Pharmacological effects of β-d-mannuronic acid (M2000) on miR-146a, IRAK1, TRAF6 and NF-κB gene expression, as target molecules in inflammatory reactions.
BACKGROUND
Impaired expression and function of microRNAs (miRNAs) are involved in the pathogenesis of many autoimmune and inflammatory diseases. Moreover, there is a close relationship between TLRs and miRNAs and impairment in regulating their expression which can play a vital role in the immunopathogenesis of many inflammatory reactions. This research aimed to study the pharmaceutical effects of M2000 (β-d-mannuronic acid) on the expression of miR-146a and its two target molecules (IRAK1 and TRAF6), and the transcription factor NF-κB in the HEK-Blue hTLR2 cell line.


METHODS
The cytotoxicity of M2000 was assessed by the MTT assay, and the qRT-PCR technique was employed in the presence and absence of M2000 treatment to measure gene-expression levels of miR-146a, IRAK1, TRAF6, and NF-κB.


RESULTS
MTT assay indicated that M2000 (before the concentration of 500μg/ml) had no cytotoxic effect on HEK-Blue hTLR2 cells. Our results showed that M2000 at low and high doses (5 and 25μg/well) could significantly reduce gene expression levels of miR-146a (p<0.01). Furthermore, it was found that this medication at two different doses could considerably decrease IRAK1 and TRAF6 gene expression (p<0.001). Moreover, this study revealed that expression level of NF-κB also significantly declined at these two doses (p<0.01).


CONCLUSIONS
This study for the first time shows that M2000 as a novel NSAID with immunosuppressive properties is able to modify TLR signaling through suppressing the adaptor molecules IRAK1 and TRAF6, the transcription factor NF-κB and miR-146a as a new therapeutic approach.
Department of Cellular and Molecular Biology, Kish International Campus, University of Tehran, Kish, Iran; School of Biology, University College of Science, University of Tehran, Tehran, Iran.
Seyed Shahabeddin
SS
Mortazavi-Jahromi
Department of Cellular and Molecular Biology, Kish International Campus, University of Tehran, Kish, Iran; School of Biology, University College of Science, University of Tehran, Tehran, Iran.
Mehdi Malek
MM
Jamshidi
Department of Cellular and Molecular Biology, Kish International Campus, University of Tehran, Kish, Iran; School of Biology, University College of Science, University of Tehran, Tehran, Iran.
Ali
A
Farazmand
Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran.
Zahra
Z
Aghazadeh
Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.
Mehdi
M
Yousefi
Department of Immunology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran. Electronic address: mirshafiey@tums.ac.ir.
Abbas
A
Mirshafiey
D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000008:administration & dosage / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects*; D057809:HEK293 Cells; D006603:Hexuronic Acids / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D007166:Immunosuppressive Agents / Q000008:administration & dosage / Q000494:pharmacology; D007249:Inflammation / Q000188:drug therapy* / Q000473:pathology; D053592:Interleukin-1 Receptor-Associated Kinases / Q000235:genetics; D047908:Intracellular Signaling Peptides and Proteins; D035683:MicroRNAs / Q000235:genetics; D016328:NF-kappa B / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D048029:TNF Receptor-Associated Factor 6 / Q000235:genetics
D000894:Anti-Inflammatory Agents, Non-Steroidal; D006603:Hexuronic Acids; D007166:Immunosuppressive Agents; D047908:Intracellular Signaling Peptides and Proteins; C542542:MIRN146 microRNA, human; D035683:MicroRNAs; D016328:NF-kappa B; D048029:TNF Receptor-Associated Factor 6; C000604754:Tifab protein, human; C008324:mannuronic acid; C507768:IRAK1 protein, human; D053592:Interleukin-1 Receptor-Associated Kinases
M2000; Non-steroidal anti-inflammatory drug (NSAID); Target molecules (IRAK1 and TRAF6); miR-146a; β-d-mannuronic acid