Fig 1: HSF1 activity was regulated by substrate stiffness(A) The illustration of the experiment device for substrate stiffness stimuli to cultured cells.(B) Fluorescent immunostaining of phosphorylated HSF1 at Serine 326 in MDCK cells under substrate stiffness stimuli. Scale bar: 10 μm.(C) The ratio of phosphorylated HSF1 at Serine 326 in nuclei and cytoplasm (N/C) in (B) (n = 9). Data are expressed as mean ± SEM, ∗p < 0.05, ∗∗p < 0.01.(D–I) Fluorescence intensity of phosphorylated HSF1 at Serine 326 and DAPI along the lines in the magnified figures in (B).
Fig 2: Immediate activation of HSF1 by fluid shear stress(A) The illustration of the flow chamber device for fluid shear stress experiments.(B) Western blot results of phosphorylated HSF1 at Serine 326 or 303 in MDCK cells after shear stress stimuli for different time.(C) Statistical analysis of the results in (B) (n = 3). The ratio of phosphorylated HSF1 at Serine 326 in nuclei and cytoplasm (N/C) in (B) (n = 10 ). Data are expressed as mean ± SEM, p<0.05.(D) Luciferase reporter assay for HSE activity in MDCK cells treated with shear stress for 1 h. (n = 3). Data are expressed as mean ± SEM, p < 0.05.(E) Fluorescent immunostaining of phosphorylated HSF1 at Serine 326 in MDCK cells 30 min after shear stress. Scale bar: 25 μm.(F) The ratio of phosphorylated HSF1 at Serine 326 in nuclei and cytoplasm (N/C) in (E). Data are expressed as mean ± SEM, p < 0.01.
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