OBJECTIVETo investigate the effect of hydroxycamptothecin (HCPT) on the proliferation, cell cycle and apoptosis of human lung carcinoma cell line A549.

METHODSThe growth of A549 cells exposed to HCPT was observed by staining with acridine orange/ethidium bromide dye. Agarose gel electrophoresis was performed to detect DNA fragmentation of the apoptotic cells. The cell cycle distribution of the exposed cells was analyzed using flow cytometry, and cell apoptosis was examined with annexin V-FITC/PI staining. RT-PCR was used to investigate Bcl-2 gene expression changes in the exposed cells.

RESULTSAgarose gel electrophoresis of the DNA from HCPT-treated cells showed a DNA ladder, and typical apoptotic appearance of the exposed cells was observed under fluorescence microscope. Treatment of A549 cells with 1 µmol/L HCPT for 24 h resulted in a cell apoptosis rate of 18.11%, significantly higher than the rate in control cells (0.09%, P<0.05). The treatment also caused a significant reduction of Bcl-2 mRNA expression by 70% (P<0.05).

CONCLUSIONHCPT can significantly inhibit the proliferation, induce apoptosis, and down-regulate Bcl-2 gene expression in human lung carcinoma cell line A549, suggesting the involvement of Bcl-2 gene in the inhibitory effect of HCPT on A549 cells.

Camptothecin ; analogs & derivatives ; pharmacology ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Down-Regulation ; Gene Expression Regulation, Neoplastic ; drug effects ; Genes, bcl-2 ; Humans ; Transfection

OBJECTIVETo study the expression changes of apoptosis related genes induced by cerulenin in multiple myeloma cell line U266 and explore its molecular mechanism.

METHODSThe expression changes of 96 apoptosis related genes were analyzed by superArray cDNA in U266 cells treated with cerulenin (20 microg/ml) for 12 h. Semi-quantitative RT-PCR was used to confirm the representative expression changes genes, Rip2, caspase 9 and TRAF2.

RESULTSAfter treated with cerulenin for 12 h, 44 apoptosis related genes expression in the U266 cells were changed, among which 41 were over 2 fold increase and 3 over 2 fold decrease. The expression of caspase 9 was increased markedly, indicating that mitochondria pathway played a key role in cerulenin inducing apoptosis and TRAF2 expression change suggested that nuclear factor (NF) participates in cerulenin inducing apoptosis.

CONCLUSIONThe death acceptor signaling pathway and the death acceptor non-dependence signaling pathway co-regulate cerulenin inducing apoptosis in U266 cells. Mitochondria pathway played the key role and nuclear factor (NF) participates in the apoptosis process.

Apoptosis ; drug effects ; genetics ; Cell Line, Tumor ; Cerulenin ; pharmacology ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; Multiple Myeloma ; genetics ; metabolism ; pathology ; Signal Transduction ; drug effects

OBJECTIVE: To explore the pathogenesis of gastric cancer through a bioinformatic approach to provide evidence for the prevention and treatment of gastric cancer. METHODS: The differentially expressed genes (DEGs) in gastric cancer and normal gastric mucosa in GSE79973 dataset were analyzed using GEO2R online tool. GO analysis and KEGG pathway enrichment analysis of the DEGs in DAVID database were performed. The protein interaction network was constructed using STRING database, and the key genes (Hub genes) were screened and their functional modules were analyzed using Cytoscape software. The GEPIA database was used to validate the Hub genes, and the Target Scan database was used to predict the microRNAs that regulate the target genes; OncomiR was used to analyze the expressions of the microRNAs in gastric cancer tissues and their relationship with the survival outcomes of the patients. RESULTS: A total of 181 DEGs were identified in gastric cancer, and 10 hub genes were screened by the protein- protein interaction network. Functional analysis showed that these DEGs were involved mainly in protein digestion and absorption, PI3K-Akt signaling pathway, ECM-receptor interaction and platelet activation signal pathway. GEPIA database validation showed that COL1A1 was highly expressed in gastric cancer tissues and was associated with a poor prognosis of patients with gastric cancer. MiR-129-5p was found to bind to the 3'UTR of COL1A1 mRNA, and compared with that in normal tissues, miR-129-5p expression was obviously down-regulated in gastric cancer tissues, and was correlated with the prognosis of the patients. CONCLUSIONS COL1A1 under regulation by MiR-129-5p is a potential therapeutic target for gastric cancer.
Collagen Type I ; drug effects ; Computational Biology ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; MicroRNAs ; therapeutic use ; Phosphatidylinositol 3-Kinases ; Stomach Neoplasms ; drug therapy

This study was purposed to explore the effects of baicalin on proliferation and apoptosis of adriamycin-resistant human myeloid leukemia cell line HL-60/ADR. HL-60/ADR cells were in vitro cultured and its proliferation inhibition was detected by MTT assay. The cell apoptosis was tested by Annexin V FITC/PI double staining analysis, DNA fragmentation and TUNEL labeling method. The expressions of c-myc and bcl-2 mRNA were detected by RT-PCR, and the protein expressions of C-MYC, BCL-2, caspase-3 precursor (procaspase-3), PARP and BAD were determined by Western blot. The results showed that baicalin could remarkably inhibited the HL-60/ADR cell proliferation, the cell doubling time was 48 hours, with an IC50 value of 28 micromol/L. Apoptosis occurred in dose dependent manner (20, 40, 80 micromol/L), and cell apoptosis in earlier and later stages could be detected by Annexin V FITC/PI double staining analysis, DNA fragmentation and TUNEL labeling method. The expressions of c-myc and bcl-2 mRNA in baicalin-treated cells decreased in a time-dependent manner (12, 24, 48 hours). Meanwhile, protein expressions of C-MYC, BBL-2, procaspase-3 and PARP (116 kD) were down-regulated in a time-dependent manner, while the expression of PARP (85 kD) and BAD were up-regulated. It is concluded that the baicalin efficiently induces proliferative inhibition and apoptosis in HL-60/ADR cells. All of above related genes and proteins may be involved in these processes.
Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; drug effects ; Flavonoids ; pharmacology ; Gene Expression Regulation, Neoplastic ; HL-60 Cells ; Humans

This study was purposed to investigate the changes of apoptosis-related gene expression in T lymphocytic leukemia JM cells induced with matrine, and its possible mechanism. JM cells was induced with 0.4 mg/ml matrine for 4 days, the total RNA was extracted from JM cells before and after matrine induction, the differential expression of apoptosis-related genes were screened with cDNA Expression Array Kit, the expression change of a part of gene was checked by Western blot. The results indicated that after induction of JM cells with matrine, differential expression of 31 genes were found by gene chip hybridization, the expression of caspase 8 was up-regulated more than 5 times. Western blot analysis showed that the up-regulation of caspase 8 gene expression positively correlated with induction time. It is concluded that differential expressions of many apoptosis-related genes in JM cells can be induced by matrine, in which gene expression of caspase 8 is up-regulated notably.
Alkaloids ; pharmacology ; Apoptosis ; drug effects ; genetics ; Caspase 8 ; metabolism ; Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Leukemia ; genetics ; Quinolizines ; pharmacology ; Up-Regulation

OBJECTIVETo detect the methylation status of 5'CpG island in the core promotor of maspin gene in RKO human colorectal cell line,and to explore the transcription regulation of DNA 5'CpG island demethylation on maspin tumor suppressor gene and its effect on the growth of cancer cell.

METHODSThe status of 5 'CpG island methylation of maspin gene in RKO human colorectal cell line was analyzed using methylation specific polymerase chain reaction (MSP). After treated with a specific demethylating agent, 5-Aza-2'-deoxycytidine, reverse transcription polymerase chain reaction (RT- PCR) was used to examine maspin gene expression. Cell proliferation was evaluated using MTT assay,distribution of cell cycle and rate of apoptosis were determined using flow cytometry.

RESULTSThe 5'CpG island methylation in the core promotor of maspin gene was detected in RKO human colorectal cell line. After treatment with three different concentration of 5-aza-2'-deoxycytidine, the expression of maspin mRNA increased 10.89, 16.91, 23.97 times respectively. MTT array showed the proliferation activity of RKO cell line was obviously reduced after 5-aza-2'-deoxycytidine treatment. The cells were arrested in G(0)/G(1) phase,and the apoptosis rates were 5.17%, 8.71% and 11.23% respectively compared with control group.

CONCLUSIONThe 5'CpG island methylation is probably responsible for maspin expression silencing in RKO human colorectal cell line, 5-aza-2'-deoxycytidine may effectively cause demethylation and inhibit the growth of tumor cell by reactivating the gene transcription silenced by aberrant hypermethylation.

Azacitidine ; analogs & derivatives ; pharmacology ; Cell Line, Tumor ; CpG Islands ; drug effects ; DNA Methylation ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Serpins ; genetics ; Transcription, Genetic ; drug effects

The study was aimed to investigate the anti-myeloma molecular mechanism of thalidomide (TLD) by detecting gene expression profiles of human myeloma cell line RPMI8226 treated with thalidomide. cDNA microarray were used to detect thousands of gene expression in gene chip. Two cDNA probes were prepared through reverse transcription from mRNA of RPMI8226 cells untreated and treated with TLD. These two probes were labeled with Cy3 and Cy5 fluorescence dyes respectively, then hybridized with cDNA microarray containing 1152 different human genes. The genes with differential expression in RPMI8226 cells treated with TLD for 72 hours were screened by scanning and analysis of computer software, and their functions were explored. The results showed that after co-culture of RPMI 8226 cells with TLD in 100 micromol/L concentration for 72 hours, 22 genes with differential expression were screened. Among these genes, the expressions of 4 genes were down-regulated including rpl32 gene, scya3 gene, mmp1 gene and igbp1 gene. Eighteen genes were up-regulated including wars gene, tubb4q gene, ube1l gene, txnrd1 gene and fyb gene. The study indicated that (1) wars gene encoding tryptophanyl-tRNA synthetase was up-regulated by TLD, while mmp1 gene encoding matrix metalloprotein 1 was down-regulated, they may be related to the inhibition of angiogenesis caused by TLD. (2) scya3 gene encoding macrophage inflammatory protein-1alpha and igbp1 gene encoding immunoglobulin binding protein 1 were down-regulated by TLD, they may play a role in the inhibition of cell proliferation caused by TLD. (3) tubb4q gene encoding tubulin beta4, ube1l gene encoding ubiquitin-activating enzyme E1-like protein and txnrd1 gene encoding thioredoxin reductase 1 were up-regulated by TLD, they may involve in apoptosis of RPMI8226 cells induced by TLD. (4) fyb gene encoding Fyn-binding protein was up regulated by TLD which associated with killing MM cells. It is concluded that 22 differentially expressed genes are involved in protein synthesis and degradation, cell signal transduction, cytoskeletal movement, immune modulation, cell metabolism, regulation of anti-oncogene and cell apoptosis, which relate directly or indirectly to molecular mechanisms of anti-myeloma effects induced by TLD.
Cell Line, Tumor ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Multiple Myeloma ; genetics ; Oligonucleotide Array Sequence Analysis ; Thalidomide ; pharmacology

Antineoplastic Agents, Phytogenic ; pharmacology ; therapeutic use ; Apoptosis ; drug effects ; Cyclin-Dependent Kinases ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Gene Expression ; drug effects ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Neoplasms ; drug therapy ; metabolism ; Protein-Tyrosine Kinases ; metabolism ; Signal Transduction

BACKGROUNDThe treatment of patients with small cell lung cancer (SCLC) is based on chemotherapy. However, the treatment is limited by the development of drug resistance. Emodin has been shown to exhibit an anti-cancer effect. But the molecular mechanism remains unclear. This study was conducted to investigate the effect of emodin on the gene expression profile changes in SCLC NCI-H446 cells.

METHODSNCI-H446 cells were treated with emodin and cell viability was determined by MTT assay. Cell apoptosis was determined by both flow cytometry and caspase-3 activity assay. The effect of emodin on the gene expression profile of NCI-H446 cells was analyzed using cDNA microarray. Semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) was used to validate the microarray results.

RESULTSEmodin suppressed viability, induced apoptosis and changed cell cycle of NCI-H446 cells. Among the 1262 genes, 10 genes were up-regulated and 8 genes were down-regulated more than 2 folds in NCI-H446 cells when compared with the control cells after treatment with emodin for 12 hours, while 12 genes were up-regulated and 24 genes were down-regulated after treatment with emodin for 24 hours. These genes were involved in metabolism, signal transduction, transcription regulation, cytoskeleton organization, immune response, transport, protein synthesis, cell cycle control, cell adhesion and RNA processing. The RT-PCR results were consistent with those obtained by the microarray.

CONCLUSIONSEmodin affects the expression of genes involved in various cellular functions and plays important roles in cell apoptosis, tumor metastasis and chemotherapy-resistance, which suggests emodin might become an effective chemopreventive or chemotherapeutic agent for SCLC.

Apoptosis ; drug effects ; Carcinoma, Small Cell ; drug therapy ; metabolism ; pathology ; Cell Line, Tumor ; Cell Survival ; drug effects ; Emodin ; pharmacology ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Lung Neoplasms ; Oligonucleotide Array Sequence Analysis ; Reverse Transcriptase Polymerase Chain Reaction

The study was designed to examine the effect of glucose on the expression of c-myc gene in cultured RINm5F cells. After monolayer culture was established in RPMI 1640 media supplemented with 10% fetal calf serum (FCS), the cells were cultured in various concentrations of glucose and 1 or 10% FCS for another 24 hours. A mRNA was extracted from the cultured cells by a single step method, and Northern analysis was done to detect RNA band. A 0.5 kilobase single band was detected as c-myc mRNA. The expression of c-myc gene mRNA was reduced with increased concentration of glucose with 1% FCS. However, supplementation of 10% FCS abolished the effect of glucose on expression of c-myc gene. These findings suggested that glucose in conjunction with other growth promoting factors played an important role in expression of oncogene and cell growth in RINm5F cells.
Animals ; Cell Line ; Gene Expression Regulation, Neoplastic/drug effects ; Genes, myc/*drug effects ; Glucose/*pharmacology ; Insulinoma/*genetics ; Pancreatic Neoplasms/*genetics ; Rats ; Tumor Cells, Cultured