Fig 1.
hBD2 expression of human-derived corneocytes affected by UV exposure and aging.
Concentrations of hBD2 were determined by ELISA assay. The hBD2 expression of (a) hBD2 expression of total corneocytes between the elderly and young adult groups (n = 42). (b) All groups (elderly adult dorsal hand (Elderly-H), elderly adult abdomen (Elderly-A), young adult dorsal hand (Young-H), and young adult abdomen (Young-A)), elderly adult (c) dorsal hand and (d) abdomen (elderly male n = 10 and female n = 11). In both elderly and young adult groups n = 21. Data are presented as the mean. p values for the differences between groups were calculated with the Mann-Whitney test (without outliners).
Fig 2.
Repeated low energy UVB irradiation-induced increase of SA markers and hBD2 expression in NHEK.
NHEK were treated with repeated UVB radiation exposures (5 mJ/cm2) at scheduled exposure times (0 to 7 times) and the time interval between exposures was 30 min. (a) Viability, (b) induction of hBD2 expression, (c) β-gal activity and (d) SA-protein markers at 64 h after each repeated exposure to UVB on NHEK. The degree of cell senescence was quantified as the percentage of SA-β-gal positive cells and expressed as a percentage of the 0 exposure group. hBD2 expression was measured using whole cell lysates by ELISA assay. Data are presented as the mean ± SEM of three independent experiments (n = 3). *, p<0.05, control vs. UVB treatment group. ns, no significance.
Fig 3.
ROS scavenging effect of adiponectin in UVB exposed NHEK.
UVB exposure was repeated 6 times and the time interval between exposures was 30 min. Intracellular ROS levels were measured by detecting the fluorescence intensity of the oxidant-sensitive florescent probe DCFH-DA. The florescence intensity was recorded in the presence or absence of adiponectin (10 μg/ml) at 485 nm/ 535 nm. Normalizing of the ROS florescence intensity was calculated using the PI fluorescence measurement intensity ratio. Data are presented as the mean ± SEM of three independent experiments (n = 3). *, p<0.05; **, p<0.005 vs. NC. #, p<0.05, vs. UVB treated group. ns, no significance.
Fig 4.
The protective effect of adiponectin pre-treatment on repeated UVB exposure induced (a) SA-β-gal activity, (b) SA-markers, and (c) hBD2 expression. NHEK pre-treatment of adiponectin for 24 h, and then treated with 6 repeated UVB exposures. After 64 h, the degree of cell senescence was quantified as the percentage of SA-β-gal positive cells and expressed as a percentage of NC cells. Protein expression levels were analyzed by Western blot. hBD2 expression was measured using the whole cell lysates by ELISA assays (representative fluorescence images of hBD2 expression levels in the presence or absence of adiponectin (10 μg/ml), showed as photographs). Data are presented as the mean ± SEM of three independent experiments (n = 3). *, p<0.05; **, p<0.005, vs. NC. #, p<0.05, vs. UVB treated group.
Fig 5.
The inhibitory effect of adiponectin on hBD2 expression signaling molecules on repeated UVB exposed NHEK.
(a) The time dependent phosphorylation of MAPKs induced by repeated UVB exposure. Adiponectin attenuated the phosphorylation of (b) JNK/SAPK, ERK and p38 MAPK phosphorylation (the relative expression levels were quantified and presented in graphical form) and (c) c-Fos and c-Jun protein expression. The UVB induced upregulation of hBD2 was suppressed through (d) the AP-1 components c-Fos and c-Jun. NHEK were treated with both SP600125 and SB203580 inhibitors before UVB exposure. Protein expression levels were analyzed by Western blot. And the relative (e) mRNA expression of c-Fos, c-Jun and hBD2 are represented in graphical form (fold change compared with NC cells). Data are presented as the mean ± SEM of three independent experiments (n = 3). *, p<0.05, vs. NC. #, p<0.05, vs. UVB treated group.