Doxorubicin induced differential response of lysosome-associated proteins in human cardiomyocytes
Abstract Body: Background Doxorubicin (Dox) is an effective anticancer drug, but its use is limited by dose-dependent cardiotoxicity involving oxidative stress, cell death, and lysosomal dysfunction in cardiomyocytes. Although lysosomal damage has been linked, the specific changes in lysosome-associated proteins in cardiomyocytes are poorly understood. We examined key lysosomal proteins to identify molecular intermediates and therapeutic targets of doxorubicin-induced lysosomal dysfunction.
Methods Human AC16 cardiomyocytes were treated with 1 µM doxorubicin or solvent for 16 hours. To evaluate changes in expression levels and post-translational modifications of lysosomal proteins caused by DOX treatment, we conducted Western blot analysis and assessed band mobility as an indicator of post-translational modification.
Results No significant changes were observed in LAMP1, LIMP2, or GCase, indicating preserved lysosomal membrane markers, lipid turnover, and membrane homeostasis. SMURF1, an E3 ubiquitin ligase promoting lysophagy, was decreased, suggesting impaired clearance of damaged lysosomes. Mucolipin-1, a lysosomal Ca+2 channel facilitating lysosome-autophagosome fusion, was increased, indicating compensatory lysosomal trafficking and fusion activation. ATP6V1A decreased slightly, potentially impairing lysosomal acidification. TFEB, a key transcription factor for lysosomal and autophagy genes, showed reduced expression and faster migration, implying post-translational modifications affecting activity and localization. CD63 levels remained stable but bands shifted higher, indicating altered maturation impacting vesicular trafficking. CTSD levels increased, suggesting redistribution rather than functional upregulation, consistent with lysosomal loss of CTSD after damage.
Conclusion Doxorubicin differentially regulates lysosome-associated proteins in cardiomyocytes. These findings suggest that DOX disrupts lysophagy, lysosomal signaling, membrane transport, and protease processing rather than causing a uniform loss of lysosomes. More comprehensive validation of these changes is warranted to identify a set of therapeutic targets for mitigating doxorubicin-induced cardiotoxicity.
Chin, Jack
(
New York Institute of Technology College of Osteopathic Medicine
, New York City , New York , United States )
Martinez, Sabrina
(
New York Institute of Technology
, Old Westbury , New York , United States )
Kobayashi, Tamayo
(
New York Institute of Technology College of Osteopathic Medicine
, New York City , New York , United States )
Liang, Qiangrong
(
New York Institute of Technology College of Osteopathic Medicine
, New York City , New York , United States )
Kobayashi, Satoru
(
New York Institute of Technology College of Osteopathic Medicine
, New York City , New York , United States )
