Research Paper Volume 11, Issue 7 pp 2003—2019

Genistein antagonizes gliadin-induced CFTR malfunction in models of celiac disease

Figure 3. Genistein prevents P31-43 induced Beclin 1 downregulation and plasma membrane CFTR disposal. (A-D) Treatment of Caco-2 cells with P57-68 or P31-43 (3h) or with P31-43 after the optional pre-treatment (20 min) with genistein or VX-770. (A) Immunoblot of total lysate with anti-Beclin 1 antibody (left) and densitometric analysis of protein levels relative to β-actin (right) (n=3 independent experiments). Means±SD of triplicates of independent experiments; ***p<0.001 untreated or P57-68 challenged cells versus P31-43 treatment, °°p<0.01 P31-43 versus genistein+P31-43 or ##p<0.01 P31-43 versus VX-770+P31-43 (ANOVA, Bonferroni post-hoc test). (B) Immunoblot of membrane protein fractions with anti-SQSTM1 Ab and anti-flotillin as a control (left), and relative densitometric analysis of immunoblot (right) (n=3 independent experiments). Means±SD of triplicates of independent experiments; ***p<0.001 untreated or P57-68 versus P31-43 treatment, °°p<0.01 P31-43 versus genistein+P31-43 or ##p<0.01 P31-43 versus VX-770+P31-43 (ANOVA, Bonferroni post-hoc test). (C-D) Immunoblot of membrane protein fractions with anti-CFTR (C) or Ezrin (D) and anti-flotillin as a control (left) and relative densitometric analysis of immunoblot (right) (n=3 independent experiments). Means±SD of triplicates of independent experiments; **p<0.01 or ***p<0.001 untreated or P57-68 challenged cells versus P31-43 treatment, °°p<0.01 P31-43 versus genistein+P31-43 (ANOVA, Bonferroni post-hoc test). Data information: The blots are representative of one experiment for group of treatment.