title: Overexpression of hsa-miR-148a promotes cartilage production and inhibits cartilage degradation by osteoarthritic chondrocytes.
abstract: OBJECTIVE
  Hsa-miR-148a expression is decreased in Osteoarthritis (OA) cartilage, but its functional role in cartilage has never been studied. Therefore, our aim was to investigate the effects of overexpressing hsa-miR-148a on cartilage metabolism of OA chondrocytes.
  
  
  METHODS
  OA chondrocytes were transfected with a miRNA precursor for hsa-miR-148a or a miRNA precursor negative control. After 3, 7, 14 and 21 days, real-time PCR was performed to examine gene expression levels of aggrecan (ACAN), type I, II, and X collagen (COL1A1, COL2A1, COl10A1), matrix metallopeptidase 13 (MMP13), a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and the serpin peptidase inhibitor, clade H (heat shock protein 47), member 1 (SERPINH1). After 3 weeks, DNA content and proteoglycan and collagen content and release were determined. Type II collagen was analyzed at the protein level by Western blot.
  
  
  RESULTS
  Overexpression of hsa-miR-148a had no effect on ACAN, COL1A1 and SERPINH1 gene expression, but increased COL2A1 and decreased COL10A1, MMP13 and ADAMTS5 gene expression. Luciferase reporter assay confirmed direct interaction of miR-148a and COL10A1, MMP13 and ADAMTS5. The matrix deposited by the miR-148a overexpressing cells contained more proteoglycans and collagen, in particular type II collagen. Proteoglycan and collagen release into the culture medium was inhibited, but total collagen production was increased.
  
  
  CONCLUSIONS
  Overexpression of hsa-miR-148a inhibits hypertrophic differentiation and increases the production and deposition of type II collagen by OA chondrocytes, which is accompanied by an increased retention of proteoglycans. Hsa-miR-148a might be a potential disease-modifying compound in OA, as it promotes hyaline cartilage production.
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands. Electronic address: l.a.vonk@umcutrecht.nl.
authors.forename: L A
authors.initials: LA
authors.lastname: Vonk
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: A H M
authors.initials: AH
authors.lastname: Kragten
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands; Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
authors.forename: W J A
authors.initials: WJ
authors.lastname: Dhert
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands; Tissue Regeneration, MIRA Institute, University Twente, Enschede, The Netherlands.
authors.forename: D B F
authors.initials: DB
authors.lastname: Saris
authors.affiliation: Department of Orthopaedics, University Medical Center Utrecht, Utrecht, The Netherlands.
authors.forename: L B
authors.initials: LB
authors.lastname: Creemers
mesh_terms: D051722:ADAM Proteins / Q000096:biosynthesis / Q000235:genetics; D000071116:ADAMTS5 Protein; D000368:Aged; D000369:Aged, 80 and over; D002358:Cartilage, Articular / Q000378:metabolism / Q000473:pathology*; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D024043:Collagen Type II / Q000096:biosynthesis / Q000235:genetics; D024121:Collagen Type X / Q000096:biosynthesis / Q000235:genetics; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D008297:Male; D053509:Matrix Metalloproteinase 13 / Q000096:biosynthesis / Q000235:genetics; D035683:MicroRNAs / Q000096:biosynthesis / Q000502:physiology*; D008875:Middle Aged; D020370:Osteoarthritis, Knee / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D011509:Proteoglycans / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics
chemical_list: D024043:Collagen Type II; D024121:Collagen Type X; C550570:MIRN148 microRNA, human; D035683:MicroRNAs; D011509:Proteoglycans; D012333:RNA, Messenger; D051722:ADAM Proteins; D000071116:ADAMTS5 Protein; C121532:ADAMTS5 protein, human; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13
keywords: Cartilage; Chondrocytes; MicroRNA; Osteoarthritis