CaMKII orchestrates endoplasmic reticulum stress and apoptosis in doxorubicin‐induced cardiotoxicity by regulating the IRE1α/XBP1s pathway

Figures

• 

  FIGURE 1. CaMKII activity and the expression of IRE1α/XBP1s in Dox‐treated mouse hearts. (A) Schematic representation of the experimental procedure. (B) Representative immunoblots showing p‐CaMKII, CaMKII, p‐PLB, PLB, IRE1α, XBP1s and GAPDH (internal control). (C–F) Statistical analysis of p‐CaMKII/CaMKII, p‐PLB/PLB, IRE1α and XBP1s, each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. **p < 0.01 versus Control.

• 

  FIGURE 2. Effects of CaMKII inhibition with KN‐93 on cardiac function and structure in Dox‐treated mouse hearts. (A) Representative images of echocardiograms of hearts. (B–F) Group results of EF, FS, heart rate, LVIDd and LVPWd. (G) The left panel shows images with HE staining; the right panel shows images with Masson's staining (Scale bar: 50 μm). (H) Quantitative analysis of collagen volume. (I) LDH activity. (J) Survival curves. The data are presented as the mean ± SEM, n = 6. **p < 0.01 versus Control, ## p < 0.01 versus Dox.

• 

  FIGURE 3. Effects of CaMKII inhibition with KN‐93 on ER stress and apoptosis in Dox‐treated mouse hearts. (A) Representative electron microscope images of the ER in the different groups (magnification: 15,000×). Arrows indicate ER. (B) Quantification of ER luminal width. (C) Representative TUNEL staining images in the different groups (Scale bar: 50 μm). (D) Apoptotic rate. (E) Representative immunoblots of PERK, GRP78, caspase‐3, cleaved caspase‐3 and GAPDH (internal control). (F–I) Statistical analysis of PERK, GRP78, caspase‐3, cleaved caspase‐3 and cleaved caspase‐3/caspase‐3; each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. **p < 0.01 versus Control, ## p < 0.01 versus Dox.

• 

  FIGURE 4. Effects of CaMKII inhibition with KN‐93 on IRE1α/XBP1s in Dox‐treated mouse hearts. (A) Representative immunoblots of p‐CaMKII, CaMKII, p‐PLB, PLB, IRE1α, XBP1s and GAPDH (internal control). (B–E) Statistical analysis of p‐CaMKII/CaMKII, p‐PLB/PLB, IRE1α and XBP1s. each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. **p < 0.01 versus Control, ## p < 0.01 versus Dox.

• 

  FIGURE 5. 4μ8C inhibited the effects of CaMKII inhibition on cardiac function and structure in Dox‐treated mouse hearts. (A) Representative images of echocardiograms of the hearts. (B–F) EF, FS, heart rate, LVIDd and LVPWd. (G) The left panel shows images with HE staining. The right panel shows images with Masson's staining (Scale bar: 50 μm). (H) Quantitative analysis of collagen volume. (I) LDH activity. (J) Survival curves. The data are presented as the mean ± SEM, n = 6. ## p < 0.01 versus Dox, ^^ p < 0.01 versus KN‐93 + Dox.

• 

  FIGURE 6. 4μ8C prevented the effects of CaMKII inhibition on ER stress and apoptosis in Dox‐treated mouse hearts. (A) Representative electron microscope images of the ER in the different groups (magnification: 15000×). Arrows indicate ER. (B) Quantification of ER luminal width. (C) Representative TUNEL staining images in the different groups (Scale bar: 50 μm). (D) Apoptotic rate. (E) Representative immunoblots of PERK, GRP78, caspase‐3, cleaved caspase‐3 and GAPDH (internal control). (F–I) Statistical analysis of PERK, GRP78, caspase‐3, cleaved caspase‐3 and cleaved caspase‐3/caspase‐3; each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. ## p < 0.01 versus Dox. ^^ p < 0.01 versus KN‐93 + Dox.

• 

  FIGURE 7. 4μ8C blocked the effects of CaMKII inhibition on IRE1α/XBP1s in Dox‐treated mouse hearts and rhADAMTS13 affects CaMKII activation in Dox‐treated AC16 cardiomyocytes. (A) Representative immunoblots of p‐CaMKII, CaMKII, p‐PLB, PLB, IRE1α, XBP1s and GAPDH (internal control). (B–E) Statistical analysis of p‐CaMKII/CaMKII, p‐PLB/PLB, IRE1α and XBP1s. Each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. ## p < 0.01 versus Dox, ^^ p < 0.01 versus KN‐93 + Dox. (F) Representative immunoblots of ADAMTS13, TSP1 and GAPDH (internal control). (G and H) Statistical analysis of ADAMTS13 and TSP1. Each protein was normalized to GAPDH. (I) Representative immunoblots of ADAMTS13, TSP1, p‐CaMKII, CaMKII and GAPDH (internal control). (J–L) Statistical analysis of ADAMTS13, TSP1 and p‐CaMKII/CaMKII. Each protein was normalized to GAPDH. The data are presented as the mean ± SEM, n = 6. **p < 0.01 versus Control, ## p < 0.01 versus Dox.