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J Cell Physiol. 2018 May;233(5):4056-4067. doi: 10.1002/jcp.26121. Epub 2017 Nov 16.

Characterization and assessment of potential microRNAs involved in phosphate-induced aortic calcification.

Author information

1
Laboratory of Cancer Biology and Molecular Immunology, Faculty of Sciences I, Lebanese University, Hadath, Beirut, Lebanon.
2
Institute of Molecular and Supramolecular Chemistry and Biochemistry (ICBMS), UMR CNRS 5246, University of Lyon 1, Bâtiment Raulin, Villeurbanne Cedex, France.
3
Institut de Biologie et de Médecine Moléculaires, Université Libre de Bruxelles, Gosselies, Belgium.
4
Department of Biochemistry and Molecular Genetics, Faculty of Medicine, American University of Beirut, Beirut, Lebanon.
5
INSERM-U1149, CNRS-ERL8252, Centre de Recherche sur l'Inflammation, and the Sorbonne Paris Cité, Laboratoire d'Excellence Inflamex, Faculté de Médecine, Site Xavier Bichat, Université Paris Diderot, Paris, France.
6
Cardiovascular Physiology Lab, Department of Anatomy, Cell Biology and Physiology, Faculty of Medicine, American University of Beirut, Beirut, Lebanon.
7
College of Medicine, Qatar University, Doha, Qatar.

Abstract

Medial artery calcification, a hallmark of type 2 diabetes mellitus and chronic kidney disease (CKD), is known as an independent risk factor for cardiovascular mortality and morbidity. Hyperphosphatemia associated with CKD is a strong stimulator of vascular calcification but the molecular mechanisms regulating this process remain not fully understood. We showed that calcification was induced after exposing Sprague-Dawley rat aortic explants to high inorganic phosphate level (Pi , 6 mM) as examined by Alizarin red and Von Kossa staining. This calcification was associated with high Tissue-Nonspecific Alkaline Phosphatase (TNAP) activity, vascular smooth muscle cells de-differentiation, manifested by downregulation of smooth muscle 22 alpha (SM22α) protein expression which was assessed by immunoblot analysis, immunofluorescence, and trans-differentiation into osteo-chondrocyte-like cells revealed by upregulation of Runt related transcription factor 2 (Runx2), TNAP, osteocalcin, and osteopontin mRNA levels which were determined by quantitative real-time PCR. To unravel the possible mechanism(s) involved in this process, microRNA (miR) expression profile, which was assessed using TLDA technique and thereafter confirmed by individual qRT-PCR, revealed differential expression 10 miRs, five at day 3 and 5 at day 6 post Pi treatment versus control untreated aortas. At day 3, miR-200c, -155, 322 were upregulated and miR-708 and 331 were downregulated. After 6 days of treatment, miR-328, -546, -301a were upregulated while miR-409 and miR-542 were downregulated. Our results indicate that high Pi levels trigger aortic calcification and modulation of certain miRs. These observations suggest that mechanisms regulating aortic calcification might involve miRs, which warrant further investigations in future studies.

KEYWORDS:

aorta; calcification; inorganic phosphate; microRNAs; trans-differentiation

PMID:
28776684
DOI:
10.1002/jcp.26121
[Indexed for MEDLINE]

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