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. 2022 May 14;2022:1260423. doi: 10.1155/2022/1260423

Figure 11.

Figure 11

Overexpression of TP53INP2 promoted cell apoptosis via caspase-8/TRAF6 pathway in ACHN cells. (a) and (b) Both mRNA levels of caspase-8 and TRAF6 were detected with qRT-PCR in TP53INP2-treated ACHN cells. (c) and (d) The protein levels of both caspase-8 and TRAF6 were evaluated with western blot. (e) Overexpression of TP53INP2 upregulated both expressions of caspase-8 and TRAF6, which was verified with immunofluorescence. (f) ACHN cells expressing FLAG-TRAF6 and TP53INP2 were immunoprecipitated with FLAG resin, and immunocomplexes were subjected to western blot analysis with anti-FLAG, anti-TP53INP2, and anti-caspase-8 antibody. (g) and (h) TRAF6 silence was transiently transfected into the ACHN cell lines. The relative mRNA level and protein level of TRAF6 was estimated with qRT-PCR and western blot. (i) and (k)–(m) Protein levels of cleaved caspase-3, cleaved caspase-8, and cleaved PARP were measured in cells administrated with TP53INP2, siTRAF6, and TP53INP2+siTRAF6. (j) and (n) Flow cytometry was accomplished for the detection on the apoptosis rate in different groups. Data were presented as mean ± SD of three or four independent experiments. ∗P < 0.05; ∗∗P < 0.01; ∗∗P < 0.001.