Evaluation of arjunolic acid against Brucella melitenis and in vitro cytotoxic study of lung adenocarcinomic cell line (A549)

Brucellosis, a neglected tropical disease of zoonotic nature, is caused by the genus Brucella, specifically by Brucella abortus and B. melitensis in cattle and humans, respectively. Arjunolic acid (AA) is a triterpenoid, isolated from Terminalia arjuna (Roxb.) Wight & Arn., a medicinally important plant used to treat various diseases in the Indian system of medicine. Here, we tried to evaluate AA for its antibacterial activity on Brucella and the in vitro cytotoxicity assay on human lung adenocarcinomic alveolar basal epithelial cell line (A549). Also, we assessed the synergistic effect of arjunolic acid and Tarenna asiatica (L.) Kuntze ex K.Schum. on B. melitensis. AA displayed a considerable antibacterial activity [zone of inhibition (9 mm) with a minimum inhibitory concentration of 30 ?g/mL] against B. melitensis. The rate of cell death for the cancer cells were at 100 ?g/mL concentration of AA was 82% which indicates that AA shows significant membrane disruption to cancer cells. The estimated IC50 of AA against the A549 cell line was 139.90 ?g/mL. The highest synergistic activity was exhibited forming a zone of inhibition measuring 10mm when arjunolic acid and AqE of T. asiatica was added in the concentration of 1:1, respectively.

Effects of irbesartan combined with 5-fluorouracil on the proliferation and ERK/PPARγ signaling pathway of Lewis lung cancer cells / 中国药房

OBJECTIVE To explore the effects of irbesartan（Irb）combined with 5-fluorouracil（5-FU）on the proliferation and extracellular signal-regulated kinase （ERK）/peroxidase proliferator-activated receptor γ（PPARγ）signaling pathway of Lewis lung cancer cells. METHODS Lewis lung cancer cells from mice were divided into normal control （NC）group，Irb low-dose （LD）group（1×10－3 mmol/L），Irb high-dose （HD）group（1×10－1 mmol/L），5-FU group （10 μmol/L），Irb LD+ 5-FU group （Irb 1×10－3 mmol/L+5-FU 10 μmol/L）and Irb HD+ 5-FU group （Irb 1×10－1 mmol/L+5-FU 10 μmol/L）. MTT method was used to measure the activity of cell proliferation in each group. Plate colony formation experiment was used to determine the number of cell colonies formed in each group ；Western blot method was used to detect the expression levels of proliferating cell nuclear antigen （PCNA），p53，ERK1/2，p-ERK1/2 and PPAR γ protein in each group. RESULTS Compared with the NC group ，the cell proliferation activity ，the number of colonies formed and the protein levels of PCNA ，p-ERK1/2，and PPARγ were significantly reduced in the other five groups ，and the protein level of p 53 was significantly increased （P＜0.05）；the protein expression of ERK1/2 had no significant difference （P＞0.05）. The changes of above indexes in Irb LD+ 5-FU group and Irb HD+ 5-FU group were more significant than Irb LD group ，Irb HD group and 5-FU group （P＜0.05）. CONCLUSIONS Irb combined with 5-FU can inhibit the proliferation of Lewis lung cancer cell ，and the effect is better than that of the two alone. The mechanism may be related to the inhibition of ERK/PPARγ signal pathway.

In vitro cytotoxicity assay of Fucoidan extracted from Turbinaria conoides against cancer cell lines MCF7, A549, and normal cell line L929

Abstract The main aim of the study is to quantify the cytotoxic property of the Fucoidan extracted from the Turbinaria conoides using the MTT assay with the standard fucose. Fucoidan was extracted using the soaked water method and it was determined using the HPLC procedure the obtained Test sample Fucoidan extracted from the Turbinaria conoides and standard fucose was subjected to the cytotoxicity assay against the MCF7 Human breast cancer cell line, A549 lung cancer cell line, and L929 normal mouse fibroblast cell line. From the results it was found that the Test sample showed good IC50 value for MCF7 cell line then A549 with an increasing concentration 24 hours incubation at 37°C The IC50 for MCF7 was 115.21 µg/ml and A549 396.46µg/ml and the Fucoidan extract was checked for its cytotoxicity against the normal mouse fibroblast cell line L929, Fucoidan was found non-lethal to the L929 mouse fibroblast normal cell line. Standard fucose also gave a significant result towards MCF7 and against the L929. This indicates that the Fucoidan extracted from Tubinaria conoides shows better anticancer potential in it. Hence its application can be further extended in the pharmacological fields.

circ-PRKDC affects the proliferation, apoptosis and radiosensitivity of lung cancer cells by regulating miR-505-3p / 中华放射肿瘤学杂志

Objective:To investigate the effect of circ-PRKDC on lung cancer cell proliferation, apoptosis and radiosensitivity and its molecular mechanism.Methods:Normal lung epithelial cells BEAS-2B and lung cancer cell lines NCI-H1299, NCI-H2170, NCI-H1975 were cultured. NCI-H1299 cells were divided into the si-NC, si-PRKDC, pcDNA-NC, pcDNA-PRKDC, miR-NC, miR-505-3p, anti-miR-NC, anti-miR-505-3p, si-PRKDC+ anti-miR-NC and si-PRKDC+ anti-miR-505-3p groups. RT-qPCR was used to detect the expression levels of circ-PRKDC and miR-505-3p. Western blot was employed to measure the protein expression. MTT was used to detect cell proliferation. Flow cytometry was utilized to detect cell apoptosis. Plate clone formation assay was conducted to detect the cell radiosensitivity. Dual luciferase reporter assay was performed to analyze the targeting relationship between circ-PRKDC and miR-505-3p.Results:Compared with normal lung epithelial cells BEAS-2B, the expression levels of circ-PRKDC in the lung cancer cell lines NCI-H1299, NCI-H2170 and NCI-H1975 were significantly up-regulated (3.65, 3.10, 2.67 vs. 1.00, all P<0.05), whereas those of miR-505-3p were significantly down-regulated (0.42, 0.50, 0.54 vs. 1.02, all P<0.05). After low expression of circ-PRKDC, the expression level of CyclinD1 was significantly down-regulated (0.42 vs. 0.81, P<0.05), whereas those of Cleaved-caspase-3(0.71 vs. 0.33, P<0.05) and γ-H 2AX (0.89 vs. 0.46, P<0.05) were significantly up-regulated, the cell A value was significantly decreased (0.413 vs. 0.839, P<0.05), cell apoptosis rate was significantly increased (20.35 vs. 6.21, P<0.05), cell survival fraction was significantly decreased ( P<0.05), and β-catenin expression was significantly down-regulated (0.35 vs. 0.73, P<0.05). After high expression of miR-505-3p, the expression level of CyclinD1 was significantly down-regulated (0.34 vs. 0.83, P<0.05), those of Cleaved-caspase-3(0.65 vs. 0.32, P<0.05) and γ-H 2AX (0.96 vs. 0.45, P<0.05) were significantly up-regulated, the cell A value was significantly decreased (0.386 vs. 0.851, P<0.05), the apoptosis rate was significantly increased (16.38 vs. 6.20, P<0.05), and the cell survival fraction was significantly decreased ( P<0.05). Compared with miR-NC, the luciferase activity of miR-505-3p group transfected with circ-PRKDC wild-type reporter plasmid was significantly decreased (0.44 vs. 1.00, P<0.05). Down-regulation of miR-505-3p could reverse the effect of low expression of circ-PRKDC on the proliferation, apoptosis, radiosensitivity and β-catenin expression of NCI-H1299 cells. Conclusion:Low expression of circ-PRKDC may inhibit lung cancer cell proliferation, promote cell apoptosis and enhance cell radiosensitivity by up-regulating miR-505-3p, which is probably associated with the Wnt/β-catenin signaling pathway.

Effect and possible mechanism of Salvianolic acid B on radiosensitivity of non-small cell lung cancer / 中华放射肿瘤学杂志

Objective:To evaluate the effect of Salvianolic acid B on the radiosensitivity of human non-small cell lung cancer cells and investigate its possible mechanism.Methods:Non-small cell lung cancer cells A549 and H1299 were cultured in vitro. The toxicity of Salvianolic acid B on non-small cell lung cancer cells was detected by MTT assay. The effect of Salvianolic acid B on the radiosensitivity was assessed by clone formation assay. Transwell chamber assay was used to evaluate the effect of Salvianolic acid B on the migration of tumor cells. Western blot was employed to detect the expression levels of OTUD7B, MMP-2, MMP-9, E-cadherin, Akt and p-Akt regulated by Salvianolic acid B. Results:Salvianolic acid B (5 μmol/L) could inhibit the proliferation of A549 and H1299 cells. Clone formation assay showed that Salvianolic acid B increased the radiosensitivity of A549 and H1299 cells, with a radiosensitization ratio of 1.45 and 1.38, respectively. Transwell chamber assay indicated that the ability of cell migration was significantly inhibited by Salvianolic acid B ( P<0.05). Western blot revealed that the expression levels of OTUD7B in A549 and H1299 cells were induced by irradiation in a time-dependent manner. Salvianolic acid B could down-regulate the expression levels of MMP-2, MMP-9 and p-Akt, whereas up-regulate the expression level of E-cadherin by down-regulating the expression level of OTUD7B. Conclusions:Salvianolic acid B can enhance the radiosensitivity of A549 and H1299 cells. The possible mechanism is that Salvianolic acid B down-regulates the expression level of OTUD7B induced by irradiation and inhibits the epithelial-mesenchymal transition process of tumor cells.

Morinda citrifolia (Noni) Juice Suppresses A549 Human Lung Cancer Cells via Inhibiting AKT/Nuclear Factor-κ B Signaling Pathway / 中国结合医学杂志

OBJECTIVE@#To study the mechanism of the anti-tumor effect of Morinda citrifolia (noni).@*METHODS@#The influences of noni juice on cell proliferation, apoptosis, invasion, migration and the activity of AKT/nuclear factor- κ B (NF- κ B) signaling pathway in A549 human lung cancer cells were detected by MTT, cell counting kit-8, colony formation, Annexin V/PI double labeling, transwell, scratch test and immunoblotting assay, respectively. A549 cells were inoculated into the right axilla of nude mice, followed by noni juice treatment. The body weight of the nude mice was weighed, and the tumor volume and weight were measured. Cell proliferation and expression of apoptosis-related proteins were measured by immunohistochemistry, and the activity of NF- κ B signaling pathway was measured by immunoblotting.@*RESULTS@#The in vitro studies showed that noni juice inhibited the A549 cells proliferation, migration and invasion. Noni juice also promoted cells apoptosis in A549 cells. Immunoblotting assay showed that the phosphorylation level of AKT, p50, and STAT3 proteins was inhibited to different extents after noni juice treatment. The in vivo studies showed that noni juice effectively suppressed tumor formation of A549 cells in nude mice. Noni juice treatment inhibited the expression of Ki67, PCNA, and Bcl-2 protein in the tumor; while promoted the expression of caspase-3 protein. Additionally, we also found that noni juice treatment could restrain the activity of AKT/NF- κ B signaling pathway in the tumor tissue.@*CONCLUSION@#Noni juice inhibited the proliferation of A549 lung cancer cells, induced apoptosis, and inhibited cell invasion and migration via regulating AKT/NF- κ B signaling pathway.

Effects of microRNAs on invasion, migration and apoptosis of lung cancer cell lines / 解剖学报

Objective To investigate the effects of microRNAs(miRNAs) on invasion, migration and apoptosis of lung cancer cell lines. Methods Target cells were infected with hsa-mir-933, hsa-mir-4700-3p, hsa-mir-3144-3p, hsamir-3972, and hsa-mir-548a-5p. Cell apoptosis was measured using flow cytomertry, Transwell invasion assay, cell migration were analyzed by lineation,miRNA was quantified using Real-time PCR. Results Compared with the vector and control, in hsa-mir-933, hsa-mir-4700-3p, hsa-mir-3144-3p, hsa-mir-397 and hsa-mir-548a-5p group the apoptosis significantly increased,cell invasion ability significantly decreased,cell migration ability significantly decreased, miRNAs expression significantly increased. Conclusion Five microRNAs can promote the apoptosis of lung cancer cell lines, change the high invasiveness of lung cancer cell lines, inhibit the migration of lung cancer cell lines, and increase the expression of corresponding miRNAs.

Effects of Periplaneta americana Extract YS-F on the Proliferation and Apoptosis of Human Non-small Cell Lung Cancer A 549 Cells / 中国药房

OBJECTIVE：To investigate the effects of Periplaneta americana extract on the proliferation and apoptosis of human non-small cell lung cancer A 549 cells as well as its possible mechanism. METHODS ：The dry bodies of P. americana were soaked with 90％ ethanol and eluted with gradient water-methanol by polyamide column chromatography. The 20％，30％，40％， 50％，60％，70％，80％，90％ methanol elution sites （YS-A-H）were obtained. MTT method was used to screen the active site ， and the inhibition rate of different doses of active site was detected. Flow cytometry was adopted to detect cell apoptosis ，cell cycle and mitochondrial membrane potential of cells after treated with different doses of active site. RESULTS ：Half inhibition concentrations of YS-A-H were （95.25±8.42），（129.93±7.24），（221.28±12.68），（275.39±14.87），（276.76±16.32），（31.90± 5.34），（163.15±6.97），（122.81±8.36）μg/mL，respectively. YS-F had the strongest activity. After treated with 3，9，27，81 μg/mL YS-F for 24，48，72 h，cell proliferation inhibitory rate was increased significantly at different time points ；after treated for 48，72 h，that was significantly higher than same group after treated for 24 h；after 72 h treatment ，that was significantly higher than same group after 48 h treatment （P＜0.01）. There was no significant effect of 24 h treatment of 3 μg/mL YS-F and 72 h treatment of 9 μg/mL YS-F on the percentage of cells in the late stage of necrosis，24 h treatment of 3 μg/mL YS-F on the percentage of cells in G2/M phase and 48 h treatment of 3 μg/mL YS-F on the reduction rate of mitochondrial membrane potential（P＞0.05）. The percentage of cells in the early stage of apoptosis ，the late stage of apoptosis and the early stage of necrosis ，the late stage of necrosis，as well as the percentage of cells in the Sub-G 0/G1 and S phase at each time point were significantly increased in other different doses groups ，while the percentage of cells in G 0/G1 and G 2/M phase was decreased significantly （P＜0.01）. In each dose group，the percentage of cells in the early stage of apoptosis ，the late stage of apoptosis and the early stage of necrosis ，the late stage of necrosis （except for the percentage of cells in the late stage of necrosis treated with YS-F 9 μg/mL for 72 h）and the percentage of cells in Sub-G 0/G1 phase，G2/M phase （except for YS-F 27，81 μg/mL for 48 h）after treated for 48，72 h were significantly higher than same group after 24 h of treatment ；the percentage of cells in G 0/G1 phase，S phase and G 2/M phase （except for YS-F 9 μg/mL for 48 h）after treated for 48，72 h were significantly lower than same group after 24 h of treatment （P＜0.01）；the percentage of cells in the early stage of apoptosis ，the late stage of apoptosis and the early stage of necrosis ，the late stage of necrosis （except for the percentage of cells in the late stage of apoptosis and early stage of necrosis when treated with YS-F 27 μg/mL for 72 h，the percentage of cells in the late stage of necrosis when treated with YS-F 3，9 μg/mL for 72 h were decreased significantly ）and the percentage of cells in S phase （except for YS-F 3 μg/mL for 72 h）and Sub-G 0/G1 phase after treated for 72 h were significantly higher than same group after 48 h of treatment ，while the percentage of cells in G 0/G1 and G 2/M phase were significantly lower than same group after 48 h of treatment （P＜0.01）. After treated with YS-F 9，27，81 μg/mL for 48 h，the reduction rate of cell mitochondrial membrane potential was increased significantly ；YS-F 27，81 μg/mL groups were significantly higher than YS-F 9 μg/mL group，and YS-F 81 μg/mL group was significantly higher than YS-F 27 μg/mL group. CONCLUSIONS：YS-F can inhibit the proliferation and promote the apoptosis of A 549 cells by preventing cell transformation from S phase to G 2/M phase ，and reducing mitochondrial membrane potential ，in time-dependent or dose-dependent manner.

In vitro antitumor effect of cucurbitacin E on human lung cancer cell line and its molecular mechanism / 中国天然药物

Cucurbitacin E (CuE) is previously reported to exhibit antitumor effect by several means. In this study, CuE acted as a tyrosine kinase inhibitor interfering with the epidermal growth factor receptor/mitogen-activated protein kinase (EGFR/MAPK) signaling pathway and subsequently induced apoptosis and cell cycle arrest in non-small-cell lung cancer (NSCLC) cell line A549. The apoptosis regulators, cleaved Caspases-3 and Caspases-9, were observed to be increased with the treatment of CuE. The activated transcription factor STAT3 and the apoptosis inhibitor protein survivin were also observed to be reduced. The cell cycle regulators, CyclinA2, cylinB1, CyclinD1 and CyclinE, were also investigated and the results suggested that the cell cycle was arrested at G1/G0 phase. Treatment of CuE also altered the existence status of most of the participants in the EGFR/MAPK signaling. Phosphorylation of EGFR enhanced significantly, leading to the alteration of members downstream, either total amount or phosphorylation level, notably, MEK1/2 and ERK1/2. Moreover, the results of molecular simulation brought an insight on the interaction mechanism between CuE and EGFR. In summary, CuE exhibited anti-proliferative effect against A549 cells by targeting the EGFR/MAPK signaling pathway.

Study on Synthesis and Anti-tumor Activity of 6 Kinds of Tectorigenin Mannich Base Derivatives / 中国药房

OBJECTIVE： To conduct structural modification of tectorigenin to search for new compounds with anti-tumor activity. METHODS： Tectorigenin was used as a lead compound， and then added into amine reagents as ethanolamine， methylamine， ethylamine， dimethylamine， diethylamine， n-propylamine and formaldehyde solution. Tectorigenin Mannich base derivatives were synthesized by mannich reaction with as the lead compound. The structures of the derivatives were identified according to IR， UV， MS and NMR data. Solubility of tectorigenin and its derivatives were investigated by solubility test method. MTT assay was used to investigate the inhibitory effects of tectorigenin and its derivatives on the proliferation of human colon cancer cell line HCT116， human lung cancer cell line A549 and human hepatoma cell line HepG2， and half inhibitory concentration （IC50） was calculated. The inhibition rate of tectorigenin and its derivatives （100 mg/kg） on H22 hepatoma-bearing mice in vivo was studied. RESULTS： Totally of 6 kinds of tectorigenin mannich base derivatives were synthesized， such as 8-（N-hydroxyethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-methyl）-methyleneamino-5，7，4′-trihydroxy-6- methoxyisoflavone， 8-（N， N-diethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N， N-dimethyl）-methyleneamino- 5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-ethyl）-methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone， 8-（N-propyl）- methyleneamino-5，7，4′-trihydroxy-6-methoxyisoflavone （compounds 1-6 in turn）. Compared with tectorigenin， the water solubility of six derivatives was significantly improved， and the solubility was 5-20 times higher than that of tectorigenin. IC50 of compounds 1， 3 and 5 to HCT116 cells were （34.82±3.27）， （16.21±4.13）， （33.12±3.25） μmol/L， which were stronger than that of tectorigenin [（45.23±5.74） μmol/L]； IC50 of compounds 1， 3 and 5 to A549 cells were （37.05±5.74）， （26.88±4.52）， （30.13±6.23） μmol/L， which were stronger than that of tectorigenin [（53.24±6.34） μmol/L]； IC50 of compounds 1， 3 and 5 to HepG2 cells were （23.74±1.45）， （18.96±2.34）， （30.95±2.87） μmol/L， which were stronger than that of tectorigenin [（48.98±2.58） μmol/L]. Compounds 1， 3 and 5 showed higher inhibition rates （55.51％， 57.20％ and 49.15％） than tectorigenin （33.05％） on H22 hepatoma-bearing mice， respectively. The other three compounds had no obvious advantage over tectorigenin in anti-tumor activity. CONCLUSIONS： In this study， compounds 1， 3 and 5 of six tectorigenin mannich base derivatives synthesized in this study have stronger antitumor activity than tectorigenin.

Silencing LncRNA HOXD-AS1 enhances the radiosensitivity of H460 cells by regulating miR-454-3p expression / 中华放射肿瘤学杂志

Objective To explore the effect of LncRNA HOXD-AS1 on the radiosensitivity of H460 cells by targeting the regulation of miR-454-3p expression. Methods H460 cells were irradiated with 0, 2, 4, 6 and 8 Gy, and the expression levels of HOXD-AS1 and miR-454-3p were detected by qRT-PCR. Cell apoptosis rate were measured by flow cytometry experiments. Cell cloning experiments were used to detect cell radiosensitivity. The protein expression levels of Caspase-3, Cyclin D1 and γ-H2AX were detected by Western blot. Results The expression of HOXD-AS1 in H460 cells was significantly increased after X-ray irradiation ( P<0.05) , while the expression of miR-454-3p was significantly decreased ( P<0.05) . Silencing HOXD-AS1 significantly promoted apoptosis, increased radiosensitivity of lung cancer cells, promoted Caspase-3, γ-H2AX protein expression, and inhibited Cyclin D1 expression. HOXD-AS1 could target the expression of miR-454-3p. Conclusion Silencing HOXD-AS1 can promote the apoptosis of lung cancer cells and inhibit cell survival by targeting miR-454-3p to increase the radiosensitivity of lung cancer cells.

Synergistic Effect of Flavonoids from Artocarpus heterophyllus Heartwoods on Anticancer Activity of Cisplatin Against H460 and MCF-7 Cell Lines

Artocarpus heterophyllus has been used as traditional medicine. This plant is one of the sources of flavonoid. Flavonoid compounds possessed a wide range of biological properties including anticancer. This study was performed to investigate the cytotoxic effect of flavonoids from A. heterophyllus on H460 and MCF-7 cell lines. The interaction of flavonoids and cisplatin against tested cancer cells was also evaluated. MTT assay was used to determine the cytotoxic effect of flavonoid. Isobologram analysis was selected to evaluate the synergistic effect between flavonoid and cisplatin, their interaction was then confirmed using AO/PI staining method. Amongst of flavonoid compounds, artocarpin exhibited strong cytotoxic effect on both MCF-7 and H460 cell lines with IC₅₀ values of 12.53 µg/mL (28.73 µM) and 9.77 µg/mL (22.40 µM), respectively. This compound enhanced anticancer activity of cisplatin against H460 and MCF-7. The combination produced a synergistic effect on H460 and MCF-7 cell lines with a combination index (CI) values of 0.2 and 0.18, respectively. The AO/PI stained demonstrated that the combination of artocarpin and cisplatin caused morphological changes that indicated apoptosis. Moreover, artocarpanone also significantly increased cytotoxic effect of cisplatin compared to its single concentration with CI below than 1. This result suggested the potency of flavonoid named artocarpin to enhance the anticancer activity of cisplatin on H460 and MCF-7 cell lines.

Construction of lung tumor cells using RFP and Fluc dual-labeled system / 国际生物医学工程杂志

Objective To establish a human lung cancer cell line that can stably express firefly luciferase (Fluc) and red fluorescent protein (RFP) gene so as to lay the foundation for the further establishment of a live-imaging lung cancer xenograft model in nude mice and therapeutic research.Methods The lentiviral vector pHBLV-FlucRFP containing luciferase and red fluorescent protein was constructed and then transfected into 293T cells for virus packaging.The complete virus was used to infect human lung cancer cell lines A549,H1975 and human B-cell lymphoma cell line K562.The stable cell lines were obtained by puromycin selection.Fluorescence microscopy and quantitative PCR were used to confirm the RFP and Fluc expression.Results The lentiviral vector pHBLV-FlucRFP was successfully constructed.Cancer cell line A549,H1975 and K562 stably expressing Fluc and RFP was obtained.The real-time quantitative PCR results showed that the relative expression of Fluc gene in the three stable infected cells was much higher than that in the corresponding wild-type cells,and the differences were statistically significant(all P＜0.05).Conclusion The human lung cancer cell line A549,H1975 and human B-cell lymphoma cell line K562 with dual expression of RFP and Fluc were obtained,which provided a new model of fluorescent cells for in vivo imaging of immunodeficient mouse models such as nude mice.

Antioxidant and cytotoxic activity of Tecoma stans against lung cancer cell line (A549)

ABSTRACT Human have been constantly using plants and plant products to overcome many diseases. The antioxidant property of the plant sources is studied to obtain an efficacious drug against cancer. The objectives of the present study is to evaluate the antioxidant and cytotoxic activity of the Tecoma stans extracts against lung cancer cell line in comparison with vincristine drug. The antioxidant activity was studied using the standard DPPH assay and the cytotoxic activity using MTT assay. DPPH assay results show that methanolic extract of T. stans in higher concentration show better antioxidant potential than the standard L-ascorbic acid. They exhibited strong antioxidant potential at 20 µg/mL concentration. The absorbance at 517 nm showed that in the range of 0.201-0.0203 compared to that of absorbance of ascorbic acid at 0.023.Cytotoxic activity was studied using MTT assay which showed that the increase in concentration of extract increases the cell death. At 100µg/mL concentration there is an increased cytotoxic activity, i.e., 99% of cell inhibition. The results of antioxidant and anticancerous activity may be positively correlated.

miR-575 Promotes the Proliferation and Invasion of Non-small Cell Lung Cancer Cell by Negatively Regulating BLID Expression / 现代生物医学进展

Objective:To explore the mechanisms of regulation of miR-575 on the proliferation and invasion properties of non-small cell lung cancer cell (NSCLC).Methods:Real-time PCR was selected to detect the expression of miR-575 and BLID in differ ent NSCLC cell lines.CCK-8 assay was processed to measure the alternations of A549 cell proliferation at different time points after transfection of miR-575 mimic and miR-575 inhibitor.The invasion ability of A549 cells was evaluated by transwell.The targeting of BLID by miR-575 was predicted by Targetcan software and verified by dual-Luciferase assay.BLID protein expression level was detected by western blot.Results:miR-575 highly expressed in NSCLC cell lines,including A549,SPC-A1,H1299,H1650 (P＜0.001),miR-575 mimic could efficiently elevated the expression ofmiR-575 in A549 cells (P＜0.001),and strengthened the proliferation and invasion ability of NSCLC cells (P＜0.05),while,transfection of miR-575 inhibitor could down-regulate the expression ofmiR-575,and also inhibit the proliferation and invasion ability of NSCLC cells (P＜0.01).Targetscan software predicted that BLID might be the target gene of miR-575,and dual-luciferase assay revealed that miR-575 could obviously decrease the luciferase reaction of wild type BLID 3'UTR (P＜0.01),besides,miR-575 could down-regulate the protein expression ofBLID (P＜0.01).Real-time PCR results showed that NSCLC cell lines had lower level of BLID mRNA expression compared with 16HBE control cells (P＜0.001),and restore of BLID could markedly inhibited cell proliferation and invasion ability (P＜0.05),which could be reversed by miR-575 co-tranfection (P＜0.01).Conclusion:In NSCLC cells,the expression ofmiR-575 could promote cell proliferation and invasion ability by directly regulating downstream target tumor-suppressor gene BLID expression.

Study of mechanisms of hesperidin onanti-lung cancer effect based on microfluidic chip technology / 中国药理学通报

Aim To investigate the effect of hesperidin on human lung cancer cell A549 and the possible mechanism.Methods The cell apoptosis and necrosis of A549 treated with hesperidin were measured by the Hoechst 33342/PI fluorescent dye based on microfluidic chip technology.Cell cycle and apoptosis rate were evaluated by flow cytometry(FCM).The expressions of the related genes were detected through the real-time fluorescent quantitative PCR technology(RT-PCR) including VEGF, PI3K and PTEN.The protein expressions of Bcl-2, Cyclin B1, PI3K, Akt and PTEN were detected by Western blot after hesperidin intervention.Results The proliferation of A549 cells was significantly inhibited by hesperidin in a dose-dependent manner.FCM results showed that hesperidin could not only influence the G0/G1 phase and S phase, but also promote the apoptosis of lung cancer cells.Meanwhile, the apoptosis and necrosis rate was increased from(6.7±0.6)% to(27.9±1.1)% compared with that of control group(P<0.05).From the level of molecular, the gene expressions of VEGF and PI3K were decreased, while the PTEN was increased after hesperidin stimulation.Western blot results showed that the expression of protein Bcl-2, Cyclin B1 and Akt were decreased, which all showed close relationship with cell apoptosis, cell cycle and PI3K-Akt signaling pathway.The expression of PI3K was increased, but the change of PTEN was not statistically significant compared with that of control group.Conclusion Hesperidin induces lung cancer cell apoptosis through PI3K-Akt signaling pathway, which blocks cancer cell division and destroys the balance of related protein expression.

Effects of rhTIMP-3 combined with DDP on growth inhibition and apoptosis of lung cancer cell A549 / 西安交通大学学报(医学版)

Objective To study the effects of restructuring tissue inhibitor of matrix metatloproteinase-3 (rhTIMP-3) in combination with cisplatin (DDP) on the growth and apoptosis of A549 lung cancer cell line.Methods We made individual and combined use of different concentrations of rhTIMP-3 and DDP on A549 cells.Methyl thiazoyl terazolium (MTT) colorimetry was used to analyze cell growth inhibition,and flow cytometry technique was used to determine the cell cycle distribution and apoptosis rate.Results rhTIMP-3 and DDP both could inhibit the proliferation of A549 cells.rhTIMP-3 exerted its effect in the time-and concentration-dependent manners while DDP did so in the concentration-dependent manner;both induced the apoptosis of A549 cells.rhTIMP-3 could make the cells stay in S and G2/M phases,and DDP made the cells stay in S phase.The combination of them obviously strengthened the inhibition of A549 cell growth,and had obvious synergy in inducing apoptosis.Conclusion Both rhTIMP-3 and DDP can inhibit the proliferation of A549 cells and induce their apoptosis.The combined use of them not only can increase the inhibition of cell growth but also has synergy in inducing cell apoptosis.

The anti-tumor effect and underlying mechanism of micheliolide on H460 lung cancer by notch4 signal pathway / 医学研究生学报

Objective Previous studies have found that micheliolide(MCL) could improve the sensitivity of breast cancer cells to chemotherapeutic drugs such as cisplatin and induce the apoptosis of breast cancer cells.This article aims to study the proliferation inhibition effect of micheliolide on lung cancer cells H460 and its underlying mechanism.Methods Human lung cancer cell line H460 was treated with different concentrations of micheliolide(30,60,90μmol/L).Then the cell proliferation was measured by-CCK8 and plate colony formation assays.The apoptosis and the cell cycle were detected by flow cytometry.Western blot was used to determine the mechanism of how MCL affecting cancer cell H460.Results Compared with the control (275.00±7.21), the clone numbers after 30μmol/L,60μmol/L and 90μmol/L MCL treatment(199.00±5.66,166.00±1.41, 90.00±7.81) were significantly decreased (P<0.05).Meanwhile, the CCK-8 results showed that compared to the control group, A value was significantly increased after 30μmol/L MCL treatment for 72h and 96h, and 60μmol/L or 90μmol/L MCL treatment for 48h,72h and 96 h (P<0.05).Compared with the control apoptotic ratio [(2.90±0.03)％], the ratio of early and late apoptotic cells after 30μmol/L, 60μmol/L and 90μmol/L MCL treatment [(5.23±0.76)％, (9.06±0.47)％, (19.00±0.64)％] were significantly increased (P<0.05).Compared with the control group, the ratio of G2/M phase cells[(12.52±0.88) ％ ,(17.22±0.43)％, (19.84±0.31)％] was gradually increased after 30μmol/L, 60μmol/L and 90μmol/L MCL treatment, and there was statistically significant difference after 60μmol/L and 90μmol/L MCL treatment (P<0.05).The ratio of S+G1/G0 phase cells[(87.53±1.06)％ ,(82.94±0.67)％ ,(79.79±0.21)％] was gradually decreased after 30μmol/L, 60μmol/L and 90μmol/L MCL treatment, and there was statistically significant difference after 60μmol/L and 90μmol/L MCL treatment (P<0.05).The expression level of notch4 was significantly decreased after 30μmol/L, 60μmol/L and 90μmol/L MCL treatment (P<0.05), while the expression level of cleaved caspase3 was significantly upregulated (P<0.05).Conclusion MCL exerted an inhibitory effect on lung cancer cell H460.

Liver kinase B1 gene enhances radiosensitivity of lung cancer H460 cells:an in vivo study / 中华放射肿瘤学杂志

Objective To investigate the effect of liver kinase B1(LKB1) on the radiosensitivity of subcutaneous xenograft tumor of lung cancer H460 cells in nude mice.Methods Human lung cancer H460 cells were implanted into female nude mice (BALB/c-nu) to establish a subcutaneous xenograft tumor model of lung cancer.A total of 24 female nude mice in which the model was successfully established were equally and randomly divided into four groups:pEGFP-Ctrl plasmid (empty vector plasmid) group, irradiation (IR)+pEGFP-Ctrl plasmid group, pEGFP-LKB1 plasmid (overexpressing LKB1) group, and IR+pEGFP-LKB1 plasmid group.The growth of xenograft tumors was observed and the tumor inhibition rate and enhancement factor (EF) were calculated.The expression of LKB1 in each group was measured by immunohistochemistry and Western blot to analyze the relationship between LKB1 and radiosensitivity.Results Compared with the pEGFP-Ctrl plasmid group, the IR+pEGFP-Ctrl plasmid group, pEGFP-LKB1 plasmid group, and IR+pEGFP-LKB1 plasmid group showed varying degrees of inhibition of tumor growth, particularly in the IR+pEGFP-LKB1 plasmid group, and the tumor inhibition rates were 31.30%, 14.78%, and 43.48%, respectively.The EF of LKB1 in the IR+pEGFP-LKB1 plasmid group was 1.18.The immunohistochemistry and Western blot showed that LKB1 could be effectively expressed in the pEGFP-LKB1 plasmid group and IR+pEGFP-LKB1 plasmid group, but not in the other two groups.Conclusions The subcutaneous xenograft tumor model of human lung cancer H460 cells has been successfully established in nude mice.LKB1 has a radiosensitizing effect on the subcutaneous xenograft tumor of lung cancer H460 cells in nude mice.

Effects of PM2.5 exposure on autophagy and apoptosis in A549 cells / 实用医学杂志

Objective To investigate the effect of PM2.5 airborne particulate matter with a mean diameter of less than 2.5μm exposure on autophagy and explore the links between autophagy and apoptosis in human lung cancer cells(A549). Methods A549 cells were exposed to 100μg/mL PM2.5 with or without 3-MA(autophagy inhibitor)for various periods of 0、2、4、12 or 24 hrs. Autophagy in A549 cells was assessed by determining the lev-el of LC3(a known autophagy marker)using confocal microscopy. The level of microtubule-associated protein 1 light chain 3(LC3) Ⅱ conversion and Bax (a pro-apoptotic protein) was detected by western blotting. Results The expression of LC3 and the ratio of LC3-II/LC3-I in A549 cells was significantly increased and Bax was signifi-cantly decreased following exposure to PM2.5100 μg/mL for 24 h in a time-dependent manner(P < 0.05). After treated with 100 μg/mL PM2.5,the formation of LC3 in A549 cells as evidenced by the intensity of intracellular fluorescence was significantly increased ,and autophageosomes were observed around nucleus in A549 cells. Fur-thermore blockage of autophagy by 3-MA led to a significant increase in the pro-apoptotic protein Bax. Conclu-sion PM2.5 exposure induces autophagy which may protect against apoptosis induced by PM2.5 in A549 cells.