Qiaoyuan Yang

MicroRNA-622 functions as a tumor suppressor by targeting K-Ras and enhancing the anticarcinogenic effect of resveratrol

Aberrant expression of microRNA (miRNA) has been previously demonstrated to play an important role in a wide range of cancer types and further elucidation of its role in the mechanisms underlying tumorigenesis, anticarcinogenesis and potential chemotherapeutics is warranted. We chose the anti-benzo[a]pyrene-7,8-diol-9,10-epoxide-transformed human bronchial epithelial cell line 16HBE-T to study miRNAs involved in anticarcinogenesis. In resveratrol-treated cells, we found that miR-622 was upregulated, whereas it was downregulated in 16HBE-T cells, suggesting that miR-622 potentially acts as a tumor suppressor. Increasing the level of miR-622 by transient transfection-induced inhibition of proliferation and G(0) arrest in 16HBE-T cells and the lung cancer cell line H460 as demonstrated by cell viability and cell cycle analysis. MiR-622 dramatically suppressed the ability of 16HBE-T cells to form colonies in vitro and to develop tumors in nude mice. According to bioinformatics analysis, K-Ras messenger RNA was predicted as a putative miR-622 target; this was confirmed by western blot and luciferase reporter assays. Cell growth retardation was inhibited upon knockdown of K-Ras and an increase in the level of miR-622 in 16HBE-T cells. Furthermore, miR-622 inhibitor partially impaired the growth of 16HBE-T cells as demonstrated by luciferase reporter activity and K-Ras protein expression in 16HBE-T cells. In summary, miR-622 functions as a tumor suppressor by targeting K-Ras and impacting the anticancer effect of resveratrol. Therefore, miR-622 is potentially useful as a clinical therapy. MiR-622 impacts the K-Ras signal pathway and the potentially anticarcinogenic or chemotherapeutic properties warrant further investigation.

Genetic variations in miR-27a gene decrease mature miR-27a level and reduce gastric cancer susceptibility

MicroRNAs (miRNAs) are noncoding RNAs that function as post-transcriptional regulators of tumor oncogenes and suppressors. Single-nucleotide polymorphisms (SNPs) in miRNA genes are a novel class of genetic variations in the human genome that are currently being identified and investigated in human cancers. In this study, we aimed to investigate whether SNPs in the miR-27a gene affect miR-27a expression and alter susceptibility to gastric cancer. Therefore, we conducted a case–control population study and the allele and genotype frequencies for polymorphism rs11671784 in miR-27a gene were examined in the study population. As a result, we found that the G/A polymorphism in the miR-27a gene exhibited a significant effect on gastric cancer risk. Compared with GG homozygotes, subjects who were GA heterozygotes or AA homozygotes exhibited a decreased risk of gastric cancer. The G/A polymorphism impaired the processing of pre-miR-27a to mature miR-27a, resulting in significantly reduced expression of mature miR-27a and an increased level of its target HOXA10. Furthermore, we confirmed these findings in in vitro studies by overexpressing pre-miR-27a carrying G or A allele. It provided further evidence demonstrating that allelic difference of rs11671784 is linked to gastric tumorigenesis. In summary, our results indicate that the G/A polymorphism in miR-27a gene (rs11671784) decreases miR-27a expression, reduces gastric cancer risk and plays a role in gastric tumorigenesis. This is the first study to address the role and function of miR-27a polymorphism rs11671784 in gastric cancer. These results could be useful to assess individual susceptibility of gastric cancer and will improve our understanding of the potential contribution of miRNA SNPs to cancer pathogenesis.

Alteration of serum miR-206 and miR-133b is associated with lung carcinogenesis induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone.

The alteration of microRNA (miRNA) expression plays an important role in chemical carcinogenesis. Presently, few reports have been published that concern the significance of circulating miRNAs in lung carcinogenesis induced by environmental carcinogens. The purpose of this study was to identify serum miRNAs that could be associated with lung carcinogenesis induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). Male F344 rats were systemically administered with NNK. The rat serum differential expression profiles of miRNAs were analyzed by small RNA solexa sequencing. Using quantitative real-time PCR, the differentially expressed serum miRNAs were identified in each individual rat. Serum miR-206 and miR-133b were selected for further identification in rat serum at different stages of lung carcinogenesis; we detected the levels of serum miR-206 and miR-133b in lung cancer tissues induced by NNK. NNK causes significant alteration of serum miRNA expression. Compared to the control group, serum miR-206 and miR-133b were significantly up-regulated in the early stage of NNK-induced lung carcinogenesis. miR-206 and miR-133b exhibited low-expression in lung cancer tissues. Our results demonstrate that lung carcinogen NNK exposure changes the expression of serum miRNAs. Serum miR-206 and miR-133b could be associated with lung carcinogenesis induced by NNK.

A novel long noncoding RNA AK001796 acts as an oncogene and is involved in cell growth inhibition by resveratrol in lung cancer.

Lung cancer is the most common form of cancer throughout the world. The specific targeting of long noncoding RNAs (lncRNAs) by resveratrol opened a new avenue for cancer chemoprevention. In this study, we found that 21 lncRNAs were upregulated and 19 lncRNAs were downregulated in lung cancer A549 cells with 25 μmol/L resveratrol treatment determined by microarray analysis. AK001796, the lncRNA with the most clearly altered expression, was overexpressed in lung cancer tissues and cell lines, but its expression was downregulated in resveratrol-treated lung cancer cells. By monitoring cell proliferation and growth in vitro and tumor growth in vivo, we observed a significant reduction in cell viability in lung cancer cells and a slow growth in the tumorigenesis following AK001796 knockdown. We also found that AK001796 knockdown caused a cell-cycle arrest, with significant increases in the percentage of cells in G0/G1 in lung cancer cells. By using cell cycle pathway-specific PCR arrays, we detected changes in a number of cell cycle-related genes related to lncRNA AK001796 knockdown. We further investigated whether AK001796 participated in the anticancer effect of resveratrol and the results showed that reduced lncRNA AK001796 level potentially impaired the inhibitory effect of resveratrol on cell proliferation. To our knowledge, this is the first study to report the changes in an lncRNA expression profile induced by resveratrol in lung cancer.

LncRNA-DQ786227-mediated cell malignant transformation induced by benzo(a)pyrene.

It has recently been found that the new class of transcripts, long non-coding RNAs (lncRNAs), are pervasively transcribed in the genome. LncRNAs are a large family of non-coding RNAs and regulate many protein-coding genes. Growing evidence indicates that lncRNAs may play an important functional role in cancer biology. Emerging data have shown that lncRNAs are closely related to the occurrence and development of lung cancer. However, the function and mechanism of lncRNAs in lung cancer remain elusive. Here, we investigated the role of a novel lncRNA in transformed human bronchial epithelial cells induced by benzo(a)pyrene. After establishing the transformed cell model using the BEAS-2B cell line in vitro, we found that expression of lncRNA-DQ786227 was high and changed during the transformation of BEAS-2B cells. Silencing of lncRNA-DQ786227 expression in malignant transformed BEAS-2B cells led to inhibition of cell proliferation and colony formation, and increased apoptosis. LncRNA-DQ786227 dramatically promoted the ability of BEAS-2B-T cells to form colonies in vitro and develop tumors in nude mice. These findings revealed that lncRNA-DQ786227 acts as an oncogene in malignantly transformed BEAS-2B cells induced by benzo(a)pyrene. The identification of lncRNA could provide new insight into the molecular mechanisms of chemical carcinogenesis.

Deregulation of Serum MicroRNA Expression Is Associated with Cigarette Smoking and Lung Cancer

Lung cancer is the leading cause of cancer-related death and cigarette smoking is the main risk factor for lung cancer. Circulating microRNAs (miRNAs) are considered potential biomarkers of various cancers, including lung cancer. However, it is unclear whether changes in circulating miRNAs are associated with smoking and smoking-related lung cancer. In this study, we determined the serum miRNA profiles of 10 nonsmokers, 10 smokers, and 10 lung-cancer patients with miRCURY LNA microRNA arrays. The differentially expressed miRNAs were then confirmed in a larger sample. We found that let-7i-3p and miR-154-5p were significantly downregulated in the sera of smokers and lung-cancer patients, so the serum levels of let-7i-3p and miR-154-5p are associated with smoking and smoking-related lung cancer. The areas under receiver operating characteristic curves for let-7i-3p and miR-154-5p were approximately 0.892 and 0.957, respectively. In conclusion, our results indicate that changes in serum miRNAs are associated with cigarette smoking and lung cancer and that let-7i-3p and miR-154-5p are potential biomarkers of smoking-related lung cancer.

Oncogenic role of long noncoding RNA AF118081 in anti-benzo[a]pyrene-trans-7,8-dihydrodiol-9,10-epoxide-transformed 16HBE cells.

Lung cancer is the leading cause of cancer deaths and remains an important public health problem worldwide. Long noncoding RNAs (lncRNAs) are newly identified regulators of tumorigenesis and tumor progression. However, the role of lncRNAs in lung cancer induced by environmental carcinogens remains largely unknown. In this study, an lncRNA microarray was used to compare the expression profiles of malignantly transformed 16HBE cells (16HBE-T) induced with anti-benzo[a]pyrene-trans-7,8-diol-9,10-epoxide (anti-BPDE) and normal 16HBE cells (16HBE-N). Using quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR), lncRNA AF118081 was identified as the most significantly overexpressed lncRNA in 16HBE-T cells, lung cancer cells, and patient samples. Cell proliferation, colony formation, apoptosis, migration, and invasion were assayed in 16HBE-T cells following the knockdown of lncRNA AF118081 with small interfering RNA. AF118081 knockdown inhibited cell growth and tumor invasion. An in vivo (nude mouse) model was then used to assay tumor growth, and the downregulation of AF118081 clearly suppressed tumor growth, consistent with the results of the in vitro assays. Together, these findings identify a new oncogenic lncRNA, lncRNA AF118081, in malignantly transformed 16HBE cells. This enhances our understanding of lncRNAs as important regulatory elements in chemical carcinogenesis and potential targets of lung cancer therapies.

Altered miRNA Expression Profiles and miR-1a Associated With Urethane-Induced Pulmonary Carcinogenesis

MicroRNAs (miRNAs) are small RNA molecules that regulate posttranscriptional gene expression. Previous research has suggested that aberrant miRNA expression often plays a critical role in many types of cancer, including lung cancer. However, the exact miRNAs that are involved in pulmonary carcinogenesis remain unclear. We investigated the miRNA-based molecular changes that occur in urethane-induced carcinogenicity and identified specific miRNA deregulation in pulmonary carcinogenesis induced by urethane. In this study, we used a lung cancer model in which Balb/c mice were exposed to urethane via ip injection once a week for four consecutive weeks. The mice were then killed in weeks 6, 12, or 24. Two small RNA libraries were constructed with the total RNA from the lung tumor and normal adjacent lung tissues of the urethane-injected mice collected in week 24. Using Solexa sequencing, we identified a plethora of differentially expressed miRNAs and predicted nine novel miRNAs. Further analysis demonstrated the sustainable downregulation of miR-1a in the lung tissues in lung carcinogenesis induced by urethane. The levels of miR-1a were also reduced in the serum. Our findings indicate that urethane exposure alters the expression of a cluster of miRNAs. The simultaneous downregulation of miR-1a in lung tissues and serum in urethane-induced pulmonary carcinogenesis suggests that miR-1a is associated with tumorigenesis.

Identification of a long non-coding RNA NR_026689 associated with lung carcinogenesis induced by NNK

Long non-coding RNAs (lncRNA) are thought to be important epigenetic regulators involved in the development of a variety of cancers. Alterations in lncRNA expression are associated with exposure to chemical carcinogens. However, it is still unclear whether lncRNA expression during lung carcinogenesis is induced by chemical carcinogens. In this study, using NNK-induced rat lung cancer model established by our previous study, we determined the lncRNA expression profiles, and an alteration in lncRNA expression was observed in lung cancer tissues and blood in the NNK treatment group. Using quantitative reverse-transcription PCR (qRT-PCR), five differentially expressed lncRNAs were further detected and validated. We identified a novel lncRNA, NR_026689, which showed increased expression in lung cancer tissues induced by NNK and the alteration of lncRNA NR_026689 was specifically observed in lung tissue. The level of NR_026689 was determined and significantly increased in rat whole blood at the 10th and 20th week after NNK treatment to evaluate it as a potential early marker for lung cancer. Together, these findings suggest that lncRNA NR_026689 may be a potential early biomarker for lung cancer and is associated with lung carcinogenesis induced by NNK.

Silencing of lncRNA AFAP1-AS1 suppressed lung cancer development by regulatory mechanism in cis and trans

Although the long noncoding RNA AFAP1-AS1 has been shown to be involved in various types of cancer, its involvement in lung cancer remains poorly understood. In the current study, we found that AFAP1-AS1 was substantially over expressed in lung cancer tissues and cell lines. In addition, AFAP1-AS1 expression level was proven to be associated with the malignant features of lung cancer. Knockdown of AFAP1-AS1 significantly suppressed cell proliferation by increasing cell apoptosis and G0/G1 phase retardation of cell cycle in lung cancer cells. Furthermore, AFAP1-AS1 knockdown could suppress tumor growth of lung cancer in BALB/c nude mice. We also identified that AFAP1-AS1 silencing could influence the expression of AFAP1 and KRT1 on mRNA and protein level by cis and trans regulatory mechanism. Moreover, the oncogenic activities of AFAP1-AS1 on cell proliferation are partially mediated by KRT1. In summary, these findings demonstrate that AFAP1-AS1 plays an essential role in promoting lung cancer development in vitro and vivo. It indicated that AFAP1-AS1 is a promising prognostic predictor for patients with lung cancer.