Abstract
Claudin-2 is highly expressed in lung adenocarcinoma tissues and increases proliferation in adenocarcinoma cells. The chemicals that reduce claudin-2 expression may have anti-cancer effects, but such therapeutic medicines have not been developed. We found that azacitidine (AZA), a DNA methylation inhibitor, and trichostatin A (TSA) and sodium butyrate (NaB), histone deacetylase (HDAC) inhibitors, decrease claudin-2 levels. The effect of AZA was mediated by the inhibition of phosphorylated Akt and NF-κB. LY-294002, an inhibitor of phosphatidylinositol 3-kinase (PI3K), and BAY 11-7082, an NF-κB inhibitor, decreased claudin-2 levels. The reporter activity of claudin-2 was decreased by AZA and LY-294002, which was blocked by the mutation in a putative NF-κB-binding site. NF-κB bound to the promoter region of claudin-2, which was inhibited by AZA and LY-294002. AZA is suggested to decrease the claudin-2 mRNA level mediated by the inhibition of a PI3K/Akt/NF-κB pathway. TSA and NaB did not change phosphorylated Akt and NF-κB levels. Furthermore, these inhibitors did not change the reporter activity of claudin-2 but decreased the stability of claudin-2 mRNA mediated by the elevation of miR-497 microRNA. The binding of histone H3 to the promoter region of miR-497 was inhibited by TSA and NaB, whereas that of claudin-2 was not. These results suggest that HDAC inhibitors decrease claudin-2 levels mediated by the elevation of miR-497 expression. Cell proliferation was additively decreased by AZA, TSA, and NaB, which was partially rescued by ectopic expression of claudin-2. We suggest that epigenetic inhibitors suppress the abnormal proliferation of lung adenocarcinoma cells highly expressing claudin-2.
Highlights
Claudin-2 is highly expressed in lung adenocarcinoma tissues and increases proliferation in adenocarcinoma cells
We recently reported that the inhibition of mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK)/c-Fos and phosphatidylinositol-3 kinase (PI3K)/Akt pathways decreases claudin-2 expression in A549 cells [18]
Effect of AZA on the Expression of Junctional Protein in A549 Cells—We examined the effect of AZA on the expression of junctional proteins in A549 cells
Summary
Effect of AZA on the Expression of Junctional Protein in A549 Cells—We examined the effect of AZA on the expression of junctional proteins in A549 cells. The primer pairs amplifying NF-B binding and the upstream sites showed PCR bands using input samples These results indicate that NF-B may have a key role in the decrease in claudin-2 expression by AZA. In the same range of concentration, both TSA and NaB increased the acetylation levels of histone H3 (Fig. 6) These results indicate that the expression of claudin-2 may be down-regulated by histone acetylation in A549 cells. In the ChIP assay, the binding of c-Fos and NF-B on the promoter region of claudin-2 was not inhibited by TSA and NaB (Fig. 8C) These results indicate that histone acetylation may not be directly involved in the transcriptional regulation of claudin-2. WST-1 activity was significantly inhibited by AZA, TSA, and NaB treatments, which was recovered by the ectopic expression of claudin-2 in human bronchial BEAS-2B. These results indicate that epigenetic inhibitors partially suppress proliferation in A549 cells mediated by the decrease in claudin-2 expression
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
