Figure 4. Probiotics treatment increases expression of tight junctions in the intestine of older obese mice.

(A and B) The mRNA expression of tight junction proteins like Zonulin-1 (Zo1) (A) and Occludin (Ocln) (B) were significantly increased in colon of probiotics fed older mice (n = 8) compared with their controls (n = 6). (C and D) Western blot analysis shows that Zo1 protein expression was significantly increased, while Ocln showed marginal increase in the colon tissues of probiotic-treated older mice (n = 7) compared with their controls (n = 6). (E and F) Global gene expression using RNAseq analysis revealed that probiotic feeding significantly increased around 856 genes while it decreased 1053 genes that were distinctly clustered in the probiotic-treated (n = 7) group versus controls (n = 6). (G) Pathway analysis of deferentially expressed genes (DEGs) shows that cell adhesion and cytokine (immune) pathways were more affected by probiotics treatment compared with their controls. Values are mean of n = 6–7 each group, and data are shown as mean ± SEM. **P < 0.01 and ***P < 0.001. Student t test (A, B, D) and random forest analysis (E) were used, as well as hierarchical clustering between samples using hclust, with diagrams drawn with ggplot2 (F) and differential expression of genes (DEGs) (G) were completed using R programs.