylori (H. pylori) infection plays a key role in the development of gastric carcinomas. Our previous studies showed that aquaporin 3 (AQP3) is overexpressed in gastric carcinoma and promotes the migration and proliferation of human gastric carcinoma cells, suggesting that AQP3 may be a potentially important determinant of gastric carcinoma. However, the role of AQP3 in H. pylori carcinogenesis is unknown.MethodsThe AQP3 protein and H. pylori were detected in human gastric tissues by immunohistochemistry and modified Giemsa staining respectively. AQP3 knockdown was obtained by small interfering (si) RNA. Western blot assays and RT-PCR were used to evaluate the change of AQP3 in the human gastric cancer AGS and SGC7901 cell lines after co-culture with H. pylori. Sprague Dawley rats were orally inoculated with H. pylori to establish a rat model colonized by H. pylori.ResultsThe present study found that AQP3 expression correlated with H. pylori infection status in gastric cancer tissues and corresponding normal mucosa, and H. pylori co-culture upregulated AQP3 expression in human gastric adenocarcinoma cells in vitro via the extracellular signal-regulated kinase signaling pathway. H. pylori infection also increased AQP3 expression in gastric mucosa colonized by H. pylori in a Sprague Dawley rat model.ConclusionsThese findings provide further information to understand the mechanism of H. pylori carcinogenesis and a potential strategy for the treatment of H. pylori-associated gastric carcinoma.

Inhibition of focal adhesion kinase induces apoptosis in human gastric carcinoma cells (SGC-7901)

Focal adhesion kinase (FAK), a non-receptor tyrosine kinase protein, acts as an early modulator of integrin signaling cascade, regulating basic cellular functions. In transformed cells, unopposed FAK signaling has been considered to promote tumor growth, progression and metastasis. The aim of this study was to assess the role of FAK in human gastric carcinoma cells. SGC-7901 cells were transfected with PGPU6/GFP/shNC (shNC), PGPU6/GFP/FAK-299 (shRNA-299), respectively. Expression of FAK was detected by real-time PCR and Western blots. MTT assay was used to examine changes in cell proliferation. Cell apoptosis was analyzed by flow cytometry. The expression of caspase-3, -9 was measured by Western blots. The expression of FAK in SGC-7901 cells significantly decreased in shRNA-299 group contrast to the control group (P < 0.01). Cells proliferation was inhibited by shRNA-299 and shRNA-299 + cisplatin, and the effects were clearly enhanced when cells treated with the anticancer agents. The level of cell apoptosis in shRNA-299 and shRNA-299 + cisplatin group was higher than in the control group (P < 0.01). The current data support evidence that down-regulation of FAK could induce human gastric carcinoma cells (SGC-7901) apoptosis through the caspase-dependent cell death pathway. Inhibition of the kinases may be important for therapies designed to enhance the apoptosis in gastric carcinoma.

Daidzein Induced Apoptosis via Down-Regulation of Bcl-2/Bax and Triggering of the Mitochondrial Pathway in BGC-823 Cells

Daidzein belongs to the group of isoflavones, found in a wide variety of plant-derived foods, especially in soybeans and soy-based foods. In this study, the effect of daidzein on human gastric carcinoma cells (BGC-823) and its mechanism were investigated. MTT assay was applied in the detection of the inhibitory effects of daidzein on cell proliferation. Hoechst-propidium iodide staining and flow cytometry were used to examine the apoptosis as well as the mitochondrial transmembrane potential. Western blotting was performed to detect the expression of apoptosis-associated proteins: cleaved PARP, cleaved caspase-9, cleaved caspase-3, Bcl-2, and Bax. Daidzein significantly inhibited the growth and proliferation of human gastric carcinoma cells (BGC-823) in a concentration- and time-dependent manner. Furthermore, it was found that an insult of daidzein to BGC-823 cells caused them to die by disruption of mitochondrial transmembrane potential, demonstrated not only by staining dead cells for phosphatidylserine but also by the up-regulation (cleaved PARP, cleaved caspase-9, cleaved caspase-3, Bax) and down-regulation (Bcl-2) of proteins associated with apoptosis and survival; whereas, the pan-caspase inhibitor z-VAD-fmk could partially rescue cells against damage of daidzein. Taken together, the results of this study demonstrate that daidzein significantly induces apoptosis via a mitochondrial pathway. Specifically, daidzein induced a change in the Bax/Bcl-2 ratios and activation of caspases-3 and -9 and the cleavage of PARP. Therefore, daidzein has the potential for use as a therapeutic agent for the treatment of gastric carcinoma.

The effects of transforming growth factor-β1 induced apoptosis of human gastric carcinoma cells related to Caspases

Objective To investigate the effects of transforming growth factor-β1 (TGF-β1) induced apoptosis.Methods Apoptosis proportion of human gastric SGC7901 carcinoma cells induced by TGF-β1 was tested by flow cytometry,blocking Caspase-8 by RNA interference (RNAi) ,the expression of Caspase-8 was analyzed by Western blotting.Results The apoptosis rate of SGC7901 cells obviously increased with the treatment of TGF-β1,an active fragment of caspase-8 started to occur 12 h after TGF-β1 treatment,whereas the caspase-9 active fragment occurred 24 h after TGF-β1 treatment. After RNAi against Caspase-8,the expression of Caspase-8 was decreased,11.2％ of control group at 24 h,and then can not be detected at 48 h,72 h,re-emergence at 96 h,9.2％ of control group.Conclusion TGF-β1 induced the apoptosis of SGC7901 cells via Caspases pathway,caspase-8 may be the first caspase activated;Suppression of caspase-8 by RNAi can effectively block the TGF-β1/Caspases signal pathway,and subsequently inhibits the apoptosis of SGC7901 cells. Key words: Gastric carcinoma; Transforming growth factor-β1; Caspase; Apoptosis

ABT-737 Synergizes with Arsenic Trioxide to Induce Apoptosis of Gastric Carcinoma Cells In Vitro and In Vivo

This study investigated the potential synergistic effects of two inducers of apoptosis: the small molecule ABT-737 and arsenic trioxide (ATO). Human gastric carcinoma cell lines SGC-7901 and MGC-803 were used to determine the effects of ABT-737 and ATO (alone or in combination) on cell proliferation and apoptosis in vitro. In vivo effects of these drugs were investigated in SGC-7901 solid tumours, grown in immunodeficient mice. ABT-737 and ATO inhibited proliferation and induced apoptosis in SGC-7901 and MGC-803 cells in concentration- and time-dependent manners, and showed a synergistic effect. ABT-737 disturbed the binding of B cell lymphoma (Bcl)-2 homologous antagonist killer and Bcl-extra large; ATO downregulated myeloid cell leukaemia (Mcl)-1 protein and upregulated Mcl-1short, the short splicing variant. ABT-737 and ATO significantly suppressed SGC-7901 xenograft growth, synergistically inhibited tumour growth and induced apoptosis in vivo. This study provides preclinical evidence that ABT-737 and ATO synergize to induce apoptosis of gastric carcinoma cells, suggesting that further investigation of these agents (as potential treatments for gastric cancer) is warranted.

In vitro effect of quercetin on human gastric carcinoma: Targeting cancer cells death and MDR

The benefits of plant polyphenols as chemotherapeutic agents are of great interest due to their possible anti-cancerogenic activities. Results available up to now suggest that flavonoid quercetin induces lethal effect in many types of tumours and may sensitize resistant cells to drugs. The aim of our study was to examine the effect of quercetin on human gastric carcinoma cells and to determine mode of its action. Parental EPG85-257P cell line and its daunorubicin-resistant variant EPG85-257RDB were used as cell models. Our data revealed that quercetin exerted antiproliferative impact on studied cells (with IC50 value of 12μM after 72h), mainly through induction of apoptosis. In sensitive cells cytostatic drug and flavonoid had synergistic effects, in EPG85-257RDB cells quercetin acted as a chemosensitizer. Its impact on resistance mechanism involved decrease of P-glycoprotein expression, inhibition of drug transport and downregulation of ABCB1 gene expression. The results demonstrate that quercetin may be considered as a prospective drug to overcome classical resistance in gastric cancer cells.

Inhibition of human gastric carcinoma cell growth in vitro by a polysaccharide from Aster tataricus

A water-soluble polysaccharide (WATP), with a molecular weight of 6.3×104Da, was isolated from Aster tataricus. According to gas chromatography (GC) analysis, WATP was composed of galactose, glucose, fucose, rhamnose, arabinose and mannose with molar ratios of 2.1:1.3:0.9:0.5:0.3:0.6. The effects of WATP on cell proliferation and apoptosis in human gastric cancer SGC-7901 cells were examined. MTT assay showed that WATP had a perfectly tumor growth inhibitory activity on SGC-7901 cells, but no cytotoxicity on SGC-7901 and primary human polymorphonuclear (PMN) cells analyzed using LDH assay. Flow cytometry analysis indicated that WATP could significantly induce apoptosis of SGC-7901 cells. Furthermore using Rh123 and Fluo-3 as fluorescent probes, respectively, it was found that mitochondrial transmembrane potential (ΔΨm) of treatment groups was significantly lower than that in un-treatment group and the concentration of calcium in cells exposed to WATP for 24h was increased in a dose dependent manner compared with unexposed group. These results suggest that WATP induces apoptosis of SGC-7901 cells through calcium- and ΔΨm-dependent pathways, indicating that it is potentially useful as a natural anti-cancer agent.

βig-h3 supports gastric cancer cell adhesion, migration and proliferation in peritoneal carcinomatosis

βig-h3 is an extracellular matrix protein and its expression is highly induced by transforming growth factor (TGF-β). It has also been suggested to play an important role in the growth and invasion of colon and pancreatic cancers. In the present study, we demonstrated that βig-h3 is expressed in mesothelial cells, especially in patients with advanced gastric cancer. The positive rate of βig-h3 was significantly higher in cases with a more invasive and advanced serous-type, with visible peritoneal metastasis, and in peritoneal lavage cytological examination (PLC) (+) and peritoneal lavage fluid CEA mRNA(+) subgroups (p<0.05). Our study also showed that the expression of βig-h3 gradually increased with increasing TGF-β1 concentrations in vitro in a time-dependant manner. In addition, βig-h3 also induced human gastric carcinoma cell line (SGC-7901) cell adhesion in a dose-dependent manner and significantly increased cell migration and proliferation. The results suggest that βig-h3 expression in peritoneal mesothelial cells in gastric cancer patients is a marker of the biological behavior of gastric cancer and plays an important role in the process of peritoneal carcinomatosis.

Anti-invasive effects of decitabine, a DNA methyltransferase inhibitor, through tightening of tight junctions and inhibition of matrix metalloproteinase activities in AGS human gastric carcinoma cells

The DNA methyltransferase inhibitor decitabine, 5-Aza-2'-deoxycytidine, possesses anti-metabolic and anticancer activities in various cancer cells. However, the biochemical mechanisms underlying decitabine-induced inhibition of invasiveness and metastasis have not been thoroughly studied. In this study, we investigated the effect of decitabine on the correlation between tightening of tight junctions (TJs) and anti-invasive activity in AGS human gastric cancer cells. Our data indicated that the inhibitory effects of decitabine on cell motility and invasiveness were associated with increased tightness of the TJ, which was demonstrated by an increase in transepithelial electrical resistance (TER). Immunoblotting results indicated that decitabine repressed the levels of the claudin proteins, major components of TJs that play a key role in the control and selectivity of paracellular transport. Furthermore, matrix metalloproteinase (MMP)-2 and -9 activity in the AGS cells was dose-dependently inhibited by treatment with decitabine, and this was correlated with a decrease in mRNA and protein expression. In addition, these effects were related to inactivation of the phosphoinositide 3-kinase (PI3K)/Akt pathway in AGS cells. In conclusion, this study suggests that TJs and MMPs are critical targets of decitabine-induced inhibition of invasiveness in AGS human gastric cancer cells.

Crosstalk between endoplasmic reticulum stress and oxidative stress in apoptosis induced by α-tocopheryl succinate in human gastric carcinoma cells

α-Tocopheryl succinate (α-TOS) has been shown to be a potent apoptosis inducer and growth inhibitor in a variety of cancer cells. Our previous studies showed the important role of endoplasmic reticulum (ER) stress and reactive oxygen species (ROS) generation in the apoptosis induced by α-TOS. However, the relationship of oxidative stress with ER stress is still controversial. The objective of the present study was to investigate the interplay between the two stress responses induced by α-TOS in SGC-7901 human gastric cancer cells. In response to α-TOS, cytological changes typical of apoptosis, induction of glucose-regulated protein 78 (GRP78) and CCAAT/enhancer-binding protein (C/EBP) homologous protein transcription factor (CHOP), and activation of caspase-4 were observed. And the antioxidant N-acetyl-l-cysteine inhibited induction of both GRP78 and CHOP by α-TOS transcriptionally and translationally. Furthermore, knocking down CHOP by RNA interference decreased ROS generation, increased glutathione level and induced glutathione peroxidase mRNA expression in α-TOS-treated cells, whereas catalase and superoxide dismutases mRNA expression were not altered. The results imply that α-TOS induces ER stress response through ROS production, while CHOP perturbs the redox state of SGC-7901 cells treated with α-TOS.

Transcriptional silencing of the inhibin-α gene in human gastric carcinoma cells

Although inhibin was first identified as a hormone regulating pituitary FSH secretion, it was later recognized to act as a tumor suppressor in the gonad and adrenal glands. Recently, the alpha subunit of this dimeric hormone (inhibin‑α) was reported to be involved in prostate tumorigenesis. To identify additional roles outside the reproductive axis, we investigated inhibin‑α gene activity and subsequent cell fate in human gastric cancer cells. The results were as follows: all the gastric cancer cells had at least one of a set of abnormalities including hypermethylation of the promoter, mutation of the 5'‑UTR or allelic imbalance including LOH in the inhibin‑α gene. Hypermethylation of the promoter and mutation of the 5'‑UTR in inhibin‑α were observed in SNU‑1, SNU‑5 and SNU‑484 cells. LOH was observed in AGS, KATO III, SNU‑5, SNU‑484 and SNU‑668 cells. Treatment with 5‑AzaC, a demethylating agent, induced demethylation of the inhibin‑α promoter in the SNU‑1, SNU‑5 and SNU‑484 cell lines, with the CpG5 site being strongly influenced by 5‑AzaC. In addition, inhibin‑α mRNA and protein were maintained at low levels in most of the gastric cancer cell lines. These low levels of mRNA and protein expression could be increased in most lines by treatment with 5‑AzaC. These increased inhibin‑α expression levels seemed to be closely associated with apoptosis and suppression of cell growth. Taken together, our results reveal that the inhibin‑α gene is transcriptionally silenced in human gastric cancer cells, and that reactivation of the gene suppresses their growth characteristics. This suggests that inhibin‑α may have a more general tumor suppressor activity outside the reproductive axis.