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. 2017 Aug;140(2):584–587.e8. doi: 10.1016/j.jaci.2016.11.049

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© 2017 The Authors

This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

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Fig E2 — A, HMGB1 protein expression in ASM cells was assessed by immunofluorescence. B, Representative immunofluorescent staining of cell nuclei (DAPI, upper panel), HMGB1 (middle panel), and merged DAPI and HMGB1 (lower panel). C, ASM from subjects with asthma and control subjects was shown to express different isoforms of RAGE by PCR. PCR products were isolated by gel purification, sequenced, and identified as full-length membrane-bound RAGE (RAGE), or soluble isoforms of RAGE (sRAGE 1-3). M = molecular weight marker, C = healthy control, A = asthma. D, Representative flow cytometry histograms showing cell-surface RAGE ( FITC fluorescence, upper panel) and TLR4 (Alexa Fluor 488 fluorescence, lower panel) expression in ASM cells. Open histograms represent isotype controls; dark gray and light gray shaded histograms are RAGE or TLR4-stained cells, respectively. E, ASM cell-surface receptor expression was expressed as gMFI (fold change antibody/isotype control) for healthy control subjects (●) and subjects with asthma () for RAGE or TLR4 (P = .84 and P = .99, healthy control vs patient with asthma, respectively, unpaired t test). DAPI, 4′,6′-Diamidino-2 phenylindole; FITC, fluorescein isothiocyanate.

Inline graphic — A, HMGB1 protein expression in ASM cells was assessed by immunofluorescence. B, Representative immunofluorescent staining of cell nuclei (DAPI, upper panel), HMGB1 (middle panel), and merged DAPI and HMGB1 (lower panel). C, ASM from subjects with asthma and control subjects was shown to express different isoforms of RAGE by PCR. PCR products were isolated by gel purification, sequenced, and identified as full-length membrane-bound RAGE (RAGE), or soluble isoforms of RAGE (sRAGE 1-3). M = molecular weight marker, C = healthy control, A = asthma. D, Representative flow cytometry histograms showing cell-surface RAGE ( FITC fluorescence, upper panel) and TLR4 (Alexa Fluor 488 fluorescence, lower panel) expression in ASM cells. Open histograms represent isotype controls; dark gray and light gray shaded histograms are RAGE or TLR4-stained cells, respectively. E, ASM cell-surface receptor expression was expressed as gMFI (fold change antibody/isotype control) for healthy control subjects (●) and subjects with asthma () for RAGE or TLR4 (P = .84 and P = .99, healthy control vs patient with asthma, respectively, unpaired t test). DAPI, 4′,6′-Diamidino-2 phenylindole; FITC, fluorescein isothiocyanate.