1. Activation of protease-activated receptor 2 leads to impairment of keratinocyte tight junction integrity 
Peter Nadeau, BS, Mason Henehan, BS, Anna De Benedetto, MD 
Journal of Allergy and Clinical Immunology 
Volume 142, Issue 1, Pages e7 (July 2018)
DOI: /j.jaci
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2. Fig 1
PAR2 activation reduced keratinocytes barrier integrity. PHKs treated with SLIGKV-NH2 (PAR2 agonist, 100 μmol/L) showed (A) significantly reduced TEER at 96 and 120 hours (mean ± SEM; *P < .01 and **P < .001; n = 9) as well as (B) reduced area under the curve of TEER measured daily from 48 to 144 hours (mean ± SEM; P = .03; n = 5). Additionally, (C) permeability to Na-fluorescein at 96-, 120-, and 144-hour time points was increased in PAR2-treated PHKs as it compared to the control (mean ± SEM; *P < .01; n = 8-9). REV, Reverse peptide.
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
Copyright © 2018 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3. Fig 2
PAR2 activation reduced and disrupted claudin-1 and occludin membrane staining patterns. PHKs were treated with SLIGKV-NH2 (PAR2) or with REV, representative images of costaining (n = 4 experiments) for claudin-1 (red), occludin (green), and merge images collected at 120 hours postdifferentiation are shown; 4′-6-diamidino-2-phenylindole, dihydrochloride (blue, insets). Images were acquired at some setting for each target/time points. Confocal microscope: Nikon A1RMPsi (Melville, NY) (magnification 20×).
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
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4. Fig E1
Reduced expression of CLDN1, OCLD, and ZO-1 at the mRNA level was observed in PAR2-treated PHKs as it compared to the control (REV); blue dotted line indicates the REV control value for each gene. Samples were collected at 72 hours postdifferentiation (mean ± SEM; *P < .05 and **P < .001; n = 7). RVU, Relative value unit.
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
Copyright © 2018 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5. Fig E2
PAR2 activation reduced and disrupted occludin and ZO-1 membrane staining patterns. PHKs were treated with SLIGKV-NH2 (PAR2 agonist) or with REV, representative images of 4 experiments for OCLD and ZO-1 at 96 hours postdifferentiation (red) are shown; 4′-6-diamidino-2-phenylindole, dihydrochloride (blue insets). Images were acquired at same setting for each target/time points. Confocal microscope: Nikon A1RMPsi (magnification 20×).
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
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6. Fig E3
Semiquantification of occludin and claudin membrane staining at 120 hours postdifferentiation was obtained using Image J (total pixel area/field). For each donor, 4 random fields (20×) were captured (mean ± SEM, *P < .05; n = 4). Confocal microscope: Nikon A1RMPsi.
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
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7. Fig E4
PAR2 activation is associated in reducing keratinocyte terminal differentiation. PHKs were treated with PAR2 peptide or REV (100 μmol/L) during differentiation. All genes were measured by quantitative RT-PCR, normalized to housekeeping gene (18s), and shown as RVU fold over control. Samples were collected 72 hours postdifferentiation. (REV = 1; n = 4, mean ± SEM; *P < .05). FLG, Filaggrin; LOR, loricrin; KLK, kallikreins.
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
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8. Fig E5
PAR2 activation is associated in increasing IL-8. PHKs were treated with PAR2 agonist peptide or REV (100 μmol/L) during differentiation. IL-8 was measured by ELISA in the cell culture supernatant collected 48 hours posttreatment. *P < .05.
Journal of Allergy and Clinical Immunology  , e7DOI: ( /j.jaci )
Copyright © 2018 American Academy of Allergy, Asthma & Immunology Terms and Conditions