BACKGROUNDTetrahydrobiopterin (BH4) is an essential cofactor of nitric oxide synthases (NOSs) for the synthesis of nitric oxide (NO). BH4 therapy can reverse the disease-related redox disequilibrium observed with BH4 deficiency. However, whether BH4 exerts a protective effect against radiation-induced damage to cardiomyocytes remains unknown.

METHODSClonogenic assays were performed to determine the effects of X-ray on H9c2 cells with or without BH4 treatment. The contents of lactate dehydrogenase (LDH), superoxide dismutase (SOD), and malondialdehyde (MDA) in H9c2 cells were measured to investigate oxidative stress levels. The cell cycle undergoing radiation with or without BH4 treatment was detected using flow cytometry. The expression levels of proteins in the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/P53 signaling pathway, inducible NOS (iNOS), and endothelial NOS (eNOS) were examined using Western blotting.

RESULTSX-ray radiation significantly inhibited the growth of H9c2 cells in a dose-dependent manner, whereas BH4 treatment significantly reduced the X-ray radiation-induced growth inhibition (control group vs. X-ray groups, respectively, P< 0.01). X-ray radiation induced LDH release, apoptosis, and G0/G1 peak accumulation, significantly increasing the level of MDA and the production of NO, and decreased the level of SOD (control group vs. X-ray groups, respectively, P < 0.05 or P < 0.01). By contrast, BH4 treatment can significantly reverse these processes (BH4 treatment groups vs. X-ray groups, P < 0.05 or P < 0.01). BH4 reversed the X-ray radiation-induced expression alterations of apoptosis-related molecules, including B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein, and caspase-3, and molecules of the PI3K/Akt/P53 signaling pathway. BH4 enhanced the production of NO in 2 Gy and 4 Gy radiated groups by upregulating eNOS protein expression and downregulating iNOS protein expression.

CONCLUSIONSBH4 treatment can protect against X-ray-induced cardiomyocyte injury, possibly by recoupling eNOS rather than iNOS. BH4 treatment also decreased oxidative stress in radiated H9c2 cells.

Animals ; Antioxidants ; metabolism ; Apoptosis ; drug effects ; Biopterin ; analogs & derivatives ; pharmacology ; Cell Cycle ; drug effects ; Cell Line ; Enzyme-Linked Immunosorbent Assay ; L-Lactate Dehydrogenase ; metabolism ; Myocytes, Cardiac ; cytology ; drug effects ; radiation effects ; Rats ; Signal Transduction

OBJECTIVETo study the radiosensitizing effect of resveratrol on hypopharyngeal carcinoma cell line FADU in vitro.

METHODSHypopharyngeal carcinoma cell line FADU was cultured in in vitro DMEM. Its inhibition on cell proliferation was detected using cytotoxicity test (MTT assay). The cell survival curve was drawn using clone formation to obtain sensitive enhancement ratio (SER). Changes of the cell cycle and cell apoptosis were analyzed using flow cytometry (FCM).

RESULTSResults of MTT showed the inhibition of resveratrol on FADU cells increased along with its concentrations (P < 0.05). Results of clone formation indicated the surviving fraction at 2 Gy (SF2) was 0.717 ± 0.062 in the irradiation group, and 0.426 ± 0.035 in the resveratrol plus irradiation group (with SER ranged 1.684 ± 0.178) with statistical difference (P = 0.007). Results of FCM showed that after radiation of 4 Gy radiation, cells at G2/M phase arrest increased, but cells at G1 decreased. After radiation of resveratrol for 24 h, cells at G1 decreased, but cells at G2/M phase and S phase arrest increased. When 4 Gy radiation combined resveratrol was used, cells at G2/M phase arrest significantly increased, but cells at G1 significantly decreased. The apoptosis rate was 1.94% ± 1.65% in the control group, 4.56% ± 0.92% in the irradiation group, 2.03% ± 1.46% in the resveratrol group, and 23.11% ± 7.22% in the resveratrol plus irradiation group. There was statistical difference between the resveratrol plus irradiation group and the rest 3 groups (P < 0.05).

CONCLUSIONResveratrol could enhance the radiosensitivity of hypopharyngeal carcinoma FADU cells in vitro possibly by inducing cell apoptosis and causing changes in the cell cycle distribution.

Apoptosis ; Carcinoma, Squamous Cell ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; Cell Survival ; Head and Neck Neoplasms ; Humans ; Hypopharyngeal Neoplasms ; drug therapy ; Radiation Tolerance ; Radiation-Sensitizing Agents ; therapeutic use ; Stilbenes ; therapeutic use

BACKGROUND/AIMS: Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and it has a poor prognosis and few therapeutic options. Radiotherapy is one of the most effective forms of cancer treatment, and P53 protein is one of the key molecules determining how a cell responds to radiotherapy. The aim of this study was to determine the therapeutic efficacy of iodine-131 in three human HCC cell lines. METHODS: Western blotting was used to measure P53 expression. The effects of radiotherapy with iodine-131 were assessed by using the clonogenic assay to evaluate cell survival. Flow cytometry was carried out to examine the effects of iodine-131 on cell death, oxidative stress, reduced intracellular glutathione expression, the mitochondrial membrane potential, and the cell cycle. RESULTS: The P53 protein was not expressed in Hep3B2.1-7 cells, was expressed at normal levels in HepG2 cells, and was overexpressed in HuH7 cells. P53 expression in the HuH7 and HepG2 cell lines increased after internal and external irradiation with iodine-131. Irradiation induced a decrease in cell survival and led to a decrease in cell viability in all of the cell lines studied, accompanied by cell death via late apoptosis/necrosis and necrosis. Irradiation with 131-iodine induced mostly cell-cycle arrest in the G0/G1 phase. CONCLUSIONS: These results suggest that P53 plays a key role in the radiotherapy response of HCC.
Apoptosis/*radiation effects ; Blotting, Western ; Carcinoma, Hepatocellular/metabolism/pathology/radiotherapy ; Cell Line, Tumor ; Cell Survival/drug effects ; G1 Phase Cell Cycle Checkpoints/radiation effects ; *Gamma Rays ; Glutathione/metabolism ; Hep G2 Cells ; Humans ; Iodine Radioisotopes/chemistry/pharmacology/therapeutic use ; Liver Neoplasms/metabolism/pathology/radiotherapy ; Phosphorylation ; Reactive Oxygen Species/metabolism ; Tumor Suppressor Protein p53/*metabolism

BACKGROUNDCaffeine suppresses ataxia telangiectasia and Rad3 related and ataxia telangiectasia mutated (ATM) activities; ATM is the major kinase for DNA damage detection. This study aimed to investigate the effects of caffeine on DNA damage responses in cells from the bladder cancer cell line RT4 those were exposed to ionizing radiation (IR).

METHODSImmunofluorescent staining was performed to investigate changes in the proteins involved in DNA damage responses with or without caffeine. A mouse xenograft model was used to study the effects of caffeine on the DNA damage responses. Western blotting was used to investigate the effects of caffeine pretreatment on the ATM-Chk2-p53-Puma axis, while real-time polymerase chain reaction (RT-PCR) assessed changes in messenger RNA levels of p53 and downstream targets responding to IR. Finally, terminal deoxynucleotidyl transferase-dUTP nick end labeling assay. Western blotting and colony formation assay were used to measure the effects of caffeine on radiation-related apoptosis. All of the data were analyzed with a two-tailed Student's t-test.

RESULTSImmunofluorescent staining showed that caffeine pretreatment profoundly suppressed the formation of γH2AXand p53-binding protein 1 foci in RT4 cells in response to irradiation. Cellular and animal experiments suggested that this suppression was mediated by suppression of the ATM-Chk2-p53-Puma DNA damage-signaling axis. RT-PCR indicated caffeine also attenuated transactivation of p53 and p53-inducible genes. The colony formation assay revealed that caffeine displayed radioprotective effects on RT4 cells in response to low-dose radiation compared to the radiosensitization effects on T24 cells.

CONCLUSIONCaffeine may inhibit IR-related apoptosis of bladder cancer RT4 cells by suppressing activation of the ATM-Chk2-p53-Puma axis.

Animals ; Apoptosis ; drug effects ; radiation effects ; Caffeine ; pharmacology ; Cell Cycle Proteins ; metabolism ; Cell Line, Tumor ; Checkpoint Kinase 2 ; metabolism ; Humans ; Immunohistochemistry ; Male ; Mice ; Mice, Nude ; Radiation, Ionizing ; Real-Time Polymerase Chain Reaction ; Signal Transduction ; drug effects ; Tumor Suppressor Protein p53 ; metabolism ; Urinary Bladder Neoplasms ; radiotherapy

OBJECTIVE: To study the radiation-sensitizing function and preliminary mechanism of paclitaxel in radiation-resistant nasopharyngeal carcinoma cells. METHOD: X-ray dose fractionated irradiation technology to build radiation-resistant subline of nasopharyngeal carcinoma; CNE-2S1 was treated with paclitaxel alone or combined with radiation therapy, while control group treated with radiation therapy; cell colony formation assay was used to observe sensitizing effect of paclitaxel on radiotherapy; flow cytometry analysis was used to analyze cell cycle distribution and apoptosis ratio of different treatment groups; immunoblotting was used to analyze SHP-1 expression levels of different treatment groups. RESULT: Nasopharyngeal carcinoma cells resistant to radiation was successfully established; cell colony formation assay showed that paclitaxel has obvious sensitizing effect on radiotherapy; FACS results showed that: CNE-2S1 treated by paclitaxel were arrested in G2M phase; paclitaxel and radiotherapy treatments significantly improved the CNE-2S1 apoptosis ratio; Western blot results showed that paclitaxel and combined radiotherapy can reduce the CNE-2S1 cells SHP-1 expression levels. CONCLUSION Paclitaxel enhanced radiation therapy for nasopharyngeal carcinoma cells resistant to radiation, and SHP-1 may be involved in this progress.
Apoptosis ; drug effects ; Carcinoma ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Humans ; Nasopharyngeal Carcinoma ; Nasopharyngeal Neoplasms ; pathology ; Paclitaxel ; pharmacology ; Protein Tyrosine Phosphatase, Non-Receptor Type 6 ; metabolism ; Radiation Tolerance ; drug effects

OBJECTIVETo study radiation-enhancing effects on human gastric cancer MKN28 cell line and underlying mechanisms of β-elemene.

METHODSInhibition of MKN28 cell proliferation at different concentrations of β-elemene was assessed using the methyl thiazolyl blue colorimetric method (MTT method), with calculation of IC50 value and choice of 20% of the IC50 as the experimental drug concentration. Irradiation group and β-elemene+irradiation group were established, and the cell survival fraction (SF) was calculated from flat panel colony forming analysis, and fitted by the 'multitarget click mathematical model'. Draw the survival curve and get the radiobiological parameters D0, Dq, SF2, N and SER. Flow cytometry (FCM) was used to detect changes in the cell cycle and cell apoptosis rates was detected by Annexin-V/PI assay.

RESULTSβ-elemene exerted inhibitory effects on proliferation of gastric cancer MKN28 cells, with an IC50 of 45.6 mg/L and we chose 8 mg/L as the experimental concentration. The cell survival fraction of MKN28 cells with irradiation decreased significantly after treated with β-elemene; D0, Dq decreased, SER = 1.3. After combined treatment of β-elemene+irradiation, the results of FCM showed that cells could be arrested in the G2/M phase and the cell apoptosis increased significantly.

CONCLUSIONSβ-elemene can enhance the radiosensitivity of gastric cancer MKN28 cell line. Mechanistically, β-elemene mainly influences the cell cycle distribution of MKN28 cells by inducing G2/M phase arrest, inhibits the repair of sublethal damage and induces cell apoptosis to enhance the killing effects of radioactive rays.

Apoptosis ; drug effects ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cell Survival ; drug effects ; Humans ; Radiation Tolerance ; drug effects ; Sesquiterpenes ; pharmacology ; Stomach Neoplasms ; pathology

OBJECTIVETo investigate the effect of curcumin (Cur) on radiosensitivity of nasopharyngeal carcinoma cell CNE-2 and its mechanism.

METHODThe effect of curcumin on radiosensitivity was determined by the clone formation assay. The cell survival curve was fitted by Graph prism 6. 0. The changes in cell cycle were analyzed by flow cytometry (FCM). The differential expression of long non-coding RNA was detected by gene chip technology. Part of differentially expressed genes was verified by Real-time PCR.

RESULTAfter 10 micro mol L-1 Cur had worked for 24 h, its sensitization enhancement ratio was 1. 03, indicating that low concentration of curcumin could increase the radiosensitivity of nasopharyngeal carcinoma cells; FCM displayed a significant increase of G2 phase cells and significant decrease of S phase cells in the Cur combined radiation group. In the Cur group, the GUCY2GP, H2BFXP, LINC00623 IncRNA were significantly up-regulated and ZRANB2-AS2 LOC100506835, FLJ36000 IncRNA were significantly down-regulated.

CONCLUSIONCur has radiosensitizing effect on human nasopharyngeal carcinoma CNE-2 cells. Its mechanism may be related to the changes in the cell cycle distribution and the expression of long non-coding IncRNA.

Cell Cycle ; drug effects ; radiation effects ; Cell Line, Tumor ; Cell Survival ; drug effects ; radiation effects ; Curcumin ; pharmacology ; Gene Expression Regulation, Neoplastic ; drug effects ; radiation effects ; Humans ; RNA, Long Noncoding ; genetics ; Radiation Tolerance ; drug effects

OBJECTIVETo study the effect of dihydroartemisinin (DHA) combined irradiation on the apoptosis of human lung cancer GLC-82 cells and to study its mechanism.

METHODSThe growth inhibition rate of GLC-82 cells acted by different concentrations DHA was detected using MTT assay at 24, 48, and 72 h, respectively. Clone forming test was used. With multi-target single-hit model, the radiosensitization effect was assessed by calculating sensitizing enhancement ratio (SER).The effect of DHA combined irradiation on the apoptosis of GLC-82 cell cycle distribution and apoptosis were measured by flow cytometry. The protein expression of p53, p21, Bcl-2, and Bax were detected by Western blot.

RESULTSDifferent concentrations DHA (4, 8, 16, 32, 64, and 128 μg/mL) had cytotoxicity on GLC-82 cells. The IC50 for 24, 48, and 72 h was 38.25,20.58, and 10.36 μg/mL, respectively, in obvious dose- and time-dependent manner. The growth inhibition rate was more significantly increased than that of the blank control group (P < 0.01, P<0.05). DHA had sensitization enhancement effect on GLC-82 cells, with SER of 1.4. DHA combined irradiation could obviously change the structure of GLC-82 cells cell cycle and induce apoptosis (with the apoptosis rate of 21.5%), which was significantly different from that of the blank control group (P < 0.05). Western blot showed the expression of p53 and p21 protein could be increased by DHA combined irradiation, and the expression of Bcl-2 protein down-regulated (P <0.01, P <0. 05).

CONCLUSIONSDHA had stronger cytotoxicity and radiosensitization on GLC-82 cells. Its mechanisms might lie in making the arrest of GLC-82 cells' growth at G0/G1 phase, decreasing the ratio of cells at S phase, restoring the function of p53, decreasing the expression of Bcl-2 protein, and inducing apoptosis in GLC-82 cells.

Apoptosis ; drug effects ; Artemisinins ; pharmacology ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Down-Regulation ; drug effects ; Flow Cytometry ; Humans ; Lung Neoplasms ; metabolism ; Neoplasm Proteins ; metabolism ; Radiation-Sensitizing Agents ; pharmacology ; Tumor Cells, Cultured ; bcl-2-Associated X Protein ; metabolism

OBJECTIVE: To determine an approach enriching cancer stem cells from laryngeal cancer cell line. To investigate whether laryngeal cancer stem cells in chemoradiotherapy have the characteristic of resistance. METHOD: CD133+ cells and CD133- cells was detected and isolated from Hep-2 cell line by fluorescence activated cell sorting technology. The cytotoxicities of cisplatin and radiation were investigated by cell counting kit-8(CCK-8) assay. The apoptosis and cell cycle was analyzed with flow cytometry. RESULT: CD133+ cells accounted for a fraction of (2.43 +/- 0.77)% in Hep-2 cell line. CD133+ cells have a more obvious characteristics of cancer stem cells. Different cisplatin and radiation concentrations of for two cell have inhibition, in a certain concentration range and the dosage dependence. Cisplatin and radiation had synergistic inhibitory effects with CD133- cells on the growth of two cell. Moreover, cell cycle arrest at G0/G1 phase and more apoptosis was induced by synergistic combination. Different concentrations of cetuximab for Hep-2 cells have inhibition, in a certain concentration range and time and the dosage dependence. The half maxial inhibitory concentration (IC50) of cetuximab to Hep-2 cells on 24 h was 1 036.84 microg/L. Cisplatin and radiation had synergistic inhibitory effects with cetuximab on the growth of Hep-2 cell line. Moreover, cell cycle arrest at G0/G1 phase and more apoptosis was induced by synergistic combination. CONCLUSION Compared with CD133- cells, CD133+ cells subpopulation exhibited extraordinary cancer stem.
AC133 Antigen ; Antibodies, Monoclonal, Humanized ; pharmacology ; Antigens, CD ; analysis ; Antineoplastic Agents ; pharmacology ; Apoptosis ; Cell Cycle ; Cell Line, Tumor ; Cetuximab ; Chemoradiotherapy ; Cisplatin ; pharmacology ; Drug Resistance, Neoplasm ; Flow Cytometry ; Glycoproteins ; analysis ; Humans ; Laryngeal Neoplasms ; therapy ; Neoplastic Stem Cells ; drug effects ; radiation effects ; Peptides ; analysis ; Radiation Tolerance

OBJECTIVETo explore the radiosensitizing effect of erlotinib on human lung adenocarcinoma cell line A549 cells and the related mechanisms.

METHODSThe inhibitory effect of erlotinib on A549 cells was assessed by MTT assay, and its IC50 concentration was calculated. The radiosensitization was evaluated by the method of clone forming assay. Flow cytometry was used to analyze the effect of erlotinib on cell cycle and apoptosis.

RESULTSThe growth of A549 cells was inhibited after the cells were exposed to erlotinib for 48 hours. Moreover, the inhibitory rates increased with the increase of erlotinib concentrations, and IC50 was 19.26 µmol/L. In contrast to the irradiation alone group, the survival rates of the cells in erlotinib plus irradiation groups decreased, and erlotinib enhanced the radiosensitivity of the A549 cells. This effect was further increased as cells were exposed to erlotinib for a longer time. In the irradiation alone group and the two groups exposed to erlotinib for 24 hours and 48 hours before irradiation, D0 values were 3.01 Gy, 2.58 Gy and 2.45 Gy respectively, and Dq values were 2.16 Gy, 1.94 Gy and 1.61 Gy, respectively. In the last two groups, SERD0 values were 1.17 and 1.23, respectively. The flow cytometry analysis showed that erlotinib induced G2/M phase arrest and increased the apoptosis rate in A549 cells. With the increase of exposure time, the effects were more significant.

CONCLUSIONSErlotinib inhibits the A549 cell growth and enhances the radiosensitivity of A549 cells in vitro. The radiosensitizing mechanisms might be related to inhibiting repair of sublethal injury and inducing G2/M phase arrest and apoptosis.

Adenocarcinoma ; pathology ; Apoptosis ; drug effects ; radiation effects ; Cell Cycle ; drug effects ; radiation effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; radiation effects ; Dose-Response Relationship, Drug ; Erlotinib Hydrochloride ; Humans ; Lung Neoplasms ; pathology ; Particle Accelerators ; Quinazolines ; administration & dosage ; pharmacology ; Radiation Tolerance ; drug effects ; Radiation-Sensitizing Agents ; administration & dosage ; pharmacology