re 5a), a trend circumstances inin untreated animals (Figure5a), a trend that was also seen for Claudin-2 exwas also noticed for Claudin-2 pression (Figure 5d, p 0.05). Nevertheless, overall, in our in vivo the Selenof Selenof expression (Figure 5d, p 0.05). However, general, in our in vivo model,model, the genotype showed showed little to no effect on mRNA expression of tight junction proteins genotype little to no impact on mRNA expression of tight junction proteins Claudin-1 (Cldn-1), 2 (Cldn-2) and 15 (Cldn-15). Western blot analyses Western blot analyses claudin-2 overClaudin-1 (Cldn-1), 2 (Cldn-2) and 15 (Cldn-15). showed low expression ofshowed low all, and no visible variations in protein visible variations in protein expression for expression of claudin-2 overall, and no expression for Claudin-1 or Claudin-3 (Figure 5g) or Claudin-2 (Figure (Figure 5g) WT and KO (Figure 5h) amongst WT and KO mice. It Claudin-1 or Claudin-35h) involving or Claudin-2mice. It should be noted that mRNA expression be noted that mRNA expression of those tight junction genes in AOM/DSS-treated should of those tight junction genes in AOM/DSS-treated animals, interestingly, showed a positiveinterestingly, showed a positivewith important influence on expression of with animals, correlation with dietary selenium, correlation with dietary selenium, Cldn-2 (p = 0.0016) and on expression of Cldn-2 (p = 0.0016) and PDE3 custom synthesis Cldn-15 (p = 0.0008). significant impact Cldn-15 (p = 0.0008). In addition to tight junction genes, we also evaluated the mRNA expression of genes normally related with adherens junctions and also other barrier integrity functions in control animals’ colon scrapes and in colon tumor tissues (Figure S8). Dietary selenium levels appeared to have an effect on mRNA expression with the transmembrane glycoprotein epithelial cell adhesion molecule (EpCAM), Nectin cell adhesion molecule (Nectin)-2, membrane-associated carbonic anhydrase four (Car4), plus the secreted glycoprotein mucin two (Muc2) in either WT or KO mice, or both. Interestingly, Selenof -genotype didn’t look to drastically affect mRNA expression of your α1β1 Accession investigated genes in colons of mice, except for Epcam, which was substantially decrease in tumors of Selenof-KO mice compared to WT mice, but only at higher selenium levels. Having said that, even though gene expression of tight junction and adherens junction genes weren’t considerably altered among Selenof-KO mice and their WT littermates, the drastically elevated size of goblet cells in KO mice recommend structural changes relevant to colon tumorigenesis.Int. J. Mol. Sci. 2021, 22, 10651 Int. J. Mol. Sci. 2021, 221,10 of 19 10 ofFigure five. Expression of tight junction Claudin genes. mRNA expression was measured with qPCR Figure 5. Expression of tight junction Claudin genes. mRNA expression was measured with qPCR in (a,c,e) colon scrapes of control mice and (b,d,f) colon tumors ofof AOM/DSS-treated mice. Mean in (a,c,e) colon scrapes of manage mice and (b,d,f) colon tumors AOM/DSS-treated mice. Mean (N = 4) + SEM, 2-way ANOVA, followed by Tukey’s post hoc analyses to examine KO vs. WT by diet; (N = four) + SEM, 2-way ANOVA, followed by Tukey’s post hoc analyses to evaluate KO vs. WT by diet plan; letters indicate statistically substantial variations. Protein expression of (g) Claudin-1 and Claudinletters indicate statistically substantial differences. Protein expression of (g) Claudin-1 and Claudin-3, three, and (h) Claudin-2 in colon scrapes of handle mice on selenium-specific diets was asse
