Tight Junction Protein Expression in Caco-2 Cells

The relative expression of tight junction proteins was determined as described by Yu et al. [26 (link)]. Human colon cell line Caco-2 was purchased from American Type Culture Collection (Manassas, VA, USA). Caco-2 cells were inoculated with 2 × 10⁵ cells/well in 6 well plates for 48 h, and the cells were treated with DSS (2.5%) to initiate intestinal barrier damage. Meanwhile, 100 μL yeast strains (10⁷ CFU/mL) were co-cultured with cells for 12 h. Treated Caco-2 cells were washed twice and separated from the culture plates.
Total RNA extraction and cDNA reverse transcription were performed using a FastPure Cell Total RNA Isolation Kit and an HiScript III RT SuperMix for qPCR, respectively (Vazyme Co., Ltd., Nanjing, China). RT-qPCR was conducted using a Fast SYBR Green Master Mix with a StepOne™ real-time PCR system with a BioRad-CFX384 machine (Bio-Rad Co., Ltd., Hercules, CA, USA). The primer sequences are listed in Table S1. The relative expression level of relative genes was calculated by 2^−ΔΔCT.