ABSTRACT
Objective:To evaluate the effect of compound UC2288 on the radiosensitivity of CNE-2R cell line and nude mouse transplanted tumor.Methods:The UC2288 concentration was referenced to previous experimental results (IC 50=12.20 μmol/L). The effect of UC2288 combined with 2, 4, 6, 8 Gy X-ray irradiation on the radiosensitivity of CNE-2R cell line was detected by clone formation experiment. The effect of UC2288 combined with 2, 4, 6, 8 Gy X-ray irradiation on the proliferation of CNE-2R cell line was determined by CCK8 assay. The nude mouse model of transplanted tumor was constructed with CNE-2R cell line. The radiosensitivity of transplanted tumor of UC2288 combined with 2 Gy/fraction X-ray irradiation for three consecutive days was evaluated. Results:The experimental concentration of UC2288 was 8 μmol/L. The clonality of CNE-2R cell line was reduced under UC2288 combined with X-ray 2, 4, 6, and 8 Gy irradiation, andthe radiosensitizationratio was 1.60. The proliferation of CNE-2R cell line was significantly decreased under UC2288 combined with X-ray 2, 4, 6, and 8 Gy irradiation. UC2288 inhibited the growth of transplanted tumor in nude mice, and the inhibitory effect was strengthened with the extension of observation time, and the most obvious effect was observed at 16 d. ( P<0.01). Theradiosensitizationratio was 4.33. The proliferation of CNE-2R cell line was decreased under UC2288 combined with X-ray irradiation. Conclusion:UC2288 can increase the radiosensitivity of nasopharyngeal carcinoma radioresistant cell line CNE-2R.
ABSTRACT
@#[Abstract] Objective: To explore the role of adhesion molecule with Ig like domain 2 (AMIGO2) in the proliferation of nasopharyn‐ geal carcinoma (NPC) cells and its mechanisms. Methods: A total of 10 NPC tissue samples and 10 normal nasopharyngeal epithelial tissue samples collected at Fujian Cancer Hospital during September 2017 and November 2017 were used for this study; in addition, NPC cell lines (CNE-1, CNE-2, SUNE-1, 6-10B, C666-1) and human immobilized nasopharyngeal epithelial cell line NP69 were also collected. The relative expression of AMIGO2 mRNAin above mentioned tissues and cell lines was detected by qPCR. Lentivirus vectors were constructed to interfere AMIGO2 mRNA expression, and qPCR was used to verify its interference efficiency. CCK-8 method, Clonal formation and Flow cytometry were performed to evaluate the effect of AMIGO2 interference on proliferation, clone formation and apoptosis of NPC cells. The influence of AMIGO2 interference on PI3K/AKT/mTOR signaling pathway and proliferation related molecular markers in NPC cells was assessed by Western blotting. Results: The results of qPCR showed that AMIGO2 was highly expressed in NPC tissues, CNE-2, and SUNE-1 cells (all P<0.01). The interference efficiency of AMIGO2 in CNE-2 and SUNE-1 cells could reach over 50%. The interfering of AMIGO2 expression significantly inhibited the proliferation and clone formation of CNE-2 and SUNE-1 cells (all P<0.01), promoted cell apoptosis (all P<0.01), reduced the phosphorylated protein expression levels of PI3K, AKT and mTOR in SUNE-1 cells (all P<0.01), as well as down-regulated the protein expressions of survivin and PCNA (all P<0.01). Conclusion: AMIGO2 may promote the proliferation as well as inhibit apoptosis of NPC cells by activating the PI3K/AKT/mTOR signaling pathway, suggesting that AMIGO2 may be a potential target for NPC therapy.
ABSTRACT
@# Objective: To explore the molecular mechanism of miR-17-5p regulating the proliferation and invasion of nasopharyngeal carcinoma cells by regulating the expression of breast cancer metastasis suppressor 1 like (BRMS1-like or BRMS1L) gene. Methods:A total of 40 cases of nasopharyngeal carcinoma tissues and corresponding paracancerous tissues resected from nasopharyngeal carcinoma patients, who were admitted to the General Hospital of Pingdingshan Shenma Medical Group during January 2014 to December 2017, were included in this study; in addition, nasopharyngeal carcinoma cell lines CNE 2, HONE 1, C666-1 and nasopharyngeal immortalized epithelial cell line NP69 were also collected for this study. The expression of miR-17-5p in nasopharyngeal carcinoma tissues and cell lines was detected by qPCR. The targeted relationship between BRMS1L and miR-17-5p was predicted by the StarBase and verified by the Dual luciferase reporter gene assay. Effects of transfection of miR-17-5p mimics and inhibitors on the expression of BRMS1Lin CNE2 cells were detected by WB assay. CCK-8, Transwell and Flow cytometry were used to detect the effects of miR-17-5p/BRMS1L axis on the proliferation, migration, invasion and apoptosis of CNE 2 cells. Results: miR-17-5p was highly expressed in nasopharyngeal carcinoma tissues and cell lines (P<0.05 or P<0.01). Knockdown of miR-17-5p significantly inhibited proliferation, invasion and migration of CNE2 cells but promoted apoptosis (P<0.05 or P<0.01); miR-17-5p targeted BRMS1Land down-regulated its expression. Over-expression of BRMS1Lsignificantly inhibited the proliferation, invasion and migration of CNE2 cells but promoted apoptosis (all P<0.01); while simultaneous over-expression of miR-17-5p and BRMS1L reversed the above effects (all P<0.01). Conclusion: miR-17-5p promoted proliferation, invasion, migration and inhibited apoptosis of CNE 2 cells by down-regulating the expression of BRMS1L.
ABSTRACT
BACKGROUND: The purpose of the present study was to investigate the role of the methylation status of the DACT1 gene on the invasion and metastasis of nasopharyngeal carcinoma cells. METHODS: The levels of methylation and expression of the DACT1 gene in nasopharyngeal carcinoma tissues and CNE2 cells were determined by methylation-specific PCR and RT-PCR, respectively. CNE2 cells were treated with 5-aza-2-deoxycytidine, and the variation in the methylation status of the DACT1 gene was detected, as well as the influence of methylation on invasiveness of nasopharyngeal carcinoma cells. RESULTS: The DACT1 gene was hyper-methylated in 44 of 62 cases of nasopharyngeal carcinoma. The DACT1 gene was hyper-methylated in 32 of 38 cases of nasopharyngeal carcinoma with lymph node metastasis, and the DACT1 gene was hyper-methylated in 7 of 24 cases of nasopharyngeal carcinoma without lymph node metastasis. The DACT1 mRNA level was weakly expressed or not expressed in all nasopharyngeal carcinoma tissues with hyper-methylated DACT1 genes; however, the DACT1 mRNA level was highly expressed in nasopharyngeal carcinoma tissues with low expression of the methylated DACT1 gene. The DACT1 gene was hyper-methylated and not expressed in CNE2 cells that did not have 5-aza-2-deoxycytidine treatment. After 5-aza-2-deoxycytidine treatment, the DACT1 gene was demethylated and the expression of DACT1 was restored. Moreover, the invasion ability was inhibited in CNE2 cells treated with 5-aza-2-deoxycytidine. CONCLUSION: The expression of DACT1 was related to the methylation status. High expression of DACT1 may inhibit the invasion and metastasis of nasopharyngeal carcinoma cells.
Subject(s)
Humans , Male , Female , Nuclear Proteins/genetics , Nasopharyngeal Neoplasms/pathology , DNA Methylation/genetics , Adaptor Proteins, Signal Transducing/genetics , Nasopharyngeal Carcinoma/secondary , Nuclear Proteins/metabolism , Nasopharyngeal Neoplasms/genetics , Promoter Regions, Genetic , DNA Methylation/physiology , Adaptor Proteins, Signal Transducing/metabolism , Nasopharyngeal Carcinoma/genetics , Neoplasm Invasiveness , Neoplasm Proteins/metabolismABSTRACT
@# Objective: To investigate the effect of licorice on radio-sensitization of nasopharyngeal carcinoma CNE-2 cells and its mechanism. Methods: The radio-resistant nasopharyngeal carcinoma cell line (CNE-2-RR) was constructed and cultured in vitro. MTT assay was used to detect the effect of different concentrations of licorice on the proliferation activity of nasopharyngeal carcinoma cells. The changes of autophagosome in CNE-2-RR cells after licorice treatment were observed by transmission electron microscopy (TEM). Western blotting was used to detect the effect of licorice on the level of autophagy protein in CNE-2-RR cells. Single cell gel electrophoresis (comet assay) was used to detect the DNAdamage and repair of different groups of CNE-2-RR cells. Flow cytometry was used to detect the apoptosis rate of CNE-2-RR cell line. Results: Low-radiation resistant CNE-2-RR cell line was successfully constructed; MTT assay showed that 20 mmol/L licorice exhibited highest inhibition on CNE-2-RR cells (58.86 ± 5.02)%. Transmission electron microscopy showed increased autophagicbody and abnormal mitochondria and nuclei morphology in CNE-2-RR cells after treatment. Western blotting showed that autophagic protein LC3-II level was increased and LC3-I level was decreased in CNE-2-RR cells (P < 0.05). The results of single cell gel electrophoresis showed that the length of comet tail distance of CNE-2-RR cells after licorice treatment was higher than that of the control group (P<0.05), indicating weakened repair ability of DNA damage. Conclusion: Licorice enhances the radio-sensitivity of CNE-2-RR cells by influencing autophagy and DNArepair ability.
ABSTRACT
Objective To investigate the effect of resveratrol on autophagic flux of nasopharyngeal carcinoma CNE-2 cells, and to explore the underlying mechanism. Methods Nasopharyngeal carcinoma CNE-2 cells were divided into control group and resveratrol group. Cells in control group were normally cultured at 37℃and received no further treatment. Resveratrol group was added 40 μmol/L resveratrol 2 h before cells were culture at 37 ℃. Western blot analysis was performed to detect protein expressions of LC3B, p62, Beclin-1, phospho-mTOR (p-mTOR) and phospho-S6 (p-S6). The autophagic flux was detected under the confocal laser scanning microscopy through different color spots, after cells were transfected with adenovirus encoding GFP-mRFP-LC3. Results (1) The protein expression of LC3B was significantly increased and the protein expression of p62 was significantly decreased in resveratrol group compared with those of control group (P<0.05). There was no significant difference in Beclin-1 expression between two groups. (2) Compared to control group, expressions of p-mTOR and p-S6 were significantly decreased in resveratrol group (P<0.05). (3) Compared to control group, the red mRFP puncta were significantly increased, and the yellow GFP puncta were significantly decreased in resveratrol group (P<0.05). Conclusion Resveratrol promotes the autophagic flux of nasopharyngeal carcinoma CNE-2 cells, and the effects are possibly dependent on the activation of mTOR pathway-related proteins.
ABSTRACT
Objective To detect the effects of miR-143 on proliferation, migration and invasion of human nasopharyngeal cancer CNE-2Z cells.Methods The expression of miR-143 in NP69 cells, CNE-1 cells and CNE-2Z cells were detected by using real-time PCR.The overexpression of miR-143 in CNE-2Z cells was constructed through infecting lentivrius, and the level of miR-143 was determined by using real-time PCR.Cell viability was detected by using a CCK-8 kit according to the manufacturer's instruction at indicated time points.The effectiveness of overex-pression of miR-143 on migration and invasion of CNE-2Z cells were detected by using Transwell cell migration and invasion assay.Results The expression level of miR-143 in CNE-2Z was significantly lower than those in NP69 and CNE-1 (P<0.01).The miR-143 over-expressed CNE-2Z cell was successfully established.The cell viability in CNE-2Z/ miR-143 group was significantly decreased compared with CNE-2Z and CNE-2Z/miR-NC (P<0.01).Overexpression of miR-143 inhibited cell migration and invasion of CNE-2Z cells significantly(P<0.01).Conclusion miR-143 might inhibit cell proliferation, migration and invasion in human nasopharyngeal cancer CNE-2Z cells, indicating its important role in diagnosis, treatment and prognosis of nasopharyngeal cancer.
ABSTRACT
Objective To investigate the effect of curcumin on proliferation and apoptosis of human nasopharyngeal carcinoma (NPC) cell line CNE-2.Methods The CNE-2 cells were treated by by different concentrations (0,10,20,40,60 μmol/L) of curcumin.The proliferation activity of CNE-2 cells was detected by MTT assay,the cell cycle and apoptosis rate of CNE-2 were detected by using the flow cytometry (FCM),and the apoptosis was observed by Hoechest33258 fluorescence staining.Results Curcumin could significantly inhibit the proliferation of CNE-2 cells,moreover which was increased with curcumin concentration increase,the inhibitory rate of CNE-2 cells showed an increasing trend (P＜0.05),the half inhibitory concentrations (IC50) of curcumin acting on CNE-2 cells at 24,48,72 h were (23.54 ± 0.36),(18.31 ± 0.42) and (8.56 ± 0.37) μmol/L respectively.Curcumin could significantly inhibit the proliferation effect of CNE-2 cells,showing the apparent concentration and time dependence.The FCM detection results showed that in treating CNE-2 cells by 0,10,20,40,60 μmol/L of curcumin,the apoptosis rate was increased with the curcumin concentration increase;the fluorescence staining results showed that CNE-2 cells without curcumin treatment were round or oval,the cell nucleis were uniform in size,chromatin distribution showed the homogeneous light blue fluorescence;after 24 h of 10 μmol/L curcumin treatment,the CNE-2 cell body was shrunk and cell nuclear chromatin was condensed,showing granular bright blue fluorescence;after 24 h of 20 μmol/L curcumin treatment,the cell body was shrunk,nuclear was condensed,chromatin was uneven,apoptotic bodies appeared,and even the nuclear fragmentation appeared;after 24 h of 40 μmol/L and 60 μmol/L curcumin treatment,the number of apoptotic cells was increased,a large number of nuclear fragmentation appeared.Conclusion Curcumin has a significant inhibitory effect on the proliferation of NPC cell line CNE-2,moreover promotes the apoptosis of CNE-2 cells.
ABSTRACT
Objective To study the growth difference and possible mechanism between nasopharyngeal carcinoma (NPC) cell line CNE-2 and its subclone S-18.Methods CNE-2 and S-18 cells were cultured in vitro.6 x 105 cells/mouse were xenografted subcutaneously in the back of nude mice.The volumes of rumors were measured on the 3 rd,7 th,10 th,14 th day after grafting.Mice were sacrificed on the 14 th day and tumors were isolated and weighed.RNA from tumor tissues were extracted and transcriptional levels of HSP27 and NF-K B were detected.Results (1) S-18,instead of CNE-2,grew to form tumor mass 7 days after xenografting subcutaneously;both cell lines formed tumor mass 10 days after xenografting,however,the volumes of S-18 tumors [(223.13 ± 21.32) mm3,10 th day;(420.25 ± 24.52) mm3,14 th day] were significant bigger than CNE-2tumors [(113.70±11.70) mm3,10thday;(279.86±25.78) mm3,14thday];The weights of S-18 umors were significantly higher than CNE-2 tumors on the 14 th day after xenografting;(2) The transcriptional levels of HSP27 and NF-KB in S-18 tumor were significantly higher than in CNE-2 tumor.Conclusion Xenografted S-18 NPC grows faster than Xenografted CNE-2 NPC.HSP27 and NF-κ B are probably involved in the regulation of growth in NPC.
ABSTRACT
Objective:To explore the proliferation inhibition and radiosensitization of Biyanqing granule on nasopharyngeal carci -noma cell line CNE-2 in vitro.Methods:CNE-2 cells were cultured in vitro.The inhibition of Biyanqing granule on the proliferation of CNE-2 cell was evaluated by MTT assay .Radiosensitization was explored by clone formation assay , and cell cycle and apopotosis were observed by flow cytometry ( FCM) .Results:Biyanqing granule could inhibit the proliferation of CNE-2 in a time-and dose-dependent manner.The IC50 in 24, 48 and 72 h was 70.79, 60.13 and 51.63 mg· ml-1(calculated according to the weight of all medicinal ma-terial), respectively.The colony formation assay showed that Biyanqing granule combined with radiation could significantly reduce the colony formation of CNE-2 cells.With the concentration increase of the main drug , the colony formation of CNE-2 cells was reduced . The number of colony formation in the negative control group , the radiation group , 10 mg· ml-1 and 20 mg· ml-1 Biyanqing combined with radiation groups (calculated according to the weight of all medicinal material ) was significant different (P<0.05).With the main drug concentration increasing , the percentage of G 2/M phase and apoptotic cells were both increased , and compared with the con-trol group, the difference was significant (P<0.05).Conclusion:Biyanqing granule can not only inhibit CNE-2 cells but also block CNE-2 cells in G2/M to improve the radiosensitization of CNE-2 cells.
ABSTRACT
objective To research whether the combination of chrysin and camptothecin can promote the apoptosis of human nasopharyngeal carcinoma cell line CNE2 and to explore the molecular mechanism of the combinative effect. Methods: CNE2 cells were pretreated with designed dose of chrysin (10|, 20, and 40 μmol/L) for 2 h, then treated with camptothecin (1 μg/mL) for 24 h. The morphologic changes were observed under inversed microscope and the cell viability was measured using MTT. The activity of caspase-3 and PARP, which was regarded as the protein marks of apoptosis, was determined by Western blotting. Then the cells were treated with chrysin for different time and the time course of apoptosis inhibitory protein, Bcl-xL was also detected using Western blotting. Results: Increases of cell death were observed in the group with combined chrysin and camptothecin, but no obvious cell death could be found in chrysin, camptothecin alone, and control groups; The data of cell viability supported this results; With the enhance of pretreatment dose of chrysin, the cell viability decreased. There were the significant differences between the combined groups and the control one (P<0.05), and between the combined groups and both the chrysin and camptothecin groups separately (P<0.05). Chromatin condensation, which was the indication of apoptosis, could be observed when the cells were stained with Hochest 33342; The proprotein of caspase-3 and PARP degraded and there were the dose-dependent and time-dependent effect. The pan-caspase inhibitor Z-VAD-fmk could inhibit the apoptosis of CNE2 cells which were treated with the combination of chrysin and camptothecin, according to the cell viability and the activation of caspase-3 and PARP; The time-dependent down-regulation in the apoptosis inhibitory protein Bcl-xL could be observed. Conclusion: The cotreatment of chrysin and camptothecin could promote the apoptosis of CNE2 and the down-regulation of apoptosis inhibitory protein Bcl-xL played an important role in the combinative effect.
ABSTRACT
Objective:To investigate the inhibitory effect and its possible molecular mechanisms of MicroRNA-34a(miR-34a) on the human nasopharyngeal carcinoma CNE-2 cell line subcutaneous xenograft tumor in nude mice.Methods: The human nasopharyngeal carcinoma CNE-2 cell line was cultured in vitro.miR-34a and Scrambled miRNA recombinant plasmids were successfully established and stably transfected into CNE-2 cells.Fifteen six-week-old male nude mice were divided randomly into three groups:miR-34a group(5 mice) ,Scrambled miRNA group(5 mice) ,Blank control group(5 mice).Different CNE-2 cells were subcuta-neously injected on the back near right lower limb.Tumor volumes were examined every 7 days.Mice were executed on the 35 days,and the eventual average tumor volumes and weights were examined.Total RNA and protein were isolated from tumors,and the expression of miR-34a,CDK6,and Bcl-2 mRNA and protein were determined by qRT-PCR and western blot,respectively.Results: The relative expressions of miR-34a was significantly up-regulated in miR-34a transfected group compared to Scrambled miRNA transfected group (P (849.62±101.32) mm3 ,respectively,and the eventual average tumor weights in miR-34a group,Scrambled miRNA group and blank control group were(0.81±0.13)g,(1.47±0.21)g and(1.58±0.37)g,respectively.Both the eventual average tumor volumes and weights in miR-34a group were lower compared to the other two groups(P<0.05).qRT-PCR results revealed that the expression of miR-34a in miR-34a transfected group was significantly higher than in the other two groups,while the mRNA and protein expression of CDK6 and Bcl-2 were lower than the other two groups ( P<0.05 ) .Conclusion: miR-34a may inhibit the growth of human nasopharyngeal carcinoma CNE-2 cell line subcutaneous xenograft tumor in nude mice by down-regulating CDK6 and Bcl-2.
ABSTRACT
Objective To observe the effects of over expression and inhibition expression of SAA protein on biological behavior of nasopharyngeal carcinoma CNE2 cells.Methods pcDNA3.1 (+)-SAA-CNE2 cell lines of high expression and pGPU6/GFP/Neo-SAA-CNE2 cell lines of interference expression of SAA protein in vitro.These two cells constructed by transfection of pcD-NA3.1(+)-SAA plasmid of SAA high expression and pGPU6/GFP/Neo-SAA plasmids of SAA inhibition expression respectively, plasmids of which were previously successfully reconstructed by the research group.Cell cycle of these two cells was analyzed by flow cytometry with PI staining.The ability of cell proliferation was inspected by plate cloning-forming test.Results Flow cytome-try showed that with the increase of expression of SAA protein,it had effect on promoting CNE2 cell division.Plate cloning-forming test showed that SAA protein can improve proliferation of the CNE2 cells.Conclusion SAA protein has the effect on promoting proliferation of human nasopharyngeal carcinoma CEN2 cell and migration in vitro.
ABSTRACT
Aim Toexploretheinhibitioneffectof triptolide on nasopharynx cancer, and the mechanism. Methods Theinhibitionofcellproliferationwasde-tected by MTT assay;the cell apoptosis was analyzed by flow cytometry with propidium iodide staining. The ex-pressions of glucose regulated protein 78 ( GRP-78 ) , Akt and pAkt in cells were examined by Western blot;the effect of triptolide on reactive oxygen species ( ROS) accumulation was detected by ROS Fluorescent Probe-DHE.Results MTTassayshowedthatthe growth of nasopharynx cancer was inhibited by triptol-ide , and the inhibition occurred in a dose and time-de-pendent manner following triptolide treatment in CNE-2Z nasopharynx cancer cells. Propidium iodide staining revealed that the apoptosis of CNE-2 Z cells was in-duced remarkably by triptolide. After CNE-2Z cells treated with 25, 50,100 nmol·L-1 of triptolide for 24 h, the apoptosis rate was 14%,26. 9% and 34. 4% re-spectively. Western blot experiment showed that the expression of GRP-78 had no significant change follow-ing triptolide treatment in CNE-2 Z nasopharynx cancer cells for 24 h, but the expression and the phosphoryla-tion level of Akt were strikingly decreased. The experi-ment of ROS uncovered that CNE-2 Z nasopharynx cancer cells increased generation of ROS after treat-ment with triptolide for 4 hours, and acted cells in a dosedependentmanner.Conclusions Triptolidecan inhibit the growth of CNE-2 Z nasopharynx cancer cells in a dose and time-dependent maner. The mechanism may be related with the point that triptolide can induce oxidative stress, incease ROS, inhibit the expression and the phosphorylation level of Akt,then promote the apoptosis of CNE-2Z cells.
ABSTRACT
Objective Naringenin has a vast prospect of application because of its important biological activities .This study aims to explore the influence of different concentrations of naringenin on the growth of nasopharyngeal carcinoma CNE 2 cells and its ac-tion mechanisms. Methods Using the MTT method, we measured the effects of naringenin on the growth of CNE 2 cells after treated at the concentrations of 0, 0.005, 0.01, 0.02, 0.04, 0.06, 0.08, 0.1, 0.2, 0.4, and 0.8 mg/mL for 24, 48, and 72 hours.At 48 hours, we observed changes in the cycle of the cells treated with naringenin at 0, 0.02 and 0.04 mg/mL by flow cytometry, in the ap-optosis of the cells by Hochest 33258 staining and flow cytometry , in the level of reactive oxygen species ( ROS) in the cells by DCFH-DA staining, and in the mRNA expressions of C-fos, Bax and Bcl-2 by qPCR. Results Compared with the control group , naringe-nin at 0.02 and 0.04 mg/mL induced a low-level rise of ROS in the CNE2 cells (MFI:5186 ±183.50 and 5508 ±155.37, P<0.05), up-regulated the expression of C-fos (P<0.05 or P<0.01), and promoted the proliferation of the cells .However, naringenin at relatively high concentrations of 0.2 and 0.4 mg/mL significantly elevated the level of ROS (MFI:10758 ±179.82 and 11241 ±1 114.45, P<0.01), up-regulated the expression of Bax (P<0.01), down-regulated that of Bcl-2 (P <0.01), induced the apoptosis (P<0.01 ) and suppressed the proliferation of CNE 2 cells. Conclusion Within a concentration range of 0.005-0.8 mg/mL, naringenin may have two-way effects on the growth of CNE2 cells, a carcinogenic effect at a relatively low dose and a good anticancer effect at a relatively high dose .
ABSTRACT
OBJECTIVE: To explore the influence and the mechanisms of different dose naringenin on nasopharyngeal carcinoma CNE2 cells growth. METHODS: Different concentrations of naringenin (0, 5, 10, 20, 40, 60, 80, 100, 200, 400, 800 μg · mL-1) were used to detect their effects on CNE2 cells growth for 24 to 72 h by MTT Method. High concentrations of naringenin (0, 200, 400 μg · mL-1) for 48 h were used to detect apoptosis by Hochest 33 258 staining. High concentrations of naringenin (0, 200, 400 μg · mL-1) for 48 h were used to detect the cell cycle by flow cytometry. Different concentrations of naringenin (0, 40, 200 μg · mL-1) for 24 and 48 h were used to detect the level of reactive oxygen species (ROS) by flow cytometry, and mRNA expression levels of P53 and C-fos were determined by qPCR. RESULTS: After different concentrations of naringenin administration, CNE2 cells were changed growth rate. Relatively low dose can induce the low level rise of ROS, the low level rise of ROS can induce the up-regulation of the expression levels of c-fos (P < 0.01) and the down-regulation of the expression levels of P53(P < 0.01), so relatively low dose can promote CNE2 cells growth rate. However, relatively high dose can induce the high level rise of ROS, the high level rise of ROS can induce the up-regulation of the expression levels of P53(P < 0.01) and the down-regulation of the expression levels of C-fos (P < 0.01), so relatively high dose can induce apoptosis, also possess the effect of S and G2/M phase blockage(P < 0.01), significantly suppress CNE2 cells growth rate. CONCLUSION: Different dose naringenin has two-way effects on CNE2 cells growth, relatively low dose has carcinogenic effect, relatively high dose has good anticancer effect.
ABSTRACT
Objective To investigate the effect of lobaplatin combined with irradiation on human nasopharyngeal cancer cell line CNE2,and to illuminate its mechanism of radiosensitization.Methods MTT assay was used to detect the outcome of lobaplatin and irradiation on CNE2 cell proliferation.Clonogenic assay was applied to testify the radiosensitization effect of lobaplatin on the cells.Flow cytometry was used to check the cell cycle distribution and cell apoptosis.Western was used to detect the expression of Bcl-2,Bax and cleaved Caspase-3.Results The proliferation of CNE2 cells was reduced by lobaplatin in a dose-dependent manner.50IC of lobaplatin on CNE2 cells and lobaplatin combined with 4 Gy irradiation was 1.610 μmol/L and 0.077 μmol/L,respectively.The radiosensitization ratio of the combination group was over 3.Within 24 h of drug treatment,the percent of cells in G2/M phase increased with the concentration of lobaplatin.When the concentration of lobaplatin increased to 6 μmol/L,the cells of combination group were arrested at S phase.The apoptosis rate of lobaplatin (5 μmol/L) group,radiotherapy(4 Gy)group and combination group was 15.6％,11.3％ and 61.8％,respectively.Western blot showed that the expressions of Bax and cleaved Caspase-3 increased but Bcl-2 decreased in the combination group.Conclusion Lobaplatin could increase radiosensitization of human nasopharyngeal cancer cell line CNE2,probably by depressing Bcl-2 but enhancing Bax expression and hence activating Bcl-2/Bax-Caspase signaling pathway.
ABSTRACT
Objective To investigate the effect of cetuximab (C225) combined with radiation on the expressions of Ku80 and ATM in CNE-2 nude mice xenograft tumor model.Methods The CNE2 nude mice xenograft tumor model was established and divided into control group,conventional radiotherapy (CRT) group,simulated intensity modulated radiation therapy (sIMRT) group,C225 group,CRT + C225 group and sIMRT + C225 group with 8 mice in each group.The dose of radiation was 20 Gy,and 1mg C225 per mice in abdominal cavity was applied for the drug treatment.The transcriptional levels of Ku80 and ATM were assayed by RT-PCR and Western blot.Results Compared with control group,the transcriptional and translational levels of Ku80 and ATM in CRT group,sIMRT group,CRT + C225 group and slMRT + C225 group were decreased (P ＜ 0.05).Compared with CRT group,the down-regulation of transcriptional and translational ATM level in sIMRT group was obvious (P ＜ 0.05),and further strengthened in the CRT + C225 group and sIMRT + C225 groups (P ＜0.05),but there was no significant difference of ATM expression between sIMRT + C225 group and CRT group (P ＞ 0.05).Conclusions C225 alone can restrain the transcriptional and translational level of ATM,and the transcriptional and translational level of Ku80 and ATM are also restrained after megadose.ATM is down-regulated by sIMRT but up-regulated by the combination of C225 and radiation.
ABSTRACT
STGC3 is a potential tumor suppressor that inhibits the growth of the nasopharyngeal carcinoma cell line CNE2; the expression of this protein is reduced in nasopharyngeal carcinoma compared with normal nasopharyngeal tissue. In this study, we investigated the tumor-suppressing activity of STGC3 in nude mice injected subcutaneously with Tet/pTRE-STGC3/CNE2 cells. STGC3 expression was induced by the intraperitoneal injection of doxycycline (Dox). The volume mean of Tet/pTRE-STGC3/CNE2+Dox xenografts was smaller than that of Tet/pTRE/CNE2+Dox xenografts. In addition, Tet/pTRE-STGC3/CNE2+Dox xenografts showed an increase in the percentage of apoptotic cells, a decrease in Bcl-2 protein expression and an increase in Bax protein expression. A proteomic approach was used to assess the protein expression profile associated with STGC3-mediated apoptosis. Western blotting confirmed the differential up-regulation of prohibitin seen in proteomic analysis. These results indicate that overexpression of STGC3 inhibits xenograft growth in nude mice by enhancing apoptotic cell death through altered expression of apoptosis-related proteins such as Bcl-2, Bax and prohibitin. These data contribute to our understanding of the function of STGC3 in human nasopharyngeal carcinoma and provide new clues for investigating other STGC3-associated tumors.
Subject(s)
Animals , Male , Genes, Tumor Suppressor , Nasopharyngeal Neoplasms , Neoplasm Proteins , Apoptosis , Electrophoresis , Mice, NudeABSTRACT
Background and purpose:Diallyl disulfide (DADS) is an oil-soluble organosulfur compound found in garlic. It can induce apoptosis in various cancer cell lines in vitro. This study investigated the effect of apoptosis of nasopharyngeal cancer cells line CNE2 induced by diallyl disulfide (DADS) and its molecular mechanisms. Methods:The growth inhibition and apoptosis of CNE2 cells and the protein levels of Bcl-2, Bax and Caspase 3 were examined by MTT assay, light microscopy, flow cytometry, DNA agarose gel electrophoresis and Western blot, respectively.Results:MTT assay showed that 50,100,200,400 ?mol/L DADS significantly inhibited proliferation of CNE2 cells at 48 hr, its inhibition ratio were 3.66%, 14.28%, 29.66%, 61.28%, respectively, in dose-dependent manner (P