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. 2013 Aug 22;23(1):34–43. doi: 10.1089/scd.2013.0038

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Copyright 2014, Mary Ann Liebert, Inc.

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FIG. 3. — Phenotypic profile of nontreated and treated KCs, at passage 3, assessed by (i) flow cytometry and (ii, iii) immunocytochemistry. (i) Representative flow cytometry histograms and data regarding the percentage of cells positive for K5, K14, and K19 confirming the high expression of early differentiation associated markers K5, K14 by all of the groups, and a (*) significantly higher expression of K19 after treatment in comparison to the nontreated cells. Filled areas in the flow cytometry histograms represent the negative population for the respective marker. (ii) (A–P) Immunofluorescence micrographs illustrating the higher expression of early differentiation-associated markers K5 (E, I, M), p63 (F, J, N), and β1-integrin (G, K, O) in treated KCs, when compared with nontreated ones (A–C). Conversely, late differentiation-associated marker involucrin was clearly expressed in nontreated cells (D) and collagen IV (H) and ROCKi independently treated KCs (L), but almost absent in the collagen IV and ROCKi combined treated KCs (P). A few k10 positive cells (arrowheads) were observed in collagen IV (E) and ROCKi (I) independently treated KCs and none in the nontreated cells (A) and in the collagen IV and ROCKi combined treated KCs (M). Scale bar corresponds to 50 μm. DAPI was used as nuclear staining. (iii) Graphical representations of p63 and involucrin positive cells quantified after immunocytochemistry. *P<0.05, **P<0.01, n=4.

FIG. 3. — Phenotypic profile of nontreated and treated KCs, at passage 3, assessed by (i) flow cytometry and (ii, iii) immunocytochemistry. (i) Representative flow cytometry histograms and data regarding the percentage of cells positive for K5, K14, and K19 confirming the high expression of early differentiation associated markers K5, K14 by all of the groups, and a (*) significantly higher expression of K19 after treatment in comparison to the nontreated cells. Filled areas in the flow cytometry histograms represent the negative population for the respective marker. (ii) (A–P) Immunofluorescence micrographs illustrating the higher expression of early differentiation-associated markers K5 (E, I, M), p63 (F, J, N), and β1-integrin (G, K, O) in treated KCs, when compared with nontreated ones (A–C). Conversely, late differentiation-associated marker involucrin was clearly expressed in nontreated cells (D) and collagen IV (H) and ROCKi independently treated KCs (L), but almost absent in the collagen IV and ROCKi combined treated KCs (P). A few k10 positive cells (arrowheads) were observed in collagen IV (E) and ROCKi (I) independently treated KCs and none in the nontreated cells (A) and in the collagen IV and ROCKi combined treated KCs (M). Scale bar corresponds to 50 μm. DAPI was used as nuclear staining. (iii) Graphical representations of p63 and involucrin positive cells quantified after immunocytochemistry. *P<0.05, **P<0.01, n=4.