Influence of AVG and CAE on the epidermal development in human epidermal equivalents.

<p>HPEKs were treated with AVG or CAE (10 μg/mL or 100 μg/mL). Frozen sections of skin equivalent model were then subjected to hematoxylin and eosin (HE) staining (A) or immunofluorescent staining for loricrin (LOR, green) and keratin 15 (K15, red) (B). The blue signals indicate nuclear staining. Scale bars, 20 μm. (C) The graphs indicate the mean ± SEM values for thickness of the whole epidermis or granular layer of the skin equivalent in micrometers. N = 20. **P<0.01, *P<0.05.</p

Influence of AVG and CAE on the proliferation of human epidermal keratinocytes.

<p>(A) HPEKs were treated with AVG or CAE (10 μg/mL or 100 μg/mL) for 48 h, and subjected to the cell proliferation assay. The graphs indicate the mean ± SEM values for proliferation rate (fold increase over control) from 20 independent experiments. **P<0.01, *P<0.05. (B) HPEKs were treated with 10 μg/mL AVG or CAE for 12 h, and subjected to the EdU incorporation assay. The graphs indicate the mean ± SEM values for percentage of EdU-positive (EdU⁺) cells from 5 independent experiments. **P<0.01. (C, D) Human full-thickness skin equivalents were treated with 10 μg/mL AVG or CAE for 3 days, and subjected to immunofluorescent staining against Ki67 (C) and p63 (D). The blue signals indicate nuclear staining. The dotted lines indicate the boundary between the epidermis and the dermis. Scale bars, 50 μm.</p

Influence of AVG and CAE on the wound healing in human epidermal keratinocytes.

<p>(A) HPEKs were treated with 10 μg/mL AVG or CAE, and subjected to the scratch wound healing assay over 8 h. White dotted lines indicate wound margins. The graph indicates the mean ± SEM values for wound closure rates of the group from 9 independent experiments. *P<0.05. (B) AVG or CAE (10 μg/mL) was directly applied onto a superficial incisional wound in which only the epidermis had been removed from human full-thickness skin equivalents and cultured at the air-liquid interface for 3 days. Representative images of hematoxylin and eosin staining of each group are shown. Arrowheads indicate wound margins.</p

Influence of AVG and CAE on the expression of adhesion molecules in human epidermal keratinocytes.

<p>(A-C) HPEKs were treated with 10 μg/mL AVG or CAE for 2 days, and subjected to flow cytometry analysis (A), q-PCR analysis (B), and western blot analysis (C). (A) Representative histograms are shown. The graphs indicate the mean ± SEM values for median fluorescent intensity (MFI) from 6 independent experiments. **P<0.01, *P<0.05. (B) The graphs indicate the mean ± SEM values for relative expression from 4 independent experiments. (C) The extracted proteins were immunoblotted with the indicated antibodies. Graphs indicate the relative band intensities as determined by ImageJ software and plotted as the means of 3 independent experiments.</p

Chemical constituents of AVG and CAE.

<p>Chemical constituents of AVG and CAE.</p

Influence of AVG and CAE on the differentiation in human epidermal keratinocytes.

<p>(A, B) HPEKs were treated with 0, 1, 10, 100, 1000, 2000 μg/mL AVG or CAE for 2 days. (A) Representative phase-contrast images of each group. Scale bars, 400 μm. (B) Gene expression levels of <i>SPRR2A</i>, <i>SPRR1B</i>, and <i>IVL</i> were quantified by q-PCR analysis. The graphs indicate the mean ± SEM values for relative expression from 5 independent experiments. **P<0.01, *P<0.05.</p