CBL knockdown protects cardiomyocytes against hypoxia-reoxygenation injury by downregulating GRB2 expression

Area of Science:

• Cardiovascular Biology
• Molecular Medicine
• Cellular Physiology

Background:

• Myocardial ischemia-reperfusion injury (MIRI) is a critical complication following myocardial infarction and interventional surgery.
• The precise molecular mechanisms underlying MIRI pathogenesis remain incompletely understood, necessitating the identification of novel therapeutic targets.
• Understanding cellular responses to hypoxia and reoxygenation is crucial for developing effective MIRI treatments.

Purpose of the Study:

• To investigate novel molecular targets for MIRI treatment.
• To elucidate the underlying mechanism of action involving Cbl proto-oncogene (CBL) and growth factor receptor-bound protein 2 (GRB2) in MIRI.
• To explore the role of CBL in regulating cardiomyocyte proliferation, oxidative stress, and apoptosis under hypoxic conditions.

Main Methods:

• Quantitative PCR and western blotting to assess CBL expression in patient samples and an in vitro MIRI model (H9c2 cells subjected to hypoxia/reoxygenation).
• Cell Counting Kit-8, western blotting, and TUNEL assay to evaluate proliferation, oxidative stress, and apoptosis in H/R-induced cells.
• Co-immunoprecipitation (Co-IP) assay to confirm the interaction between CBL and GRB2; GRB2 overexpression studies to assess its role.

Main Results:

• CBL expression was significantly upregulated in patients with ischemic cardiomyopathy and in H/R-induced H9c2 cells.
• Genetic silencing of CBL using small interfering RNA (siRNA) promoted proliferation and oxidative stress while inhibiting apoptosis in H/R-induced cells.
• Overexpression of GRB2 reversed the protective effects of CBL knockdown, indicating GRB2 acts downstream of CBL in mediating these cellular responses.

Conclusions:

• CBL knockdown promotes cardiomyocyte proliferation and antioxidant capacity while inhibiting apoptosis under hypoxic conditions, primarily by downregulating GRB2 expression.
• The CBL/GRB2 signaling pathway represents a potential therapeutic target for preventing or treating myocardial ischemia-reperfusion injury.
• These findings provide novel insights into the molecular mechanisms governing MIRI, paving the way for targeted therapeutic interventions.