The tumor suppressor gene RBM5 inhibits lung adenocarcinoma cell growth and induces apoptosis

ABSTRACT: BACKGROUND: The loss of tumor suppressor gene (TSG) function is a critical step in the pathogenesis of human lung cancer. RBM5 (RNA-binding motif protein 5, also named H37/LUCA-15) gene from chromosome 3p21.3 demonstrated tumor suppressor activity. However, the role of RBM5 played in the occurrence and development of lung cancer is still not well understood. METHOD: Paired non-tumor and tumor tissues were obtained from 30 adenocarcinomas. The expression of RBM5 mRNA and protein was examined by RT-PCR and Western blot. A549 cell line was used to determine the apoptotic function of RBM5 in vitro. A549 cells were transiently transfected with pcDNA3.1-RBM5. AnnexinV analysis was performed by flow cytometry. Expression of Bcl-2, cleaved caspase-3, caspase-9 and PAPP proteins in A549 lung cancer cells and the A549 xenograft BALB/c nude mice model was determined by Western blot. Tumor suppressor activity of RBM5 was also examined in the A549 xenograft model treated with pcDNA3.1-RBM5 plasmid carried by attenuated Salmonella typhi Ty21a. Result The expression of RBM5 mRNA and protein was decreased significantly in adenocarcinoma tissues compared to that in the non-tumor tissues. In addition, as compared to the vector control, a significant growth inhibition of A549 lung cancer cells was observed when transfected with pcDNA3.1-RBM5 as determined by cell proliferation assay. We also found that overexpression of RBM5 induced both early and late apoptosis in A549 cells using AnnexinV/PI staining as determined by flow cytometry. Furthermore, the expression of Bcl-2 protein was decreased, whereas the expression of cleaved caspase-3, caspase-9 and PARP proteins was significantly increased in the RBM5 transfected cells; similarly, expression of decreased Bcl-2 and increased cleaved caspase-3 proteins was also examined in the A549 xenograft model. More importantly, we showed that accumulative and stable overexpression of RBM5 in the A549 xenograft BALB/c nude mice model significantly inhibited the tumor growth rate in vivo as compared to that in the control. CONCLUSION: Our study demonstrates that RBM5 can inhibit the growth of lung cancer cells and induce apoptosis both in vitro and in vivo, which suggests that RBM5 might be used as a potential biomarker or target for lung cancer diagnosis and chemotherapy. Moreover, we propose a novel animal model set up in BALB/c nude mice treated with attenuated Salmonella as a vector carrying plasmids to determine RBM5 function in vivo.     

Heat shock protein B8 promotes proliferation and migration in lung adenocarcinoma A549 cells by maintaining mitochondrial function

Heat shock protein B8 (HSPB8) impacts on tumor proliferation and migration of malignancy. However, the role of HSPB8 in lung adenocarcinoma (LUAC) remains unclear. The aim of this study, therefore, was to clarify whether HSPB8 could bring benefits to proliferation and migration of LUAC and its underlying mechanisms. The expression of HSPB8 was first evaluated by immunohistochemistry in 35 LUAC samples. Then, A549 lung adenocarcinoma cells were transfected with pcDNA-HSPB8 or si-HSPB8 to induce HSPB8 overexpression and silence. Cellular activity was evaluated with a Cell Counting Kit-8 (CCK-8) assay. Cell proliferation and migration were observed by EdU assay and scratch assay. Mitochondria-specific reactive oxygen species (mtROS) and membrane potential were measured using MitoSOX Red probe and JC-1 staining. Superoxide dismutase (SOD) activities and malondialdehyde (MDA) level were measured using commercial kits, respectively. HSPB8 protein, mitochondrial fusion protein MFN2 and mitochondrial fission protein p-Drp1/Drp1 were measured using western blot. Compared with the normal tissues, the expression of HSPB8 protein was higher in LUAC tissues and upregulation of HSPB8 protein was related to tumor size and tumor location. Furthermore, HSPB8 overexpression aggravated cell proliferation and migration of A549 cells. Mechanistically, HSPB8 suppressed mitochondrial impairment, leading to promoting the progress of A549 lung adenocarcinoma cells. These data demonstrate that HSPB8 plays an important role in progression of LUAC and may be a new target to treat LUAC.

     

TRB3 interacts with ERK and JNK and contributes to the proliferation,apoptosis, and migration of lung adenocarcinoma cells

Tribbles homolog 3 (TRB3) has been accounted for regulation of a few cell processes through interaction with other significant proteins. The molecular mechanisms underlying TRB3 in tumorigenesis in lung adenocarcinoma have not been entirely elucidated. The present study is aimed at determining the function and fundamental mechanisms of TRB3 in lung adenocarcinoma progression. TRB3 was highly expressed in A549 and H1299 cells and lung adenocarcinoma tissues compared with human bronchial epithelial cells (HBEpC) and adjacent normal lung tissues. Hypoxia significantly upregulated the expression of TRB3 protein in A549 and H1299 cells in a time-dependent way. Gene expression profiling interactive analysis data analysis indicated that patients with lung adenocarcinoma with excessive expression of TRB3 mRNA had fundamentally shorter survival time. TRB3 knockdown in A549 cells can inhibit cell proliferation and migration, and promote cell apoptosis. TRB3 knockdown reduced the expression of p-ERK and p-JNK, but did not affect the expression of p-P38 MAPK. TRB3 overexpression enhances the malignant transformation abilities of HBEpC such as cell proliferation, migration and colony formation, which could be reversed by U0126 and SP600125. TRB3 overexpression promotes the phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) but was not affected by U0126 and SP600125. The results of coimmunoprecipitation experiments indicated that TRB3 binds directly to ERK and JNK. This study suggests that TRB3 has a potentially carcinogenic role in lung adenocarcinoma by binding to ERK and JNK and promoting the phosphorylation of ERK and JNK. TRB3 can be a possible therapeutic focus for lung adenocarcinoma.     

ALKBH5-m6A-FOXM1 signaling axis promotes proliferation and invasion of lung adenocarcinoma cells under intermittent hypoxia

Obstructive sleep apnea (OSA) is closely associated with cancer progression and cancer-related mortality. N6-methyladenosine (m⁶A) is involved in the process of intermittent hypoxia (IH) promoting tumor progression. However, it is unclear how m⁶A regulates the development of lung adenocarcinoma under IH. In this study, we found that ALKBH5 was elevated in lung adenocarcinoma cells and subcutaneous tumors in mice under IH, which was associated with decreased m⁶A levels in these cells and tissues. Next, we knocked out ALKBH5 in a human lung adenocarcinoma cell line under IH, and we found that the proliferation and invasion of these cells were significantly inhibited. Mechanistic analysis showed that under IH, knockout of ALKBH5 in lung adenocarcinoma cells upregulated the level of m⁶A in Forkhead box M1 (FOXM1) mRNA and decreased the translation efficiency of FOXM1 mRNA, resulting in downregulation of the FOXM1 protein. The FOXM1 protein is elevated in lung adenocarcinoma cells and subcutaneous tumor tissues of mice under IH. By knocking out FOXM1 in lung adenocarcinoma cells under IH, proliferation and invasion of these cells were inhibited, and overexpression of FOXM1 partially restored the inhibition of growth and invasion of lung adenocarcinoma cells due to ALKBH5 knockout. Collectively, our findings demonstrate that the m⁶A demethylase ALKBH5 affects the proliferation and invasion of lung adenocarcinoma cells under IH by downregulating m⁶A modification on FOXM1 mRNA and by promoting FOXM1 expression.     

Smac调控Caspase-3对耐药肺腺癌细胞株生物活性及相关信号的研究

摘要 目的：探讨Smac基因调控Caspase-3表达对紫杉醇耐药肺腺癌细胞株生物活性及经典凋亡信号通路的作用机制。方法：取构建好的耐药A549细胞,将其分为A549细胞（LC）组、A549细胞+Smac-NC（SN）组、A549细胞+Smac抑制剂（SI）组、A549细胞+Smac激动剂（SM）组、A549细胞+Caspase-3-NC（CN）组、A549细胞+Caspase-3抑制剂（CI）组、A549细胞+Caspase-3激动剂（CM）组、A549细胞+Smac激动剂+Caspase-3激动剂（MM）组;Real-time PCR法检测正常肺上皮细胞及4种肺腺癌细胞系中Smac、Caspase-3表达水平,将阴性对照、Smac、Caspase-3类似物转染至紫杉醇耐药肺腺癌细胞株,MTT法检测细胞增殖,流式细胞仪检测细胞凋亡,免疫印迹法检测经典凋亡信号通路表达,并分析Smac与Caspase-3的相关性。结果：肺腺癌细胞系中的Smac、Caspase-3 mRNA表达量显著低于正常肺上皮细胞系BEAS-2B（P＜0.05）,其中A549的Smac、Caspase-3 mRNA值最小（P＜0.05）,因此选取其作为此次实验细胞;LC组与SN组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与SN组相比,SI组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显降低（P＜0.05）,增殖率、Bcl-2表达明显升高（P＜0.05）,与SI组相比,SM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;LC组与CN组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与CN组相比,CI组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显降低（P＜0.05）,增殖率、Bcl-2表达明显升高（P＜0.05）,与CI组相比,CM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;SM组与CM组相比,细胞增殖率、凋亡率及Caspase-3、Bcl-2、Bax、Cyto-C蛋白表达基本无差异（P＞0.05）,与CM组相比,MM组细胞凋亡率及Caspase-3、Bax、Cyto-C蛋白表达明显升高（P＜0.05）,增殖率、Bcl-2表达明显降低（P＜0.05）;Smac与Caspase-3呈现正相关（r=0.470,P=0.002）,组间具有显著差异。结论：Smac基因可显著改善紫杉醇耐药肺腺癌细胞株细胞生物活性,并激活经典凋亡信号通路,其作用机制可能与调控Caspase-3表达有关。     

The high expression of miR-31 in lung adenocarcinoma inhibits the malignancy of lung adenocarcinoma tumor stem cells

Therapies for lung adenocarcinoma (LUAD) are mainly limited by drug resistance, metastasis or recurrence related to cancer stem cells (CSCs) with high proliferation and self-renewing. This research validated that miR-31 was over-expressed in LUAD by the analysis of generous clinical samples data. And the results of clinical data analysis showed that high expression of miR-31 was more common in patients with worse prognosis. The genes differentially expressed in LUAD tissues compared with normal tissues and A549CD133⁺ cells (LUAD CSCs) compared with A549 cells were separately screened from Gene Expression Profiling Interactive Analysis and GEO datasets. The target genes that may play a role in the regulation of lung adenocarcinoma was screened by comparison between the differential genes and the target genes of miR-31. The functional enrichment analysis of GO Biological Processes showed that the expression of target genes related to cell proliferation was increased, while the expression of target genes related to cell invasion and metastasis was decreased in LUAD tissues and A549CD133⁺ cells. The results suggested that miR-31 may have a significant inhibitory effect on the differentiation, invasion, metastasis and adhesion of LUAD CSCs, which was verified in vivo and in vitro experiments. Knock down of miR-31 accelerated xenograft tumor growth and liver metastasis in vivo. Likewise, the carcinogenicity, invasion and metastasis of A549CD133⁺ CSCs were promoted after miR-31 knockdown. The study validated that miR-31 was up regulated in LUAD and its expression may affect the survival time of patients with lung adenocarcinoma, which indicated that miR-31 may have potential value for diagnosis and prognosis of LUAD. However, the inhibitory effect of miR-31 on tumorigenesis, invasion and metastasis of lung adenocarcinoma CSCs suggested its complexity in the regulation of lung adenocarcinoma, which may be related to its extensive regulation of various target genes.     

Overexpression of geranylgeranyl diphosphate synthase contributes to tumour metastasis and correlates with poor prognosis of lung adenocarcinoma

This study aimed to evaluate the biological role of geranylgeranyl diphosphate synthase (GGPPS) in the progression of lung adenocarcinoma. GGPPS expression was detected in lung adenocarcinoma tissues by qRT‐PCR, tissue microarray (TMA) and western blotting. The relationships between GGPPS expression and the clinicopathological characteristics and prognosis of lung adenocarcinoma patients were assessed. GGPPS was down‐regulated in SPCA‐1, PC9 and A549 cells using siRNA and up‐regulated in A549 cells using an adenoviral vector. The biological roles of GGPPS in cell proliferation, apoptosis, migration and invasion were determined by MTT and colony formation assays, flow cytometry, and transwell and wound‐healing assays, respectively. In addition, the regulatory roles of GGPPS on the expression of several epithelial‐mesenchymal transition (EMT) markers were determined. Furthermore, the Rac1/Cdc42 prenylation was detected after knockdown of GGPPS in SPCA‐1 and PC9 cells. GGPPS expression was significantly increased in lung adenocarcinoma tissues compared to that in adjacent normal tissues. Overexpression of GGPPS was correlated with large tumours, high TNM stage, lymph node metastasis and poor prognosis in patients. Knockdown of GGPPS inhibited the migration and invasion of lung adenocarcinoma cells, but did not affect cell proliferation and apoptosis. Meanwhile, GGPPS inhibition significantly increased the expression of E‐cadherin and reduced the expression of N‐cadherin and vimentin in lung adenocarcinoma cells. In addition, the Rac1/Cdc42 geranylgeranylation was reduced by GGPPS knockdown. Overexpression of GGPPS correlates with poor prognosis of lung adenocarcinoma and contributes to metastasis through regulating EMT.     

Knockdown of LncRNAZFAS1 suppresses cell proliferation and metastasis in non-small cell lung cancer

ABSTRACT

To evaluate the effects of LncRNAZFAS1 on cell proliferation and tumor metastasis in non-small cell lung cancer (NSCLC), we detected the expression level of LncRNAZFAS1 in NSCLC-related tissues and cells. qRT-PCR results revealed that LncRNAZFAS1 in tumor tissues was significantly higher than that in normal lung tissue, especially significantly up-regulated in stage III / IV and in metastatic NSCLC tissues. LncRNAZFAS1 expression was dramatically up-regulated in 4 NSCLC-related cells (A549, SPC-A1, SK-MES-1, and NCI-H1299), with having the highest expression level in A549 cells. Furthermore, we implemented a knockdown of LncRNAZFAS1 in A549 cells, and the results of CCK8 and Transwell assays suggested that knockdown of LncRNAZFAS1 significantly inhibited NSCLC cell proliferation and metastasis. Next, we constructed a tumor xenograft model to evaluate the effect of LncRNAZFAS1 on the NSCLC cell proliferation in vivo. The results indicated that knockdown of LncRNAZFAS1 dramatically inhibited A549 cells proliferation and repressed tumor growth. Additionally, knockdown of LncRNAZFAS1 drastically weakened the expressions of MMP2, MMP9 and Bcl-2 proteins, whereas noticeably strengthened the expression of BAX protein. Our results altogether suggest that knockdown of LncRNAZFAS1 has a negative effect on the proliferation and metastasis of NSCLC cell, which implying LncRNAZFAS1 is a potential unfavorable biomarker in patients with NSCLC.     

Choline Transporters in Human Lung Adenocarcinoma： Expression and Functional Implications

Choline is an essential nutrient for cell survival and proliferation, however, the expression and function of choline transporters have not been well identified in cancer. In this study, we detected the mRNA and protein expression of organic cation transporter OCT3, carnitine/cation transporters OCTN 1 and OCTN2, and choline transporter-like protein CTL1 in human lung adenocarcinoma cell lines A549, H 1299 and SPC-A-1. Their expression pattern was further confirmed in 25 human primary adenocarcinoma tissues. The choline uptake in these cell lines was significantly blocked by CTL1 inhibitor, but only partially inhibited by OCT or OCTN inhibitors. The efficacy of these inhibitors on cell proliferation is closely correlated with their abilities to block choline transport. Under the native expression of these transporters, the total choline uptake was notably blocked by specific PI3K/AKT inhibitors. These results describe the expression of choline transporters and their relevant function in cell proliferation of human lung adenocarcinoma, thus providing a potential ＂choline-starvation＂ strategy of cancer interference through targeting choline transporters, especially CTL1.     

Protein arginine methyltransferase 5 is essential for growth of lung cancer cells

PRMT5 (protein arginine methyltransferase 5) is an enzyme that catalyses transfer of methyl groups from S-adenosyl methionine to the arginine residues of histones or non-histone proteins and is involved in a variety of cellular processes. Although it is highly expressed in some tumours, its direct role in cancer growth has not been fully investigated. In the present study, in human lung tissue samples we found that PRMT5 was highly expressed in lung cancer cells, whereas its expression was not detectable in benign lung tissues. Silencing PRMT5 expression strongly inhibited proliferation of lung adenocarcinoma A549 cells in tissue culture, and silencing PRMT5 expression in A549 cells also abolished growth of lung A549 xenografts in mice. In vitro and in vivo studies showed that the cell growth arrest induced by loss of PRMT5 expression was partially attributable to down-regulation of fibroblast growth factor receptor signalling. These results suggest that PRMT5 and its methyltransferase activity is essential for proliferation of lung cancer cells and may serve as a novel target for the treatment of lung cancer.     

Endotoxin induces proliferation of NSCLC in vitro and in vivo: role of COX-2 and EGFR activation

Lung cancer is frequently complicated by pulmonary infections which may impair prognosis of this disease. Therefore, we investigated the effect of bacterial lipopolysaccharides (LPS) on tumor proliferation in vitro in the non-small cell lung cancer (NSCLC) cell line A549, ex vivo in a tissue culture model using human NSCLC specimens and in vivo in the A549 adenocarcinoma mouse model. LPS induced a time- and dose-dependent increase in proliferation of A549 cells as quantified by MTS activity and cell counting. In parallel, an increased expression of the proliferation marker Ki-67 and cyclooxygenase (COX)-2 was detected both in A549 cells and in ex vivo human NSCLC tissue. Large amounts of COX-2-derived prostaglandin (PG)E₂ were secreted from LPS-stimulated A549 cells. Pharmacological interventions revealed that the proliferative effect of LPS was dependent on CD14 and Toll-like receptor (TLR)4. Moreover, blocking of the epidermal growth factor receptor (EGFR) also decreased LPS-induced proliferation of A549 cells. Inhibition of COX-2 activity in A549 cells severely attenuated both PGE₂ release and proliferation in response to LPS. Synthesis of PGE₂ was also reduced by inhibiting CD14, TLR4 and EGFR in A549 cells. The proliferative effect of LPS on A549 cells could be reproduced in the A549 adenocarcinoma mouse model with enhancement of tumor growth and Ki-67 expression in implanted tumors. In summary, LPS induces proliferation of NSCLC cells in vitro, ex vivo in human NSCLC specimen and in vivo in a mouse model of NSCLC. Pulmonary infection may thus directly induce tumor progression in NSCLC.     

Id 基因在多种肺癌细胞中的表达及意义

目的：研究Id基因在肺癌和永生化支气管上皮细胞中的表达,探讨其在肺癌细胞中表达的意义。方法：利用半定量RT—PCR和Western blot方法检测多种肺癌和永生化支气管上皮细胞中Id1—Id4 mRNA和Id1—Id4蛋白的表达。结果：A549、NCI—H460、NCI—H446、SK—MES—1、Anip973中Id1-Id3 mRNA均高表达,Id1相对表达较强;而AGZY和MP-184中未表达Id1-Id3 mRNA;腺癌细胞均表达了Id4 mRNA,而NCI-H446、SK—MES—1未表达Id4mRNA。A549,NCA—H460,NCA—H446,SK—MES—1,Anip973中Id1,Id2,Id3蛋白均高表达,A549,NCA—H446,Anip973中Id2的表达高于NCA—H460,SK—MES—1;A549,NCA—H460,Anip973有出的高表达,NCI—H446,SK—MES—1无Id4的表达,Id1-Id4在AGZY和MP-184中均耒表达。结论：4种Id基因均作为癌基因在肺癌的发生发展中发挥作用,Id1,Id2,Id3与肺癌细胞的恶性程度以及增殖和转移密切相关,Id4可做为肺腺癌的检测标志物。     

Akt2 and nucleophosmin/B23 function as an oncogenic unit in human lung cancer cells

The signaling network of protein kinase B(PKB)/Akt has been implicated in survival of lung cancer cells. However, understanding the relative contribution of the different isoform of Akt network is nontrival. Here, we report that Akt2 is highly expressed in human lung adenocarcinoma cell line A549 cells. Suppression of Akt2 expression in A549 cells results in notable inhibition of cell poliferation, soft agar growth, and invasion, accompanying by a decrease of nucleophosmin/B23 protein. Overexpression of Akt1 restores cancerous growth of A549 cells in B23-knockdown (KD) cells while Akt2 overexpression did not restore proliferating potential in cells with downregulated B23, thus suggesting Akt2 requires B23 to drive proliferation of lung cancer cell. Loss of functional Akt2 and B23 has similar defects on cell proliferation, apoptotic resistance and cell cycle regulation, while loss of Akt1 has less defects on cell proliferation, survial and cell cycle progression in A549 cells. Moreover, overexpression of B23 rescues the proliferative block induced as a consequence of loss of Akt2. Thus our data suggest that Akt2/B23 functions as an oncogenic unit to drive tumorigenesis of A549 lung cancer cells.     

Ku80 is differentially expressed in human lung carcinomas and upregulated in response to irradiation in mice

Based on the role of Ku80 in mediating radiation-induced DNA repair, we investigated Ku80 expression in human lung cancers of different pathological types and evaluated the effect of radiotherapy on Ku80 expression levels in a mouse model. We used immunohistochemistry and real-time PCR to determine Ku80 protein and mRNA levels, respectively. We inoculated nude mice with A549 cells and subjected the tumor-bearing mice to varying doses of irradiation. Lung carcinoma tissue exhibited higher Ku80 mRNA and protein levels when compared with normal tissue. Among the tumor subtypes, lung adenocarcinoma and lung squamous carcinoma showed higher levels of Ku80 protein and mRNA, compared with small-cell lung carcinoma. There was a dose-dependent and time-dependent increase in Ku80 mRNA levels in nude mice that were inoculated with A549 cells and exposed to varying doses of irradiation. Ku80 may play an important role in the DNA damage response pathway. Higher Ku80 levels in lung squamous carcinoma and adenocarcinoma may explain their lower radiosensitivity when compared with small-cell lung carcinoma. Ku80 expression levels could be useful in predicting radiosensitivity of lung tumors and inhibition of Ku80 may be an interesting target to improve radiosensitivity in lung cancer patients.     

Nuclear distribution of claudin-2 increases cell proliferation in human lung adenocarcinoma cells

Claudin-2 is expressed in human lung adenocarcinoma tissue and cell lines, although it is absent in normal lung tissue. However, the role of claudin-2 in cell proliferation and the regulatory mechanism of intracellular distribution remain undefined. Proliferation of human adenocarcinoma A549 cells was decreased by claudin-2 knockdown together with a decrease in the percentage of S phase cells. This knockdown decreased the expression levels of ZONAB and cell cycle regulators. Claudin-2 was distributed in the nucleus in human adenocarcinoma tissues and proliferating A549 cells. The nuclear distribution of ZONAB and percentage of S phase cells were higher in cells exogenously expressing claudin-2 with a nuclear localization signal than in cells expressing claudin-2 with a nuclear export signal. Nuclear claudin-2 formed a complex with ZO-1, ZONAB, and cyclin D1. Nuclear distribution of S208A mutant, a dephosphorylated form of claudin-2, was higher than that of wild type. We suggest that nuclear distribution of claudin-2 is up-regulated by dephosphorylation and claudin-2 serves to retain ZONAB and cyclin D1 in the nucleus, resulting in the enhancement of cell proliferation in lung adenocarcinoma cells.     

MiR‐376a suppresses the proliferation and invasion of non‐small‐cell lung cancer by targeting c‐Myc

It has been reported that miR‐376a is involved in the formation and progression of several types of cancer. However, the expression and function of miR‐376a is still unknown in non‐small cell lung carcinomas (NSCLC). In this study, the expression of miR‐376a in NSCLC tissues and cell lines were examined by real‐time PCR, the effects of miR‐376a on cell proliferation, apoptosis and invasion were evaluated in vitro. Luciferase reporter assay was performed to identify the targets of miR‐376a. The results showed that miR‐376a was significantly downregulated in NSCLC tissues and cell lines. Restoration of miR‐376a in NSCLC cell line A549 significantly inhibited cell proliferation, increased cell apoptosis and suppressed cell invasion, compared with control‐transfected A549 cells. Luciferase reporter assay showed that c‐Myc, an oncogene that regulating cell survival, angiogenesis and metastasis, was a direct target of miR‐376a. Over‐expression of miR‐376a decreased the mRNA and protein levels of c‐Myc in A549 cells. In addition, upregulation of c‐Myc inhibited miR‐376a‐induced inhibition of cell proliferation and invasion in A549 cells. Therefore, our results indicate a tumor suppressor role of miR‐376a in NSCLC by targeting c‐Myc. miR‐376a may be a promising therapeutic target for NSCLC.     

Critical roles of mucin-1 in sensitivity of lung cancer cells to tumor necrosis factor-alpha and dexamethasone

Lung cancer is the leading cause of death from cancer. Mucins are glycoproteins with high molecular weight, responsible for cell growth, differentiation, and signaling, and were proposed to be correlated with gene heterogeneity of lung cancer. Here, we report aberrant expression of mucin genes and tumor necrosis factor receptors in lung adenocarcinoma tissues compared with normal tissues in GEO datasets. Mucin-1 (MUC1) gene was selected and considered as the target gene; furthermore, the expression pattern of adenocarcinomic cells (A549, H1650, or H1299 cells) was validated under the stimulation with tumor necrosis factor-alpha (TNFα) or dexamethasone (DEX), separately. MUC1 gene interference was done to A549 cells to show its role in sensitivity of lung cancer cells to TNFα and DEX. Results of our experiments indicate that MUC1 may regulate the influence of inflammatory mediators in effects of glucocorticoids (GCs), as a regulatory target to improve therapeutics. It shows the potential effect of MUC1 and GCs in lung adenocarcinoma (LADC), which may help in LADC treatment in the future.