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. 2020 May 18;16(12):2159–2169. doi: 10.7150/ijbs.45933

Figure 4.

Figure 4

MiR-4516 negatively regulates OTX1 expression by binding to its 3'-UTR. (A) Bioinformatics analysis was performed to predict the potential targets of miR-4516 using online database TargetScan, miRDB, and TarBase. (B) PANC-1 cells were transfected with miR-4516 or miR-NC. Potential target gene expression levels were analyzed by qPCR 48 hours later. (C) The putative binding sequences between miR-4516 and WT 3'-UTR of OTX1 was shown. (D) PANC-1 or AsPC-1 cells were transfected with luciferase reporter vector containing WT or mutated 3'-UTR of OTX1, together with miR-4516 or miR-NC. The relative luciferase activity was analyzed 48 hours later. (E, F) AsPC-1 cells were transfected with miR-NC, miR-4516 mimics, miR-4516 inhibitor or inhibitor negative control. 48 hours later, the mRNA expression (E) and protein expression (F) of OTX1 were analyzed by qPCR and western blot. (G) The expression of OTX1 in pancreatic cancer tissues and adjacent non-tumor tissues was analyzed by immunohistochemical staining. (H) The relative expression of OTX1 in pancreatic cancer tissues and adjacent non-tumor tissues was analyzed by qPCR. *, P < 0.05; **, P < 0.01.