Journal: The Journal of investigative dermatology

Article Title: The δ-opioid receptor affects epidermal homeostasis via ERK-dependent inhibition of transcription factor POU2F3.

doi: 10.1038/jid.2014.370

Figure Lengend Snippet: Figure 2. d-Opioid receptor (DOPr) overexpression inhibits keratinocyte proliferation. (a) Proliferation curves of DOPr-overexpressing and control cells, in either vehicle control medium (0.001% DMSO) or 100 nM SNC80- containing medium, were obtained from the images captured hourly with a 10x objective lens by the Incucyte machine. The graph depicts the meanþ / SEM percentage confluence per field of view of a representative experiment (n ¼ 3 per culture condition). Phase-contrast images of control and DOPr-overexpressing N/TERT-1 cultures were captured at 20x magnification, 5 days after plating the same amount of N/TERT-1 cells into standard culture vessels. Bar ¼ 20mm. (b) The percentage confluence after 5 days (120 hours) of culture is displayed as mean þ / SEM of three independent experiments. One-way ANOVA with the Newman-Keuls post hoc test, *Po0.05, **Po0.01. GFP, green fluorescent protein.

Article Snippet: SNC80 and naltrindole were from Tocris Biosciences (Bristol, UK), PD98059 from Promega (Madison, WI), and the solvent for SNC80 and PD98059, DMSO, as well as Met-enkephalin from Sigma-Aldrich (Singapore).

Techniques: Over Expression, Control

Journal: The Journal of investigative dermatology

Article Title: The δ-opioid receptor affects epidermal homeostasis via ERK-dependent inhibition of transcription factor POU2F3.

doi: 10.1038/jid.2014.370

Figure Lengend Snippet: Figure 3. d-Opioid receptor (DOPr) overexpression and activation impairs keratinocyte differentiation. Cells were grown to confluence and differentiation induced by removal of growth factors under the influence of methionine5 (Met)-enkephalin for up to 10 days. Differentiation marker gene expression was analyzed by quantitative real-time PCR. Data are represented after normalization to RPL13a expression as mean ±SEM (n ¼ 3 keratin intermediate filament (KRT10), n ¼ 4 KRT1, n ¼ 5 involucrin (IVL), loricrin (LOR), and filaggrin (FLG)). Expression levels of (a) KRT10, (b) KRT1, (c) IVL, (d) LOR, and (e) FLG are shown after normalization to vehicle-treated control cells. Two-way analysis of variance, ***Po0.001, **Po0.01, and *Po0.05. (f, g) Whole-cell lysates from confluent cultures (day 0) and day 1, 4, and 7 of differentiation with and without addition of 100 nM Met-enkephalin (f) or SNC80 (g) were analyzed by immunoblot for KRT10 and KRT1 expression, with equal loading verified by glyceraldehyde-3-phosphate dehydrogenase (GAPDH). GFP, green fluorescent protein.

Article Snippet: SNC80 and naltrindole were from Tocris Biosciences (Bristol, UK), PD98059 from Promega (Madison, WI), and the solvent for SNC80 and PD98059, DMSO, as well as Met-enkephalin from Sigma-Aldrich (Singapore).

Techniques: Over Expression, Activation Assay, Marker, Gene Expression, Real-time Polymerase Chain Reaction, Expressing, Control, Western Blot

Journal: The Journal of investigative dermatology

Article Title: The δ-opioid receptor affects epidermal homeostasis via ERK-dependent inhibition of transcription factor POU2F3.

doi: 10.1038/jid.2014.370

Figure Lengend Snippet: Figure 4. POU domain, class 2, transcription factor 3 (POU2F3) is a target of d-opioid receptor (DOPr)-mediated signaling. Keratinocytes were grown to confluence and differentiation induced by removal of growth factors. POU2F3 expression was analyzed by quantitative real-time PCR. Relative quantity is represented after normalization to RPL13a and the respective vehicle control as reference. (a) POU2F3 expression under influence of methionine5 (Met)- enkephalin during 7 days of differentiation. Graphs show mean ±SEM (n ¼ 5). Two-way analysis of variance, ***Po0.001. (b) Keratinocytes were differentiated for 24hours in the presence of 100 nM Met-enkephalin, 100 nM SNC80, 10mM naltrindole, or vehicle control. Graph represents mean ±SEM (n ¼ 4). t-Test, *Po0.05. (c) Keratinocytes were differentiated for 24hours in the presence of 100 nM Met-enkephalin, 100 nM SNC80, 20mM of extracellular signal-regulated kinase (ERK) inhibitor PD98059, or vehicle control. The graph represents mean ±SEM (n ¼ 4). t-Test, *Po0.05 and ***Po0.001. GFP, green fluorescent protein.

Article Snippet: SNC80 and naltrindole were from Tocris Biosciences (Bristol, UK), PD98059 from Promega (Madison, WI), and the solvent for SNC80 and PD98059, DMSO, as well as Met-enkephalin from Sigma-Aldrich (Singapore).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Control