Hepatocellular carcinoma (HCC) is a major cause of cancer-related mortality in part due to its high resistance to chemotherapeutic drugs. The anti-apoptotic Mcl-1 expression has been reported as a resistance factor in various types of tumors. Here, we investigated the expression of Mcl-1 in hepatoma cells and HCC tissues and its relationship with p53, and analyzed the possibility of the gene as a molecular target for HCC therapy. HCC specimens of 30 patients were examined by immunohistochemistry for Mcl-1 and p53 expression. Mcl-1 expression in hepatoma cell lines was measured by RT-PCR and Western blotting. The suppression of Mcl-1 by RNA interference or specific phosphatidylinositol-3 kinase (PI3K) inhibitor, LY294002, was evaluated as monotherapy, and it was combined with mitomycin C (MMC) in treating hepatoma cell line HepG2. Cell viability and apoptosis were assessed by MTT and FACS analysis. Finally, changes of Mcl-1 or p53 expression in various hepatoma cell lines were examined after transfection with Mcl-1 siRNA, the Mcl-1 expression plasmid, or the wide-type p53 expression plasmid, respectively. Mcl-1 protein was remarkably enhanced in HCC tissues as compared with adjacent non-tumor liver tissues. In addition, Mcl-1 was prominently expressed in HepG2 and Hep3B cells, weakly in SMMC7721 cells, and not in L02 cells. P53 protein was also overexpressed in HCC tissues and there was a significant correlation between the expression of p53 and Mcl-1. Silencing Mcl-1 by RNAi or LY294002 downregulated Mcl-1 expression and led to decreased cell viability and increased apoptosis. Combination of MMC and Mcl-1 RNAi or LY294002 exhibited a significant chemosensitizing effect. The expression of p53 was not influenced by Mcl-1 siRNA in HepG2 cells or transfection with the Mcl-1 expression plasmid in L02 cells. Furthermore, the expression of Mcl-1 in Hep3B cells was also not significantly changed after transfection with the wild-type p53 expression plasmid. It is concluded that Mcl-1 is overexpressed in HCC tissues. The mechanisms by which silencing Mcl-1 sensitizes hepatoma cells towards chemotherapy may be not attributed to the upregulated expression of p53 but the dysfunction of p53 through Mcl-1/p53 interaction. Mcl-1 may be a potential target of gene therapy for HCC.
Adenoma, Liver Cell ; drug therapy ; genetics ; pathology ; Apoptosis ; drug effects ; Biomarkers, Tumor ; biosynthesis ; genetics ; Chromones ; administration & dosage ; Gene Expression Regulation, Neoplastic ; drug effects ; Hep G2 Cells ; Humans ; Liver Neoplasms ; drug therapy ; genetics ; pathology ; Morpholines ; administration & dosage ; Myeloid Cell Leukemia Sequence 1 Protein ; biosynthesis ; genetics ; RNA, Small Interfering ; genetics ; Transfection ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics

OBJECTIVETo re-analyze the data published in order to explore plausible biological pathways that can be used to explain the anti-aging effect of curcumin.

METHODSMicroarray data generated from other study aiming to investigate effect of curcumin on extending lifespan of Drosophila melanogaster were further used for pathway prediction analysis. The differentially expressed genes were identified by using GeneSpring GX with a criterion of 3.0-fold change. Two Cytoscape plugins including BisoGenet and molecular complex detection (MCODE) were used to establish the protein-protein interaction (PPI) network based upon differential genes in order to detect highly connected regions. The function annotation clustering tool of Database for Annotation, Visualization and Integrated Discovery (DAVID) was used for pathway analysis.

RESULTSA total of 87 genes expressed differentially in D. melanogaster melanogaster treated with curcumin were identified, among which 50 were up-regulated significantly and 37 were remarkably down-regulated in D. melanogaster melanogaster treated with curcumin. Based upon these differential genes, PPI network was constructed with 1,082 nodes and 2,412 edges. Five highly connected regions in PPI networks were detected by MCODE algorithm, suggesting anti-aging effect of curcumin may be underlined through five different pathways including Notch signaling pathway, basal transcription factors, cell cycle regulation, ribosome, Wnt signaling pathway, and p53 pathway.

CONCLUSIONGenes and their associated pathways in D. melanogaster melanogaster treated with anti-aging agent curcumin were identified using PPI network and MCODE algorithm, suggesting that curcumin may be developed as an alternative therapeutic medicine for treating aging-associated diseases.

Aging ; drug effects ; genetics ; Animals ; Cell Cycle ; drug effects ; genetics ; Curcumin ; pharmacology ; Drosophila Proteins ; genetics ; metabolism ; Drosophila melanogaster ; drug effects ; genetics ; Gene Expression Regulation ; drug effects ; Gene Regulatory Networks ; drug effects ; Genes, Insect ; Protein Biosynthesis ; drug effects ; genetics ; Protein Interaction Maps ; drug effects ; genetics ; Receptors, Notch ; genetics ; metabolism ; Ribosomes ; drug effects ; metabolism ; Signal Transduction ; drug effects ; genetics ; Tumor Suppressor Protein p53 ; metabolism ; Wnt Signaling Pathway ; drug effects ; genetics

Ac-Phe-Lys-PABC-DOX (PDOX) is a smart doxorubicin (DOX) prodrug designed to decrease toxicities while maintaining the potent anticancer effects of DOX. This study was aimed at elucidating the effectiveness and toxicities of DOX and PDOX in patient-derived MCF-7 breast cancer cells in vitro. The MCF-7 cells were exposed to both PDOX and DOX, and cytotoxicities, cell cycle and P53/P21 signaling alterations were studied. Abundant cathepsin B was found in the MCF-7 cells, and treatment with PDOX and DOX triggered dose- and time-dependent cytotoxicity and resulted in a significant reduction in cell viability. The IC50 of PDOX and DOX was 3.91 and 0.94 μmol/L, respectively. Both PDOX and DOX caused an up-regulation of the P53/P21-related signal pathway, and PDOX significantly increased expression of P53 and caspase 3, and arrested the cell cycle at the G1/G2 phase. As compared with DOX, PDOX reduced toxicities, and it may have different action mechanisms on breast cancer cells.
Antibiotics, Antineoplastic ; pharmacology ; Breast Neoplasms ; drug therapy ; metabolism ; pathology ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; biosynthesis ; Doxorubicin ; analogs & derivatives ; pharmacology ; Drug Screening Assays, Antitumor ; methods ; Female ; G1 Phase ; drug effects ; G2 Phase ; drug effects ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Oligopeptides ; pharmacology ; Signal Transduction ; drug effects ; Tumor Suppressor Protein p53 ; biosynthesis

OBJECTIVETo observe the effect of Angelica sinensis polysaccharides (ASP) on the length of telomere, the activity of telomerase and the expression of P53 protein in mice hematopoietic stem cells (HSCs), and explore ASP's potential mechanism for regulating HSC aging.

METHODC57BL/6J mice were randomly divided into the normal group, the aging group and the intervention group. The aging group was radiated with X ray to establish the mice aging HSC model. The intervention group was orally administered with ASP during X-ray irradiation, while the normal group was orally administered with NS. Their HSCs were isolated by immunomagnetic beads. Cell cycles analysis and senescence-associated beta-galactosidase (SA-beta-Gal) staining were used to detect changes in aging HSCs. The expression of P53 was determined by western blot analysis. The length of telomere and the vitality of telomerase were analyzed by southern blot and TRAP-PCR, respectively.

RESULTCompared with the normal group, X-ray irradiation could significantly increase the cell ratio of in HSC G1 stage, rate of SA-beta-Gal positive cells and expression of P53 protein, and reduce the length of telomere and the vitality of telomerase. Compared with the aging group, ASP could significantly inhibit the cell ratio of in HSC G1 stage and the increase in the number of SA-beta-Gal positive cells, down-regulate the expression of P53 protein, and increase the length of telomere and the vitality of telomerase in HSCs.

CONCLUSIONASP could antagonize X-ray-induced aging of HSCs, which may be related to the increase in the length of telomere and the activity of telomerase, as well as the down-regulation of the expression of P53 protein.

Angelica sinensis ; chemistry ; Animals ; Cell Cycle ; drug effects ; physiology ; Cellular Senescence ; drug effects ; physiology ; Female ; Hematopoietic Stem Cells ; cytology ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Plant Extracts ; chemistry ; pharmacology ; Plants, Medicinal ; chemistry ; Polysaccharides ; isolation & purification ; pharmacology ; Telomerase ; biosynthesis ; metabolism ; Telomere ; drug effects ; metabolism ; Tumor Suppressor Protein p53 ; biosynthesis ; metabolism

As an essential metal for sustaining life, iron is involved in a number of metabolic processes, including DNA synthesis, electron transport, oxygen delivery, and so on. Iron metabolism involves the absorption, transport, and use of iron and is strictly regulated. Numerous studies have found a positive correlation between iron storage and the risk of tumors, such as colorectal carcinoma, hepatic cancer, renal carcinoma, lung cancer, and gastric cancer. In tumor cells, iron metabolism changes by several mechanisms, such as regulating the growth of tumor cells by transferrin, accelerating the uptake of iron by the overexpressions of transferrin receptors 1 and 2 (TfR1 and TfR2), synthesizing or secreting ferritin by some malignant tumor cells, and upregulating the level of hepcidin in patients with cancer. Some advances on diagnosis and treatment based on iron metabolism have been achieved, such as increasing the transfection and target efficiency of transferrin-polyethylenimine (PEI), inducing cell apoptosis by beta-guttiferin through interacting with TfR1.
Animals ; Antibiotics, Antineoplastic ; pharmacology ; Antigens, CD ; genetics ; metabolism ; Antimicrobial Cationic Peptides ; biosynthesis ; genetics ; Apoptosis ; Cell Proliferation ; Doxorubicin ; pharmacology ; Ferritins ; metabolism ; physiology ; Hepcidins ; Humans ; Interleukin-18 ; pharmacology ; Iron ; metabolism ; physiology ; Neoplasms ; metabolism ; pathology ; RNA, Messenger ; metabolism ; Receptors, Transferrin ; genetics ; metabolism ; Transferrin ; metabolism ; physiology ; Tumor Suppressor Protein p53 ; pharmacology

To study the inhibitory effect of a recombinant adenoviral vector carrying human IL-24 gene on SGC-7901 human gastric cancer cell. We infected the SGC-7901 gastric cancer cells with Ad blank adenovirus at various multiplicity of infection (MOIs) to find the optimal infective dose. The SGC-7901 tumor cells were infected with Ad-IL-24 at the optimal MOI in the following experiments. Adenovirus-mediated IL-24 transcription expression in SGC-7901 cells was examined by RT-PCR. The growth-suppressing effect of Ad-IL-24 on SGC-7901 tumor cells was assessed by MTT assay. Apoptosis and cell cycle of SGC-7901 tumor cells infected with Ad-IL-24 was evaluated by flow cytometer (FCM), respectively. The karyomorphology of apoptotic SGC-7901 tumor cells was examined using Hoechst33258 staining under fluorescence microscopy. The expression of apoptosis-related genes was future determined by semi-quantification RT-PCR; We demonstrated that the MOI of 100 was the optimal infective dose in the study on adenovirus-mediated IL-24 gene transfer into SGC-7901 gastric cancer cell; IL-24 gene mediated by adenovirus could successfully transcribe in SGC-7901 tumor cells; Ad-IL-24 could significantly inhibit SGC-7901 tumor cell growth and induce apoptosis, it also can up-regulate the express of bax, caspase-3 and p53 whilst down-regulate the bcl-2 expression. Thus, adenovirus-mediated IL-24 expression had marked anti-tumor effect in suppressing SGC-7901 human gastric cancer cell growth and inducing apoptosis, which may be closely associated with its up-regulation of bax/bcl-2, caspase-3 and p53.
Adenoviridae ; genetics ; metabolism ; Apoptosis ; genetics ; Caspase 3 ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Humans ; Interleukins ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; Recombinant Proteins ; biosynthesis ; genetics ; Stomach Neoplasms ; genetics ; pathology ; Transfection ; Tumor Suppressor Protein p53 ; genetics ; metabolism ; Up-Regulation ; bcl-2-Associated X Protein ; genetics ; metabolism

Animals ; Cell Culture Techniques ; methods ; Cell Line ; Cell Proliferation ; Hepatocytes ; cytology ; metabolism ; Humans ; Liver ; cytology ; Liver Diseases ; therapy ; Liver, Artificial ; Mice ; Recombination, Genetic ; Simian virus 40 ; genetics ; Telomerase ; genetics ; metabolism ; Transfection ; Tumor Suppressor Protein p53 ; biosynthesis

OBJECTIVE: To investigate the effect of human papillomavirus types 16E6 on the sensitivity of chemotherapy for cervical carcinoma in different p53 genotype cell lines. METHODS: The apoptosis rates of each group were detected by AO/EB, immunofluorescence and Annexin V/PI stained methods. The expressions of protein HPV16E6 and p53(mt) after the treatments of different concentration of DDP were detected by Western blot. HPV16E6 mRNA in C33A, C33A-E6, C33A-P, and CaSki cell lines under different DDP treatments were detected by RT-PCR. RESULTS: AO/EB and Annexin V/PI stained tests showed that the apoptosis rates of C33A, C33A-E6, C33A-P, and CaSki cells were significantly increased when DDP concentration increased. Western blot showed that the HPV16E6 protein could be detected only in C33A-E6 and CaSki cell lines. The expression of HPV16E6 protein in C33A-E6 and CaSki cell lines gradually decreased and was hardly detected with increased dosage of DDP and the prolonged treatment time (P<0.01), and slightly increased in C33A-E6 and Caski cell lines without the treatment, but there was no significant difference between them (P>0.05). Protein p53(mt) persistently expressed in C33A-E6, C33A, and C33A-P cell lines following the increased dosage of DDP and the prolonged treatment time(P>0.05), while it couldn't be found in CaSki cell line. RT-PCR showed that without DDP intervention, there was no significant difference of HPV16E6 mRNA in C33A-E6 and CaSki cell lines within 24 h.The HPV16E6 mRNA in C33A-E6 cell line expressed much higher than that in CaSki (P<0.05), and HPV16E6 mRNA of 2 cell lines expressed much higher at 48 h than at 24 h (P<0.05).The expression of HPV16E6 mRNA in C33A-E6 and CaSki cell lines gradually decreased with the increased DDP and prolonged treatment time (P<0.01), while there was no significant difference between C33A-E6 and CaSki cell lines under the same DDP concentration (P>0.05). CONCLUSION Effect of HPV16E6 on the sensitivity of chemotherapy for cervical carcinoma cell lines is not markedly related with the different p53 genotype forms(p53(mt)/p53wt ). HPV16E6 may affect the proliferation and sensitivity of chemotherapy in C33A cell line through other mechanism.
Antineoplastic Agents ; pharmacology ; Apoptosis ; genetics ; Cell Line, Tumor ; Cisplatin ; pharmacology ; Female ; Genotype ; Human papillomavirus 16 ; genetics ; Humans ; Oncogene Proteins, Viral ; biosynthesis ; genetics ; Papillomavirus Infections ; virology ; Repressor Proteins ; biosynthesis ; genetics ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics ; Uterine Cervical Neoplasms ; pathology ; virology

In present study, we investigated the mechanism of regulating HIF-1alpha expression by hydroxysafflor yellow A (HSYA) in Eahy 926 cell line under 1% O2 hypoxia. Eahy 926 cells were incubated with HSYA (100, 10 and 1 micromol x L(-1)) under hypoxia for the indicated time after treatment. Cell proliferation rate was detected using MTT assays. VHL and p53 location and protein expression were analyzed by immunocytochemical stain. HIF-1alpha, VHL and p53 mRNA expression were detected by RT-PCR. Protein expression of HIF-1alpha, VHL and p53 were assayed by Western blotting method. HSYA at 100 micromol x L(-1) increased Eahy 926 cells proliferation rate under hypoxia. HIF-1alpha mRNA and protein expression were up-regulated in the presence of HSYA. VHL, p53 mRNA and protein expression decreased significantly after 8 hours of treatment under hypoxia. HSYA protected Eahy 926 cells from hypoxia, and up-regulated HIF-1alpha expression partially via its inhibition of VHL and p53 expression.
Carthamus tinctorius ; chemistry ; Cell Hypoxia ; Cell Line ; Cell Proliferation ; drug effects ; Chalcone ; analogs & derivatives ; isolation & purification ; pharmacology ; Endothelial Cells ; cytology ; metabolism ; Flowers ; chemistry ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; biosynthesis ; genetics ; Plants, Medicinal ; chemistry ; Quinones ; isolation & purification ; pharmacology ; RNA, Messenger ; metabolism ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics ; Umbilical Veins ; cytology ; Up-Regulation ; Von Hippel-Lindau Tumor Suppressor Protein ; biosynthesis ; genetics

To study the anti-proliferation effect of lambda-carrageenan oligosaccharides (lambda-CO) on human umbilical vein endothelial cells (HUVECs) and expression of apoptotic relevant genes, the influence of lambda-CO on HUVECs proliferation was measured by MTT assay; apoptotic rate, cell cycle distribution and the level of active caspase-3 of HUVECs were analyzed using flow cytometry; the mRNA level of apoptosis related genes was determined by RT-PCR. At a high concentration of 1 mg x mL(-1), lambda-CO significantly inhibited the endothelial cell proliferation. Annexin-V FITC/PI double stain assay showed that when treated with 0, 0.8, 1 mg x mL(-1) of lambda-CO for 24 h, cell apoptotic rates were (1.67 +/- 1.6)%, (11.48 +/- 2.4)% and (13.81 +/- 2.2)%, respectively, when treated for 48 h, cell apoptotic rates were (2.02 +/- 2.3)%, (13.84 +/- 1.9)% and (38.72 +/- 2.5)%, respectively, cell cycle assay showed the decrease of cells in G0/G1 phase, and increase in S phase. Furthermore, we observed the level of active caspase-3 increased in a dose-dependent manner at 24 th and 48 th. RT-PCR results indicated that mRNA of TNFalpha, p53, caspase-8 and caspase-3 in cells increased after treated with lambda-CO. lambda-CO induce apoptosis of HUVECs in a dose-dependent way and arrests cells at S phase, which mainly due to the up-regulation of apoptotic genes such as TNFalpha, p53, caspase-8, caspase-3 and increase the level of active caspase-3.
Angiogenesis Inhibitors ; pharmacology ; Apoptosis ; drug effects ; Carrageenan ; pharmacology ; Caspase 3 ; genetics ; metabolism ; Caspase 8 ; biosynthesis ; genetics ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; drug effects ; Humans ; Oligosaccharides ; pharmacology ; RNA, Messenger ; metabolism ; Tumor Necrosis Factor-alpha ; biosynthesis ; genetics ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics ; Umbilical Veins ; cytology