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. 2011 Dec 1;7(12):e1002396. doi: 10.1371/journal.pgen.1002396

Figure 5. Compromised ex vivo formation and function of HPCs in Ogna keratinocytes.

Figure 5

(A) Immunolocalization (double labeling) of ITGα6 and plectin in primary keratinocytes isolated from newborn mice. In wild-type (+/+) keratinocytes ITGα6 and plectin show codistribution in densely clustered HPCs (arrowheads) contrasting the more diffuse distribution in Ogna keratinocytes. Bar, 20 µm. (B) Column diagram showing proportions (%) of wild-type (+/+) and mutant keratinocytes having formed (clustered), or lacking (diffuse) HPCs. Data are shown as mean values from cell counts (>100/genotype) in randomly chosen optical fields from three independent experiments ±95% CI. *** P<0.001, two-way ANOVA with Bonferroni post test. (C) Immunofluorescence microscopy images (contrast–enhanced by conversion to grey scale and inversion of contrast) of keratin 5 filament networks in hyperosmotic shock-treated (+urea) and non-treated keratinocytes. Note more advanced network collapse in Plec Ogna/+ (arrow) compared to wild-type keratinocytes. Bar, 20 µm. (D) Comparison of cell numbers re-populating scratch wounds (n = 15) 16 hours after wound infliction. Box and whisker plots as in Figure 2. *** P<0.001, unpaired two-tailed t-test. (E) Migration velocities of keratinocytes overexpressing wild-type and Ogna-P1a. Immortalized Plec +/+ keratinocytes, transfected with either full-length wild-type (WT) P1a (n = 30 cells), or Ogna P1a (n = 34 cells), or empty (GFP-expressing) vector (mock, n = 30 cells), were monitored for migration over a period of 20 hours. Box and whisker plots as in Figure 2). *** P<0.001, one-way ANOVA with Tukey post test for multiple comparisons.