Osteopontin stimulates cell cycle in neonatal cardiomyocytes and improves infarct repair

Introduction: Osteopontin (OPN) is a macrophage-derived protein which is upregulated following tissue injury. While OPN regulates cell adhesion, spreading and migration, its role in myocardial regeneration is unknown.

Objective: To determine the role of OPN in heart regeneration and repair.

Methods and results: Isolated neonatal cardiomyocytes were treated with a serum-free medium for 24 hours. Immunofluorescent staining showed that these cells expressed the OPN receptor CD44. MTT colorimetric assays showed that OPN preserved the number of cardiomyocytes. Staining for propidium iodide, a marker for apoptosis and necrosis, showed that OPN does not affect cell survival. Staining for phosphohistone-3 (pH3), a marker of nucleus division, showed that OPN increased pH3 expression in cardiomyocytes and non-cardiomyocytes. Immunoblotting revealed that OPN induced the phosphorylation of cell-cycle activity proteins, known as yes-associated protein (YAP)1, and extracellular signal-regulated kinase (ERK)2, while decreasing the phosphorylation of the large tumor-suppressor kinase (LATS)1/2, a YAP1 inhibitor. Gene expression analysis revealed that OPN upregulated the transcriptional enhancer factor TEF-1 (TEAD1), which interacts with YAP1, and the connective tissue growth factor (CTGF), which is a downstream target of YAP1. Furthermore, OPN significantly upregulated cyclin B1 and cyclin-dependent kinase 1 (CDK1) both regulators of the mitotic (M) phase. Finally, echocardiography and postmortem histology showed that a single injection of OPN to adult mice improved cardiac remodeling and function after myocardial infarction (MI).

Conclusions: OPN stimulates cell-cycle activity in neonatal cardiomyocytes and improves infarct repair. Our findings suggest a new therapy for heart repair after MI.