Journal: Scientific reports
Article Title: Tension as a key factor in skin responses to pollution.
doi: 10.1038/s41598-023-42629-6
Figure Lengend Snippet: Figure 6. Ozone exposure upregulates protein levels of COX2 and MMP9, while the presence of tension affects the timing of the response. Ex vivo human skin biopsies were cultured with (tension—Ten) or without (non-tension—NT) tension (Ten Bio models) and then exposed for 4 h to either air or 0.4 ppm of O3. Samples were collected directly after exposure (T0), after 3 h (T3), 6 h (T6) and 24 h (T24). (a) Levels of COX2, green fluorescence staining represents COX2 immunoreactivity. Original magnification ×40. (b) Semi-quantification of the COX2 immunofluorescence intensities performed by ImageJ. (c) Levels of MMP9, red fluorescence staining represents MMP9 immunoreactivity. Original magnification ×40. (d) Semi-quantification of the MMP9 immunofluorescence intensities performed by ImageJ. Results are expressed as arbitrary units ± standard deviation. *p-value < 0.05 by ANOVA.
Article Snippet: Antigen retrieval was achieved via heat-based epitope retrieval with 10 mM sodium citrate buffer (AP-9003-500, Thermo), pH 6.0 with 0.05% Tween 20, at a sub-boiling temperature in a water bath set at 95 °C for 8 min. After cooling for 20 min, the sections were washed 2 X 5 min with 1% Phosphate Buffered Saline (PBS) and then blocked with 5% Bovine Serum Albumin (BSA) in PBS for 45 min. After blocking, sections were then incubated overnight with primary antibodies prepared in 2% BSA/PBS for the following markers: LL-37 (sc-166770, Santa Cruz, dilution 1:50), hBD2 (ab63982, AbCam, dilution 1:500), hBD3 (sc-59495, Santa Cruz, dilution 1:50), 4HNE (AB5605, Millipore, dilution 1:500), AhR (NB 100–128, Novus, 1:150), COX2 (NB 100–868, Novus, 10 μg/ml), MMP9 (NBP2-13173, Novus, 1:200).
Techniques: Ex Vivo, Cell Culture, Fluorescence, Staining, Immunofluorescence, Standard Deviation