Gastric Cancer Cell Line MGC-803 Research Articles

GRB7 plays a promoting role in the progression of gastric cancer

BackgroundGastric cancer is a clinically common tumor, showing an upward trend of both incidence and mortality. GRB7 has been identified as a vital regulator in tumor progression. This study aims to uncover the biological function of GRB7 in gastric cancer process.Methodsimmunohistochemical (IHC) staining using a tissue microarray (TMA), quantitative reverse transcription PCR (qRT-PCR) and Western blotting were performed to detect the expression of genes. Furthermore, gastric cancer cell lines AGS and MGC-803 were transfected with short hairpin RNAs against GRB7. The biological function of GRB7 in gastric cancer cells were examined by CCK-8, flow cytometry, wound healing and Transwell assays. Then, in vivo tumor formation assay was conducted to explore the effects of GRB7 on tumor growth. Finally, expression levels of proteins related to cell functions were determined by Western blotting. Coimmunoprecipitation (CoIP) assay was performed to assess the protein-protein interaction.ResultsGRB7 was up-regulated in gastric cancer tissues and cell lines, and its expression was inversely proportional to survival of gastric cancer patients. Moreover, GRB7 knockdown inhibited proliferative, migratory abilities, as well as promoted cell apoptosis in gastric cancer cells. Further study suggested that GRB7 silencing could suppress gastric cancer tumor growth in vivo. Furthermore, our study uncovered an important interaction between GRB7 and MyD88. Silencing MyD88 was observed to alleviate the malignant phenotypes promoted by GRB7 in gastric cancer cells.ConclusionsTogether, this study provided evidence that GRB7 may be an effective molecular targets for the treatment of gastric cancer.

Microvesicles from bone marrow-derived mesenchymal stem cells promote Helicobacter pylori-associated gastric cancer progression by transferring thrombospondin-2

BackgroundOur previous study found that bone marrow-derived mesenchymal stem cells (BMSCs) promote Helicobacter pylori (H pylori)-associated gastric cancer (GC) progression by secreting thrombospondin-2 (THBS2). Extracellular vesicles (EVs) are important carriers for intercellular communication, and EVs secreted by BMSCs have been shown to be closely related to tumor development. The aim of this study was to investigate whether BMSC-derived microvesicles (MVs, a main type of EV) play a role in H. pylori-associated GC by transferring THBS2.MethodsBMSCs and THBS2-deficient BMSCs were treated with or without the supernatant of H. pylori for 12 h at a multiplicity of infection of 50, and their EVs were collected. Then, the effects of BMSC-derived MVs and THBS2-deficient BMSC-derived MVs on the GC cell line MGC-803 were assessed by in vitro proliferation, migration, and invasion assays. In addition, a subcutaneous xenograft tumor model, a nude mouse intraperitoneal metastasis model, and a tail vein injection metastasis model were constructed to evaluate the effects of BMSC-derived MVs and THBS2-deficient BMSC-derived MVs on GC development and metastasis in vivo.ResultsBMSC-derived MVs could be readily internalized by MGC-803 cells. BMSC-derived MVs after H. pylori treatment significantly promoted their proliferation, migration and invasion in vitro (all P < 0.05) and promoted tumor development and metastasis in a subcutaneous xenograft tumor model, a nude mouse intraperitoneal metastasis model, and a tail vein injection metastasis model in vivo (all P < 0.05). The protein expression of THBS2 was significantly upregulated after H. pylori treatment in BMSC-derived MVs (P < 0.05). Depletion of the THBS2 gene reduces the tumor-promoting ability of BMSC-MVs in an H. pylori infection microenvironment both in vitro and in vivo.ConclusionOverall, these findings indicate that MVs derived from BMSCs can promote H. pylori-associated GC development and metastasis by delivering the THBS2 protein.16ZaMehzoqZcatB8td6PJeVideo

Forkhead Box M1 Regulates the Proliferation,Invasion,and Drug Resistance of Gastric Cancer Cells via circ_NOTCH1

Objective To investigate the impacts of forkhead box M1(FOXM1)on the proliferation,invasion,and drug resistance of gastric cancer cells by regulating the circular RNA circ_NOTCH1.Methods Western blotting and real-time quantitative PCR were performed to determine the expression of FOXM1 protein and circ_NOTCH1,respectively,in the gastric cancer tissue,para-carcinoma tissue,human normal gastric mucosa epithelial cell line GES-1 and gastric cancer cell lines MGC-803,HGC-27,and BGC-823.BGC-823 cells were classified into the following groups:control,short hairpin RNA FOXM1(sh-FOXM1)and negative control(sh-NC),small interfering RNA circ_NOTCH1(si-circ_NOTCH1)and negative control(si-NC),and sh-FOXM1+circ_NOTCH1 overexpression plasmid(sh-FOXM1+pcDNA-circ_NOTCH1)and sh-FOXM1+negative control(sh-FOXM1+pcDNA).CCK-8 assay and clone formation assay were employed to measure the cell proliferation,and Transwell assay to measure cell invasion.After treatment with 1.0 mg/L adriamycin for 48 h,the cell resistance in each group was analyzed.Western blotting was employed to determine the expression levels of FOXM1,proliferating cell nuclear antigen(PCNA),Bax,multi-drug resistance-associated protein 1(MRP1),and multi-drug resistance gene 1(MDR1).RNA pull-down and RNA immunoprecipitation were employed to examine the binding of circ_NOTCH1 to FOXM1 protein.Results Compared with those in the para-carcinoma tissue,the expression levels of FOXM1 protein and circ_NOTCH1 in the gastric cancer tissue were up-regulated(all P<0.001).Compared with GES-1 cells,MGC-803,HGC-27,and BGC-823 cells showed up-regulated expression levels of FOXM1 protein and circ_NOTCH1(all P<0.001).Compared with the control group and sh-NC group,the sh-FOXM1 group with down-regulated expression of FOXM1 protein and circ_NOTCH1 showed decreased optical density value,clone formation rate,cell invasion number,and cell viability,down-regulated expression of PCNA,MRP1,and MDR1,and up-regulated expression of Bax protein in BGC-823 cells(all P<0.001).Compared with the control group and the si-NC group,the si-circ_NOTCH1 group with down-regulated expression of circ_NOTCH1 showed decreased optical density value,clone formation rate,cell invasion number,and cell viability,down-regulated expression of PCNA,MRP1,and MDR1,and up-regulated expression of Bax protein in BGC-823 cells(all P<0.001).Compared with sh-FOXM1 group and sh-FOXM1+pcDNA group,the sh-FOXM1+pcDNA-circ_NOTCH1 group with up-regulated expression of circ_NOTCH1 showed increased optical density value,clone formation rate,cell invasion number,and cell viability,up-regulated expression of PCNA,MRP1,and MDR1,and down-regulated expression of Bax protein(all P<0.001).FOXM1 protein was able to interact with circ_NOTCH1.Conclusion Interference with FOXM1 may inhibit the proliferation,invasion,and drug resistance of gastric cancer cells by silencing circ_NOTCH1 expression.

Long non-coding RNA ABHD11-AS1 promotes glycolysis in gastric cancer cells to accelerate tumor progression

To explore the role of long non-coding RNA ABHD11-AS1 in regulation of glycolysis in gastric cancer cells and its molecular mechanism. The null plasmid pcDNA-Vector and the overexpression plasmid pcDNA-ABHD11-AS1 were transfected into human gastric cancer cell lines MKN45 and MGC803 with low ABHD11-AS1 expression, and the changes in cell proliferation, colony formation, migration and invasion were examined using CCK-8 assay, colony formation assay and Transwell assay. Glucose uptake and lactate production of the cells were detected to assess the changes in glycolytic activity. The LncMAP database was used to identify potential transcription factors regulated by ABHD11-AS1, and the candidate transcription factor was determined by literature review, and the result was verified using Western blotting. Transfection with pcDNA-ABHD11-AS1 significantly increased ABHD11-AS1 expression in MGC803 and MKN45 cells, which exhibited obviously accelerated cell proliferation (P<0.05), increased colony formation rate and enhanced cell migration and invasion abilities (P<0.01). ABHD11-AS1 overexpression obviously promoted glycolysis in MGC803 and MKN45 cells (P<0.05). Analysis of the database suggested that ABHD11-AS1 may regulate the classical glycolysis-related gene c-Myc in gastric cancer cells. Western blotting demonstrated that the expression of c-Myc increased significantly after upregulating ABHD11-AS1 in gastric cancer cells. ABHD11-AS1 promotes glycolysis in gastric cancer cells by upregulating c-Myc to accelerate gastric cancer progression.

Synthesis and anticancer properties of celastrol derivatives involved in the inhibition of VEGF

In this study, fourteen celastrol derivatives (1–14) were synthesised by esterification of celastrol at the 29th position. The in vitro anticancer activity of them was determined by the MTT assay. All the synthetic compounds showed significant antiproliferative activity against six cancer cells, with IC50 of the submicron molar level. The most promising compound (2) blocked the cell cycle in the G2 phase and inhibited the expression of VEGF and MMP-9 in gastric cancer cell line MGC-803. In flow cytometry analysis, compound 2 induced cancer cell apoptosis in a dose-dependent manner. In the mouse tumour xenograft model, compound 2 showed significant anti-tumour activity in vivo at the dosage of 2.5 mg/kg and 1 mg/kg, with a higher inhibition rate than 5-FU (10 mg/kg). What’s more, the anticancer mechanism involved in the inhibition of VEGF and the toxicity evaluation of compound 2 were also investigated.

Targeting Snail1 by CRISPR/Cas9 System Inhibits the Proliferation and Migration of Human Gastric Cancer Cells

The zinc-finger transcriptional repressor Snail1 affects cancer progression by controlling the epithelial cell-mesenchymal transition. The RNA-guided clustered regularly interspaced short palindromic (CRISPR) with a CRISPR-associated nuclease 9 (Cas9) nuclease system has been extensively used for gene editing. Here, we used two distinct sgRNAs to successfully target Snail1 in the gastric cancer cell line MGC803 with the CRISPR/Cas9 system. Furthermore, we discovered that Snail1 knockout reduced the proliferation and migration of MGC803 cells.

MiR-28-5p's Targeting of GAGE12I Inhibits Proliferation, Migration, and Invasion of Gastric Cancer in Vitro.

GAGE12I is a tumor metastasis-promoting factor, which can induce gastric cancer cells to invade and migrate. We investigated the effect of miR-28-5p targeting GAGE12I on proliferation, invasion, and migration of human gastric cancer cell lines SGC-7901, AGS, and MGC-803. The expression levels of miR-28-5p and GAGE12I were detected by real-time PCR and western blot, respectively. Cell proliferation, migration, and invasion were measured by MTT and Transwell chamber. The interaction between miR-28-5p and GAGE12I was investigated by bioinformatics analysis and luciferase assay. Results showed that the expression of miR-28-5p in human gastric cancer cell lines was lower than that in normal gastric epithelial cells (P < 0.05). Overexpression of miR-28-5p suppressed cell proliferation, invasion, and migration (P < 0.05). GAGE12I was confirmed as a target of miR-28-5p. Cell proliferation, invasion, and migration were decreased in cells transfected with shGAGE12I compared with those of the scrambled group (P < 0.05). Collectively, miR-28-5p negatively regulated GAGE12I and reduced the proliferation, invasion, and migration of gastric cancer cells.

Discovery of novel coumarin-indole derivatives as tubulin polymerization inhibitors with potent anti-gastric cancer activities

Novel coumarin-indole derivatives were designed, synthesized and evaluated as tubulin polymerization inhibitors targeting the colchicine binding site. Among these compounds, compound MY-413 displayed the most potent inhibitory activities against gastric cancer cell line MGC-803 with an IC50 value of 0.011 μM. Furthermore, the IC50 values of compound MY-413 was less than 0.1 μM for other 17 cancer cell lines and less than 0.05 μM for other 8 cancer cell lines. Compound MY-413 effectively inhibited the tubulin polymerization (IC50 = 2.46 μM) by binding to the colchicine site. Screening for the inhibitory effects of compound MY-413 on 61 kinases, it was found that compound MY-413 could inhibit MAPK pathways-related kinases. Because of the inhibitory effects of compound MY-413 on tubulin polymerization and MAPK signaling pathway, compound MY-413 induced cell apoptosis, arrested the cell cycle in the G2/M phase, induced the inhibition of cell proliferation and migration in gastric cancer cells MGC-803 and HGC-27. In addition, compound MY-413 could significantly inhibit tumor growth in MGC-803 xenograft tumor models with tumor growth inhibition (TGI) rates of 70% (15 mg/kg) and 80% (30 mg/kg) without obvious toxicity. Consistent with the in vitro results, compound MY-413 also inhibited MAPK signaling pathway, and induced apoptosis and proliferation inhibition in vivo. In conclusion, this work indicated that compound MY-413 was a promising lead compound for the further investigation as a potential anti-gastric cancer agent.

Construction of a competing endogenous RNA network and identification of potential regulatory axes in gastric cancer chemoresistance

BackgroundEmerging evidence highlights the multifunctional role of noncoding RNAs (ncRNAs) in gastric cancer (GC) chemoresistance. However, the comprehensive expression profile and competing endogenous RNAs (ceRNAs) regulatory network of GC chemoresistance remain unanswered. MethodsThe whole-transcriptome sequencing (RNA sequencing) was performed to comprehensively analyze the differentially expressed (DE) lncRNAs, miRNAs and mRNAs in cisplatin-resistant cells MGC-803/DDP and GC cells MGC-803. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were conducted to investigate the biological functions implicated with the DEncRNAs. Then, the cisplatin-resistant-related ceRNA network and potential regulatory axes were constructed by bioinformatic analysis. ResultsWe successfully generated cisplatin-resistant GC cell line MGC-803/DDP. Differential expression analysis showed that a total of 1936 DElncRNAs, 2194 DEmRNAs and 174 DEmiRNAs were identified. Functional enrichment analysis indicated that those DEncRNAs were mainly involved in neuroactive ligand-receptor interaction, drug metabolism and Hippo signaling pathway. Subsequently, the cisplatin-resistant-related ceRNA network was constructed with the widely accepted vital chemo-resistant-related genes and signaling pathways. In addition, two constructed regulatory axes (include FAM66C/miR-129–5p/7 mRNAs and SFTA1P/miR-206/FN1 or NRP1) were successfully validated by the Genomic Data Commons (GDC) GC data. ConclusionsThe novel ceRNA network and the potential regulatory axes may provide the most comprehensive view of GC chemoresistance to date. Our findings uncovered potential biomarkers for prognostic prediction and novel therapeutic targets for reversing cisplatin resistance in GC.

LSD1 inhibitors from the roots of Pueraria lobata

One new 6a,11a-dehydropterocarpan derivative, 6-O-methyl-anhydrotuberosin (1), one new 6a-hydroxypterocarpan, (6aR,11aR,11bR)-hydroxytuberosone (7), and seven known compounds including two 6a,11a-dehydropterocarpans (2 and 4), two coumestans (3 and 5), one isoflavonoid (6) and two other phenolic compounds (8 and 9) were isolated from the roots of Pueraria lobata. The structures of the isolated compounds were elucidated with spectroscopic and spectrometric methods (1 D and 2DNMR, HRESIMS). Compounds 1, 2, 4–5 showed potent LSD1 inhibitory activities with IC50 values ranging from 1.73 to 4.99 μM. Furthermore, compound 2 showed potent cytotoxicity against gastric cancer cell lines MGC-803 and BGC-823, and lung cancer cell lines H1299 and H460.

Zinc finger protein 852 is essential for the proliferation, drug sensitivity, and self-renewal of gastric cancer cells.

Exploring cellular and molecular mechanisms responsible for gastric cancer growth, survival, self-renewal, and metastasis helps develop efficacious therapeutic strategies. In this study, the expression and function of zinc finger protein 852 (ZNF852) in human gastric cancer cell lines were characterized. ZNF852 was upregulated in gastric cancer cell lines relative to normal gastric epithelial cell line GES-1. When the ZNF852 gene was ablated in gastric cancer cell line MGC-803 using the CRISPR/Cas9-encoding lentivirus, the proliferation of MGC-803 was suppressed. ZNF852 deficiency also resulted in the inhibition of MGC-803 sphere formation, along with decreases in SRY-box 2 (SOX2), octamer-binding transcription factor 4 (OCT4), and Nanog homeobox (NANOG), suggesting that ZNF852 sustains self-renewal of MGC-803 cells. Furthermore, ZNF852 deficiency increased oxaliplatin-induced MGC-803 cell death, implying the role of ZNF852 in drug sensitivity. Subcutaneous infusion of MGC-803 cells into nude mice illustrated the same effects of ZNF852 on the proliferation and self-renewal of gastric cancer cells. Similar effects of ANF852 were also seen in gastric cancer cell line SNU-1. Interestingly, ZNF852 deficiency caused downregulation of epidermal growth factor receptor (EGFR) on gastric cancer cells. In summary, this study uncovers the positive regulatory role of ZNF852 in gastric cancer growth and maintenance. ZNF852 could be a potential therapeutic target for inhibiting gastric cancer initiation or progression.

Effects of long-chain noncoding RNA Linc00673 overexpression on proliferation and apoptosis of gastric cancer cell line MGC-803

Objective: To investigate the effects of long-chain noncoding RNA Linc00673 overexpression on proliferation and apoptosis of gastric cancer cells and its mechanisms. Methods: The recombinant lentivirus expressing plasmid pLVX-Linc00673 and the control empty plasmid pLVX-NC were packaged and amplified in 293T cells, and the recombinant lentivirus was transfected into gastric cancer cell line MGC-803 to establish a cell line stably overexpressing Linc00673. The expression of Linc00673 gene was detected by real-time fluorescence quantitative PCR. The growth and proliferation of cells were observed by MTT assay and clone formation assay. Cell cycle and apoptosis were detected by flow cytometry. The expressions of cell cycle related regulatory genes were detected by qPCR. The expressions of key molecules in the PI3K/Akt signaling pathway and tumor proliferation related proteins were detected by Western blot. Results: The expressions of Linc00673 in gastric cancer cell line MGC-803, BGC-823 and AGS were significantly higher than that in normal gastric mucosa cell line GES-1 (P＜0.05). MGC-803 cell line with stable overexpression of LINC00673 was established, and the expression level of LincC00673 was 200 times higher than that of the control empty carrier group. Overexpression of Linc00673 promoted proliferation of MGC-803 cells (P＜0.05) and clone formation (P＜0.05), inhibited cell apoptosis and affected the G1→S phase progression of cell cycle (P＜0.01). Overexpression of Linc00673 could affect the expressions of cell cycle regulatory gene CCNG2, P19 and CDK1 in MGC-803. Western blot showed that Linc00673 overexpression not only promoted the expressions of the key molecule pAkt in PI3K / Akt signaling pathway and its downstream target NF-κ B and Bcl-2 protein, but also up regulated the expressions of tumor related factors β-catenin and EZH2 proteins. Conclusion: Overexpression of Linc00673 may promote proliferation and inhibit apoptosis of MGC-803 cells through PI3K/Akt signaling pathway.

Arsenic trioxide inhibits the activity of SphK1 by decreasing the level of phosphatidylserine and phosphatidic acid in the human gastric cancer cell line MGC-803

Sphingosine kinase 1 (SphK1) is an important synthetase during the synthesis of sphingosine-1-phosphate (S1P) from sphingosine (Sph). Previous studies demonstrated that arsenic trioxide (As2O3) could reduce the level of S1P in human gastric cancer cell line MGC-803, indicating that As2O3 may inhibit the activity of SphK1. In this study, the effect of As2O3 on the SphK1 activation pathway was investigated. Western blot and quantitative real-time PCR analysis were used to evaluate the changes in protein and mRNA levels. The multi-dimensional mass spectrometry-based shotgun lipidomics method (MDMS-SL) was used for the quantitative detection of phosphatidylserine (PS) and phosphatidic acid (PA). The results revealed that As2O3 did not affect the protein and mRNA expression of SphK1 in the MGC-803 cells. However, As2O3 increased the levels of p-ERK1/2 and CIB1 in the SphK1 activation pathway and decreased the levels of PS and PA in the MGC-803 cells. The outcomes suggested that As2O3 may enhance the activity of SphK1 by increasing the levels of p-ERK1/2 and CIB1 and decrease the activity of SphK1 by decreasing the levels of PS and PA. It was suggested that the inhibition effect is stronger and resulting in an overall decrease in the activity of SphK1.

Arsenic trioxide inhibits the activity of SphK1 by decreasing the level of phosphatidylserine and phosphatidic acid in the human gastric cancer cell line MGC-803

Sphingosine kinase 1 (SphK1) is an important synthetase during the synthesis of sphingosine-1-phosphate (S1P) from sphingosine (Sph). Previous studies demonstrated that arsenic trioxide (As2O3) could reduce the level of S1P in human gastric cancer cell line MGC-803, indicating that As2O3 may inhibit the activity of SphK1. In this study, the effect of As2O3 on the SphK1 activation pathway was investigated. Western blot and quantitative real-time PCR analysis were used to evaluate the changes in protein and mRNA levels. The multi-dimensional mass spectrometry-based shotgun lipidomics method (MDMS-SL) was used for the quantitative detection of phosphatidylserine (PS) and phosphatidic acid (PA). The results revealed that As2O3 did not affect the protein and mRNA expression of SphK1 in the MGC-803 cells. However, As2O3 increased the levels of p-ERK1/2 and CIB1 in the SphK1 activation pathway and decreased the levels of PS and PA in the MGC-803 cells. The outcomes suggested that As2O3 may enhance the activity of SphK1 by increasing the levels of p-ERK1/2 and CIB1 and decrease the activity of SphK1 by decreasing the levels of PS and PA. It was suggested that the inhibition effect is stronger and resulting in an overall decrease in the activity of SphK1.

Lactate induces aberration in the miR-30a\u2013DBF4 axis to promote the development of gastric cancer and weakens the sensitivity to 5-Fu

BackgroundGastric cancer (GC) is one of the most common malignancies, molecular mechanism of which is still not clear. Aberrant expression of tumor-associated genes is the major cause of tumorigenesis. DBF4 is an important factor in cancers, although there is yet no report on its function and molecular mechanism in GC.MethodsThe expression of DBF4 in tumor tissues or cells of GC was detected by qRT-PCR and western blotting. Gastric cancer cell line MGC-803 and AGS were transfected with DBF4 siRNA or overexpression vector to detect the function of DBF4 in proliferation, migration and the sensitivity to 5-Fu with CCK-8 assay, colony formation assay, transwell assay, and wound healing assay. miR-30a was found to be the regulator of DBF4 by online bioinformatics software and confirmed with qRT-PCR, western blot and dual-luciferase reporter assays.ResultsIn our study, increased expression of DBF4 in GC tissues was first identified through The Cancer Genome Atlas (TCGA) and later confirmed using specimens from GC patients. Furthermore, functional experiments were applied to demonstrate that DBF4 promotes cell proliferation and migration in GC cell lines, moreover weakens the sensitivity of MGC803 and AGS cells to 5-Fu. We further demonstrated that miR-30a showed significantly lower expression in GC cells and inhibited the expression of DBF4 through 3ʹ-UTR suppression. Furthermore, rescue experiments revealed that the miR-30a-DBF4 axis regulated the GC cell proliferation, migration and the sensitivity to 5-Fu. The important composition in tumor microenvironment, lactate, may be the primary factor that suppressed miR-30a to strengthen the expression of DBF4.ConclusionsTaken together, our study was the first to identify DBF4 as a regulator of cell proliferation and migration in GC. Furthermore, our study identified the lactate-miR-30a-DBF4 axis as a crucial regulator of tumor progression and the tumor sensitivity to 5-Fu, which maybe serve useful for the development of novel therapeutic targets.

FTIR microspectroscopic study of biomacromolecular changes in As2O3 induced MGC803 cells apoptosis

It is well-known that As2O3 has significant anticancer effects, however, little is known regarding its mechanism for treating gastric cancer. Thus, we investigated biomacromolecular (DNA, proteins and lipids) changes of human gastric cancer cell line MGC803 to further understand As2O3-induced apoptosis. Conventional methods showed the increase of the apoptosis rate, the decrease of mitochondrial membrane potential (MMP), the accumulation of reactive oxygen species (ROS) and the changes of apoptotic proteins, etc. Fourier transform infrared (FTIR) microspectroscopy sensitively recognized overall biomacromolecular changes caused by the above: Peak-area ratios indicated the content/structure changes in DNA, proteins and lipids. Principle component analysis (PCA) revealed significant changes in intracellular DNA concentration and structure. This study suggests that As2O3 may exert anti-gastric cancer effect by altering intracellular biomacromolecules especially DNA.

AnnexinA5 Might Suppress the Phenotype of Human Gastric Cancer Cells via ERK Pathway.

Gastric cancer is one of the most fatal diseases around the world. However, the mechanism of the development of gastric cancer is still not clarified. In addition, the anticancer drugs have cytotoxicity with different degrees. AnnexinA5, a member of the annexin family, has a great binding ability with the membrane phospholipid in a calcium dependent manner and is involved in the development of various cancers. This study aims to explore the influence of annexinA5 on human gastric cancer cells and whether it has the potential to be an auxiliary treatment to gastric cancer. In this study, the role of annexinA5 was detected from both the endogenous and the exogenous aspects on the gastric cancer cell lines MGC-803 and MKN-45. The cells were divided into a knockdown group in which RNA interference technique was used to suppress annexinA5 expression and a protein-supplementing group in which annexinA5 protein was added in the culture supernatant. After the suppression ratio of RNA interference was determined and the IC50 of annexinA5 protein was decided respectively, the cells’ proliferation was detected by MTT assay, colony formation assay, and the expression of PCNA. FCM assay and PI staining methods were applied to test cell apoptosis and necrosis. To investigate whether ANXA5 influence cell metastasis, wound healing assay and transwell assay were employed. To further detect the mechanism of annexinA5 action, the signal pathway was examined with Western Blot method. When ANXA5 gene was knocked down, cell proliferation and metastasis were promoted, while cell apoptosis was suppressed. On the other hand, after the annexinA5 protein was applied to the gastric cancer cells, cell proliferation and metastasis were inhibited, while cell apoptosis and necrosis were promoted. AnnexinA5 played its role via ERK signal pathway. ANXA5 acted as tumor suppressor gene in the gastric cancer by suppressing ERK signal pathway and has the potentiality to be an auxiliary anticancer agent.

Effects of Birinapant on Proliferation and Invasion of MGC-803 Gastric Cancer Cells and Mechanism Underlying These Effects.

We studied the effects of birinapant, a mimetic of the second mitochondria-derived activator of caspase (SMAC), on invasion and proliferation of MGC-803 gastric cancer cells and the molecular mechanisms underlying these processes. The expression of cellular inhibitor of apoptosis 1 (cIAP1) and TNF receptor-associated factor 3 (TRAF3) in gastric cancer cell line MGC-803 and normal gastric mucosa GES-1 cells were analyzed by Western blotting and cell immunofluorescence assay. After pretreatment of MGC-803 cells with birinapant, a Transwell invasion assay was used to evaluate the cell invasion ability. MGC-803 cells were implanted under the skin of BALB/c nude mice. The tumors were removed 10 days later and its size was measured. Protein expression of proliferating cell nuclear antigen (PCNA) in the subcutaneous tumors was analyzed by immunohistochemical method. In addition, the expression of cIAP1, TRAF3, pNF-κB, and NF-κB in control and birinapant-treated cells was compared by Western blotting and the rate of cell apoptosis was evaluated by flow cytometry. In untreated MGC-803 gastric cancer cells, the expression of cIAP1 was higher and the expression of TRAF3 was lower than in normal gastric mucosa cell line GES-1. Pretreatment with birinapant inhibited the invasion and proliferation of MGC-803 cells and promoted cell apoptosis. Birinapant also promoted the expression of TRAF3 and inhibited the expression of cIAP1 and pNF-κB in MGC-803 cells. Thus, birinapant inhibited the expression of cIAP1, prevented degradation of TRAF3, and suppressed invasion and proliferation of MGC-803 cells by promoting cell apoptosis.

Bufalin inhibits peritoneal dissemination of gastric cancer through endothelial nitric oxide synthase-mitogen-activated protein kinases signaling pathway.

Peritoneal dissemination threatens the survival of patients with gastric cancer (GC). Bufalin is an extract of traditional Chinese medicine, which has been proved to have anticancer effect. The target of bufalin in suppressing gastric cancer peritoneal dissemination (GCPD) and the underlying mechanism are still unclear. In this research, GC cell line MGC-803 and high-potential peritoneal dissemination cell line MKN-45P were treated with bufalin or L-NAME. Malignant biological behavior and protein level of GC cell lines were detected with MTT, wound healing, transwell, adhesion, and western blotting. Bioinformatics analysis and patient tissues were used to verify the role of endothelial nitric oxide synthase (NOS3) in GC. Mice model was used to assess the effect of bufalin and role of NOS3 in vivo. We found that bufalin inhibited the proliferation, invasion, and migration in GC cell lines. NOS3, which was an independent prognostic factor of GC patients, was predicted to be a potential target of bufalin. Further experiments proved that bufalin reduced the phosphorylation of NOS3, thereby inhibiting the mitogen-activated protein kinase (MAPK) signaling pathway, and ultimately suppressed GCPD by inhibiting EMT process. In conclusion, NOS3 was a potential therapeutic target and prognostic biomarker of GC. Bufalin could suppress GCPD through NOS3-MAPK signaling pathway, which provided more evidence support for intraperitoneal perfusion of bufalin to treat GCPD.

Monobenzone, a Novel and Potent KDM1A Inhibitor, Suppresses Migration of Gastric Cancer Cells.

Lysine-specific demethylase1 (KDM1A) is generally highly expressed in various cancer tissues, and promotes the initiation and development of cancers via diverse cellular signaling pathways. Therefore, KDM1A is a promising drug target in many cancers, and it is crucial to find effective KDM1A inhibitors, while none of them has entered into market. With the help of compound library, monobenzone, a local depigmentor using as a treating over-pigmentation in clinic, was characterized as an effective KDM1A inhibitor (IC50 = 0.4507 μM), which may competitively inhibit KDM1A reversibly. Further cellular study confirmed that monobenzone could inhibit the proliferation of gastric cancer cell lines MGC-803 and BGC-823 with IC50 as 7.82 ± 0.55 μM and 6.99 ± 0.51 μM, respectively, and erase the substrate of KDM1A, H3K4me1/2 and H3K9 me2, and inhibit the migration of gastric cancer cell by reversing epithelial–mesenchymal transition (EMT). As the structure of monobenzone is very simple and small, this study provides a novel backbone for the further optimization of KDM1A inhibitor and gives monobenzone potential new application.