Asporin Reduces Adult Aortic Valve Interstitial Cell Mineralization Induced by Osteogenic Media and Wnt Signaling Manipulation In Vitro

Figure 6

Human recombinant Asporin (rhAsp) treatment decreases matrix mineralization in recombinant Wnt3a- (rhWnt3A-) treated osteogenic (OS) media-induced avian adult AVICs in culture. (a–c) Bright-field microscopic images (×10) of alizarin-stained OS-induced AVICs show increased matrix mineralization in rhWnt3a-treated OS-induced AVICs compared to OS-induced AVICs. Moreover, decreased matrix mineralization is detected in rhWnt3A+rhAsp-treated OS-induced AVICs (arrowheads). (d) Quantitative assay of Alizarin Red S shows that absorbance of rhWnt3A-treated OS-induced AVICs is upregulated (0.16) compared to only OS-induced AVICs (0.093), and absorbance of rhWnt3A+rhAsp-treated OS-induced AVICs is downregulated (0.065) compared to rhWnt3A-treated OS-induced AVICs (, ). (e) (I and VI): here, qRT-PCR data show that Asporin decreased by 0.73 fold in rhWnt3A-treated and increased by 3.43 fold in rhWnt3A+rhAsp OS-induced AVICs (I). Osteopontin and Runx2 mRNA expressions increased by 1.44 fold and 0.89 fold in rhWnt3A-treated and decreased by 0.07 fold and 1.23 fold in rhWnt3A+rhAsp-treated OS-induced AVICs (II and III). β-Catenin and Wnt3A mRNA increased by 2.2 fold and 1.73 fold in rhWnt3A-treated and decreased by 0.33 fold and 0.26 fold in rhWnt3A+rhAsp OS AVICs. Also, Dkk1 decreased by 0.71 fold in rhWnt3A-treated and increased by 1.44 fold in rhWnt3A+rhAsp-treated OS-induced AVICs. Statistical significance was determined by Student’s -test, where denotes , denotes , and denotes and .

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