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. Author manuscript; available in PMC: 2023 Feb 1.
Published in final edited form as: Gastroenterology. 2021 Oct 21;162(2):439–453. doi: 10.1053/j.gastro.2021.10.016

Figure 5. TSPAN12 deficiency affects endothelial functions.

Figure 5.

A, Representative qPCR (left) and western blot (right) of TSPAN12 after TSPAN12 downregulation by shRNA in HEsMEC. Numbers above TSPAN12 band represent the signal intensity normalized by HSP90 and relative to the control (shCtrl) set as 1. B-C, The effect of TSPAN12 gene silencing on the endothelial functions of barrier integrity (TEER [B] and FITC-dextran flux measurements [C]). Data were normalized to the control (shCtrl) set as 1. D, Heat diagram representing RNA-sequencing analysis of the TSPAN12-deficient cells and control cells. Differentially expressed genes (DEGs) were identified by filtering on TPM>1, moderated t-test with FDR P < .05. E, Gene ontology analysis of DEGs in the TSPAN12-deficient cells. F, Schematic of media swap experiments. G, Representative western blots demonstrating levels of collagen I and α-SMA in esophageal fibroblasts of media swap experiments. H, EDN1 (endothelin-1) expression in HEsMECs (shCtrl, shTSPAN12). qPCR (left) and ELISA of supernatants (right). I, Representative immunofluorescence staining of esophageal biopsy sections from normal control individuals and patients with fibrostenotic EoE; TSPAN12 (magenta), collagen (cyan), and VWF (red) with DAPI–stained nuclei (blue). White arrows indicate blood vessels, and white broken line indicates basal membrane. For A-C, G, and H, data are the means ± SEM of three independent experiments performed in duplicate. *P < .05, **P < .01, ***P < .001, and ****P < .0001 using the Mann-Whitney U test (H), one-way ANOVA test followed by a Sidak’s multiple comparisons test (B-C), or compared with shCtrl using one-way ANOVA test followed by a Tukey’s multiple comparisons test (A, G). DAPI, 4ʹ,6-diamidino-2-phenylindole; EoE, eosinophilic esophagitis; EDP, EoE Diagnostic Panel; HEsMEC, human esophageal microvascular endothelial cells; TEER, transendothelial electrical resistance; FITC, fluorescein isothiocyanate; FDR, false-discovery rate; shCtrl, shRNA non-silencing control; shTSPAN12, shRNA silencing TSPAN12; VWF, von Willebrand factor.