Article Title: Cardiac troponin I autoantibody induces myocardial dysfunction by PTEN signaling activation
Figure Lengend Snippet: cTnImAb1 treatment increases ENO1 expression followed by activation of PTEN and suppression of Akt signaling in mouse hearts. a. Heat map from PathScan® Akt Signaling Antibody Array kit analysis showing changes in 16 phosphorylated proteins. b & c. Statistical analysis of heat map data in a and volcano plot showing that cTnImAb1 treatment significantly increased PTEN Ser380 phosphorylation in the mouse heart. 1. PTEN Ser380, *p = .023, ( t -test). 2. GSK-3b Ser9, * p = .038, ( t -test). 3. PTEN Ser380, * p = .031 ( t -test). d&e. Immunoblot analysis showed that cTnImAb1 treatment significantly increased ENO1 expression and PTEN Ser380 phosphorylation but suppressed Akt Thr308 phosphorylation. β-actin served as a loading control. e is the statistical analysis of d . ENO1 * p = .002, pPTEN/PTEN/Actin, * p = .003, pAkt/Akt/Actin, * p = .003. (ANOVA analysis). f-h. cTnImAb1 treatment (50 μg/ml) induced changes in ENO1 expression and phosphorylation of PTEN Ser380 and Akt Thr308 at different time points in cultured cardiomyoyctes. cTnImAb1 increased the p-Ser380-PTEN/total PTEN ratio at 8 h but decreased the pThr308-AKT/total Akt ratio at 8–12 h. Mouse IgG treatment did not alter pSer380 PTEN but significantly increased the pThr308-Akt/total Akt ratio at 24–36 h. f shows representative blot images of three independent experiments. g and h represent ImageJ-based densitometry analysis of relative changes in p-Ser380-PTEN/total PTEN ratio ( G, 8 h * p = .020, 12 h * p = .024, t -test) and pThr308-AKT/total Akt ( H , 12 h * p = .018, 24 h * p = .001, 36 h * p = .001, t -test). i-l. Knockdown of ENO1 significantly suppressed cTnImAb1–induced alteration in PTEN/Akt signaling. Cultured cardiomyocytes were transfected with siRNA targeting Eno1 and then subjected to cTnImAb1 treatment (50 μg/ml) for 4–48 h as indicated. Mock-siRNA was used as a control. Depletion of Eno1 suppressed the p-PTEN Ser380/total PTEN ratio and increased the pAkt-Thr308/total Akt ratio at 4, 8, and 12 h, respectively. i shows representative blot images of three independent experiments. j represents ImageJ-based densitometry analysis of relative changes in ENO1 protein levels ( J ,48 h * p = .000, t -test). k and l represent ImageJ-based densitometry analysis of relative changes in p-Ser380-PTEN/total PTEN ratio ( k, 48 h * p = .000, t -test) and pThr308-AKT/total Akt ( l, 6 h * p = .000, 12 h * p = .000, t -test).
Article Snippet: Mouse cardiac tissue total protein Akt signaling was screened using the PathScan® Akt Signaling Antibody Array Kit (Cell Signal, 9474, USA).
Techniques: Expressing, Activation Assay, Ab Array, Western Blot, Cell Culture, Transfection