Background -Heart failure (HF) is a leading cause of mortality and morbidity, and the search for novel therapeutic approaches continues. In the monogenic disease mucopolysaccharidosis (MPS) VI, loss of function mutations in arylsulfatase B (ASB) leads to myocardial accumulation of chondroitin sulfate (CS) glycosaminoglycans (GAGs), manifesting as a myriad of cardiac symptoms. Here, we studied changes in myocardial CS in non-MPS failing hearts, and assessed its generic role in pathological cardiac remodeling. Methods -Healthy and diseased human and rat left ventricles were subjected to histological and immuno-staining methods to analyze for GAG distribution. GAGs were extracted and analyzed for quantitative and compositional changes using Alcian Blue assay and liquid chromatography mass spectrometry. Expression changes in 20 CS-related genes were studied in three primary human cardiac cell types and THP-1 derived macrophages under each of 9 in vitro stimulatory conditions. In two rat models of pathological remodeling induced by transverse aortic constriction (TAC) or isoprenaline infusion, recombinant human arylsulfatase B (rhASB), clinically used as enzyme replacement therapy (ERT) in MPS VI, was administered intravenously for 7 or 5 weeks respectively. Cardiac function, myocardial fibrosis and inflammation were assessed by echocardiography and histology. CS-interacting molecules were assessed using surface plasmon resonance and a mechanism of action was verified in vitroResults -Failing human hearts displayed significant perivascular and interstitial CS accumulation, particularly in regions of intense fibrosis. Relative composition of CS disaccharides remained unchanged. Transforming growth factor β (TGFβ) induced CS upregulation in cardiac fibroblasts. CS accumulation was also observed in both the pressure-overload and the isoprenaline models of pathological remodeling in rats. Early treatment with rhASB in the TAC model, and delayed treatment in the isoprenaline model, proved rhASB to be effective at preventing cardiac deterioration and augmenting functional recovery. Functional improvement was accompanied by reduced myocardial inflammation and overall fibrosis. Tumor necrosis factor α (TNFα) was identified as a direct binding partner of CS GAG chains, and rhASB reduced TNFα-induced inflammatory gene activation in vitro in endothelial cells and macrophages. Conclusions -CS GAGs accumulate during cardiac pathological remodeling, and mediate myocardial inflammation and fibrosis. RhASB targets CS effectively as a novel therapeutic approach for the treatment of heart failure.