Journal: Microbiology spectrum

Article Title: Phillyrin ameliorates DSS-induced colitis in mice via modulating the gut microbiota and inhibiting the NF-κB/MLCK pathway.

doi: 10.1128/spectrum.02006-24

Figure Lengend Snippet: Figure 5A illustrates that p-NF-κB p65 and p-IκB expression in the DSS group was markedly enhanced in comparison to the control group. However, the PHY group notably reversed these effects in comparison to the DSS group (Fig. 6A through C). Research has demonstrated that NF-κB can enhance MLCK transcription, increasing MLC phosphory lation and decreasing TJ protein expression (23). Furthermore, the MLCK and p-MLC protein expression in DSS was enhanced in comparison to the control group. Neverthe less, in the PHY pre-protected group, the expression of MLCK (Fig. 6A and D) and p-MLC (Fig. 6A and E) proteins was lower in comparison to the DSS group. These outcomes revealed that PHY could restore the intestinal barrier by regulating the NF-κB /MLCK/MLC signaling pathway.

Article Snippet: The protein was transferred from gel to polyvinylidene difluoride membrane (PVDF) (Millipore, Darmstadt, Germany) by wet transfer method at constant flow for 220 mA and 75 min. After the transfer, 5% skim milk powder was added, and the table was closed at room temperature for 4 h. After closure, tris-buffered saline with Tween 20 (TBST) was washed three times for 10 min each time, and then the primary antibody was incubated in primary antibodies against occludin (1:1,000; Proteintech), myosin light-chain kinase (MLCK) (1:1,000; Abcam, Cambridge, UK), phosphorylated myosin light chain (p-MLC) (1:1,000; Abcam), myosin light-chain (MLC) (1:1,000; CST, Boston, USA), p-NF-κB (1:1,000; Abcam), NF-κB (1:1,000; Abcam), p-IκB (1:1,000; CST), IκB (1:1,000; CST), claudin-3 (1:1,000; Abcam), inducible nitric oxide synthase (iNOS) (1:1,000; Proteintech), cyclooxygenase-2 (COX-2) (1:1,000; Proteintech), and β-actin (1:1,000; Proteintech) at 4°C overnight.

Techniques: Expressing, Comparison, Control