No abstract

Epithelial-mesenchymal transition is a crucial event for metastasis and could be mediated by several pathways such as phosphoinositide 3-kinase/Akt, mitogen-activated protein kinases, as well as many epigenetic regulators. Special AT-rich sequence-binding protein 2 is an epigenetic regulator involved in epithelial-mesenchymal transition and osteoblastic differentiation. It has been reported that the crosstalk between several pathways is responsible for the regulation of epithelial-mesenchymal transition in cancer cells. However, crosstalks between p38 and Akt pathways involved in epithelial-mesenchymal transition are still unknown. We recently reported that there is a crosstalk between p38 and Akt pathways in non-small-cell lung carcinoma cells, and this crosstalk is associated with E-cadherin and special AT-rich sequence-binding protein 2 expressions. Therefore, we aimed to determine whether this crosstalk has a mediator role in the regulation of epithelial-mesenchymal transition in non-small-cell lung carcinoma. Our results showed that inhibition of p38 leads to the disruption of this crosstalk via decreased expression of phosphatase and tensin homolog (PTEN) and subsequently increased activation of Akt in non-small-cell lung carcinoma cells. Then, we found that p38 inhibition upregulated special AT-rich sequence-binding protein 2 expression and reversed epithelial-mesenchymal transition in non-small-cell lung carcinoma cells. Furthermore, special AT-rich sequence-binding protein 2 knockdown abolished the effect of p38 inhibition on epithelial-mesenchymal transition in non-small-cell lung carcinoma cells. In conclusion, our results strongly indicate that the crosstalk between p38 and Akt pathways can determine special AT-rich sequence-binding protein 2 expression and epithelial character of non-small-cell lung carcinoma cells, and special AT-rich sequence-binding protein 2 is a critical epigenetic regulator for epithelial-mesenchymal transition mediated by p38 pathway in non-small-cell lung carcinoma. Our findings will contribute to illuminate the molecular mechanisms of the epithelial-mesenchymal transition process that has a critical significance for lung cancer metastasis.

Transforming growth factor-β (TGF-β) pathway plays crucial roles during the carcinogenesis and metastasis. TGF-β receptor 2 (TGFBR2) is a key molecule for the regulation of TGF-β pathway and frequently downregulated or lost in several cancer types including non-small cell lung cancer (NSCLC), and TGF-β pathway is often regulated by negative-feedback mechanisms, but little is known about the mechanism of TGFBR2 downregulation in NSCLC. Here, we found that the expression of miR-520e is upregulated in metastatic tumor tissues compared with non-metastatic ones, and its expression is inversely correlated with that of TGFBR2 in clinical samples. We also discovered that TGF-β dramatically increased the expression of miR-520e, which targeted and downregulated TGFBR2, and the suppression of miR-520e significantly impaired TGF-β-induced TGFBR2 downregulation. Chromatin immunoprecipitation–PCR experiments further showed that miR-520e is transcriptionally induced by SMAD2/3 in response to TGF-β. Our findings reveal a novel negative-feedback mechanism in TGF-β signaling and the expression level of miR-520e could be a predictive biomarker for NSCLC metastasis.

Background: Non-Small Cell Lung Cancer (NSCLC) is an aggressive cancer type due to high metastatic capacity. Nuclear Factor Kappa B (NF-κB) is a consistently active transcription factor in malignant lung cancer cells and has crucial significance in NSCLC progression. It is also implicated in the transcriptional regulation of many genes including microRNAs (miRNAs) that function as tumor suppressor or oncogene. It has been increasingly reported that several miRNAs defined as gene members are induced by NF-κB. The present study aimed to find novel miRNAs that are regulated by NF-κB. Methods: Chromatin İmmunoprecipitation Sequencing (ChIP-Seq) experiment and bioinformatic analysis were used to determine NF-κB-dependent miRNAs. Western blot analysis, quantitative real-time polymerase chain reaction (qRT-PCR), luciferase reporter gene assays were carried out to investigate the target genes of miRNAs. To determine biologic activity, transwell invasion and MTT assay were carried out on H1299 NSCLC cell line. miRNA expression level was evaluated in metastatic and non-metastatic tissue samples of NSCLC patients. Results: ChIP-Seq and qRT-PCR experiments showed that miR-548as-3p is transcriptionally regulated by NF- κB in response to Tumor Necrosis Factor-α (TNF-α) treatment. Then, we found that tumor suppressor Phosphatase and Tension homolog (PTEN) is a direct target of miR-548as-3p. Furthermore, miR-548as-3p mediates phosphatidylinositol-3-OH kinase (PI3K)/Akt pathway and NF-κB-implicated genes including Matrix Metalloproteinases 9 (MMP9), Slug and Zeb1. We further showed that miR-548as-3p increased invasiveness of NSCLC cells and was upregulated in metastatic tumor tissues compared to non-metastatic ones. Conclusion: All these findings provide a miRNAs-mediated novel mechanism for NF-κB signaling and that miR-548as-3p could be a biomarker for NSCLC metastasis.

Objective: In this study, it was aimed to investigate that PON2, in hypoxia and normoxia in relation to HIF-1α transcription factor that has a role in tumour angiogenesis. Hypoxia inducible factor 1 alpha (HIF-1α) is a transcription factor which is sensitive to hypoxia, causes initiation of angiogenesis and metastasis by providing transcription of numerous genes. As well as hypoxia several inflammatory agents such as lipopolysaccharide (LPS) can regulate the expression of HIF-1α as well as PON2. Oxidative stress is known to have role in cancer. Paraoxonase 2 which is one of the members of paraoxonase family serves as intracellular anti-oxidant.Methods: H1299, A549 ve PC14 non-small cell lung carcinoma cell lines used in this study. Cells were cultured under hypoxia and normoxia conditions with LPS stimulation. HIF-1α and PON2 mRNA expression levels measured by real-time PCR. Western blot studies were performed for protein expression. Results:In this study, it was observed that LPS treatment stimulates HIF-1α expression which increases PON2 expression in NSCLC cell line in under hxpoxia conditions. Conclusion:This study shows that PON2 is regulated by HIF-1α in hypoxia and inflammation. The relationship between hypoxia and inflammation and oxidative status of cells requires further studies.Key words: HIF-1α, PON2, LPS, Hypoxia ÖZET Amaç: Bu çalışmada tümör anjiyogenezinde önemli rol oynayan hipoksi indüklenebilir faktör 1 (HIF-1) transkripsiyon faktörünün hipoksik ve normoksik koşullarda paraoksonaz 2 (PON2) ile ilişkisinin araştırılması hedeflenmiştir. Hipoksiye duyarlı bir transkripsiyon faktör olan HIF-1α çok sayıda genin transkripsiyonunu sağlayarak anjiyogenez ve metastazın başlamasına yol açar. Hipoksinin yanı sıra lipopolisakkarit (LPS) gibi çeşitli inflamatuvar ajanlar da HIF-1 α'nın ve PON2 nin ekspresyonunu düzenleyebi-lirler. Oksidatif stres karsinogenezde önemli etiyolojik faktörlerden birisidir. Paraoksonaz ailesi üyelerinden paraoksonaz 2 (PON2) nin hücre içi antioksidan olarak görev yaptığı bilinmektedir.Yöntemler: H1299, A549 ve PC14 akciğer hücre dizileri normoksi ve hipoksi ortamlarında kültürü yapılarak ve lipopolisakkaritle indüklenerek HIF-1 ve PON2 mRNA ekspresyonları ve western blot tayinleri çalışılmıştır. Bulgular: Çalışma sonucunda LPS uygulamasının HIF-1α ekspresyonunu uyardığı ve hipoksi koşullarında akciğer hücre dizilerinde PON2 ekspresyonunun arttığı göz-lendi.Sonuç: Bu çalışmada PON 2 enzim ekspresyonunun HIF-1α aracılığıyla hipoksi ve normoksi koşullarında regüle edildiği gözlenmiş olup hipoksi, inflamasyon ve hüc-renin oksidatif durumu arasındaki ilişkinin aydınlatılması için ileri çalışmalar gerekebilir. Anahtar kelimeler: HIF-1α, PON2, LPS, Hipoksi GİRİŞBu çalışmada akciğer kanseri hücre hatları kullanı-larak, tümör anjiyogenezinde önemli bir rol oynayan hipoksi indüklenebilir faktör 1 transkripsiyon faktörünün (HIF-1α) hipoksik ve normoksik koşullarda PON 2 enzimiyle ilişkisinin araştırılması amaçlanmıştır.