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. 2021 Nov 8;6(21):e148135. doi: 10.1172/jci.insight.148135

Figure 2. Transcription of the SPINK1 gene is upregulated under hypoxia in a HIF-dependent manner.

Figure 2

(A–D) HeLa cells were cultured under the indicated oxygen conditions in the presence or absence of 5 μg/mL Act D for 24 hours (A), 100 μM deferoxamine for 48 hours (B), 2 mM DMOG for 24 hours (C), or 3 μM MG132 for 12 hours (D) and subjected to qPCR. (E–G) HeLa cells were transfected with the indicated siRNA or scramble siRNA for negative control, cultured under the indicated oxygen conditions for 24 hours, and subjected to qPCR. (H and I) After simultaneously silencing HIF-1α, HIF-2α, and HIF-3α using 2 kinds of mixtures using HIF-αs and siRNAs (mixture-1 and mixture-2), HeLa cells were cultured under the indicated oxygen conditions for 24 hours and subjected to qPCR (H) or the ELISA assay (I). (J and K) The same experiments as in H and I were conducted after silencing HIF-1β. Scramble siRNA was used as a negative control. Data are represented as mean ± SD (n = 3). Two-tailed Student’s t test (A–D). One-way ANOVA with Dunnett’s test (E–K). *P < 0.05, **P < 0.01, ***P < 0.001. SPINK1, serine peptidase inhibitor Kazal type 1; EV, empty vector; Act D, actinomycin D; DMOG, dimethyloxallyl glycine.