Effects of curcumin analogue B50 on proliferation and apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance

AIM: To compare the effects of B50, a mono-carbonyl analogue of curcumin, on the proliferation and apoptosis between homologous nasopharyngeal carcinoma cells CNE-2R and CNE-2 with different radioresistance.METHODS: The effects of B50 on cell viability and cell growth were detected by MTT assay and colony-forming experiment, respectively. The changes of cell cycle, apoptosis and mitochondrial membrane potential (MMP) were determined by flow cytometry.RESULTS: B50 inhibited the cell viability of CNE-2R cells in a time-and dose-dependent manner with the IC₅₀ of (8.06±0.14) μmol/L (24 h), (2.49±0.02)μmol/L (48 h) and (1.42±0.02) μmol/L (72 h), which was more effective than that in CNE-2 cells . The inhibitory effect of B50 on CNE-2R cell growth was more effective than that on CNE-2 cells . After treated with B50 for 48 h, the proportion of CNE-2R cells in G₂/M stage was increased from 7.1% to 34.9%, which was better than that of CNE-2 cells (from 12.4% to 35.7%). After treated with B50 for 24 h, the early apoptotic rate in CNE-2R cells was increased from 3.7% to 19.5%, which was better than that in CNE-2 cells (from 4.4% to 14.8%), and the MMP in CNE-2R cells was decreased by (43.17±3.11)%, which was better than that in CNE-2 cells .CONCLUSION: B50 is more effective on inhibiting the cell viability and cell growth, blocking the cell cycle at G₂/M stage, inducing apoptosis and decreasing MMP in CNE-2R cells than those in CNE-2 cells, indicating that B50 may enhance the radio-sensitivity of CNE-2R cells by blocking the cell cycle and inducing apoptosis through mitochondrial pathway.     

Effects of aspirin on apoptosis of homologous nasopharyngeal carcinoma cells with different radioresistance

AIM: To investigate the effects of aspirin on the apoptosis of nasopharyngeal carcinoma cell lines CNE2R and CNE2 with different radioresistance and its potential mechanism. METHODS: The effects of aspirin on the cell viability, apoptosis, and protein levels of procaspase-3, cleaved caspase-3, procaspase-9, procaspase-12, PARP and cleaved PARP, PI3K p110α, Akt, Bcl-2, Bax and p27 in the CNE2R cells and CNE2 cells were detected by the methods of MTT assay, flow cytometry and Western blot. RESULTS: Aspirin inhibited the viability of homologous nasopharyngeal carcinoma cell lines CNE2 and CNE2R (with the IC₅₀ to CNE2 cells of 6.18, 3.92 and 3.06 mmol/L for 24 h, 48 h and 72 h, respectively; and with the IC₅₀ to CNE2R cells of 7.05, 3.90 and 2.20 mmol/L for 24 h, 48 h and 72 h, respectively). After treated with aspirin for 48 h, the apoptotic rate of CNE2R cells was higher than that of CNE2 cells (P<0.05). After treated with aspirin for 48 h, the protein levels of procaspase-3, procaspase-9, procaspase-12 and PARP were decreased, the protein levels of cleaved caspase-3, cleaved PARP and p27 were increased, and the protein levels of PI3K p110α, Akt and Bcl-2/Bax were decreased. CONCLUSION: Aspirin inhibits the viability of homologous nasopharyngeal carcinoma cell lines CNE2R and CNE2 with different radioresistance. Aspirin also induces the apoptosis of CNE2 and CNE2R cells, which is more effective in CNE2R cells. The underlying mechanisms may be involved in affecting PI3K/Akt signaling pathway, Bcl-2/Bax and p27 expression.     

Curcumin analogue B67 induces apoptosis and G2/M arrest to suppress radioresistant nasopharyngeal carcinoma cells

AIM: To study the effect of curcumin analogues B67 on radioresistant nasopharyngeal carcinoma cells (CNE-2R). METHODS: The effects of B67 on the cell viability and proliferation of CNE-2R and the parent cells CNE-2 were detected by MTT assay and colony formation assay, respectively. The changes of cell cycle, apoptosis and mitochondrial membrane potential were determined by flow cytometry. The morphological changes of the cells were observed under fluorescence microscope. Node mice were subcutaneously inoculated with the cells to determine the tumorigenic ability. RESULTS: The IC₅₀ of B67 on the viability of CNE-2R cells after treatment for 24 h, 48 h and 72 h were 3.96,2.59 and 0.89 μmol/L, respectively, and those of CNE-2 cells were 8.84, 3.55 and 1.10 μmol/L,respectively. The IC₅₀ of B67 on the proliferation of CNE-2R cells after treatment for 48 h was 0.55 μmol/L, and that of CNE-2 cells was 0.73 μmol/L. After treated with B67 for 24 h, CNE-2R and CNE-2 cells at G₂/M stage increased from 5.32% to 40.01% and from 9.07% to 15.73%,respectively. After treated with B67 for 48 h, the apoptosis of CNE-2R and CNE-2 cells increased from 5.49% to 38.06% and from 4.99% to 35.74%, respectively. The mitochondrial membrane potential in CNE-2R and CNE-2 cells was decreased by 66.76% and 72.09%, respectively. After treated with B67 for 24 h, the tumorigenic rate of CNE-2R cells was 0%, while the rates of CNE-2 cells in low- and high-concentration groups were 100% and 0%, respectively.CONCLUSION: Curcumin analogue B67 exhibits enhanced suppressive activity on radioresistant nasopharyngeal carcinoma cells by inducing G₂/M-phase arrest, promoting cell apoptosis and changing mitochondrial membrane potential.     

T63 enhances radiotherapeutic sensitivity of nasopharyngeal carcinoma cells through apoptosis and G2/M arrest

AIM: To explore the antitumor effect of a curcumin analogue T63 in human nasopharyngeal carcinoma (NPC) cell lines CNE-2 and CNE-2R. METHODS: Cell viability was monitored by the methods of MTT and colony formation assay. Cell cycle distribution was detected by flow cytometry. Apoptosis was examined using the annexin V-FITC/PI staining assay. RESULTS: A growth inhibitory effect was observed with T63 treatment in a dose-dependent manner. Either T63 or ionizing radiation (IR) significantly induced G2/M arrest and apoptosis in NPC cells. In addition, T63 treatment combined with IR induced significantly higher apoptosis and G2/M arrest in NPC cells. CONCLUSION: T63 exhibits potent inhibitory activity on NPC cells and induces the radiotherapeutic sensitivity. Therefore, T63 has a potential as a preventive or therapeutic agent for treating NPC.     

Effects of tamoxifen on volume-activated Cl- currents of nasopharyngeal carcinoma cells at different stages of the cell cycle

AIM:To investigate the inhibitory effects of Cl- channel blocker, tamoxifen, on volume-activated Cl- currents of nasopharyngeal carcinoma cells (CNE-2Z cells) in G₁ and S phases. METHODS:Highly synchronous cells in G₁ phase and S phase were obtained by the serum starvation and the double-block techniques. The whole-cell patch clamp technique was used to observe the effects of tamoxifen on volume-activated Cl- currents and to analyze the anion permeability of volume-activated Cl- channels. RESULTS:47% hypotonic stimulation activated a Cl- current in the nasopharngeal carcinoma cells at the cell cycle stage G₁ phase and S phase. Tamoxifen at concentration of 10 to 30 μmol/L completely inhibited the current. However, the time needed to completely inhibit the current was dose-dependent and was different between G₁ phase and S phase. The time needed to completely inhibit the current was shorter in G₁ cells than that in S phase cells. The anion permeability sequence of the volume-activated Cl- channel was I->Cl->gluconate in both G₁ phase and S phase cells. The permeability of G₁ phase cells to I- was higher than that in S phase cells, but to gluconate was lower than that in S phase cells. CONCLUSIONS:The density of the volume-activated Cl- current, the anion permeability of the channel and the sensitivity of the current to tamoxifen were different between the CNE-2Z cells in G₁ phase and those in S phase. The results suggest that the expression of tamoxifen-sensitive, volume-activated chloride channels is differentiated at different stages of the cell cycle.     

Effects of EBV infection on growth and apoptosis of nasopharyngeal carcinoma cell line

AIM:To explore the effect of EBV infection on growth and apoptosis of nasopharyngeal carcinoma(NPC)cell line.METHODS:NPC cell line CNE1 was directly infected by Epstein Barr virus(EBV).The expression of EBV-latent membrane protein 1(EBV-LMP1)and bcl-2 were detected by immunohistochemistry method(LSAB).The growth of NPC cells was identified by MTT method.Apoptotic carcinoma cells were detected by flowcytometry analysis and the terminal deoxynucletidyl transferase-medicated dUTP-biotin nick end labeling(TUNEL)methods.RESULTS:EBV-LMP1 was positive in CNE1 infected by EBV(E-CNE1).Compared with CEN1, the growth of E-CNE1 apparently increased(P<0.01).No apoptotic carcinoma cel s were detected and bcl-2 postive cells were 2%~3%respectively in 2 kinds of NPC cells.CONCLUSION:Growth of NPC cells is enhanced by EBV infect ion and EBV-LMP1 expression, but no influence on expression of bcl-2 and apoptosis of NPC cells.     

Effects of EBV infection on growth and apoptosis of nasopharyngeal carcinoma cell line

AIM: To explore the effect of EBV infection on growth and apoptosis of nasopharyngeal carcinoma(NPC) cell line.METHODS: NPC cell line CNE1 was directly infected by Epstein Barr virus (EBV). The expression of EBV-latent membrane protein 1 (EBV-LMP1) and bcl-2 were detected by immunohistochemistry method (LSAB). The growth of NPC cells was identified by MTT method. Apoptotic carcinoma cells were detected by flow cytometry analysis and the terminal deoxynucletidyl transferase-medicated dUTP-biotin nick end labeling (TUNEL) methods. RESULTS: EBV-LMP1 was positive in CNE1 infected by EBV(E-CNE1). Compared with CEN1, the growth of E-CNE1 apparently increased (P＜0.01). No apoptotic carcinoma cells were detected and bcl-2 postive cells were 2%~3% respectively in 2 kinds of NPC cells.CONCLUSION: Growth of NPC cells is enhanced by EBV infection and EBV-LMP1 expression, but no influence on expression of bcl-2 and apoptosis of NPC cells.     

Role of Cl- in regulatory volume decrease of nasopharyngeal carcinoma cells

AIM:To clarify the role of Cl^- in regulatory volume decrease (RVD) of nasopharyngeal carcinoma cells (CNE-2Z).METHODS:Analysis of living cell images was used to detect the volume changes following exposure to hypotonic solution. Iron replacement and block of iron channels were also applied in the present study. RESULTS:Extracelluar hypotonic treatment made the cells swell and induced RVD. The RVD was correlated positively to the swelling in the range of 160-230 mOsmol/L. Substitution of gluconate for Cl^- in perfusing solutions markedly increased RVD. Depletion of cellular Cl^- abolished, and chloride channel blockers inhibited RVD. CONCLUSION:Cl^- is the key iron to establish the RVD in CNE-2Z cells. Activation of Cl^- channels and Cl^- efflux are the major mechanisms of RVD.     

Photoactivation promotes curcumin to induce apoptosis of human gastric cancer MGC-803 cells

AIM: To investigate the effect of photoactivated curcumin on apoptosis of human gastric cancer MGC-803 cells. METHODS: The effect of photoactivated curcumin on the growth inhibitory rate of gastric cancer MGC-803 cells was detected by MTT assay. The changes of nuclear morphology were observed under optical microscope with Hoechst 33258 fluorescent staining. The apoptotic rate, mitochondrial membrane potential, intracellular reactive oxygen species and Ca²⁺ level was determined by flow cytometry. The activity of caspase-3, caspase-8 and caspase-9 was detected by colorimetry. The protein levels of cytochrome C, Bcl-2, Bax and heat-shock protein 70 (HSP70) were analyzed by Western blotting. RESULTS: The growth inhibitory rate of MGC-803 cells treated with curcumin at concentration of 5.0 μmol/L was (29.74±2.30)%.Some apoptotic cells were observed under optical microscope, and the apoptotic rate was (12.54±1.75)%. The growth inhibitory rate of MGC-803 cells treated with photoactivated curcumin was (44.93±3.61)%.Significant morphological changes in the nucleus, such as chromatin condensation and apoptotic body formation, were observed under light microscope, and the apoptotic rate was (26.58±2.67)%. The cell cycle was arrested at G₀/G₁ phase. Compared with curcumin group, significant reduction in mitochondrial membrane potential,significant increase in cytochrome C, intracellular reactive oxygen species, Ca²⁺ level and the activity of caspase-3, caspase-8 and caspase-9 were observed in photoactivated curcumin group (P<0.01). Photoactivated curcumin also significantly inhibited the protein expression of Bcl-2 and HSP70 in the cells. CONCLUSION: Photoactivated curcumin enhances the apoptosis of gastric cancer MGC-803 cells by inhibiting Bcl-2 expression and promoting the mitochondrial pathway.     

Protective effect of curcumin analogue L6H4 on diaphragm of type 2 diabetic rats

AIM: To investigate the protective effect of curcumin analogue L6H4 on diaphragm of type 2 diabetic rats.METHODS: SPF male Sprague-Dawley rats (n=40) were randomly divided into 5 groups: normal control (NC) group, high fat (HF) group, high fat+L6H4 treatment (FT) group, diabetes mellitus (DM) group and DM+L6H4 treatment (DT) group. The rats in the later 4 groups were fed with high-fat diet. After 4 weeks of high-fat diet fee-ding, the rats in DM and DT groups were intraperitoneally injected with streptozotocin to induce type 2 diabetes melliutus. The rats in FT and DT groups were given L6H4 by gavage for 8 weeks. Blood glucose and blood lipid levels were detected biochemically. Fasting serum insulin (FINS) level was measured by radioimmunoassay and insulin resistance index (HOMA-IR) was calculated. Serum adiponectin (APN) level was measured by ELISA. The morphological changes of the diaphragm were observed under light and transmission electron microscopes. Lipid deposition and the activity of succinate dehydrogenase (SDH) and NADH-tetrazolium reductase (NADH-TR) were observed by enzyme histochemical staining. The content of malondialdehyde (MDA) and the activity of superoxide dismutase (SOD) in the diaphragm were measured by thiobarbituric acid method and hydroxylamine method, respectively. The protein expression of adiponectin receptor 1 (AdipoR1) in the diaphragm was determined by immunohistochemistry and Western blot. RESULTS: The levels of blood lipids, blood glucose, FINS and HOMA-IR in HF and DM groups were higher than those in NC group, but decreased after L6H4 treatment. The serum APN level in HF and DM groups was lower than that in NC group, but increased after treatment with L6H4. The muscle fibers of the diaphragm were shrunk, fat particles accumulated in the muscle fibers, and the mitochondria were slightly swollen in HF and DM groups. The diaphragmatic fibrosis was obvious in DM group. These lesions were relieved after L6H4 treatment. Compared with NC group, the level of MDA and the activity of SDH and NADH-TR in the diaphragm were increased in HF and DM groups, but decreased after treatment with L6H4. The activity of SOD and the expression of AdipoR1 in the diaphragm were lower than those in NC group, but increased after L6H4 treatment.CONCLUSION: The curcumin analogue L6H4 exerts a protective effect on diaphragm in type 2 diabetic rats. The strengthened protein expression of AdipoR1, the increased serum level of APN, and anti-lipid peroxidation may be involved in the process.     

Effect of microtubule-destabilizing protein stathmin gene silencing on proliferation and apoptosis of nasopharyngeal carcinoma cell line 5-8F

AIM: To investigate the effects of stathmin gene silencing on nasopharyngeal carcinoma cell line 5-8F. METHODS: Double-strand siRNA targeting to stathmin gene was obtained by chemical synthesis and annealing, and was sub-cloned into the vector pGenesil-1.1. The plasmid was introduced into 5-8F cells by liposome-mediated transfection. The gene expression of stathmin, and the proliferation, morphology and apoptosis of the cells were analyzed by Western blotting, MTT assay and flow cytometry. RESULTS: The cell suppression rate in stathmin gene silencing group was (53.01?1.12)%, significantly higher than that in transfection reagent group and in negative control group. The cell apoptotic rate in stathmin gene silencing group was (8.75?0.67)%, also significantly higher than that in transfection reagent group and in negative control group (P<0.05). CONCLUSION: Silencing of stathmin gene in nasopharyngeal carcinoma cells inhibits the cell proliferation and induces cell apoptosis.     

Inhibition of cell proliferation and arrest of cell cycle progression by blocking Cl- channels in nasopharyngeal carcinoma cells

AIM: The roles of Cl^-channels in regulatory volume decrease (RVD), cell proliferation and cell cycle progression in nasopharyngeal carcinoma cells (CNE-2Z) were investigated. METHODS: Image analysis of living cells was used to detect the volume changes following exposure to hypotonic solutions. Cell viability was determined by the trypan blue assay. MTT method was applied to detected cell proliferation. The effect of the blocker on the cell cycle distribution was monitored by the flow cytometry. RESULTS: 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) inhibited RVD and cell proliferation in a dose-dependent manner. NPPB at the concentration of 100 μmol/L arrested cells in G₁ phase (G₁ population increased from 54% to 71% at 48 h after treatments), but did not significantly alter cell viability. CONCLUSION: Block of chloride channels suppressed cell proliferation by arresting cells in G₁ phase. The results suggest that activation of Cl^-channels and RVD is necessary for facilitating cells to proceed to the S phase from G₁ phase and maintaining cell proliferation.     

Inhibitory effect of JIP on AP-1 activity induced by LMP1 in nasopharyngeal carcinoma cells and its mechanism

AIM: To investigate the mechanism of the AP-1 signal transduction pathway inhibited by JIP in nasopharyngeal carcinoma cells. METHODS: AP-1 activity was triggered by Dox-induced LMP1 expression in Tet-on-LMP1-HNE₂ cells (L7). The retention of phospho-JNK in the cytoplasm caused by JIP was examined with immunofluroscence assay. RESULTS: 24 h after transfection of L7 cells with the JIP expression plasmid, the translocation of activated JNK was inhibited, which resulted in the retention of phospho-JNK in the cytoplasm and down-regulation of the AP-1 activity. CONCLUSION: JIP down-regulates the activity of AP-1 through the inhibition of the translocation of JNK.     

Effect of transketolase-like protein 1 on proliferation of human nasopharyngeal carcinoma cells

AIM: To investigate the effect of transketolase-like protein 1 (TKTL1) on proliferation of human nasopharyngeal carcinoma cells in vitro. METHODS: The siRNA against TKTL1 mRNA was constructed and transfected into human nasopharyngeal carcinoma cells (CNE cell line). The activity of transketolase was detected before and after RNA interference.Real-time PCR was used to determine the mRNA expression of transketolase (TKT) gene family in the CNE cells.Flow cytometry and MTT test were used to detect the effect of anti-TKTL1 siRNA on cell proliferation and cell cycle in the CNE cells. RESULTS: The total transketolase activity was significantly decreased in the CNE cells transfected with siRNA TKTL1 construct compared with the cells transfected with control vector or untransfected CNE cells. No significant difference in the expression level of TKT and TKTL2 gene between the CNE cells transfected with siRNA TKTL1 construct and the cells transfected with control vector or untransfected CNE cells was observed (P>0.05). However, the expression level of TKTL1 gene was significantly downregulated in the CNE cells transfected with siRNA TKTL1 construct compared with the cells transfected with control vector.Cancer cells were arrested in G0/G1 phase, and cancer cell proliferation was significantly inhibited in the CNE cells transfected with siRNA TKTL1 construct. CONCLUSION: TKTL1 plays an important role in the total transketolase activity and cell proliferation of human nasopharyngeal carcinoma. TKTL1 may be considered as a potential target for novel anti-cancer therapy.     

miRNA-7-mediated Bax/Bcl-2 expression promotes apoptosis of human nasopharyngeal carcinoma CNE-1 cells

AIM: To investigate the relationship of microRNA-7 (miRNA-7) over-expression and Bax/Bcl-2 expression in human nasopharyngeal carcinoma CNE-1 cells.METHODS: The CNE-1 cells were transfected with miRNA-7 mimics using Lipofectamine 2000. The expression of miRNA-7 was detected by real-time PCR. CCK-8 assay and Hoechst 33258 staining were used to detect the cell activity and apoptosis. The expression of Bax/Bcl-2 at mRNA and protein levels was determined by real-time PCR and Western blot. RESULTS: The expression of miRNA-7 was increased significantly in the CNE-1 cells compared with negative control group and mock group (P<0.01). The activity of CNE-1 cells were extremely decreased after tansfected with miRNA-7 mimics (P<0.01). The typical apoptotic nuclear morphological changes were observed in the CNE-1 cells under the fluorescence microscope with Hoechst 33258 staining. The expression of Bax at mRNA and protein levels was significantly increased compared with the other 2 groups (P<0.01), while the Bcl-2 expression at mRNA and protein levels was significantly down-regulated (P<0.01).CONCLUSION: Over-expression of miRNA-7 significantly inhibits the growth and promotes the apoptosis of nasopharyngeal carcinoma CNE-1 cells by increasing the expression of Bax and down-regulating Bcl-2.     

Effects of PKC-α asODN and PKA-ⅠasODN on growth and proliferation of the CNE-2Z cells of nasopharyngeal carcinoma

AIM:To investigate the effects of antisense oligonucleotides (asODN) of PKC-α and PKA-Ⅰon growth and proliferation of the CNE-2Z cells.METHODS:The expression of PKC-α and PKA-Ⅰ was observed with immunohistochemistry method. The asODNs of (1)PKC-α, (2)PKA-Ⅰ, (3)PKC-α and PKA-Ⅰ, were transfected into CNE-2Z cells by lipofectin (LP), and a random sequence as a control was used. The cell growth index (GI) and the clone formation rate of CNE-2Z were detected by MTT colorimetric assay and soft agar assy, respectively.RESULTS:The expression of PKC-α or PKA-Ⅰin CNE-2Z in experimental group were both significantly lower than that of control group(P<0.05). The GI and clone formation rates of CNE-2Z cells transfected by PKC-α and PKA-ⅠasODN with concentrations ranging from 0.05 μM to 1.00 μM were lower significantly than that of control groups(P<0.05), and there was a dose-dependent relationship among them. The inhibitory effects of PKC-α and PKA-ⅠasODNs both on the cell growth index (GI) and clone formation rates were more significant than that of control group(P<0.01),and the GI were significantly lower than that of the other experimental groups(P<0.05).CONCLUSION:PKC-α asODN and PKA-ⅠasODN inhibited CNE-2Z growth and proliferation in vitro, and a synergetic inhibitory effect of PKC-α asODN and PKA-ⅠasODN was also observed.     

Effect of arctigenin on apoptosis of human nasopharyngeal carcinoma cell line CNE-1

AIM: To investigate the effect of arctigenin on the apoptosis of human nasopharyngeal carcinoma cell line CNE-1 and its potential mechanism. METHODS: The inhibition of cell viability was analyzed by CCK-8 assay. The activity of caspase-3 and caspase-9 was analyzed by caspase-3 and caspase-9 activity kit. Apoptotic cell percentage was evaluated by Annexin V-PI staining. The expression of PI3K/AKT/XIAP signal pathway-related molecules at mRNA and protein levels was analyzed by real-time PCR and Western blot. RESULTS: Arctigenin inhibited the cell activity in a dose- and time-dependent manner after treatment with arctigenin at concentrations of 10, 20, 40 and 80 μmol/L for 24 h, 48 h and 72 h (P<0.01). Arctigenin also increased the activity of caspase-3 and caspase-9 and the apoptotic rate (P<0.05), and down-regulated the mRNA and protein expression of PI3K/AKT/XIAP signal pathway-related molecules (P<0.05).CONCLUSION: Arctigenin induces the apoptosis of CNE-1 cells through PI3K/AKT/XIAP signal pathway.     

Inhibition of miR-9 expression suppresses proliferation,invasion and migration of nasopharyngeal carcinoma cells

AIM: To investigate the effects of down-regulated miR-9 expression on the proliferation, invasion and migration of nasopharyngeal carcinoma (NPC) cells. METHODS: Human NPC CNE1 and CNE2 cells were transfected with the inhibitor of miR-9 by Lipofectamine to down-regulate the expression of miR-9, and the cells transfected with an inhibitor control were also set up. The cell proliferation and cell cycle were evaluated by CCK-8 assay and flow cytometry. The cell invasion and migration abilities were detected by Transwell invasion and wound-healing assays. Immunoblotting was applied to analyze the levels of the proteins. RESULTS: Compared with control group, inhibition of miR-9 expression in the NPC cells by transfection of the miR-9 inhibitor significantly decreased the proliferation ability (P<0.05). The percentages of the cells in G0/G1 phase ［CNE2: (57.96±1.39)% vs (47.93±1.76)%, P<0.05; CNE1: (51.24±0.88)% vs (48.29±0.39)%, P<0.05］ were significantly increased. The migration distances ［CNE2: (186.50±7.94)μm vs (247.56±15.56)μm, P<0.05; CNE1: (139.06±16.73)μm vs (230.66±14.27)μm, P<0.01］ and the invasion ability of the CNE2 cells (43.00±3.17 vs 65.80±5.20, P<0.01) were also significantly inhibited. Moreover, the tumor cells transfected with the inhibitors produced lower β-catenin. CONCLUSION: Inhibition of miR-9 expression suppresses the proliferation, invasion and migration of nasopharyngeal carcinoma cells.     

Effect of tetrandrine on radiosensitivity of nasopharyngeal carcinoma cells

AIM: To investigate the mechanism of the radiosensitizing effect of maximum non-cytotoxic doses of tetrandrine (Tet) on nasopharyngeal carcinoma cell lines CNE1 and CNE2.METHODS: The cells were treated with ma-ximum non-cytotoxic doses of Tet (for CNE1 cells at 1.5 μmol/L and for CNE2 cells at 1.8 μmol/L), irradiation at 4 Gy, or combination of irradiation and maximum non-cytotoxic doses of Tet. The cell cycle distribution was analyzed by flow cytometry. The protein levels of γ-H2AX, cleaved caspase-3, p-CDC25C, CDK1, p-CDK1, cyclin B1, ERK and p-ERK were determined by Western blot.RESULTS: The expression of γ-H2AX was increased in CNE1 cells and CNE2 cells after combined treatment with irradiation and maximum non-cytotoxic doses of Tet. The percentages of CNE1 cells and CNE2 cells at G₂/M phase in irradiation group were (18.09±0.42)% and (18.48±1.32)%, respectively, which were decreased to (15.88±1.04)% and (13.80±0.82)% in combined treatment group, respectively (P<0.05). Combined treatment enhanced the increase in the protein level of cleaved caspase-3 caused by irradiation. The protein levels of p-CDC25C and p-CDK1 were increased in a dose-dependent manner by Tet treatment (P<0.05), while the expression of CDK1 showed no difference among different doses of Tet treatments. The protein levels of p-CDC25C, p-CDK1 and CDK1 showed no difference after the treatment with maximum non-cytotoxic doses of Tet. The combined treatment with irradiation and the maximum non-cytotoxic doses of Tet decreased the protein levels of p-CDC25C and p-CDK1 (P<0.05), increased the expression of cyclin B1, and had no influence on the expression of CDK1 (P<0.05). The combined treatment resulted in an increase in the protein level of p-ERK1 (P<0.05).CONCLUSION: The maximum non-cytotoxic doses of Tet enhance the DNA damage and apoptosis in CNE1 cells and CNE2 cells caused by irradiation, and the mechanism might be associated with ending of G₂/M arrest via activation of ERK/CDC25C/CDK1/cyclin B1 pathways.     

Efficacy of cytokine-induced killer cell transfusion combined with chemotherapy using gemcitabine and cisplatin for nasopharyngeal carcinoma with hepatic or pulmonary metastasis

AIM: To evaluate the recent and long-term efficacy of cytokine-induced killer(CIK) cell and transfusion plus chemotherapy with gemcitabine and cisplatin(GP) for patients of nasopharyngeal carcinoma(NPC) with hepatic or pulmonary metastasis.METHODS: From Aug. 2007 to Jul. 2008, 30 patients of nasopharyngeal carcinoma with hepatic or pulmonary metastasis were enrolled in the study and randomly divided into 3 groups. Ten patients(CIK+GP group) received CIK cell transfusion plus GP chemotherapy for 4 cycles. Another 10 patients(GP group)received the regimen of GP chemotherapy alone. The other 10 patients as well as 10 healthy volunteers did not receive any anti-cancer treatment. The early response, the changes of serum EBV-DNA by quantitative PCR detection and distribution of lympholeukocyte subset in the patients of the 3 groups were determined before and after treatment. The long-term effects were observed in a follow-up process of 2 years. RESULTS: The recent complete and partial response(CR+PR) rates were 90%, 70% and 10% in CIK+GP group, GP group and patient control group, respectively, and obviously statistical differences among these 3 groups were observed. The proportion of CD3⁺ lymphocytes in NPC patients with hepatic or pulmonary metastasis were obviously lower than that in healthy volunteer. It also descended after CIK+GP/GP therapy, but partly recovered in CIK+GP group. Furthermore, no statistical difference of CD4⁺/CD8⁺ ratio among NPC patients with hepatic or pulmonary metastasis and volunteers was found. The ratio of CD4⁺/CD8⁺ in CIK+GP group increased, but decreased in GP group after treatment. In the 2-year follow-up, the overall survival rate was 60.0%, 40.0% and 20.0% in CIK+GP, GP and control group,respectively. The Kaplan-Meier survival curves showed statistical difference. CONCLUSION: The recent and long-term effects of CIK cell transfusion combined with GP chemotherapy for patients of NPC with hepatic or pulmonary metastasis have been confirmed. The chemotherapy and biotherapy of CIK+GP improve the prognosis with synergistic effects, possibly owing to the changed proportion of CD3⁺and ratio of CD4⁺/CD8⁺.