Overexpressed Thrombospondin 2 Induced Osteogenic Differentiation of Valve Interstitial Cells via Inhibition of Akt/NF-κB Signaling Pathway to Promote Calcific Aortic Valve Disease Development

Figure 6

THBS2 mediated the OD of VICs via inhibition of Akt/NF-κB signaling pathway. (a) ALP activity of VICs transfected with shTHBS2 and treated with LY294002 was detected through colorimetry after the induction of OD. (b) Calcic nodule formation of VICs transfected with shTHBS2 and treated with LY294002 was measured by Alizarin red staining after the induction of OD (scale: 200 μm; magnification: ×100). (c) and (d) Relative mRNA expressions of osteocalcin, Runx2, SPARC, COL1A2, and COL1A1 (c) and SPP1, CTGF, MMP-2, and MMP-13 (d) in VICs transfected with shTHBS2 and treated with LY294002 were determined through qRT-PCR after the induction of OD. GAPDH was a loading control. vs. vector group; , vs. shTHBS2 group; ; vs. LY294002 group. All experiments were repeated independently at least three times. Data were performed as the . Abbreviations: THBS2: thrombospondin 2; OD: osteogenic differentiation; VICs: valve interstitial cells; ALP: alkaline phosphatase; shTHBS2: short hairpin THBS2; Runx2: Runt-related transcription factor-2; SPARC: secreted protein acidic and rich in cysteine; COL1A: collagen type I alpha; SPP1: secreted phosphoprotein 1; CTGF: connective tissue growth factor; MMP-: matrix metalloproteinase-; qRT-PCR: quantitative reverse transcription-polymerase chain reaction; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

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