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. 2022 Jan 7;14(2):282. doi: 10.3390/cancers14020282

Figure 1.

Figure 1

Inducible HPV16E6E7 knockdown drives a pleotropic anti-cancer effect in HPV16+ SCC cells. (A) Design of short hairpin RNA (shRNA) constructs targeting HPV16E6 or HPV16E7. (B) Validation of shRNA constructs to target HPV16E6 and HPV16E7, and suppress colony formation. Immunoblot for HPV16E6 and HPV16E7 (left). Uncropped immunoblots are available in File S1. Colony formation assay (right). Number of colonies as a percentage of control (CTL) count for HPV16E6E7 knockdown (E6E7 KD) induced by shRNA constructs 4 and 5. * p < 0.05; ** p < 0.01. (C) Colony formation assay. Number of colonies as a percentage of CTL. ** p < 0.01. (D) Tumorsphere formation assay. The number of tumorspheres is shown as a percentage of the CTL count. ** p < 0.01. (E) Cell cycle analysis. Black, gray, and white bars indicate the percentage of the total population in each cell cycle phase for CTL and HPV16E6E7 KD cells. (F) Apoptosis assay. Percentage of CTL and HPV16E6E7 UMSCC47 cells that are annexin V+ as measured by flow cytometry. * p < 0.05. (G) Senenscence assay. The percentage of CTL and HPV16E6E7 UMSCC47 cells that are B-galactosidase positive. ** p < 0.01.