Anti zo 1

We used the jejunal mucosa to prepare the protein samples for western blot assay, as previously described [28 (link)–30 (link)]. The jejunal segments samples were collected from the approximately middle positions in the jejunal tracts, as previously described [28 (link), 31 (link)]. The mid-jejunal segments were rinsed with PBS solution, and then the jejunal mucosa samples were gently scraped off. WCLs of the jejunal mucosa were prepared in RIPA lysis buffer (Sangon Biotech, C500005). The nuclear and cytoplasmic extraction of the intestinal epithelium was conducted using a kit (Thermo Fisher Scientific, 78833). This western blot assay was conducted as previously described [31 (link)]. Primary antibodies included the anti-E-cadherin (Cell Signaling Technology, 14472S), anti-ZO-1 (ABclonal, A11417), anti-Connexin 43 (Proteintech, 26980-1-AP), anti-Lamin B1 (Proteintech, 12987-1-AP), anti-CYP1A1 (Proteintech, 13241-1-AP), anti-AhR (Proteintech, 67785-1-Ig), anti-β-actin (Sigma, A5441), and anti-β-tubulin (Proteintech, 66240-1-Ig). Secondary antibodies included the HRP-conjugated secondary antibodies (Cell Signaling Technology, 7076S and 7074S).

The western blot was performed as described previously [62 (link)]. Proteins (20–30 μg) of the left cerebral hemisphere extracted were separated by SDS-PAGE gel electrophoresis (Proteinbio) and transferred onto polyvinylidene fluoride (PVDF) membranes (Bio-rad) by electroblotting. The membranes were blocked by 5% skim milk dissolved with Tris-buffer saline (TBS, containing 0.1% Tween-20) for 1 h at RT and then incubated with primary antibody at 4℃ overnight. These primary antibodies were used: anti-Mfsd2a (Thermo Fisher Scientific), anti-ZO-1 (Abclonal), anti-occludin (Abcam), anti-AQP4 (Abclonal), anti-albumin (Abclonal), anti-caveolin-1 (CST), anti-beta actin (Abcam). Horseradish peroxidase (HRP)-conjugated secondary antibody was incubated with the membranes at RT for 1 h. The secondary antibody was the anti-rabbit antibody (Abcam). The proteins were detected by the ECL chemiluminescence system (Vazyme), and the corresponding images were captured using a ChemiDoc Touch Imaging System (Bio-Rad). The band intensity was analyzed by ImageJ software.