Colony Formation Of A549 Cells Research Articles

Network pharmacology analysis, molecular docking integrated experimental verification reveal β-sitosterol as the active anti-NSCLC ingredient of Polygonatum cyrtonema Hua by suppression of PI3K/Akt/HIF-1α signaling pathway

Ethnopharmacological relevancePolygonatum cyrtonema Hua (Huangjing) is a Chinese herb that is considered by ancient Chinese healers to have the effect of nourishing yin and moisturizing the lungs. It is clinically used to treat diseases of the pulmonary system, including non-small cell lung cancer. However, the precise active components and underlying mechanisms of Huangjing in the context of treating NSCLC remain uncertain. Aim of the studyThis study aimed to explore the active components and mechanisms of Huangjing for the treatment of NSCLC by means of data mining, network pharmacology, and in vitro and vivo experiments. Materials and methodsFirst, the main active compounds and key targets of Huangjing were predicted by network pharmacology. The potential key targets of Huangjing were molecularly docked with the main active compounds using Pymol. In vivo, we verified whether Huangjing and its main active compound have anti-lung cancer effects. Key targets were verified by PCR and immunohistochemistry. In vitro, we verified the effects of Huangjing's main active compound on the proliferation, apoptosis, and migration of A549 cells by CCK-8, colony formation, wound healing assay, and flow cytometry. Key targets and signaling pathway were validated by PCR and Western blot. ResultsThe network pharmacology results suggested that β-sitosterol was the main active substance. TP53, JUN, AKT1, MAPK14, ESR1, RELA, HIF1A, and RXRA were potential targets of Huangjing. Molecular docking results suggested that MAPK14, HIF-1α, and RXRA docked well with β-sitosterol. In vivo tests also confirmed that Huangjing could significantly inhibit the growth of lung cancer tumors, while PCR and immunohistochemistry results suggested that the expression of HIF-1α was significantly decreased. Critically, KEGG analysis indicated that the PI3K/Akt/HIF-1α signaling pathway was recommended as one of the main pathways related to the anti-NSCLC effect of Huangjing. We conducted in vitro experiments to confirm the significant impact of β-sitosterol on the proliferation, apoptosis, migration, and colony formation of A549 cells. Furthermore, our findings indicate that a high dosage of β-sitosterol may effectively decrease the expression of HIF-1α, AKT1, JUN and RELA in A549 cells. Similarly, in vitro experiments also revealed that high doses of β-sitosterol could inhibit the PI3K/Akt/HIF-1α signaling pathway. ConclusionsWe discovered Huangjing and its main active ingredient, β-sitosterol, can reduce HIF-1α, AKT1, JUN and RELA expression and decrease non-small cell lung cancer growth through the PI3K/Akt/HIF-1α signaling pathway.

Synthesis and in-vitro study of a novel ligustrazine diselenide as a potential chemotherapy drug for lung adenocarcinoma

A novel ligustrazine diselenide, 1,2-bis ((3,5,6-trimethylpyrazin-2-yl) methyl) diselenide (Se2), for potential treatment on adenocarcinoma of lung cancer was successfully synthesized and fully characterized by various analytical approaches. Cytotoxic, antiproliferative and apoptosis-triggering mechanism of Se2 compound have been investigated through human lung adenocarcinoma (LUAD) cell line A549. The study found that Se2 significantly inhibit the proliferation of A549 cells in a dose-dependent manner. Flow cytometry showed that Se2 induced cell arrest and apoptosis in S and G2/M phase, and the apoptotic effect of Se2 were associated with the increase of caspase 3 and PARP-1 level approved by western blot assay. Further mechanism study results suggested that Se2 suppressed the migration,invasion and colony formation of A549 cells, significantly inhibited the PI3K/Akt/m-TOR signaling pathway. The study indicated that Se2 is a bioactive substance that can induce apoptosis of A549 cells in-vitro, and it is a potent candidate drug for LUAD.

CUR5g, a novel autophagy inhibitor, exhibits potent synergistic anticancer effects with cisplatin against non-small-cell lung cancer

Autophagy, a highly conserved degradation process of eukaryotic cells, has been proven to be closely related to chemoresistance and metastasis of non-small-cell lung cancer (NSCLC). Autophagy inhibitors, such as chloroquine (CQ) and its derivative hydroxychloroquine (HCQ), has been shown to mediate anticancer effects in preclinical models, especially when combined with chemotherapy. However, the vast majority of autophagy inhibitors, including CQ and HCQ, actually disrupt lysosomal or/and possibly non-lysosomal processes other than autophagy. It is therefore of great significance to discover more specific autophagy inhibitors. In this study, after screening a series of curcumin derivatives synthesized in our laboratory, we found that (3E,5E)-1-methyl-3-(4-hydroxybenzylidene)-5-(3-indolymethylene)-piperidine-4-one (CUR5g) selectively inhibited autophagosome degradation in cancer cells by blocking autophagosome-lysosome fusion. CUR5g did not affect the lysosomal pH and proteolytic function, nor did it disturb cytoskeleton. CUR5g blocked the recruitment of STX17, a soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein, to autophagosomes via a UVRAG-dependent mechanism, resulting in the inability of autophagosomes to fuse with lysosomes. CUR5g alone did not induce apoptosis and necrosis of A549 cells, but significantly inhibited the mobility and colony formation of A549 cells. More excitingly, CUR5g showed no obvious toxicity to normal HUVECs in vitro or mice in vivo. CUR5g enhances the cisplatin sensitivity of A549 cells and effectively inhibited autophagy in tumor tissues in vivo. Collectively, our study identified a new late-stage autophagy inhibitor and provided a novel option for NSCLC treatment, particular when combined with cisplatin.

SECONDARY METABOLITES OF ANTAGONISTIC BACTERIAL STRAIN Cytobacillus firmus TCP1 EXERT ANTIBACTERIAL AND ANTICANCER ACTIVITY AGAINST LUNG CANCER A549 CELLS

Marine bacteria are abundant producers of many different kinds of bioactive substances. In this study, we isolated the antagonistic marine bacterial strain Cytobacillus firmus TCP1 from soil samples collected from the shoreline and evaluated the antibacterial and anticancer effects of its crude secondary metabolites. The potent antagonistic bacterial isolate C. firmus was identified using 16S rRNA sequencing analysis. The secondary metabolites from strain C. firmus were extracted by centrifugation with ethanol, and the agar well diffusion method was used to evaluate them for antibacterial activity against respiratory tract bacterial pathogens. This study demonstrated that the metabolic extract of the TCP1 effectively inhibited the growth of respiratory tract bacterial pathogens. Moreover, it established concentration-dependent cytotoxicity on A549 lung cancer cells. The administration of metabolic extract of the TCP1 showed an effective inhibitory effect on colony formation of A549 cells, increased lipid peroxidation, and decreased the antioxidant activity in A549 cells. The results showed that the metabolic extract of the TCP1 showed effective antibacterial and anticancer activity. Therefore, upon additional clinical testing, the secondary metabolites of strain C. firmus TCP1 may be exploited to create potent antibiotics and anticancer drugs for lung cancer.

Pomolic acid inhibits proliferation of human lung carcinoma cells via induction of apoptosis and suppression of cell migration and invasion

Purpose: To investigate the anti-proliferative effect of pomolic acid on lung cancer cells (A549), and theunderlying mechanism.Methods: The viability of pomolic acid-treated A549 cells was determined by MTT and colony formation assays. Cell colony formation was monitored with acridine orange/ethidium bromide (AO/EB) staining. Protein expressions of Bax and Bcl-2 were assayed by western blotting.Results: Pomolic acid suppressed the growth of A549 cells, with an half-maximal inhibitory concentration of (IC50) of 10 μM (p < 0.05). However, the IC50 of pomolic acid for normal BEAS-2B cells was 80 μM. Pomolic acid also decreased colony formation of A549 cells. At 20 μM, the percentage of A549 colonies decreased to 14 % of control. The dose-dependent cytotoxicity of pomolic acid against A549 cells was mediated via induction of apoptosis and oxidative stress. Pomolic acid treatment enhanced the expression of Bax and decreased the expression of Bcl-2 in A549 cells. Moreover, pomolic acid inhibited the migration and invasion in A549 cells in a dose-dependent manner (p < 0.05).Conclusion: These results indicate the potent anticancer effect of pomolic acid against human lung cancer cells. Thus, pomolic acid has promising potential as a lead molecule for the development of chemotherapy.

Comparative Evaluation of Apoptosis Induction Using Needles, Bark, and Pollen Extracts and Essential Oils of Pinus eldarica in Lung Cancer Cells

Lung cancer is one of the leading causes of cancer-related mortality worldwide. Although effective clinical drugs for treating advanced stages are available, interest in alternative herbal medicines has gained momentum. Herbal extracts are potent antioxidants that reportedly inhibit the growth of various cancer cell lines. In the present study, we investigated the effects of essential oils and hexane, methanolic, and aqueous extracts, obtained from various parts (bark, needles, and pollen) of Pinus eldarica against human lung cancer (A549) cells. First, the DPPH radical scavenging activities of P. eldarica extracts and essential oils were examined, which revealed that methanolic extracts presented higher antioxidant activity than the other extracts and essential oils. Next, A549 cells were exposed to various concentrations of the extracts and essential oils for 48 h. P. eldarica extracts/essential oil-treated lung cancer cells demonstrated a significant decrease in cell proliferation, along with an induction of apoptotic cell death, particularly, the pollen hexane extract, bark essential oil, and methanolic needle extract showed superior results, with IC50 values of 31.7, 17.9, and 0.3 μg/mL, respectively. In the cell cycle analysis, treatment of A549 cells with the methanolic needle and pollen hexane extracts led to apoptosis and accumulation of cells in the sub-G1 phase. Further, exposure to the bark essential oil and methanolic needle extract decreased the cell population in the G2/M phase. Notably, treatment with the pollen hexane extract, bark essential oil, and methanolic needle extract resulted in caspase-3 activation, poly (ADP-ribose) polymerase cleavage, Bcl-2 downregulation, and Bax and p53 regulation in A549 cells. Furthermore, these extracts and essential oils decreased the migration, and colony formation of A549 cells. These findings provide experimental evidence for a new therapeutic effect of P. eldarica against human lung cancer and suggest P. eldarica as a potential chemopreventive natural resource for developing novel cancer therapeutics.

Methyltransferase-like 1 regulates lung adenocarcinoma A549 cell proliferation and autophagy via the AKT/mTORC1 signaling pathway.

Methyltransferase-like 1 (METTL1) is a transfer RNA and microRNA modifying enzyme. However, its role in lung adenocarcinoma (LUAD) remains unknown. The present study aimed to investigate the effect of METTL1 in LUAD and determine the association between METTL1 expression and prognosis of patients with LUAD. The expression profile of METTL1 in LUAD tissues was downloaded from public cancer databases and analyzed using the Gene Expression Profiling Interactive Analysis database and UALCAN online software. In addition, the association between METTL1 expression and prognosis of patients with LUAD was assessed using the Kaplan-Meier Plotter software. The effect of METTL1 in the A549 cell line was determined in vitro via overexpression and knockdown experiments. The results demonstrated that METTL1 was upregulated in LUAD tissues, and its increased expression was associated with unfavorable prognosis. Furthermore, METTL1 promoted proliferation and colony formation of A549 cells, and inhibited autophagy via the AKT/mechanistic target of rapamycin complex 1 signaling pathway. Taken together, the results of the present study suggest that METTL1 acts as an oncogene in LUAD, thus may be a potential prognostic predictor and therapeutic target for LUAD.

Cyano Enone-Bearing Triterpenoid Soloxolone Methyl Inhibits Epithelial-Mesenchymal Transition of Human Lung Adenocarcinoma Cells In Vitro and Metastasis of Murine Melanoma In Vivo.

Introduction of α-cyano α,β-unsaturated carbonyl moiety into natural cyclic compounds markedly improves their bioactivities, including inhibitory potential against tumor growth and metastasis. Previously, we showed that cyano enone-bearing derivatives of 18βH-glycyrrhetinic (GA) and deoxycholic acids displayed marked cytotoxicity in different tumor cell lines. Moreover, GA derivative soloxolone methyl (SM) was found to induce ER stress and apoptosis in tumor cells in vitro and inhibit growth of carcinoma Krebs-2 in vivo. In this work, we studied the effects of these compounds used in non-toxic dosage on the processes associated with metastatic potential of tumor cells. Performed screening revealed SM as a hit compound, which inhibits motility of murine melanoma B16 and human lung adenocarcinoma A549 cells and significantly suppresses colony formation of A549 cells. Further study showed that SM effectively blocked transforming growth factor β (TGF-β)-induced epithelial-mesenchymal transition (EMT) of A549 cells: namely, inhibited TGF-β-stimulated motility and invasion of tumor cells as well as loss of their epithelial characteristics, such as, an acquisition of spindle-like phenotype, up- and down-regulation of mesenchymal (vimentin, fibronectin) and epithelial (E-cadherin, zona occludens-1 (ZO-1)) markers, respectively. Network pharmacology analysis with subsequent verification by molecular modeling revealed that matrix metalloproteinases MMP-2/-9 and c-Jun N-terminal protein kinase 1 (JNK1) can be considered as hypothetical primary targets of SM, mediating its marked anti-EMT activity. The inhibitory effect of SM on EMT revealed in vitro was further confirmed in a metastatic model of murine B16 melanoma: SM was found to effectively block metastatic dissemination of melanoma B16 cells in vivo, increase expression of E-cadherin and suppress expression of MMP-9 in lung metastatic foci. Altogether, our data provided valuable information for a better understanding of the antitumor activity of cyano enone-bearing semisynthetic compounds and revealed SM as a promising anti-metastatic drug candidate.

Aqueous extract of Fritillariae cirrhosae induces cellular apoptosis through activation of STATs-mediated immunomodulation

Ethnopharmacological relevanceFritillariae cirrhosae (FC), referred to‘Chuan beimu'in China. As an important edible and medicinal plant, the bulbs of F.cirrhosae is used traditionally in the treatment of pulmonary diseases associated with lung heat, inflammation and tumors. In the study, we investigated the effect of aqueous extract of FC (FC-AE) and elucidated its mechanism in non-small cell lung cancer A549 cells and a xenograft model of nude mice. Materials and methodsCCK-8 and plate colony formation assay were used to evaluate the effect of FC-AE in A549 cells in vitro, and the gene expression profile of FC-AE on A549 cells was assessed by RNA sequencing system. Then, the effects of FC-AE on cell cycle and apoptosis of A549 cells were analyzed by flow cytometry. In combination with RNA-seq data, RT-PCR and western blot were used to evaluate the expression of proteins related to apoptosis and immune regulation. A xenograft model of nude mice was used to assess the effect of FC-AE in vivo. ResultsCCK-8 and plate cloning assays showed that FC-AE inhibited the proliferation and colony formation of A549 cells. A549 cells treated with FC-AE can triggered apoptosis. GO and KEGG pathway enrichment analysis of RNA-seq data showed that most of the differentially expressed genes (DEGs) were related to immune response, apoptosis and cell cycle process. Several immune and apoptotic DEGs were identified by qRT-PCR which were consistented with RNA-seq data. In nude mice, FC-AE reduced the tumor size and promoted the secretion of cytokines IL12 and IFNγ. FC-AE up-regulated the two members (STAT1 and STAT4) of STATs and their target genes (IFNγ and IL-12, respectively) protein expressions, and actively regulates Bcl-2/Bax family proteins which resulted in cellular apoptosis in A549 cells. ConclusionOur finding suggests that FC-AE mediates apoptosis through a STAT1 and STAT4-mediated co-regulatory network, which may be the key novel mechanism for its antitumor activity. The F. cirrhosa may be a promising antitumor drug for modulating immune responses to improve cancer therapy.

2-Hydroxy-3-methylanthraquinone inhibits lung carcinoma cells through modulation of IL-6-induced JAK2/STAT3 pathway

Background2-hydroxy-3-methylanthraquinone (HMA), an anthraquinone monomer in traditional Chinese medicine Hedyotis diffusa, has been reported to inhibit the growth of several types of cancer, but its effect on lung cancer has not been adequately investigated. Hypothesis/PurposeThis study aimed to test the hypothesis that HMA inhibit the growth, migration, and invasion of lung cancer cells in part via downregulation of interleukin (IL)-6-induced JAK2/STAT3 pathway. MethodsGrowth and apoptosis of lung cancer cells were quantitated by CCK-8 assay and Annexin V-FITC/PI flow cytometric analysis, respectively. Migration and invasion of A549 cells were determined by wound-healing assay and transwell invasion assay, respectively. The effect of HMA on cytokines expression in A549 cells was evaluated by the cytokine antibody array assay. Gene expression and protein levels of related molecular markers were quantitated by real time-PCR and Western blot analysis, respectively. ResultsHMA significantly inhibited IL-6-stimulated growth and colony formation of A549 cells, increased the number of apoptotic cells, and inhibited invasion associated with downregulation of expression of IL-6-induced MMP-1, MMP-2, and MMP-9 genes. IL-6 increased the levels of tyrosine phosphorylation of JAK2 and STAT3 in A549 cells, which was reversed by HMA treatment. In addition, HMA reduced the expression of a series of inflammation-related cytokines in A549 cells supernatant, including IL-6, G-CSF, IL-6R, IL-8, MCP-1, RANTES, TNF-α. ConclusionThese results suggest that HMA may inhibit the growth and invasion of lung cancer cells in part via downregulation of IL-6-induced JAK2/STAT3 pathway.

Knockdown of high mobility group box 3 impairs cell viability and colony formation but increases apoptosis in A549 human non-small cell lung cancer cells.

Previous research has linked high mobility group box 3 (HMGB3) overexpression to the malignant progression and poor prognosis of non-small cell lung cancer (NSCLC). The present study investigated the role of HMGB3 in cell survival and colony formation of NSCLC cells. Stable knockdown of HMGB3 in A549 cells was achieved by lentiviral-based shRNA interference and verified by detection of the transcriptional and translational level of HMGB3 with reverse transcription-quantitative polymerase chain reaction and western blotting, respectively. The influence of HMGB3 knockdown on A549 cell viability and apoptotic rate was evaluated by Cell Counting Kit-8 assay and flow cytometry following annexin V staining, respectively. The proliferative capacity of A549 cells with or without HMGB3 knockdown was compared by measuring their colony forming efficiency. The results of the current study revealed that HMGB3 knockdown significantly reduced cell viability and colony forming efficiency while promoting apoptosis in A549 cells, indicating that HMGB3 may be pivotal for the survival and colony formation of A549 cells, serving a notable role in NSCLC progression.

Abstract 109: Effects of trifluoperazine analogue on A549 human lung cancer cells

Abstract Although there have been great advancement in technology, molecular diagnosis, therapeutics, lung cancer is still the leading cause of cancer related mortality all over the world. Recently, some antipsychotic drugs have been shown to possess the anticancer activity. Thus the present study was designed to evaluate the anticancer effects of trifluoperazine (TFP), a commonly used antipsychotic drug, and its synthetic analogs on human lung cancer cell lines. To this end, effects of TFP and its selected analog on A549 cells were investigated in vitro as well as in vivo experiments. Synthetic TFP analogs were evaluated by the viability of A549 cells following drug treatment and compared to TFP. KCFC51, a selected TFP analog, significantly inhibited the proliferation of A549 cells. Further experiment showed that TFP and KCFC51 had activities to inhibit the migration, and anchorage dependent/independent colony formation of A549 cells. Western blot analysis revealed that TFP and KCFC51 affect the protein expression levels related to cell apoptosis and cell cycle. KCFC51 showed stronger anticancer effects in all the experiment than TFP. Flow cytometric analysis showed that KCFC51 induced sub-G1 population and reduced cell population in S and G2/M phase. In addition, the two in vivo experimental models, KCFC51 also showed powerful anticancer effect in skin xenograft tumor growth and orthotropic lung cancer development than TFP. Thus present study demonstrates that a synthetic TFP analog has anti-lung cancer activity and provides a potential therapeutic candidate for lung cancer. Citation Format: Joo Yeon Jeong, Jinhyun Ryu, Nal Ae Yoon, Seokmin Kang, Hyemin Seong, Ahmad Fudhaili, Juyeong Park, Jimin Sim, Jiyeon Lee, Soohyun Hwang, Sang Soo Kang. Effects of trifluoperazine analogue on A549 human lung cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 109. doi:10.1158/1538-7445.AM2017-109

Elevated expression of STIM1 is involved in lung tumorigenesis

This study aimed to address the potential role of STIM1 (stromal interaction molecule 1) in lung tumorigenesis. Colony formation in soft agar assay and tumorigenicity in nude mice assay were conducted. Western blot, immunohistochemistry and quantitative real-time polymerase chain reaction were used to measure the STIM1 expression. The distribution of cell cycle was detected by flow cytometry assay. Our results showed that the expression of STIM1 mRNA was significantly higher in human lung tumors than that in adjacent non-neoplastic lung tissues. Significantly increased expression of STIM1 mRNA and protein was observed in 16HBE-benzo(a)pyrene (BaP) cells and in BaP-treated mice lung tissues compared with 16HBE-control cells and the control group, respectively. Silencing STIM1 inhibited the proliferation and colony formation of A549 cells in in vitro experiments, attenuated the growth of tumor xenografts of A549 cells in in vivo experiments and induced the arrest of cell cycle in the G1 phase. The markedly decreased expression of cyclin D1 protein was observed in A549-shRNA-STIM1 cells as compared to A549-shRNA-control cells. The markedly increased expression of p21 protein was observed in A549-shRNA-STIM1 cells as compared to A549-shRNA-control cells. The expression levels of β-catenin and TGIF proteins were lower in A549-shRNA-STIM1 cells than those in A549-shRNA-control cells. In conclusion, this study indicated that the elevated expression of STIM1 might be involved in lung tumorigenesis.

Decreased MicroRNA-26a expression causes cisplatin resistance in human non-small cell lung cancer

ABSTRACTBackground: Lung cancer is the most common cancer that is caused by perturbation of regulatory pathways rather than dysfunction of a single gene. Cisplatin (CDDP; cis-diamminedichloroplatinum II) is the first member of a class of platinum-containing anti-cancer medication, which binds to DNA and triggers apoptosis. CDDP-based chemotherapy is used to treat various types of cancers. However, the efficacy of CDDP in the treatment of non-small-cell lung cancer (NSCLC) is limited by acquired drug resistance. MicroRNAs have recently emerged as key regulators of cancers, and miR-26a is one of down-regulated miRNAs in A549/CDDPres cell line. This study aimed to investigate the role of miR-26a in CDDP resistance in NSCLC as well as the underlying mechanisms.Methods: In this study, we analyzed expressional profiles of CDDP resistance-related mRNA, miRNA, and transcription factors (TF) that regulate miRNA expression in NSCLC. A549 cells were treated with CDDP, miR-26a mimic, or miR-26a inhibitor, and followed by biological analysis including drug sensitivity assay, colony formation assay, terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling (TUNEL) assay, and cell cycle analysis. Luciferase assay was used to determine the target of miR-26a. The regulation of miR-26a in Akt pathway was measured by western blot.Results: High mobility group A (HMGA) 2 was identified as the target of miR-26a. Overexpression of miR-26a in A549 cells inhibited G1-S transition, increased cell death in response to CDDP treatment, and decreased the colony formation of A549 cells. MiR-26a significantly decreased the expression of E2F1, diminished Akt phosphorylation, and downregulated Bcl2 expression. Cell growth was suppressed by inhibiting HMGA2-mediated E2F1-Akt pathway.Conclusion: MiR-26a is responsible for A549 cell sensitivity in the treatment of CDDP through regulating HMGA2-mediated E2F1-Akt pathway.

Novel taspine derivative 12k inhibits cell growth and induces apoptosis in lung cell carcinoma

Taspine is an active compound in anticancer agent development. 12k was synthesized with taspine as lead compound bearing biphenyl scaffold and showed potent anticancer activity. Here, we investigated the effect of taspine derivative 12k on A549 lung cells. We showed that 12k not only decreased significantly A549 cell viability, A549 cell colony formation but also impaired A549 cell migration. Moreover, 12k treatment blocked cell cycle progression by increasing cell number in S phase to 42.80% for 6μmol/L vs. 28.86% for control while decreasing cell number in G1 phase. Accordingly, this was associated with an increase protein expression of cyclin E and a decrease protein expression of cyclin D1, cyclin B1 and its associated CDK1 (cdc2). Meanwhile, we found that 12k induced A549 cell apoptosis, which was closely associated with the effect of the Bcl-2 family. Increase of Bad, Bak and Bax expression levels, decrease of Bcl-2 and Mcl-1 expression levels were observed. SiRNA knockdown of c-myc in A549 cells significantly attenuated tumor inhibition effects of 12k. In conclusion, our results demonstrate that 12k has an inhibitory effect on growth of A549 cell by inducing cell cycle arrest and apoptosis.

Antitumor activity of cobrotoxin in human lung adenocarcinoma A549 cells and following transplantation in nude mice.

The aim of the present study was to investigate cobra neurotoxin (cobrotoxin) activity in A549 cell lines transplanted into nude mice, and to explore its molecular mechanism. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method was used to detect the growth inhibition rate of cobrotoxin in human lung A549 adenocarcinoma cells and HFL1 lung fibroblasts. Cell colony formation assays were performed to determine the effect of cobrotoxin on A549 cell colony formation, and transmission electron microscopy was used to detect cobrotoxin autophagy. In addition, western blot analysis was performed to determine the effect of 3-methyl adenine (3-MA) activity on the inhibition of autophagy, SB203580 inhibition of the p38-mitogen-activated protein kinase (MAPK) pathway, and Beclin 1, LC3, p62, p38 and phosphorylated (p)-p38 protein expression. Nude mice were injected with human lung A549 cells, and intervention and control groups were compared with regard to tumor suppression. The MTT assay revealed that various concentrations of cobrotoxin inhibited growth of A549 cells, but not HFL1 cells. A549 cell colony formation decreased and autophagosome activity was significantly increased compared with the controls. Following 3-MA administration, SB203580 autophagosome activity decreased, and following cobrotoxin administration, Beclin 1, p-p38, and LC3-II protein expression significantly increased, whereas p62 expression significantly decreased. Following 3-MA inhibition of autophagy, Beclin 1, LC3-II and p62 expression increased. Furthermore, following SB203580 inhibition of the p38-MAPK pathway, Beclin 1, p-p38, LC3-II and p62 protein expression increased. Cobrotoxin exhibited inhibitory activity on the human lung cancer A549 cells transplanted into the nude mice, suppressing the tumor growth rate by 43.4% (cobrotoxin 40 μg/kg group). However, following the addition of 3-MA (10 mmol/kg) and SB203580 (5 mg/kg), the suppression of the tumor growth rate decreased significantly. Cobrotoxin inhibits the growth of human lung cancer A549 cells in vitro and A549 cells transplanted into nude mice. Furthermore, the induction of autophagy may be associated with the activation of the p38-MAPK pathway.

MiR-101 sensitizes A549 NSCLC cell line to CDDP by activating caspase 3-dependent apoptosis

MicroRNA-101 (miR-101) is evidently downregulated in several types of cancer, including non-small cell lung cancer (NSCLC), and is crucial in sensitizing cells to chemotherapy drugs. The aim of the present study was to investigate the correlation between miR-101 and chemosensitivity in the A549 NSCLC cell line. Here, we used the human A549 cell line for transfection with an miR-101 overexpressing vector and detected the cytotoxic acticity, proliferation and apoptosis of cis-diaminedichloroplatinum (CDDP) in A549-miR-101 and A549-mock cells. We demonstrated that overexpression of miR-101 sensitized A549 cells to CDDP, one of the most frequently used agents in curing or controlling NSCLC and enhanced CDDP-induced cell death and caspase 3-dependent apoptosis. In addition, miR-101 facilitated the inhibitory role of CDDP in A549 cell colony formation. Overall, the results of the present study demonstrated that miR-101 sensitizes the A549 NSCLC cell line to CDDP via the activation of caspase 3-dependent apoptosis.

Usnic Acid Inhibits Growth and Induces Cell Cycle Arrest and Apoptosis in Human Lung Carcinoma A549 Cells

Usnic acid (UA) is a secondary metabolite abundantly found in lichens. Some studies have shown the anticancer potential of UA; however, its efficacy and associated mechanisms are yet to be fully explored. Herein, we assessed the anticancer potency and associated molecular alterations by UA in human lung carcinoma A549 cells. UA treatment (25–100 μM) for 24 and 48 h decreased total cell number by 39–67% (P < 0.01) and 68–89% (P < 0.001), respectively, and enhanced cell death by up to twofold and eightfold (P < 0.001), respectively. UA (1–10 μM) also significantly (P < 0.001) suppressed colony formation of A549 cells. The cell growth inhibition was associated with cell cycle arrest at G0/ G1 phase. UA decreased the expression of cyclin-dependent kinase (CDK)4, CDK6, and cyclin D1 and increased the expression of CDK inhibitor (CDKI) p21/cip1 protein. While examining the cell death associated molecular changes, we observed that UA induces mitochondrial membrane depolarization and led to more than twofold increase (P < 0.01) in apoptotic cells. The apoptotic effect of UA was accompanied by enhanced poly(ADP-ribose) polymerase cleavage. This study shows that UA inhibits cell growth involving G0/G1 phase cell cycle arrest and induces cell death via mitochondrial membrane depolarization and induction of apoptosis in human lung carcinoma cells.

Identification of small molecule inhibitors of telomerase activity through transcriptional regulation of hTERT and calcium induction pathway in human lung adenocarcinoma A549 cells

High telomerase activity (TA) is detected in most cancer cells; and thus, TA inhibition by drug or dietary food components is a new strategy for cancer prevention. In this report, we examined the effects of fourteen natural or synthetic compounds on TA in human lung adenocarcinoma A549 cells. The results demonstrated that some of the tested compounds inhibited TA, being 2′-hydroxy-2,3,4′,6′-tetramethoxychalcone (HTMC) was the most potent among tested. In A549 cells, HTMC also inhibited the cell proliferation, decreased the expression of human telomerase reverse transcriptase (hTERT) and sequentially reduced the hTERT promoter. In soft agar assay HTMC treatment reduced the colony formation of A549 cells. Cellular fractionation and immunofluorescence assay demonstrated that there was no translocation of hTERT from nuclei to cytoplasm. Further studies revealed that the release of Ca 2+ was the underlying mechanism of suppressed TA and hTERT transcription in A549 cells exposed to HTMC. These in vitro data support the development of HTMC as a therapeutic agent for cancer complications.

The Lethal Effects of Nicotinamide Combined With Nonfocused Ultrasound on Human Pulmonary Carcinoma Cell Line A549 in Suspension

Objective: To explore the lethal effects of nicotinamide combined with nonfocused ultrasound (NFU) on human pulmonary carcinoma A549 cell live in suspension and provide new evidence for killing the cancer cells in hydrothorax and hydroperitonia or malignant cells infiltrating in tissues. Patients and Methods: Six groups were set in our experiment: blank control group, NA group, NFU group in 0 C, NFU group in 37 C, NA-combined NFU group in 0 C, and NA-combined NFU group in 37 C. Instant and 24-hour survival rates of A549 cells treated by different methods were determined based on the trypan blue exclusive assay, growth rates were detected by MTT after 24 hours, capability of cell colony formation was detected by clone formation test, changes in construction and form of Climbing film cells could be observed under the light microscope after 24 hours, and the percentage changes of apoptosis and the distributional percentage of the cell cycle were analyzed with flow cytometry. Results: Instant and 24-hour survival rates, growth rates, and capability of cell colony formation of A549 cells decreased more obviously in NA-combined NFU group than those in the other four groups. On the contribution of NA-combined NFU, the number of karyokinesis decreased significantly, while karyopyknosis and nuclear fragmentation increased greatly. Flow cytometry showed that the apoptotic index in NA-combined NFU group was higher than that in the NA or NFU group alone, and the cell cycle was held up in G1 stage. Conclusion: Certain concentration of NA has enhancement property on NFU and can increase the irradiated effectiveness of focused ultrasound markedly, which can provide reliable evidence for clinical application of NA as an ultrasound sensitizer.