UVRAG Deficiency Exacerbates Doxorubicin-Induced Cardiotoxicity

Figures

• 

  Figure 1. UVRAG deficiency exacerbates acute DOX-induced cardiotoxicity. (a) Survival curves of WT and UVRAG-deficient mice after acute DOX treatment were created by Kaplan-Meier method (*P < 0.05 using the Log-rank test). n = 7 mice for saline+/+, n = 8 mice for saline−/−, n = 10 mice for DOX+/+, n = 10 mice for DOX−/−. (b) Representative H&E images of degenerative vacuoles in LVs on heart sections from WT and UVRAG-deficient mice 3 or 5 days after acute DOX or vehicle treatment. Scale bar: 40 μm. (c) Quantification of degenerative vacuoles in LVs in the experiments as illustrated in (b). n = 3 mice for each group. (d) Representative images of fibrosis stained with picrosirius red in LVs on heart sections from WT and UVRAG-deficient mice 3 or 5 days after acute DOX or vehicle treatment. Scale bar: 40 μm. (e) Quantification of fibrosis in LVs in the experiments as illustrated in (d). n = 3 mice for each group. (f) Serum LDH activity in WT and UVRAG-deficient mice 3 days after acute DOX or vehicle treatment. n = 4 mice for each group. (g) Serum CK-MB activity in WT and UVRAG-deficient mice 3 days after acute DOX or vehicle treatment. n = 4 mice for each group. *P < 0.05 vs. WT + vehicle, #P…

• 

  Figure 2. UVRAG deficiency exacerbates chronic DOX-induced cardiotoxicity. (a) Survival curves of WT and UVRAG-deficient mice after chronic DOX treatment were created by Kaplan-Meier method (*P < 0.05 using the Log-rank test). n = 6 mice for saline + / + , n = 6 mice for saline−/−, n = 12 mice for DOX + / + , n = 13 mice for DOX−/−. (b) Representative H&E images of degenerative vacuoles in LVs on heart sections from WT and UVRAG-deficient mice after chronic DOX or vehicle treatment over the indicated time. Scale bar: 40 μm. (c) Quantification of degenerative vacuoles in LVs in the experiments as illustrated in (b). n = 3–5 mice for each group. (d) Representative images of fibrosis stained with picrosirius red in LVs on heart sections from WT and UVRAG-deficient mice after chronic DOX or vehicle treatment over the indicated time. Scale bar: 40 μm. (e) Quantification of fibrosis in LVs in the experiments as illustrated in (d). n = 3–5 mice for each group. (f) Serum LDH activity in WT and UVRAG-deficient mice 2 weeks after chronic DOX or vehicle treatment. n = 4 mice for each group. (g) Serum CK-MB activity in WT and UVRAG-deficient mice 2 weeks after chronic DOX or vehicle treatment. n = 4…

• 

  Figure 3. UVRAG deficiency increases ROS production and apoptosis in DOX-induced cardiotoxicity. (a) ROS levels in LVs detected using DCFH-DA 5 days after acute DOX or vehicle treatment. n = 3 mice for each group. (b) ROS levels in LVs detected using DCFH-DA at 4 weeks of DOX or vehicle treatment in chronic DOX cardiotoxicity. n = 3 mice for each group. (c) Quantification of apoptotic cells 5 days after acute DOX or vehicle treatment. n = 3 mice for each group. (d) Quantification of apoptotic cells at 4 weeks of DOX or vehicle treatment. (e) Western blot analysis of XIAP in LVs from WT and UVRAG-deficient mice 3 days after acute DOX or vehicle treatment. (f) Quantification of XIAP as illustrated in panel (e). (g) Western blot analysis of XIAP in LVs from WT and UVRAG-deficient mice at 4 weeks of chronic DOX treatment or vehicle treatment. (h) Quantification of XIAP as illustrated in panel (g). n = 3 mice for each group. *P < 0.05 vs. WT + vehicle, #P < 0.05 vs. WT + DOX.

• 

  Figure 4. Autophagic flux is impaired in acute and chronic DOX-induced cardiotoxicity. (a) Western blot detection of LC3 II and p62 protein abundance in LVs from mice 5 days after acute DOX treatment and further elicited by CQ treatment. (b) Quantification of LC3-II protein abundance as illustrated in panel (a). n = 3 mice for each group. (c) Quantification of p62 protein abundance as illustrated in panel (a). n = 3 mice for each group. (d) Western blot detection of LC3 II and p62 protein abundance in LVs from mice treated with DOX over 4 weeks and further elicited by CQ treatment. (e) Quantification of LC3-II as illustrated in panel (d). n = 3 mice for each group. (f) Quantification of p62 as illustrated in panel (d). n = 3 mice for each group. *P < 0.05 vs. vehicle. #P < 0.05 vs. CQ.

• 

  Figure 5. UVRAG deficiency aggravates impaired autophagy in DOX-induced cardiotoxicity. (a) Western blot detection of LC3 II and p62 protein levels in LVs from WT and UVRAG-deficient mice at the time points as indicated in acute DOX-treated mice. (b) Quantification of LC3 II protein abundance in the experiments as illustrated in (a). n = 3–5 mice for each group. (c) Quantification of p62 protein abundance in the experiments as illustrated in (a). n = 3–5 mice for each group. (d) Western blot detection of LC3 II and p62 protein levels in LVs from WT and UVRAG-deficient mice at 4 weeks of DOX or vehicle treatment in chronic DOX-treated mice. (e) Quantification of LC3 II protein abundance in the experiments as illustrated in (d). n = 3 mice for each group. (f) Quantification of p62 protein abundance in the experiments as illustrated in (d). n = 3 mice for each group. (g) Representative images of LC3 immunohistochemistry in LVs on heart sections from WT and UVRAG-deficient mice after acute DOX or vehicle treatment over the indicated time. Scale bar: 40 μm. (h) Quantification of LC3-positive dots in LVs in the experiments as illustrated in (g). n = 3 mice for each group. (i) Representative…

• 

  Figure 6. Intermittent fasting restores autophagy in DOX-induced cardiotoxicity. (a) Representative images of LC3 immunohistochemistry in LVs on heart sections from fed or fasted WT and UVRAG-deficient mice 5 days after acute DOX or vehicle treatment. Scale bar: 40 μm. (b) Quantification of LC3-positive dots in LVs in the experiments as illustrated in (a). n = 3 mice for each group. (c) Representative images of LC3 immunohistochemistry in LVs on heart sections from fasted WT and UVRAG-deficient mice at 4 weeks of DOX treatment in chronic cardiotoxicity. Scale bar: 40 μm. (d) Quantification of LC3-positive dots in LVs in the experiments as illustrated in (c). n = 3 mice for each group. (e) Western blot detection of LC3 and p62 in LVs from fed or fasted WT and UVRAG-deficient mice 5 days after acute DOX or vehicle treatment. (f) Quantification of LC3 II protein abundance in the experiments as illustrated in (e). n = 3 mice for each group. (g) Quantification of p62 protein abundance in the experiments as illustrated in (e). n = 3 mice for each group. (h) Western blot analysis of LC3 II and p62 in LVs from fed or fasted WT and UVRAG-deficient mice at 4 weeks of DOX treatment in chronic…

• 

  Figure 7. Intermittent fasting ameliorates pathology of DOX-induced cardiotoxicity. (a) Representative H&E images of degenerative vacuoles in LVs on heart sections from fed or fasted WT and UVRAG-deficient mice 5 days after acute DOX or vehicle treatment. Scale bar: 40 μm. (b) Quantification of degenerative vacuoles in LVs in the experiments as illustrated in (a). n = 3 mice for each group. (c) Representative images of fibrosis stained with picrosirius red in LVs on heart sections from fed or fasted WT and UVRAG-deficient mice 5 days after acute DOX or vehicle treatment. Scale bar: 40 μm. (d) Quantification of fibrosis in LVs in the experiments as illustrated in (c). n = 3 mice for each group. (e) Representative H&E images of LVs on heart sections from fed or fasted WT and UVRAG-deficient mice at 4 weeks of DOX treatment in chronic cardiotoxicity. Scale bar: 40 μm. (f) Quantification of degenerative vacuoles in LVs in the experiments as illustrated in (e). n = 3 mice for each group. (g) Representative images of fibrosis stained with picrosirius red in LVs on heart sections from fed or fasted WT and UVRAG-deficient mice at 4 weeks of DOX treatment in chronic cardiotoxicity. (h)…

• 

  Figure 8. Intermittent fasting attenuates enhanced ROS levels and increased apoptotic cell death in UVRAG-deficient mice treated by DOX. (a) Intermittent fasting rescued enhanced ROS levels in LVs from UVRAG-deficient mice measured 5 days after acute DOX treatment. n = 3 mice for each group. (b) Intermittent fasting attenuated increased apoptotic cell death in the hearts from UVRAG-deficient mice assessed 5 days after acute DOX treatment. n = 3 mice for each group. (c) Intermittent fasting rescued enhanced ROS levels in LVs from UVRAG-deficient mice at 4 weeks of DOX treatment in chronic DOX cardiotoxicity. n = 3 mice for each group. (d) Intermittent fasting attenuated increased apoptotic cell death in the hearts from UVRAG-deficient mice at 4 weeks of DOX treatment in chronic cardiotoxicity. n = 3 mice for each group. *P < 0.05 vs. WT + DOX + Fed, #P < 0.05 vs. UVRAG−/− + DOX + Fed, §P < 0.05 vs. WT + DOX + Fasted.