Figure 1

Henna and Lawsone activate AhR in HaCaT and human primary keratinocytes. (A) Chemical structures of TCDD, Phthiocol (Pht) and Lawsone (Law) and (B) in silico modeling studies predicting binding of these molecules in the AhR binding pocket. Upper panel: 2D-interaction plot (LigandScout 4.1), hydrogen-donor (green dashed), -acceptor (red dashed), hydrophobic (orange); lower panel: 3D-interaction models, hydrogen bonds (yellow dashed), potential halogen bond (green dashed). (C) Luciferase activity of AhR reporter HaCaT cells stimulated for 4 hours (h) with TCDD, Phthiocol (Pht), Henna or Lawsone (Law). (D) Dose dependent CYP1A1 expression of HEK cells stimulated for 4 h, in the presence (black dots) or absence (red dots) of the AhR inhibitor CH223191 (CH, 12 µM) normalized to control DMSO in the absence of Lawsone. (E) CYP1A1 and AHRR expression after 4 h Lawsone (10 µM) stimulation of HEK cells normalized to DMSO. Each dot represents one individual. (F-G) HEK cells were transiently transfected with AhR-siRNA (siAhR) or Scramble control (siScr) in different individuals (dots). Each color depicts results of the same individual. (F) AhR knockdown validation relative to non-transfected wild type (WT) cells. (G) CYP1A1 expression after 4 h stimulation with Lawsone normalized to DMSO. (H) 48 h CYP1A1 enzymatic activity in HEK cells treated with Lawsone (10 µM) normalized to DMSO. (I) AhR-target gene enrichment after Lawsone stimulation (10 µM) relative to TLR2 stimulation (Pam2CSK4, 300 ng/mL). Area under the curve (AUC), q-values and highly enriched genes are indicated. (C,E–H) Data from at least 3 independent experiments are shown. (D) Data from 1 representative experiment out of 2 is shown. (C) Mean + S.E.M., (D) Mean, (E–H) Floating bars, Mean Min to Max. and. (E,H) Student’s t-test, (F,G) One-way ANOVA with Fisher’s test. *P < 0.05; **P < 0.01; ***P < 0.001.