Figure 5 | Scientific Reports

Figure 5

From: Molecular hydrogen suppresses activated Wnt/β-catenin signaling

Figure 5

H2 inhibits Wnt/β-catenin signaling in chondrocytes.

(a) HCS-2/6 human chondrosarcoma cells were treated with control CM (Cont.), Wnt3a CM, or 2 μM BIO with 10% H2 or 10% N2 gas for 24 h. The Wnt/β-catenin signaling activity was measured by Topflash luciferase reporter assay (n = 9). *P < 0.05 and **P < 0.01 by Student’s t-test. (b) Human OAC cells were treated with control CM (Cont.), Wnt3a CM, or 2 μM BIO with 10% H2 or 10% N2 gas for 24 h. Representative Western blots are shown with densitometry of β-catenin/β-actin (n = 4). *P < 0.05 by Student’s t-test. (c–g) Human OAC cells (clone 1) were treated with control CM (Cont.), Wnt3a CM, or 2 μM BIO with 10% H2 or 10% N2 gas for 24 h. Expression of AXIN2 (n = 3) (c), SOX9 (n = 6) (d), MMP3 (n = 6) (e), COL2A1 (n = 3) (f) and ACAN (n = 3) (g) were quantified by qRT-PCR. *P < 0.05 by Student’s t-test. (h) Differentiated ATDC5 cells were treated with control CM (Cont.), Wnt3a CM, or 2 μM BIO with 10% H2 or 10% N2 gas for 48 h. Proteoglycans were stained with Alcian blue (upper panel) and quantified by measuring the optical intensity at 630 nm of the cell lysates (n = 9 for Cont.-treated and BIO-treated cells, n = 6 for Wnt3a-treated cells). *P < 0.05 by Student’s t-test. Non-significant P values less than 0.10 are indicated above each pair.

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