β-Adrenergic receptor (β-AR) stimulation increases extracellular ubiquitin (UB) levels, and extracellular UB inhibits β-AR-stimulated apoptosis in adult cardiac myocytes. This study investigates the role of exogenous UB in chronic β-AR-stimulated myocardial remodeling. l-Isoproterenol (ISO; 400 μg·kg(-1)·h(-1)) was infused in mice in the presence or absence of UB (1 μg·g(-1)·h(-1)). Left ventricular (LV) structural and functional remodeling was studied 7 days after infusion. UB infusion enhanced serum UB levels. In most parts, UB alone had no effect on morphometric or functional parameters. Heart weight-to-body weight ratios were increased to a similar extent in the ISO and UB + ISO groups. Echocardiographic analyses showed increased percent fractional shortening, ejection fraction, and LV circumferential stress and fiber-shortening velocity in the ISO group. These parameters were significantly lower in UB + ISO vs. ISO. Isovolumic contraction and relaxation times and ejection time were significantly lower in ISO vs. UB + ISO. The increase in the number of TUNEL-positive myocytes and fibrosis was significantly higher in ISO vs. UB + ISO. Activation of Akt was higher, whereas activation of GSK-3β and JNKs was lower in UB + ISO vs ISO. Expression of MMP-2, MMP-9, and TIMP-2 was higher in UB + ISO vs ISO. In isolated cardiac fibroblasts, UB enhanced expression of MMP-2 and TIMP-2 in the presence of ISO. Neutralizing UB antibodies negated the effects of UB on MMP-2 expression, whereas recombinant UB enhanced MMP-2 expression. UB activated Akt, and inhibition of Akt inhibited UB + ISO-mediated increases in MMP-2 expression. Thus, exogenous UB plays an important role in β-AR-stimulated myocardial remodeling with effects on LV function, fibrosis, and myocyte apoptosis.