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Objective To investigate the effect of LncRNA OIP5-AS1 on radiosensitivity of non-small cell lung cancer (NSCLC) cells and its mechanism.Methods The radiation-resistant cell A549R was established by using A549 cells irradiated by X-ray 6Gy in 5 fractions.The expression levels of OIP5-AS1 and miR-34c-5p in A549 and A549R cells were detected by qRT-PCR.OIP5-AS1 inhibitor or miR-34c-5p mimetic was transfected into A549R cells,or OIP5-AS1 overexpression plasmid was transfected into A549 cells.Cell apoptosis was detected by flow cytometry.Cell radiosensitivity was analyzed by colony formation assay.The expression levels of p-Chk2 and p-ATM proteins were measured by Western blot.Dual luciferase assay was adopted to verify the relationship between OIP5-AS1 and miR-34c-5p.Results Compared with A549 cells,the expression of OIP5-AS1 was significantly up-regulated in A549R cells (1.97±0.11 vs.1.01±0.05,P＜0.05),whereas the expression of miR-34c-5p was remarkably down-regulated (0.43±0.02 vs.1.02±0.06,P＜0.05).The expression levels of p-Chk2 and p-ATM proteins in A549R cells in the silencing OIP5-AS1 +6Gy group were significantly lower (0.43±0.03 vs.1.39±0.15,0.51 ±0.0 5 vs.1.21 ± 0.11,both P＜0.05),whereas the apoptotic rate was significantly higher than those in the silencing control + 6Gy group [(13.29± 1.25)％ vs.(28.47± 2.31)％,P＜0.05)].The expression levels of p-Chk2 and p-ATM proteins in A549 cells in overexpressing OIP5-AS1+6 Gy group were significantly higher than those in overexpression control+6 Gy group (1.23±0.13 vs.0.75±0.06,1.08±0.11 vs.0.59± 0.04,both P＜0.05).Inhibiting miR-34c-5p expression reversed the effect of silencing OIP5-AS1 on survival fraction of A549R cells (SER =1.42).OIP5-AS1 negatively regulated the expression of miR-34c-5p.Conclusion Silencing OIP5-AS1 enhances the radiosensitivity of radiation-resistant A549 cells by up-regulating the expression of miR-34c-5p,providing a potential target for radiotherapy of NSCLC cells.
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Objective@#To investigate the effect of LncRNA OIP5-AS1 on radiosensitivity of non-small cell lung cancer (NSCLC) cells and its mechanism.@*Methods@#The radiation-resistant cell A549R was established by using A549 cells irradiated by X-ray 6Gy in 5 fractions. The expression levels of OIP5-AS1 and miR-34c-5p in A549 and A549R cells were detected by qRT-PCR. OIP5-AS1 inhibitor or miR-34c-5p mimetic was transfected into A549R cells, or OIP5-AS1 overexpression plasmid was transfected into A549 cells. Cell apoptosis was detected by flow cytometry. Cell radiosensitivity was analyzed by colony formation assay. The expression levels of p-Chk2 and p-ATM proteins were measured by Western blot. Dual luciferase assay was adopted to verify the relationship between OIP5-AS1 and miR-34c-5p.@*Results@#Compared with A549 cells, the expression of OIP5-AS1 was significantly up-regulated in A549R cells (1.97±0.11 vs.1.01±0.05, P<0.05), whereas the expression of miR-34c-5p was remarkably down-regulated (0.43±0.02 vs.1.02±0.06, P<0.05). The expression levels of p-Chk2 and p-ATM proteins in A549R cells in the silencing OIP5-AS1+ 6Gy group were significantly lower (0.43±0.03 vs.1.39±0.15, 0.51±0.0 5 vs.1.21± 0.11, both P<0.05), whereas the apoptotic rate was significantly higher than those in the silencing control+ 6Gy group [(13.29±1.25)% vs. (28.47±2.31)%, P<0.05)]. The expression levels of p-Chk2 and p-ATM proteins in A549 cells in overexpressing OIP5-AS1+ 6Gy group were significantly higher than those in overexpression control+ 6Gy group (1.23±0.13 vs.0.75±0.06, 1.08±0.11 vs.0.59±0.04, both P<0.05). Inhibiting miR-34c-5p expression reversed the effect of silencing OIP5-AS1 on survival fraction of A549R cells (SER=1.42). OIP5-AS1 negatively regulated the expression of miR-34c-5p.@*Conclusion@#Silencing OIP5-AS1 enhances the radiosensitivity of radiation-resistant A549 cells by up-regulating the expression of miR-34c-5p, providing a potential target for radiotherapy of NSCLC cells.
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Objective: The objective of this study was to evaluate the cytotoxic potential of A3AR agonist (ABMECA) against human lung cancer cell line A549 by using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) assay. Methods: Adenocarcinoma cell line A549 was used to assess MTT based cells viability. In vitro cytotoxic activity was evaluated for 3 different concentration of doxorubicin and A3AR by MTT cytotoxicity assay. Cytotoxicity assay carried out for 3 consecutive days that involves culturing cells into Dulbecco’s MEM medium modified with 10% FBS for 24 h then treatment with different dose of standard and test drug with incubation period of 24 h followed by treatment with MTT for estimation of cytotoxicity and finally, optical density (OD) was measured at 570-630 nm. Results: Different concentration of doxorubicin (1, 5, 10 µM) and ABMECA (10-6M, 10-5M and 10-4M) shown dose-dependent cytotoxicity. There was a significant decrease (p<0.05) in cell viability in both doxorubicin and ABMECA concentration in a dose-dependent manner. This study may guide further for in vivo evaluation of test drug in the lung cancer model. Conclusion: A3 Adenosine Receptor agonist could be potential moiety for the treatment of lung cancer and it would require in vivo study for further research.
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Objective The biological function of E3 ubiquitin-protein ligase RNF19A in lung cancer is yet clear. This study was to investigate the effects of RNF19A on the proliferation, invasion and migration of lung cancer cells and its underlying mechanisms. Methods A549 cells were transfected with control siRNA or RNF19A siRNA for 48 hours. Then, the viability and proliferation of the cells were measured by CCK8 and BrdU incorporation assay, respectively. Immunoprecipitation and Western immunoblotting were used to detect the effect of RNF19A-TAK1 interaction on the ubiquitination of TAK1 and the effects of TAK1 and NF-κB inhibitors on the proliferation, invasion and migration of the Flag-RNF19A-mediated A549 cells. Results After 48 hours of transfection, the viability and proliferation of the A549 cells were significantly decreased in the RNF19A siRNA group as compared with the control group (P < 0.001), and so were the numbers of migrating (441.0 ± 18.63 vs 960.6 ± 37.82, P < 0.05) and invading cells (488.2 ± 26.06 vs 1120 ± 58.96, P < 0.05) and the level of TAK1 ubiquitination in the A549 cells (0.425 ± 0.01 vs 0.656 ± 0.012, P < 0.05). Over-expressed Flag-RNF19A markedly enhanced the proliferation of the cells in comparison with that of the control group (P < 0.05), and increased the numbers of migrating (1032 ± 38.86 vs 721.7 ± 26.60, P < 0.05) and invading cells (657.7 ± 13.74 vs 355.7 ± 15.51, P < 0.05), but showed no statistically significantly difference from the control in the proliferation of the cells with the addition of TAK1 and NF-κB inhibitors (P > 0.05). Conclusion RNF19A can increasing the proliferation, migration and invasion of A549 lung cancer cells, probably by enhancing TAK1 ubiquitination and NF-κB activation.
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Objective@#To investigate the effect and potential mechanism of HOXC8 on the radiosensitivity of non-small cell lung cancer cell line A549, aiming to provide novel ideas for clinical combined treatment.@*Methods@#The A549 cells with stable knockdown of HOXC8 were constructed by using lentivirus and validated by qPCR and Western blot. The radiosensitivity of A549 stable cell line was assessed by plate clone formation assay. The expression levels of TGF-β1 and the proteins in the downstream signal pathway after knockdown of HOXC8 were detected by Western blot.@*Results@#The A549 cells with stable knockdown of HOXC8 were successfully constructed. The viability and clonogenic capacity of A549 cells were significantly reduced after silencing HOXC8. Silencing HOXC8 also increased the sensitivity of A549 cells to radiotherapy and significantly inhibited the expression of TGF-β1 and p-Smad2/3 proteins in the downstream signaling pathway.@*Conclusion@#Silencing HOXC8 can increase the sensitivity of A549 cells to radiotherapy probably by inhibiting TGF-β1 signaling transduction. HOXC8 might play an important role in A549 cells.
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Objective@#To investigate the inhibitory effect of 17AAG-Cypate micelles on the non-small cell lung cancer A549 cells in nude mice and to explore its possible mechanism.@*Methods@#A549 lung adenocarcinoma tumor-bearing nude mice were established. The nude mice were treated with saline ( saline group), X-ray (X-ray group), 17AAG micelles+ X-ray (17AAG-M/X group) and 17AAG-Cypate micelles+ laser/X-ray (17AAG-Cypate-M/L+ X group), respectively. The growth of xenograft tumors in different groups was measured on a regular basis to delineate the growth curve. The expression of proliferating cell nuclear antigen (PCNA) was measured by immunohistochemistry. The microvascular density was detected. The apoptosis of xenograft tissues was observed by TUNEL staining. The expression levels of p-ERK1/2 and p-AKT were quantitatively measured by Western blot.@*Results@#Compared with the saline group, varying degrees of inhibition of tumor growth were observed in the X-ray, 17AAG-M/X-ray and 17AAG-Cypate-M/L+ X groups, particularly in the 17AAG-Cypate-M/L+ X group (all P<0.05). In all groups, the expression levels of PCNA were significantly down-regulated (all P<0.05), the microvascular density was remarkably reduced (all P<0.05) and the expression levels of p-ERK1/2 and p-AKT were considerably down-regulated (all P<0.05).@*Conclusions@#17AAG-Cypate micelles can inhibit the growth of human non-small cell lung cancer in nude mice, probably by reducing the activity of p-ERK1/2 and p-AKT, thereby weakening the activation of the MAPK-ERK and PI3K-AKT signaling pathways.
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Objective To explore the effects of lipopolysaccharide (LPS) on the expression of inflammatory genes in A549 cells line under different concentrations and different action time, this study laid the foundation for further establishment of acute respiratory distress syndrome (ARDS) cell model in the optimal concentration-time way. Methods A549 cells line was incubated routinely in 5%CO2 incubator at 37 ℃ with high glucose DMEM medium which included 10% fetal calf serum. Cells in logarithmic phase was cultured for passage, the cells was count to adjust cell density to (5-7)×105 and tile evenly in six-hole plate. Cells were cultivated for 2 days and once the cells confluence to 50%-60%, serum-free medium DMEM was changed for 12 hours cultivation. 10 mg LPS was added to 10 mL DMEM for oscillated blending to prepare 1 g/L stock solution. 0.5, 1.0 and 2.5 mL LPS stock solution was taken respectively and diluted LPS stock solution for 50 mL constant volume to prepare 0, 10, 20 and 50 mg/L LPS working solution. Then 0, 10, 20 and 50 mg/L LPS solution was added to react for 0, 1, 3 and 5 hours respectively. Reverse transcription-polymerase chain reaction (RT-PCR) was performed to examine mRNA expression of A549 cells line interleukins (IL-6, IL-1β) and tumor necrosis factor-α(TNF-α). LPS action of 10 mg/L for 0 hour was used as the time control group, LPS action of 0 mg/L for 1 hour was used as the concentration control group, and the gene expression was calculated with 2-ΔΔCt method. Results ① As to the time factor, with the same action of LPS concentration, the relative expression levels of inflammatory genes (IL-6, IL-1β and TNF-α) in A549 cells line after being treated with 10 mg/L LPS for 1 hour were significantly higher than those for 0 hour respectively [IL-6 mRNA (2-ΔΔCt): 5.71±0.42 vs. 1.00±0.00, IL-1β mRNA (2-ΔΔCt): 5.63±0.30 vs. 1.00±0.00, TNF-α mRNA (2-ΔΔCt): 5.38±0.61 vs. 1.00±0.00, all P < 0.01], and there were no significant changes in the expression levels of inflammatory genes in A549 cells line of other time groups. ② As to the concentration factor, with the same action time, the relative expression levels of inflammatory genes (IL-6, IL-1βand TNF-α) in A549 cells line after being treated with 10 mg/L LPS for 1 hour were significantly higher than with 0 mg/L for 1 hour respectively [IL-6 mRNA (2-ΔΔCt): 5.70±0.64 vs. 1.00±0.00, IL-1β mRNA (2-ΔΔCt): 6.25±0.25 vs. 1.00±0.00, TNF-α mRNA (2-ΔΔCt): 5.57±0.25 vs. 1.00±0.00, all P < 0.01], there were no significant changes in the expression levels of inflammatory genes in A549 cells line of other concentration groups. Conclusion The LPS concentration of 10 mg/L and the action time of 1 hour are the most suitable concentration-time conditions for establishing ARDS cell models of A549 cells line.
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@#[Abstract] Objective: To investigate he effect of tetracycline- (Tet-on) mediated livin RNA interference on growth of lung carcinoma xenegrafts, and find a better regulatory way to interfere the development on lung cancer. Methods: livin shRNA lentiviral vectors were constructed; and the lung cancerA549 cells were subcutaneously injected into right upper back of nude mice to establish xenegraft model. The livin shRNAlentiviral vectors were injected into xenografts to interfere the expression of livin, then tetracycline was injected intraperitoneally for the induction. The suppressive effect of Tet-on mediated livin RNA interference efficiency was investigated and lung cancer xenograft development was observed. Results: After the induction with Tet-on, livin gene expression was significantly inhibited by livin shRNAcompared with the control group and Tet-on-NC group; the xenograft volume in Tet-on- livin shRNAgroup was significantly smaller than that in control group and Tet-on-NC group ([5.31±0.86]g vs [8.22±0.63]g and [7.17±0.54] g, P<0.05). Moreover, little body toxicity was observed and no nude mice died in this study. Conclusion: The Tet-on mediated livin shRNA could suppress the growth of lung cancer development with good targeting and controllable characteristics, which might provide a potent tool for treating lung cancer with livin protein as target.
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Objective: To synthesize bio-inspired gold nanoparticles (AuNPs) using the leaf extract of Justicia adhatoda and evaluate the anti-cancer activity on human lung cancer cell line (A549). Methods: Synthesis of AuNPs was done using an aqueous leaf extract of Justicia adhatoda as a green route. The bio-synthesized AuNPs were confirmed and characterized by using various spectral studies such as UV-Vis spectrum, Scanning Electron Microscope with EDAX, Transmission Electron Microscope, Fourier Transmission Infrared Spectroscope analysis and Surface Enhanced Raman Spectroscopy. The cell viability was determined by MTT reduction assay. In addition, cytomorphology and the nuclear morphological study of A549 cell line was observed under fluorescence microscope. Results: UV-Vis spectrum showed surface plasmon resonance peak at 547 nm, scanning electron microscope and transmission electron microscope studies showed the monodispersed spherical shape and its average size in the range of 40.1 nm was noticed. Fourier Transmission Infrared Spectroscope analysis confirmed that the C=O group of amino acids of proteins had strong ability to bind with the surface of nanoparticle. Interestingly, our results also demonstrated inhibited proliferation of A549 cell line by MTT (IC50 value: 80 μg/mL). Cell morphology was observed and cell death was caused by apoptosis as revealed by propidium iodide staining. Conclusions: The current study proves the anticancer potential of bio-synthesized AuNPs. Thus, synthesized AuNPs can be used for the treatment of human lung cancer cell (A549) and it can be exploited for drug delivery in future.
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Objective: To investigate the reversal effects in A549/DDP by standard substances (SS) from Traditional Chinese Medicine. Methods: Cell proliferation assays were performed to investigate the Resistant Index of A549/DDP and its tolerance to selected SS. The working concentrations of SS, IC5 calculated by nonlinear regressions, were applied as reversal doses to investigate the effects. Results: The resistant index of A549/DDP was 31.79. Tetramethylpyrazine and Matrine were well tolerated. Berberine hydrochloride, Dauricine, Oridomin exhibited dose-depend inhibitory effects on A549/DDP cell line. The working concentrations of them could effectively reverse the resistance of A549/DDP cell line to DDP. (P＜0.01) . Conclusion: The selected standard substances from Traditional Chinese Medicine were capable to reverse the resistance of A549/DDP cell line to DDP.
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Objective: To synthesize bio-inspired gold nanoparticles (AuNPs) using the leaf extract of Justicia adhatoda and evaluate the anti-cancer activity on human lung cancer cell line (A549). Methods: Synthesis of AuNPs was done using an aqueous leaf extract of Justicia adhatoda as a green route. The bio-synthesized AuNPs were confirmed and characterized by using various spectral studies such as UV-Vis spectrum, Scanning Electron Microscope with EDAX, Transmission Electron Microscope, Fourier Transmission Infrared Spectroscope analysis and Surface Enhanced Raman Spectroscopy. The cell viability was determined by MTT reduction assay. In addition, cytomorphology and the nuclear morphological study of A549 cell line was observed under fluorescence microscope. Results: UV-Vis spectrum showed surface plasmon resonance peak at 547 nm, scanning electron microscope and transmission electron microscope studies showed the monodispersed spherical shape and its average size in the range of 40.1 nm was noticed. Fourier Transmission Infrared Spectroscope analysis confirmed that the C=O group of amino acids of proteins had strong ability to bind with the surface of nanoparticle. Interestingly, our results also demonstrated inhibited proliferation of A549 cell line by MTT (IC
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Objective To compare the growth of three different cancer cell lines on chick chorioallantoic membrane (CAM), to select the best transplanted cancer cell line for establishing a transplanted tumor model and to observe the biological characteristics.Methods The human lung cancer cell line A549, human tongue cancer cell line TCA8113 and human liver cancer cell line QGY7703 were respectively inoculated into CAM at the 7th day of age.The chick embryo survival rate, tumor survival rate, tumor formation rate and induced angiogenesis were detected and the growth characteristics of the transplanted tumor model were observed.Results Compared with the groups inoculated with A549 cells and QGY7703 cells, the tumor formation rate of TCA8113 cells was the highest (P < 0.05), to be the best cancer cell line for transplanted tumor.The optimal inoculated number of cells was 8.0×106/chick embryo, the optimal growth period of the tumor was 4~8 d, and the best experiment time was 7 d after inoculation.Conclusion The TCA-CAM transplanted tumor model of tongue squamous cell cancer is successfully established for further study of the biological characteristics and mechanisms of tumor growth, angiogenesis, invasion and metastasis, and provide a good experimental animal model for anti-tumor drug screening.
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Arisaema heterophyllum Blume is one of the three medicinal plants known as traditional Chinese medicine Rhizoma Arisaematis (RA). RA has been popularly used to treat patients with convulsions, inflammation, and cancer for a long time. However, the underlying mechanisms for RA effects are still unclear. The present study was designed to determine the cytotoxicity of agglutinin isolated from Arisema heterophyllum Blume (AHA) and explore the possible mechanisms in human non-small-cell lung cancer A549 cells. AHA with purity up to 95% was isolated and purified from Arisaema heterophyllum Blume using hydrophobic interaction chromatography. AHA dose-dependently inhibited the proliferation of A549 cells and induced G phase cell cycle arrest. AHA induced apoptosis by up-regulating pro-apoptotic Bax, decreasing anti-apoptotic Bcl-2, and activating caspase-9 and caspase-3. In A549 cells treated with AHA, the PI3K/Akt pathway was inhibited. Furthermore, AHA induced increase in the levels of ER stress markers such as phosphorylated eukaryotic initiation factor 2α (p-eIF2α), C/EBP-homologous protein (CHOP), inositol-requiring enzyme 1α (IRE1α), and phosphorylated c-Jun NH-terminal kinase (p-JNK). AHA also induced autophagy in A549 cells. Staining of acidic vesicular organelles (AVOs) and increase in the levels of LC3II and ATG7 were observed in AHA-treated cells. These findings suggested that AHA might be one of the active components with anti-cancer effects in Arisaema heterophyllum Blume. In conclusion, cytotoxicity of AHA on cancer cells might be related to its effects on apoptosis and autophagy through inhibition of PI3K/Akt pathway and induction of ER stress.
Subject(s)
Humans , A549 Cells , Agglutinins , Pharmacology , Apoptosis , Arisaema , Chemistry , Autophagy , Carcinoma, Non-Small-Cell Lung , Drug Therapy , Metabolism , Cell Line, Tumor , Drugs, Chinese Herbal , Pharmacology , Endoplasmic Reticulum Stress , MAP Kinase Signaling System , Phosphatidylinositol 3-Kinases , Genetics , Metabolism , Phosphorylation , Proto-Oncogene Proteins c-akt , Genetics , MetabolismABSTRACT
Objective To determine the effects of endostatin on the expression of vascular endothelial growth factor receptor?2 ( VEGFR?2 ) in non?small cell lung cancer cells ( human A549 lung adenocarcinoma cells and human Calu?1 lung carcinoma cells) , and to investigate the possible mechanisms underlying its radiosensitizing effect. Methods The CCK8 method was used to determine the inhibitory effect of endostatin on cell proliferation and calculate the drug concentration that caused a 20% reduction in cell proliferation within 24 h ( IC20 ) . RT?PCR and Western blot assays were used to assess the mRNA and protein expression of VEGFR?2, proteins within its related signaling pathways, and HIF?1α, respectively. The radiosensitivity of cells in each group was determined by colony formation assay;cell apoptosis and cell cycle distribution were determined by flow cytometry. Comparison of mean values between multiple samples was made by one?way analysis of variance, and comparison of mean values between two samples was made by t test. Results Endostatin significantly inhibited the proliferation of Calu?1 cells ( F=50?36,P<0?01) with an IC20 of 296?5 μg/ml;the mRNA and protein expression of VEGFR?2 and HIF?1α was also significantly inhibited in endostatin?treated Calu?1 cells ( F=25?43,10?44, all P<0?05) . Moreover, the phosphorylation of Akt, ERK 1/2, and p38 was significantly reduced in endostatin?treated Calu?1 cells ( F=2?89,0?24, 1?09, all P<0?05) . The radiosensitivity enhancement ratios for Calu?1 cells and A549 cells were 1?38 and 1?09, respectively. Endostatin significantly induced apoptosis ( F=44?15, P<0?01) and G2/M blockage ( F= 104?24, P< 0?01 ) in Calu?1 cells. Conclusions Endostatin induces apoptosis and enhances radiosensitivity in Calu?1 cells with high expression of VEGFR?2, but it has a limited impact on A549 cells with low expression of VEGFR?2.
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Objective To investigate the method for effective establishment of nude rat tumor xenograft model of human lung cancer cells A549 in order to provide the experimental basis for tumor-related interventional research in vivo. Methods A549 cell lines were subcutaneously transplanted in nude rats, then single-cell suspension or tumor tissue block were prepared when the tumor lesion was established. The single-cell suspension and tumor tissue block were transplanted into subcutaneous tissue behind ear in rats. The tumor formation rate, growth situation and cell cycle of primary xenograft tumor group, the secondary single-cell suspension group and the secondary tumor block group were evaluated. The results were analyzed. Results The tumor formation rate of the secondary tumor block group was significantly higher than that of the other two groups. The tumor cells quickly proliferated with less tumor variation. Tumor cell cycle analysis indicated that G2/M ratio of the secondary tumor block group was remarkably higher than that of the other two groups. Conclusion Transplantation with tumor tissue block can significantly increase the tumor formation rate of human lung cancer cells A549 in experimental rats. This technique is an effective method for the establishment of nude rat tumor xenograft model.
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OBJECTIVE: To study the effect of valdecoxib and pirarubicina combination on cell cycle and apoptosis of human lung cancer A549 cell line in vitro. METHODS: MTT assay was used to analyze the effect of valdecoxib and pirarubicina on the growth of human lung cancer A549 cell line. The median-effect principle was applied to determine the combination effect of valdecoxib and pirarubicina. Flow cytometry (FCM) was used to observe the cell cycle and apoptosis. The expressions of Bcl-2, Bax, and Caspase-3 were detected by western blotting. RESULTS: The inhibition ratio of A549 cell in valdecoxib and pirarubicina combination group was increased compared with their respective application, and CI<1. In valdecoxib and pirarubicina combination group, the apoptosis rate and the expression of Bax and Caspase-3 was increased, and the expression of Bcl-2 was decresed. CONCLUSION: Valdecoxib and pirarubicina combination has synergistic effect, which is partly due to the increase of the apoptosis rate.
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ObjectiveTo investigate the radiosensitizing effect of 3-(5'-hydroxy-2'-furyl)-1-benzyl indazole ( YC-1 ) on hypoxic human adenocarcinoma cell line A549.MethodsMTT assay was used to test the inhibitory effect of YC-1 on proliferation of A549 cells.Clonogenic assay was performed to determine the radiosensitizing effect of YC-1 on hopxic A549 cells.Single-hit multi-target model was used to plot survival curve and calculate sensitization enhancement ratio (SER).The cell cycle and apoptosis were measured by flow cytometry.ResultsThe proliferation of A549 cells was inhibited by YC-1 in a time-dose-dependent manner.In normoxic and hypoxic cells,the IC20 was 16.7 μmol/L and 39.2 μmol/L at 24 h,respectively.In the group of hypoxia plus YC-1,SERD0 and SERDq were 1.11 and 1.26,respectively.In hypoxia,YC-1combined with 2 Gy irradiation could induce cell apoptosis and prolong G2 + M phase arrest ( ( 30.17 ±1.21 )％ ∶ ( 15.44 ±0.96) ％,P =0.000; (21.56 ±0.47 )％ ∶ (6.16 ±0.16)％,P =0.000).Concinsions YC-1 could enhance the radiosensitivity of hypoxic A549 cells.
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Objective To explore the effect of β-element on the oadiosensitivity of transplanted tumor, and its relationship with the expression of survivin. Methods The transplanted mice model was established through the cell suspension inoculation. The mice with transplanted tumor size of 0. 8-1.0 cm3 were randomly divided into 8 groups as blank control, 25, 45 and 100 mg/kg group, irradiation group,25 mg/kg + irradiation group, 45 mg/kg + irradiation group, 100 mg/kg + irradiation group. The tumor size was measured every other day until tumor size was double, and the growth curve was obtained. The average tumor growth inhibition rate of β-element and tumor size were attained at 2,4,6 and 8 d after β-element injection. The expression of survivin was detected with immunohistochemistry. Results The nude mice model was successfully established and the growth curves were obtained according to the tumor size.Between 2 and 8 d after β-elemene injection, the variation tendency of the average tumor growth inhibition rate was consistent with the size in β-elemene treatment groups. The antitumor effect of β-elemene was in a dose-dependent manner. The values of radiosensitivity enhancement factor were 0. 84,1.24,2.04 for 25,45 and 100 mg/kg group, respectively ,and the optimal dose was 45 mg/kg. β-element had little effect on the expression of survivin, and the expression of survivin significantly enhanced in irradiation group and decreased in β-element + irradiation groups. Conclusions β-elemene could enhance the tumor radiosensitivity through inhibitiong the expression of survivin.
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Objective To investigate the effect of hydroxyapatite nanoparticles (nHAP) mediated human telomerase re-verse transcriptase (hTERT) RNA interference of A549 human lung cancer cells in vitro. Methods The nHAP were synthe-sized by the homogeneous precipitation method. The structure of the nanoparticles was observed under transmission electron mi-croscope. The nHAP were prepared using nltrasonication and Na_2CO_3 and modified with poly-L-lysine (PLL) at pH 7. 4. The transfection of pGenesil-hTERT into A549 was divided into four groups as follows: nHAP-PLL group mediated by hydroxyapatite nanoparticles modified with poly-L-lysine ( nHAP-PLL), liposome group mediated by Lipefectamine, nHAP group mediated by hydroxyapatite nanoparticles and control group. The growth ability of cells was assayed with methyl thiazolyl tetrazolium meth-od. The expression level of hTERT protein was examined by Western blotting. Flow cytometry was used to detect the apeptosis ratio of A549 cells line. Results Under transmission electron microscope, the synthesized product presented needle-like and well dispersed particles with evenly distributed sizes of (15-20) nm × (60-80) nm. The proliferation of A549 cells of nHAP-PLL group, liposome group and nHAP group were obviously inhibited as compared with the control group (P < 0.05 ).The inhibition rate of nHAP-PLL group was more than the other groups. There was a significant difference inhibition rate be-tween the nHAP-PLL group compared with the liposome group and nHAP group (P <0.05 ). The level of hTERT protein hada similar varietal tendency with the result of proliferation of each group. Flow cytometry showed the apoptasis ratio of nHAP-PLL group, liposome group, nHAP group and control group was (28.1±1.4)%, (19.2±1.3)%, (10.9±1.2)% and (0.3±0.2 ) %, respectively. There was a significant difference in apoptosis ratio between the nHAP-PLL group, liposome group and nHAP group compared with control group( P < 0.05 ). Conclusion A549 human lung cancer cells overexpreas hTERT and this may be a target for inhibiting proliferation of A549. Hydroxyapatite nanoparticles can induce apeptosis of ASA9 cells in vitro. Hydroxyapatite nanoparticles modified with poly-L-lysine can effectively combine and protect DNA and mediate gene transfection to A549, it can mediate human telomerase reverse transcriptase RNA interference of A549 cells and inhibit the pro-liferation of ,4549 in vitro.
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The low dose hyper-radiosensitivity (HRS) in human lung cancer cell line A549 was in-vestigated,the changes of ATM kinase,cell cycle and apoptosis of cells at different doses of radiation were observed,and the possible mechanisms were discussed.A549 cells in logarithmic growth phase were irradiated with 60Co γ-rays at doses of 0-2 Gy.Together with flow cytometry for precise cell sorting,cell survival fraction was measured by means of conventional colony-formation assay.The expression of ATM1981Ser-P protein was examined by Western blot 1 h after radiation.Apoptosis was detected by Hoechst 33258 fluorescent staining,and Annexin V-FITC/PI staining flow cytometry 24 h after radiation.Cell cycle distribution was observed by flow cytometly 6,12 and 24 h after ra-diation.The results showed that the expression of ATM1981Ser-P protein was observed at 0.2 Gy,followed by an increase at >0.2 Gy,and reached the peak at 0.5 Gy,with little further increase as the dose exceeded 0.5 Gy.Twenty-four h after radiation,partial cells presented the characteristic mor-phological changes of apoptosis,and the cell apoptosis curve was coincident with the survival curve.As compared with control group,the cell cycle almost had no changes after exposure to 0.1 and 0.2 Gy radiation (P>0.05).After exposure to 0.3,0.4 and 0.5 Cry radiation,G2/M phase arrest occurred 6 and 12 h after radiation (P<0.05),and the ratio of G2/M phase cells was decreased 24 h after radiation (P<0.05).It was concluded that A549 cells displayed the phenomenon of HRS/IRR.The mode of cell death was mainly apoptosis.The activity of ATM and cell cycle change may take an important role in HRS/IRR.