Figure 4

Keratinocyte differentiation induced in 3D raft cultures. (A) The panel depicts 3D raft cultures obtained from NIKS transduced with the indicated HPV16 E6 variants and vector. Micrographs include immunoftuorescence for the basal cell marker K5 (green), suprabasal cell marker K10 (red), the overlay of both and H+E staining. Rafts had been grown three times and sections were cut twice from each set of paraffin blocks to ensure reproducibility. (B) Calculation of K5 and K10 levels are shown for each HPV16 genotype and the empty vector raft as described in Materials and Methods. Values of area intensities for K5 and K10 in the basal and suprabasal regions from 4 sections are presented as mean ± standard deviation (SD). (C) Raft cultures were produced as described in (A). Micrographs showing immunoftuorescence of the basal cell marker K14 (Red) and nuclear staining of the same section (DAPI; blue) are shown. (D) Calculation of K14 levels in the basal and suprabasal regions were carried out as described in (B). Significance was tested by one way ANOVA test. A P value of < 0.05 was considered significant.