Fig. 2

Inflammation upregulates SGLT1, ACE2 and TMPRSS2 in Caco-2/TC7 cells. (A) Graphical illustration of the experimental protocol for the standard and inflamed Caco-2/TC7 cell models. Cells were differentiated in 25 mM glucose until day 7, then in 5.5 mM glucose until day 14. Cells were differentiated for 14 days from confluence and treated with dexamethasone (DEX) or 0.1% (v/v) DMSO (vehicle control) up to twice daily for 60 h, 24 h, or 4 h. For the inflamed model, a cytokine cocktail containing IL-1β (25 ng/mL) and TNF-α (50 ng/mL) was added to the basolateral (BL) once daily for 72 h. Finally, cell culture media from both apical (AP) and BL compartments were collected from the inflamed cells for IL-8 quantification. mRNA and protein were extracted from cells in both the standard and inflamed models and analysed through ddPCR and ELISA, respectively. A comparison of (B) IL-8 secretion and (C) SGLT1, (D) GLUT2, (E) ACE2 and (F) TMPRSS2 mRNA expression between standard (grey) and inflamed (blue) Caco-2/TC7 cells is shown. Total RNA was extracted and reverse transcribed, and absolute copies of SGLT1, GLUT2, ACE2 or TMPRSS2 cDNA measured by ddPCR are expressed relative to the housekeeping gene TBP. For IL-8 detection, cell culture media were processed, and IL-8 measured using ELISA and corrected for total protein measured by Bradford assay. Data are presented as mean ± SD (n = 6 technical replicates from N = 3 biological replicates). Differences between cells in the standard and inflamed environments in B-F were determined by unpaired t-test with Welch’s correction; ns—not significant, **P < 0.01, ***P < 0.001. A—created with BioRender.com (https://BioRender.com/r16s632); B-F—created with GraphPad Prism version 9.0.1.