Fig 2: TFAP2A positively regulates PD-L1 expression in cervical cancer. (A) IHC staining of PD-L1 expression and hematoxylin-eosin staining in normal and cancer tissues. (B) The association between PD-L1 and TFAP2A protein (semi-quantitative analysis using IHC staining). (C) The association between PD-L1 and TFAP2A protein in The Cancer Genome Atlas database. (D) Western blot analysis of TFAP2A and PD-L1 expression in normal (END1) and cervical epithelial (HeLa, C33a and SiHa) cell lines. (E) Reverse transcription-quantitative PCR of TFAP2A and PD-L1 expression in normal (END1) and cancer) cervical epithelial (HeLa, C33a and SiHa) cell lines. (F) Western blot analysis revealed that PD-L1 expression was reduced in TFAP2A-overexpressing cervical cancer cell lines. (G) Western blot analysis revealed that PD-L1 expression was reduced in cervical cancer cell lines subjected to TFAP2A knockdown. n=3 for cell culture analysis. *P<0.05, **P<0.01 and ***P<0.001. TFAP2A, transcription factor AP-2 alpha; PD-L1, programmed death-ligand 1; H&E, hematoxylin-eosin; IHC, immunohistochemical.

Fig 3: TFAP2A expression is increased in cervical cancer and is associated with clinicopathological features. (A-D) Analysis of TFAP2A mRNA expression in different datasets. (E) Expression of TFAP2A mRNA in six sets of paired cancer-paraneoplastic tissue samples. (F) IHC staining of TFAP2A expression and H&E staining in normal and cancer tissues. (G) Analysis of TFAP2A mRNA expression in different FIGO grades. (H) The expression level of the TFAP2A gene in different histopathology grades. (I) The expression level of TFAP2A gene in different FIGO T Stages. (J) The expression level of the TFAP2A gene according to lymph node metastasis. (K) The expression level of TFAP2A gene in association with patient survival. in each group. *P<0.05, **P<0.01 and ***P<0.001. TFAP2A, transcription factor AP-2 alpha; H&E, hematoxylin-eosin; IHC, immunohistochemical.

Fig 4: TFAP2A promotes the proliferation and migration, and inhibits the apoptosis of cervical cancer cells via PD-L1. (A) Western blot analysis of TFAP2A, PD-L1 expression in the shctr, shTFAP2A#1, shTFAP2A #1 + Vector and shTFAP2A #1 + PD-L1 groups. (B) Cell Counting Kit-8 assays revealed the growth rate of the cells. (C) Colony formation assay revealed the proliferation of the cells. (D) Transwell assay revealed the migration and invasion of cells. (E) Apoptosis detection using flow cytometric analysis revealed apoptosis rate of the cells. n=3 in each group. *P<0.05, **P<0.01 and ***P<0.001. TFAP2A, transcription factor AP-2 alpha; PD-L1, programmed death-ligand 1; sh-, short hairpin.

Fig 5: TFAP2A and PD-L1 form a positive feedback pathway, which may be regulated by HPV E6 protein. (A) Western blot analysis revealed that TFAP2A expression was reduced in PD-L1-overexpressing cervical cancer cell lines. (B) Western blot analysis revealed that TFAP2A expression was reduced in cervical cancer cell lines in which PD-L1 was knocked down. (C) Western blot analysis revealed that TFAP2A and PD-L1 expression was reduced in cervical cancer cells in which HPV E6 was knocked down. n=3 in each group. *P<0.05, **P<0.01 and ***P<0.001. TFAP2A, transcription factor AP-2 alpha; PD-L1, programmed death-ligand 1; sh-, short hairpin.