MicroRNA miR-320a modulates induction of HO-1, GCLM and OKL38 by oxidized phospholipids in endothelial cells.
OBJECTIVE
Oxidized phospholipids (OxPLs), which are highly abundant in atherosclerotic lesions, are known to induce electrophilic stress response (ESR). ESR induces cytoprotective genes via the NF-E2-related factor 2 (NRF2) transcription factor. In order to get further insight into the mechanisms of ESR, we studied the role of microRNA (miR)-320a in induction of NRF2-dependent genes by OxPLs.


METHODS
Microarray profiling and qRT-PCR methods were used for measurements of mRNA and miRNA levels. miR-320a levels were changed by transfection with synthetic oligonucleotides. Protein analysis was performed by Western blotting. The functional activity of NRF2 was measured by DNA-binding ELISA.


RESULTS
Oxidized palmitoyl-arachidonoyl-phosphatidylcholine (OxPAPC) induced miR-320a in endothelial cells. Induction of HO-1, OKL38 and GCLM mRNAs by OxPAPC and sulforaphane was attenuated upon knockdown of miR-320a. In contrast, transfection of ECs with miR-320a mimic oligonucleotide potentiated the effects of OxPAPC and sulforaphane on induction of HO-1, OKL38 and GCLM mRNAs. OxPAPC-induced p38 activation, levels of NRF2 protein and its ability to bind to consensus NRF2 DNA binding site were elevated in ECs transfected with miR-320a mimic. miR-320a positively regulated induction of VEGF mRNA by OxPAPC. Levels of miR-320a and HO-1 and OKL38 mRNAs were elevated in aortas of ApoE knockout mice fed with high fat diet. Manipulation of miR-320a level in ECs did not affect ability of OxPAPC to induce IL-8, COX-2 and MCP-1.


CONCLUSIONS
miR-320a plays important role in induction of expression of HO-1, GCLM and OKL38 upon ESR induced either by OxPAPC or sulforaphane. These observations propose a general role of miR-320a in control of ESR induced by different electrophilic agents.
Institute for Physiology, Centre for Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
Waltraud C
WC
Schrottmaier
Department of Vascular Biology and Thrombosis Research, Centre for Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria.
Olga V
OV
Oskolkova
Institute for Physiology, Centre for Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria. Electronic address: gernot.schabbauer@meduniwien.ac.at.
Gernot
G
Schabbauer
Department of Vascular Biology and Thrombosis Research, Centre for Physiology and Pharmacology, Medical University of Vienna, Vienna, Austria. Electronic address: taras.afonyushkin@meduniwien.ac.at.
Taras
T
Afonyushkin
D000818:Animals; D001057:Apolipoproteins E / Q000235:genetics; D051017:Apoptosis Regulatory Proteins; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005721:Glutamate-Cysteine Ligase / Q000235:genetics / Q000378:metabolism*; D051547:Heme Oxygenase-1 / Q000378:metabolism*; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D017879:Isothiocyanates / Q000737:chemistry*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D051267:NF-E2-Related Factor 2 / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D009841:Oligonucleotides / Q000737:chemistry; D018384:Oxidative Stress; D010100:Oxygen / Q000737:chemistry; D010743:Phospholipids / Q000737:chemistry*; D011506:Proteins / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D013454:Sulfoxides; D014162:Transfection
D001057:Apolipoproteins E; D051017:Apoptosis Regulatory Proteins; D017879:Isothiocyanates; C572916:MIRN320 microRNA, human; D035683:MicroRNAs; C541056:Mirn320 microRNA, mouse; D051267:NF-E2-Related Factor 2; C495635:NFE2L2 protein, human; C432368:OSGIN1 protein, human; D009841:Oligonucleotides; C492609:Osgin1 protein, rat; D010743:Phospholipids; D011506:Proteins; D012333:RNA, Messenger; D013454:Sulfoxides; D051547:Heme Oxygenase-1; C546365:GCLM protein, human; C560749:GCLM protein, mouse; D005721:Glutamate-Cysteine Ligase; C016766:sulforaphane; D010100:Oxygen
Electrophilic stress response; GCLM; HO-1; MicroRNA-320a; NRF2; OKL38; Oxidized phospholipids