GDF15 knockdown promotes erastin-induced ferroptosis by decreasing SLC7A11 expression.

Ferroptosis is an iron-dependent form of regulated cell death. GDF15 affects various properties of cancer cells, but the role of GDF15 in ferroptosis has not been reported. In the present study, we found that GDF15 knockdown led to decreased expression of SLC7A11, which is a key component of system Xc- and a regulator of ferroptosis, indicating that GDF15 might play important roles in ferroptosis. CCK8 assay showed that GDF15 knockdown promoted erastin-induced ferroptosis in MGC803 cells. qRT-PCR and western blotting results demonstrated that GDF15 inhibition attenuated the increased SLC7A11 expression induced by erastin. Further study revealed that GDF15 knockdown promoted the decreased level of extracellular glutamate and intracellular GSH as well as the increased level of lipid ROS in the presence of erastin in MGC803 cells. Overall, the study shows that GDF15 knockdown promotes erastin-induced ferroptosis in MGC803 cells by attenuating the expression of SLC7A11 and the function of system Xc-.

IL-1ß promotes the nuclear translocaiton of S100A4 protein in gastric cancer cells MGC803 and the cell's stem-like properties through PI3K pathway.

It has been shown that nuclear expression of S100A4 is significantly correlated with increased metastasis and reduced survival in patients with gastric cancer and many other cancers. However, the factors which could influence the nuclear contents of S100A4 in cancer cells are not clear. It has also been reported that Interleukin-1ß (IL-1ß) promotes the nuclear translocation of S100A4 in chondrocytes. Previous studies have shown that IL-1ß promotes the stemness of colon cancer cells, and S100A4 is also involved in maintaining cancer-initiating cells in head and neck cancers. We speculate that IL-1ß might promote the nuclear translocation of S100A4 protein in MGC803 gastric cancer cells and therefore enhance their stem-like properties. The results from Western-blot and qRT-PCR analysis showed that IL-1ß increased the nuclear and total cellular content of S100A4 protein and S100A4 mRNA level in MGC803 cells. LY294002, a pharmacological inhibitor of Phosphoinositide 3-kinase (PI3K) reversed the above effects. Functional studies indicated that IL-1ß promoted the colony-forming and spheroid-forming capabilities of the cells and the expression of SOX2 and NANOG gene. PI3K or S100A4 inhibition reversed the IL-1ß-mediated increase in colony and spheroid-forming capabilities of the cells. LY294002 also reversed the elevated SOX2 and NANOG expression induced by IL-1ß. Our study demonstrated that IL-1ß promote the nuclear translocation of S100A4 protein in gastric cancer cells MGC803, which are PI3K dependent, suggesting the existence of IL-1ß-PI3K-S100A4 pathway for the first time. The study also showed that IL-1ß promoted stem-like properties of the cells through the new pathway.

miR-3189-3p Mimics Enhance the Effects of S100A4 siRNA on the Inhibition of Proliferation and Migration of Gastric Cancer Cells by Targeting CFL2.

GDF15 is a downstream gene of S100A4. miR-3189 is embedded in the intron of GDF15-and coexpressed with it. miR-3189-3p functions to inhibit the proliferation and migration of glioblastoma cells. We speculated that S100A4 might regulate miR-3189-3p to affect its function in gastric cancer cells. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) showed that miR-3189-3p expression was significantly downregulated in MGC803 cells after S100A4 knockdown. Overexpression of miR-3189-3p significantly inhibited the proliferation and migration of the cells. Moreover, miR-3189-3p mimics enhanced the effects of an S100A4 siRNA on the inhibition of cell proliferation and migration. Dual luciferase reporter assays, qRT-PCR, and Western blotting verified that CFL2 is a direct target of miR-3189-3p. CFL2 mediates the regulation of miR-3189-3p on the proliferation and migration of MGC803 cells. Data mining based on Kaplan-Meier plots showed that high CFL2 expression is associated with poor overall survival and first progression in gastric cancer. These data suggested that miR-3189-3p mimics enhanced the effects of the S100A4 siRNA on the inhibition of gastric cancer cell proliferation and migration by targeting CFL2. The findings suggested that when targeting S100A4 to treat gastric cancer, consideration and correction for counteracting factors should obtain a satisfactory effect.

FAM107B is regulated by S100A4 and mediates the effect of S100A4 on the proliferation and migration of MGC803 gastric cancer cells.

FAM107B expression was decreased in stomach cancer and many other kinds of cancer. The forced expression of FAM107B in HeLa cells diminished proliferation in response to growth factors, suggesting that FAM107B might play important roles in many types of cancers. But the mechanisms underlying the decreased expression of FAM107B in cancers are not clear, the functional significance needs to be further clarified. Our previous findings from cDNA microarray showed that there are 179 differentially expressed genes after S100A4 inhibition in gastric cancer cells MGC803. FAM107B was an upregulated one among them. In the present study, we confirmed that FAM107B expression was upregulated in MGC803 cells after S100A4 inhibition by qRT-PCR. We demonstrated for the first time that FAM107B was downregulated by S100A4. The results from CCK-8 and transwell assay showed that FAM107B inhibition by siRNA led to significantly increased proliferation and migrating abilities of MGC803 cells, respectively, indicating that FAM107B plays important roles in inhibiting the proliferation and migration of MGC803 cells. The rescue experiment showed that FAM107B-siRNA transfection reversed the reduced proliferation and migration abilities induced by S100A4 inhibition in the cells. These findings suggest that, as a downstream effector, FAM107B at least partly mediates the effect of S100A4 on the proliferation and migration of MGC803 cells. In conclusion, we first provide experimental evidence suggesting that FAM107B was downregulated by S100A4 in gastric cancer MGC803 cells. And FAM107B at least partially mediates the biological effect of S100A4 in the cells.

EZH2 Mediates the Regulation of S100A4 on E-cadherin Expression and the Proliferation, Migration of Gastric Cancer Cells.

BACKGROUND/AIMS: Several reports have showed the inverse correlation between S100A4 and E-cadherin expression, but the exact molecular mechanism remained unclear. It has been reported that EZH2 mediates transcriptional silencing of E-cadherin by trimethylating lysine 27 of histone H3 (H3K27me3). Therefore, we hypothesized that EZH2 might mediate the inhibition of S100A4 on E-cadherin and further affect the functions of S100A4 in gastric cancer cells. METHODOLOGY: RT-PCR and Western Blot were used to detect the expression of EZH2 and E-cadherin after inhibiting or increasing S100A4 expression. MTT and Transwell assay were performed to detect the proliferation and migration of gastric cancer cells. RESULTS: Inhibition or overexpression of S100A4 led to decreased or increased EZH2 expression, and increased or decreased E-cadherin expression. The SET domain was important for EZH2 in rescuing the decreased proliferation and migration of the cells after S100A4 inhibition. CONCLUSION: As a novel downstream target of S100A4, EZH2 mediates the inhibition of S100A4 on E-cadherin. The SET domain is important for EZH2 in mediating the cellular function of S100A4.

The TBX1/miR-193a-3p/TGF-ß2 Axis Mediates CHD by Promoting Ferroptosis.

Congenital heart disease (CHD) is the most common noninfectious cause of death during the neonatal stage. T-box transcription factor 1 (TBX1) is the main genetic determinant of 22q11.2 deletion syndrome (22q11.2DS), which is a common cause of CHD. Moreover, ferroptosis is a newly discovered kind of programmed cell death. In this study, the interaction among TBX1, miR-193a-3p, and TGF-ß2 was tested using quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blotting, and dual-luciferase reporter assays. TBX1 silencing was found to promote TGF-ß2 messenger ribonucleic acid (mRNA) and protein expression by downregulating the miR-193a-3p levels in H9c2 cells. In addition, the TBX1/miR-193a-3p/TGF-ß2 axis was found to promote ferroptosis based on assessments of lipid reactive oxygen species (ROS) levels, Fe2+ concentrations, mitochondrial ROS levels, and malondialdehyde (MDA) contents; Cell Counting Kit-8 (CCK-8) assays and transmission electron microscopy; and Western blotting analysis of glutathione peroxidase 4 (GPX4), nuclear factor erythroid 2-related factor 2 (NRF2), heme oxygenase-1 (HO-1), NADPH oxidase 4 (NOX4), and acyl-CoA synthase long-chain family member 4 (ACSL4) protein expression. The protein expression of NRF2, GPX4, HO-1, NOX4, and ACSL4 and the level of MDA in human CHD specimens were also detected. In addition, TBX1 and miR-193a-3p expression was significantly downregulated and TGF-ß2 levels were high in human embryonic CHD tissues, as indicated by the H9c2 cell experiments. In summary, the TBX1/miR-193a-3p/TGF-ß2 axis mediates CHD by inducing ferroptosis in cardiomyocytes. TGF-ß2 may be a target gene for CHD diagnosis and treatment in children.

S100A4 protects gastric cancer cells from anoikis through regulation of αv and α5 integrin.

Detachment from the extracellular matrix induces a form of programmed cell death termed anoikis. Resistance to anoikis permits cancer cells to survive in systemic circulation and facilitates their metastasis to distant organs. It is well known that S100A4 is overexpressed in many tumors and involved in tumor metastasis, but the mechanisms of the metastasis-promoting function of S100A4 are not fully understood. We hypothesized that S100A4 might play a role in anoikis of gastric cancer cells and further affects their metastasis. To test this hypothesis, we changed the expression of S100A4 by means of RNA interference or experimental overexpression and investigated the effect on anoikis. We found that knockdown of S100A4 by RNA interference led to significantly increased anoikis, whereas overexpression of S100A4 resulted in inhibition of anoikis. Furthermore, we provide evidence that inhibition of S100A4 resulted in the downregulation of α5 and αv integrin expression. These findings suggest that S100A4 protects gastric cancer cells from anoikis by regulation of αv and α5 integrin expression, which sheds a novel light for the role of S100A4 in cancer metastasis.

Subcellular distribution of S100A4 and its transcriptional regulation under hypoxic conditions in gastric cancer cell line BGC823.

It is well known that S100A4 is overexpressed in many tumors and involved in tumor invasion and metastasis. But the regulation of it is ill understood. We previously found that hypoxia mimicking cobalt chloride (CoCl(2)) enhanced the mRNA and protein expressions of the S100A4 gene in the gastric cancer cell line BGC823. In this study we found that S100A4 also displayed increased expression in BGC823 cells after exposure to real hypoxia (2.5% O(2)) as that by CoCl(2) treatment. Moreover, S100A4 protein showed different subcellular distribution under real hypoxia compared with that by CoCl(2) treatment or in normoxic conditions. To investigate the underlying molecular mechanism by which hypoxia regulates the expression of S100A4, we analyzed the regulatory sequences of the genes by bioinformatics and found a putative hypoxia responsive element (HRE) motif in the first intron of S1004. Furthermore, luciferase reporter assay showed that it is responsive to hypoxia. Electrophoretic mobility shift assay and chromatin immunoprecipitation assays demonstrated that hypoxia-inducible factor 1 (HIF-1) binds to the functional HRE in vitro and in vivo. The results provide evidence that S100A4 is a hypoxia-inducible gene, whose transcription is stimulated at least partly through the interaction of HIF-1 and HRE located at +329 to +334 of S100A4.

Genetic alterations in the PI3K pathway in prostate cancer.

Alterations in the PIK3CA and PTEN genes were assessed in 40 prostate tumors (radical prostatectomy samples). Genetic analyses in glands of the highest Gleason pattern within each tumor revealed PIK3CA amplification in 13%, PIK3CA mutations in 3%, PTEN homozygous deletion in 13% and PTEN hemizygous deletion in 8% of the cases analyzed. Supporting the view that PTEN and PIK3CA act in the same PI3K signaling pathway, genetic alterations in the PIK3CA and PTEN genes were mutually exclusive, except in one tumor. Overall, 13 of the 40 (33%) prostate tumors had alterations in the PI3K pathway. For cases with genetic alterations, other tumor areas with lower Gleason patterns as well as non-tumorous prostate glands were also analyzed. Of nine tumors with Gleason score 7, five cases contained the same genetic alterations in tumor areas of Gleason patterns 3 and 4, whereas in another four cases, genetic alterations were detected only in tumor areas of Gleason 4 but not Gleason 3 patterns. There were no alterations in non-tumorous glands. These results suggest that genetic alterations in the PI3K pathway are common in prostate cancer, and occur mainly through PIK3CA amplification and PTEN hemizygous or homozygous deletion. Glands of Gleason pattern 3 are genetically heterogeneous, some containing the same genetic alterations observed in glands of Gleason pattern 4.

[Effect of cobalt chloride on S100A4 expression in human gastric cancer cell BGC823].

S100A4 is an important metastasis-associated gene. Researches have confirmed the close correlation between overexpression of S100A4 gene and gastric cancer's infiltration, lymph node metastasis and in vitro invasiveness of gastric cancer cells. In order to investigate the mechanism of overexpression of S100A4 gene, hypoxia mimetic cobalt chloride (CoCl2) was used to treat gastric cancer cell BGC823, and then the expression of S100A4 mRNA and protein in BGC823 cells were detected by RT-PCR, immunohistochemistry, immunofluorescence, and Western blotting analysis. After treatment with CoCl2, the expression of S100A4 mRNA and protein in BGC823 cell was increased. These results suggested that hypoxia mimetic cobalt chloride could increase the expression of S100A4 gene in gastric cancer cell BGC823.

MicroRNA-375 inhibits the proliferation, migration and invasion of kidney cancer cells by triggering apoptosis and modulation of PDK1 expression.

Kidney cancer is one of the deadly cancers and is the cause of significant number of deaths worldwide. The treatments used for the treatment of kidney cancer are limited and associated with number of side effects. Therefore, there is need for the development of new drug options or to identify novel therapeutic targets for the treatment of kidney cancer. In this study we investigated the potential of miR-375 as the therapeutic target for the treatment of Kidney cancer. The results revealed that miR-375 is significantly downregulated in the Kidney cancer cells. To investigate the role therapeutic potential of miR-375, one kidney cancer cell line (A-498) was selected for further experimentation. It was observed that overexpression of miR-375 inhibits A-498 kidney cancer proliferation by induction of apoptosis. In addition, overexpression of miR-375 causes suppression of migration and invasion of the A-498 kidney cancers cells. Bioinformatic analysis revealed PDK1 to be putative target of miR-375 in Kidney cancer cells. The western blot analysis revealed the expression of PDK1 to be significantly upregulated in Kidney cancer cells and overexpression of miR-375 in A-498 cells caused inhibition of PDK1 preventing the phosphorylation of AKT (Thr308 and Ser473). Additionally, inhibition of PDK1 had similar effects as that of miR-375 overexpression on cell proliferation of A-498 kidney cancer cells. The inhibition of miR-375 expression could not rescue the effects of PDK-1 suppression on A-498 cell proliferation. In contrary, overexpression of PKD1 in A-498 cells transfected with miR-375 mimics could nullify the effects of miR-375 on proliferation of the A-498 cells. Taken together, we conclude that miR-375 regulates cell proliferation, migration and invasion of A-498 kidney cancer cells and may prove to be an important therapeutic target.

Valsartan prevents glycerol-induced acute kidney injury in male albino rats by downregulating TLR4 and NF-κB expression.

In this study, the protective effect of valsartan against glycerol-induced acute kidney injury (AKI) in male albino rats was investigated. Valsartan is used to treat high blood pressure and congestive heart failure and can prolong lifespan following a heart attack. The rats were divided into control, AKI, AKI + valsartan 100 mg/kg bw, and AKI + valsartan 200 mg/kg bw groups. Superoxide dismutase, glutathione peroxidase, catalase, lipid peroxidation, and reduced glutathione were assessed, and histopathological, immunohistochemical and western blot analyses were performed. Valsartan supplementation in AKI rats substantially increased superoxide dismutase, catalase, glutathione peroxidase, and glutathione levels but reduced the level of lipid peroxidation. Valsartan significantly reduced the severity of the renal tubular injury, renal lesions, and necrosis. Valsartan decreased NF-κB and TLR4 mRNA expression by >50% and their protein levels by >40%. Therefore, valsartan supplementation inhibited glycerol-induced functional and pathological damage to the kidney in a concentration-dependent manner. We propose that valsartan protects rat kidney tissue by downregulating NF-κB and TLR4 expression.

Decreased expression of DICER1 in gastric cancer.

BACKGROUND: The role of epigenetics in gene expression regulation and development significantly enhances our understanding of carcinogenesis. All the tumor related genes may be the target of epigenetical or genetic regulation. We selected some epigenetically regulated genes for cDNA array analysis and observed variability in the expression of the DICER1 gene in distinct stages of gastric cancer. The aim of this study was to assess the correlation between the expression of DICER1, an epigenetically regulated gene, and gastric cancer. METHODS: To detect the expression of 506 tumor-associated genes, including DICER1, in the matched cancerous mucosa, pre-malignant lesion (adjacent mucosa), non-cancerous gastric mucosa and distant lymphocyte metastatic lesion in 3 cases of gastric cancers using cDNA array. DICER1 mRNA expression and DICER1 protein expression were further analyzed by Real-time PCR and Western blot in 32 cases of progressive gastric cancer. DICER1 protein expression was also detected in 33 early and 30 progressive gastric cancers by the immunohistochemistry (IHC) method. RESULTS: In 3 cases of gastric cancer cDNA array showed dramatically decreased expression of DICER1 in pre-malignant lesion, cancerous mucosa and distant lymphocyte metastatic lesions compared with matched noncancerous gastric mucosa, pre-malignant lesion and cancerous mucosa. Real-time PCR results showed that the expression level of DICER1 mRNA in gastric cancer was significantly down-regulated compared to normal gastric tissue (P < 0.05). The IHC assay also showed that the expression of DICER1 was significantly decreased in progressive gastric cancer. Among the 63 cases of gastric cancers, 13/33 early (39.4%) and 19/30 (63.3%) progressive cancers showed negative expression of DICER1 (50.8%). The difference in expression of DICER1 between early and progressive gastric cancers was significant (P < 0.01). The result of Western blotting showed that DICER1 protein was down-regulated significantly in advanced gastric cancer (P < 0.05). CONCLUSIONS: DICER1 expression is decreased during the progression of gastric cancer, especially in progressive gastric cancers, which indicating DICER1 may play an important role in the development of cancer and the epigenetical regulation involved.

[Effect of B-RAF-specific RNA interference on gastric cancer BGC823 cell line].

To investigate the influence of B-RAF-specific RNA interference on the proliferation and apoptosis of gastric cancer BGC823 cell line. B-RAF-siRNA and scramble-siRNA were synthesized and transfected into BGC823 cells by TransMessenger. The expression of B-RAF gene and Bcl-2 gene in BGC823 cells was detected by RT-PCR and the level of apoptosis was evaluated in transfected cells by flow cytometry. Results showed that TransMessenger could effectively transfect B-RAF-siRNA and scramble-siRNA into BGC823 cells. B-RAF-siRNA significantly inhibited the expression of B-RAF gene and Bcl-2 gene in BGC823 cells by more than 90% to 100%. B-RAF-siRNA inhibited BGC823 cell prolifera-tion and induced apoptosis (P < 0.01). In conclusion, B-RAF-siRNA can effectively inhibit the expression of B-RAF gene and Bcl-2 gene, induce cell apoptosis and inhibit the proliferation of gastric cancer BGC823 cells.

Gene expression patterns in gastric cancer.

OBJECTIVE: To identify gene expression patterns in distinct stages of intestinal-type gastric cancer(GC). METHODS: Gene expression patterns of distinct stages of intestinal-type GC samples from 3 patients were compared with cDNA microarray, which contained 576 genes. There were 506 target genes, which included 51 genes identified from our previous experiment with suppression subtractive hybridization(SSH) and other 455 genes chosen for their important roles in cancers. Hierarchical clustering was performed to clarify genes in association with distinct stages of GC. RESULTS: One hundred and eighty-one differentially expressed genes with average Cy5:Cy3 ratios higher than 2.0 or lower than 0.5 in at least one stage of GC were identified by cDNA microarray. Among them, 48 genes were up-regulated and 133 down-regulated. Hierarchical clustering analysis separated the differentially expressed genes in different stages of GC into 5 main characteristic groups. Some important differentially expressed genes in different stages of GC were identified, such as SEC23IP, LIPF, ES(BQ291520), SLC5A1, PG(encoding similar to pepsin A precursor), CXCR4, DICER1, SH3GL2, and IGF2R. CONCLUSION: The differentially expressed gene patterns and some important genes were identified, which might be useful in further study on carcinogenesis, progression and metastasis of intestinal-type GC.

RNA interference-mediated silencing of speckle-type POZ protein promotes apoptosis of renal cell cancer cells.

This study aimed to investigate the effects of silencing the speckle-type POZ protein (SPOP) gene on renal cell cancer (RCC) cells and to explore its possible mechanism. The A498 and ACHN RCC cells were transfected with small interference RNA (siRNA)-SPOP by lipofection methods. The silencing efficiency was monitored by quantitative real-time polymerase chain reaction and Western blot. The effects of SPOP silencing on cell apoptosis, cell viability, colony formation ability, cell migration ability, and chemosensitivity to Sorafenib were assessed by flow cytometry, an MTT assay, a colony formation assay, a trans-well migration assay, and a CCK-8 assay, respectively. Its effects on the expression of several cytokines were determined by a protein microarray. Relevant signaling pathways were also analyzed. Compared with the control group, the cell apoptosis rate was significantly higher; the cell viability, the colony formation, and migration ability were significantly decreased in the siRNA-SPOP group. The protein microarray screening showed that the expression of vascular endothelial growth factor receptor, matrix metallopeptidase-9, vascular cell adhesion molecule-1, and stromal cell-derived factor-1 in the siRNA group was significantly decreased and that the expression of granulocyte-macrophage colony-stimulating factor and E-cadherin was significantly increased (P<0.05). The relevant signaling pathways were the integrin-mediated cell surface interactions pathway and extracellular matrix organization signal pathway. SPOP gene silencing induced cell apoptosis, decreased cell viability, colony formation, and migration ability, and elevated the drug sensitivity in the RCC cells. A possible mechanism is that silencing SPOP induces the differential expression of E-cadherin, vascular endothelial growth factor receptor, matrix metallopeptidase-9, and vascular cell adhesion molecule, which are related to the integrin-mediated cell surface interactions and extracellular matrix organization signaling pathway.

S100A4 influences cancer stem cell-like properties of MGC803 gastric cancer cells by regulating GDF15 expression.

Many studies have revealed that S100A4 is involved in cancer progression by affecting a variety of biological functions. Our previous study showed that S100A4 influences many biological properties of gastric cancer cells; however, the underlying mechanisms are far from clear. In this study, we used cDNA microarray analysis to investigate the global alterations in gene expression in MGC803 gastric cancer cells after siRNA-mediated S100A4 inhibition. Among the total genes investigated, 179 differentially expressed genes (38 upregulated and 141 downregulated) were detected in S100A4-siRNA transfected MGC803 cells compared with NC-siRNA transfected cells. We focused on the GDF15 gene, which was significantly downregulated after S100A4 inhibition. ChIP studies showed that the S100A4 protein binds to the GDF15 promoter, implicating S100A4 in GDF15 regulation at the transcriptional level. GDF15 overexpression promoted CSC-like properties of MGC803 cells, such as spheroid and soft-agar colony forming abilities. S100A4 inhibition suppressed the CSC-like properties of the cells, whereas, GV141-GDF15 vector transfection reversed these effects. Our results suggest that S100A4 influences the CSC-like properties of MGC803 gastric cancer cells by regulating GDF15 expression.

Analysis of metastasis suppressing function of E-cadherin in gastric cancer cells by RNAi.

AIM: To study the effect of inhibited E-cadherin expression on invasion of cancer cells. METHODS: We designed the nucleotide sequence of siRNA corresponding to 5' non-coding and coding sequence of E-cadherin. 21-nucleotide dssiRNA was synthesized by in vitro transcription with Ambion Silencer TM siRNA Construction Kit. siRNA was transfected into gastric cancer MKN45 using TransMessenger transfection Kit. RT-PCR and immunofluorescent assay were used to investigate the inhibition of the expression of mutated E-cadherin. Invasive ability of cancer cells was determined by Transwell assay. RESULTS: The synthesis of E-cadherin mRNA rather than protein expression was suppressed dramatically 7 d after interference. Decreased protein expression was observed on d 10 after interference. On d 11, invasion ability was enhanced significantly. CONCLUSION: siRNA targeted at non-coding and coding sequence of E-cadherin showed significant inhibition on mRNA and protein expression. Inhibited E-cadherin expression results in increased invasion ability of cancer cells.

[Screening and analysis of associated genes in the carcinogenesis and progression of gastric cancer].

OBJECTIVE: To screen and analyze the important associated genes in different stages of gastric cancer. METHODS: Using suppression subtractive hybridization (SSH) to screen differentially expressed genes; detecting the expression of genes in different stages of gastric cancer with dot blot hybridization; and verifying the results with semi-quantitative reverse transcriptase-polymerase chain reaction(RT-PCR). RESULTS: Twenty-six differentially expressed gene fragments were obtained by means of SSH. Among them,24 were known genes, 1 was a new expressed sequence tags(EST), and 1 was a hypothetical gene. The results of dot blot hybridization demonstrated that the expressions of Annexin A2, RPS29, RPS12 etc. in dysplasia were higher than those in normal mucosa; the expressions of RPS12 etc.in early cancer were higher than those in normal mucosa;the expressions of cytochromosome C oxidase II, ferritin light chain, RPS12 etc. in advanced gastric cancer and lymph node metastases were consistently higher than those in normal mucosa. The expression of proteasome 26S subunit gene in advanced gastric cancer was higher than that in normal mucosa. The expression of RPS12 was consistently higher in different stages of gastric cancer. It was demonstrated by RT-PCR that the expression of RPS12 in gastric cancer was higher than that in normal mucosa. CONCLUSION: The authors have identified some important genes that might be involved in the carcinogenesis and progression of gastric cancer, and RPS12 may play more important roles in gastric cancer.

S100A4 interacts with mutant p53 and affects gastric cancer MKN1 cell autophagy and differentiation.

The acquired p53 mutations are the most common genetic alterations in human cancers. Mutant p53 proteins tend to accumulate, augmenting their oncogenic potential. However, the mechanisms for mutant p53 accumulation are not known. Previous studies have shown that S100A4 interacts with wild­type p53. The present study marks the first time the effect of S100A4 on mutant p53 levels in gastric cancer MKN1 cells, which harbor mutant p53V143A, and the functional consequences have been investigated. S100A4 interacted with mutant p53V143A in the cells, and S100A4 inhibition decreased mutant p53V143A levels, indicating that S100A4 promoted mutant p53 accumulation through their interaction. We also found that S100A4 inhibition altered the expression of the mutant p53V143A target genes [c-Myc and inhibitor of DNA binding 2 (Id2)]. Moreover, we demonstrated that S100A4 knockdown increased mutant p53-related autophagy and cell differentiation. In conclusion, our data suggest a novel mechanism for mutant p53V143A accumulation and add a new facet to the role of S100A4 in cancer.