Fig 2: IT-dependent induced p-HSPB1 might play a protective role against AApoAII amyloidosis. (A) Western blot analysis and quantification of phosphorylated p38 MAPK (Thr180/Tyr182), total p38 MAPK, p-HSPB1 (Ser82) and total HSPB1 protein in the livers of mice after 16-week interventions. Box and whisker plots show fold changes relative to the VS group. The boxes represent the 25th, median (black line), mean (cross) and 75th percentiles of the data; the whiskers represent the lowest (or highest) data within 1.5 interquartile range from the 25th (or 75th) percentile range. (B) Representative images of IHC for p-HSPB1 and Congo Red birefringence of amyloid deposition in the livers of mice after 16-week interventions. p-HSPB1 and amyloid deposition are indicated by orange and gray arrows, respectively. Scale bars: 50 μm. Data are mean±s.d. (VS, n=7; VI, n=8; FS, n=8; FI, n=8). #P<0.05, ##P<0.01, ###P<0.001 (one-way ANOVA with Tukey–Kramer method for comparison between groups). !!!P<0.001 (two-way ANOVA for comparison of the magnitude of changes between different groups in mice with or without amyloidosis induction).

Fig 3: p53 signaling pathway upregulated Hspb1 levels in mice with IT. (A) Western blot analysis and quantification of TP53 protein in the livers of mice after 16-week interventions. Box and whisker plots show fold changes relative to the VS group. The boxes represent the 25th, median (black line), mean (cross) and 75th percentiles of the data; the whiskers represent the lowest (or highest) data within 1.5 interquartile range from the 25th (or 75th) percentile range. (B) Immunofluorescence determination of TP53 in the liver after 16-week interventions. Scale bars: 50 μm. (C) Two-tailed Pearson's correlation analysis between TP53 and HSPB1 proteins in the livers of mice after 16-week interventions (31 female mice). (D) Predictive analyses coupled with database-deposited data (JASPAR open-access database) suggested that TP53 binds the promoter of the Hspb1 gene. Data are mean±s.d. (VS, n=7; VI, n=8; FS, n=8; FI, n=8). ##P<0.01; ###P<0.001 (one-way ANOVA with Tukey–Kramer method for comparison between groups). !!!P<0.001 (two-way ANOVA for comparison of the magnitude of changes between different groups in mice with or without amyloidosis induction).

Fig 4: A potential molecular mechanism underlying IT exhibits an anti-amyloid effect. Here, we propose a novel molecular mechanism in which the p38 MAPK-p53-HSPB1 axis could play a key role in the anti-amyloid load. The p38 MAPK-p53-HSPB1 axis showed upregulation in the livers of mice with exercise (top left) or systemic AApoAII amyloidosis alone (top right). In particular, phosphorylated HSPB1 showed an exercise-dependent increase. Exercise additively enhanced this phenomenon in the amyloid-reduced liver and greatly increased levels of phosphorylated HSPB1 outside the cell. Phosphorylated HSPB1 colocalized with amyloid depositions, which in turn could inhibit fibril formation (bottom).