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. 2022 Apr 6;2022:1835900. doi: 10.1155/2022/1835900

Figure 2.

Figure 2

USP7 knockdown inhibits ATDC5 cell proliferation and increases apoptosis and inflammatory response after 48 h chondrogenic induction under TNF-α-induced inflammation. (a) Relative USP7 mRNA expression under 20 ng/mL TNF-α. (b) Relative USP7 protein expression in USP7 knockdown and its control groups. (c) Alcian blue and toluidine blue staining in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction. Scale bars = 100 μm. (d) Growth curves in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction measured by CCK-8 assay. (e) Relative Col2a1 and Sox9 mRNA expression of in USP7 knockdown and its control groups under TNF-α stimulation after 48 h chondrogenic induction. (f) Col2a1, Sox9, Cleaved Caspase-3, Bax, and Bcl-2 protein expression of in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction. (g) Quantitative measurement of (f). (h) Relative Caspase-3 activity in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction. (i) Quantitative measurement of cell apoptosis measured by flow cytometry in the USP7-nc and USP7-sh2 groups under TNF-α-induced inflammation after 48 h chondrogenic induction. (j) TUNEL staining in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction. White arrows indicated TUNEL-positive cells. Scale bars = 50 μm. (k) Quantitative measurement of (j). (l) Relative IL-6, COX, NOS2, and MMP13 mRNA expression in USP7 knockdown and its control groups under TNF-α-induced inflammation after 48 h chondrogenic induction. (m) IL-6 expression in USP7 knockdown and its control group supernatant under TNF-α-induced inflammation after 48 h chondrogenic induction. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.