Stabilin-1 mediates beneficial monocyte recruitment and tolerogenic macrophage programming during CVB3-induced viral myocarditis.

Pathological innate and adaptive immune response upon viral infection may lead to cardiac injury and dysfunction. Stabilin-1 is a scavenger receptor that regulates several aspects of the innate immunity. Whether stabilin-1 affects the inflammatory response during viral myocarditis (VM) is entirely unknown. Here, we assess the role of stabilin-1 in the pathogenesis of VM and its suitability as a therapeutic target. Genetic loss of stabilin-1 increased mortality and cardiac necrosis in a mouse model of human Coxsackievirus B3 (CVB3)-induced myocarditis. Absence of stabilin-1 significantly reduced monocyte recruitment and strongly reduced the number of alternatively activated anti-inflammatory macrophages in the heart, enhancing a pro-inflammatory cardiac niche with a detrimental T lymphocyte response during VM. Yeast two-hybrid screening, confirmed by affinity chromatography, identified fibronectin as a stabilin-1 interacting partner. Absence of stabilin-1 specifically decreased monocyte adhesion on extracellular fibronectin in vitro. Loss of Type III repeats Extra Domain A (EDA) of fibronectin during VM also increased the mortality and cardiac necrosis as in stabilin-1 knockout mice, with reduced monocytic cardiac recruitment and increased T lymphocyte response. Collectively, stabilin-1 has an immune-suppressive role of limiting myocardial damage during VM, regulating anti-inflammatory monocyte-recruitment to the site of inflammation.

Extracellular SPARC increases cardiomyocyte contraction during health and disease.

Secreted protein acidic and rich in cysteine (SPARC) is a non-structural extracellular matrix protein that regulates interactions between the matrix and neighboring cells. In the cardiovascular system, it is expressed by cardiac fibroblasts, endothelial cells, and at lower levels by ventricular cardiomyocytes. SPARC expression levels are increased upon myocardial injury and also during hypertrophy and fibrosis. We have previously shown that SPARC improves cardiac function after myocardial infarction by regulating post-synthetic procollagen processing, however whether SPARC directly affects cardiomyocyte contraction is still unknown. In this study we demonstrate a novel inotropic function for extracellular SPARC in the healthy heart as well as in the diseased state after myocarditis-induced cardiac dysfunction. We demonstrate SPARC presence on the cardiomyocyte membrane where it is co-localized with the integrin-beta1 and the integrin-linked kinase. Moreover, extracellular SPARC directly increases cardiomyocyte cell shortening ex vivo and cardiac function in vivo, both in healthy myocardium and during coxsackie virus-induced cardiac dysfunction. In conclusion, we demonstrate a novel inotropic function for SPARC in the heart, with a potential therapeutic application when myocyte contractile function is diminished such as that caused by a myocarditis-related cardiac injury.

Osteoglycin prevents the development of age-related diastolic dysfunction during pressure overload by reducing cardiac fibrosis and inflammation.

The small leucine-rich proteoglycan osteoglycin has been implicated in matrix homeostasis in different organs, including the ischemic heart. However, whether osteoglycin modulates cardiac hypertrophy, fibrosis or inflammation in hypertensive heart disease and during aging remains unknown. Angiotensin-II-induced pressure overload increases cardiac osteoglycin expression, concomitant with the onset of inflammation and extracellular matrix deposition. Interestingly aging led to decreased cardiac levels of osteoglycin, yet absence of osteoglycin did not affect organ structure or cardiac function up to the age of 18months. However, Angiotensin-II infusion in combination with aging resulted in exaggerated cardiac fibrosis and inflammation in the osteoglycin null mice as compared to wild-type mice, resulting in increased diastolic dysfunction as determined by magnetic resonance imaging. In vitro, stimulation of bone marrow derived macrophages from osteoglycin null mice with Angiotensin-II resulted in significantly higher levels of ICAM-1 as well as pro-inflammatory cytokines and chemokines IL-1ß and MCP-1 as compared to WT cells. Further, stimulation of human cardiac fibroblasts with osteoglycin reduced cell proliferation and inhibited TGF-ß induced collagen gene expression. In mouse cardiac tissue, osteoglycin expression inversely correlated with TGF-ß expression and in cardiac biopsies of aortic stenosis patients, osteoglycin expression is significantly higher than in control biopsies. Interestingly, osteoglycin levels were higher in patients with less severe myocardial fibrosis and overall in the aortic stenosis patients osteoglycin levels negatively correlated with collagen content in the myocardium. In conclusion, osteoglycin expression is increased in the heart in response to pressure overload and its absence results in increased cardiac inflammation and fibrosis resulting in increased diastolic dysfunction.

Inhibition of MicroRNA-146a and Overexpression of Its Target Dihydrolipoyl Succinyltransferase Protect Against Pressure Overload-Induced Cardiac Hypertrophy and Dysfunction.

BACKGROUND: Cardiovascular diseases remain the predominant cause of death worldwide, with the prevalence of heart failure continuing to increase. Despite increased knowledge of the metabolic alterations that occur in heart failure, novel therapies to treat the observed metabolic disturbances are still lacking. METHODS: Mice were subjected to pressure overload by means of angiotensin-II infusion or transversal aortic constriction. MicroRNA-146a was either genetically or pharmacologically knocked out or genetically overexpressed in cardiomyocytes. Furthermore, overexpression of dihydrolipoyl succinyltransferase (DLST) in the murine heart was performed by means of an adeno-associated virus. RESULTS: MicroRNA-146a was upregulated in whole heart tissue in multiple murine pressure overload models. Also, microRNA-146a levels were moderately increased in left ventricular biopsies of patients with aortic stenosis. Overexpression of microRNA-146a in cardiomyocytes provoked cardiac hypertrophy and left ventricular dysfunction in vivo, whereas genetic knockdown or pharmacological blockade of microRNA-146a blunted the hypertrophic response and attenuated cardiac dysfunction in vivo. Mechanistically, microRNA-146a reduced its target DLST-the E2 subcomponent of the α-ketoglutarate dehydrogenase complex, a rate-controlling tricarboxylic acid cycle enzyme. DLST protein levels significantly decreased on pressure overload in wild-type mice, paralleling a decreased oxidative metabolism, whereas DLST protein levels and hence oxidative metabolism were partially maintained in microRNA-146a knockout mice. Moreover, overexpression of DLST in wild-type mice protected against cardiac hypertrophy and dysfunction in vivo. CONCLUSIONS: Altogether we show that the microRNA-146a and its target DLST are important metabolic players in left ventricular dysfunction.

The diverse functions of osteoglycin: a deceitful dwarf, or a master regulator of disease?

Small leucine-rich proteoglycans are emerging as important regulatory proteins within the extracellular matrix, where they exert both structural and nonstructural functions and hence are modulators of numerous biological processes, such as inflammation, fibrosis, and cell proliferation. One proteoglycan in particular, osteoglycin (OGN), also known as mimecan, shows great structural and functional diversity in normal physiology and in disease states, therefore making it a very interesting candidate for the development of novel therapeutic strategies. Unfortunately, the literature on OGN is confusing, as it has different names, and different transcript and protein variants have been identified. This review will give a clear overview of the different structures and functions of OGN that have been identified to date, portray its central role in pathophysiology, and highlight the importance of posttranslational processing, such as glycosylation, for the diversity of its functions.-Deckx, S., Heymans, S., Papageorgiou, A.-P. The diverse functions of osteoglycin: a deceitful dwarf, or a master regulator of disease?

Breeding Strategy Determines Rupture Incidence in Post-Infarct Healing WARPing Cardiovascular Research.

BACKGROUND: Von Willebrand A domain Related Protein (WARP), is a recently identified extracellular matrix protein. Based upon its involvement in matrix biology and its expression in the heart, we hypothesized that WARP regulates cardiac remodeling processes in the post-infarct healing process. METHODS AND RESULTS: In the mouse model of myocardial infarction (MI), WARP expression increased in the infarcted area 3-days post-MI. In the healthy myocardium WARP localized with perlecan in the basement membrane, which was disrupted upon injury. In vitro studies showed high expression of WARP by cardiac fibroblasts, which further increases upon TGFß stimulation. Furthermore, WARP expression correlated with aSMA and COL1 expression, markers of fibroblast to myofibroblast transition, in vivo and in vitro. Finally, WARP knockdown in vitro affected extra- and intracellular basic fibroblast growth factor production in myofibroblasts. To investigate the function for WARP in infarction healing, we performed an MI study in WARP knockout (KO) mice backcrossed more than 10 times on an Australian C57Bl/6-J background and bred in-house, and compared to wild type (WT) mice of the same C57Bl/6-J strain but of commercial European origin. WARP KO mice showed no mortality after MI, whereas 40% of the WT mice died due to cardiac rupture. However, when WARP KO mice were backcrossed on the European C57Bl/6-J background and bred heterozygous in-house, the previously seen protective effect in the WARP KO mice after MI was lost. Importantly, comparison of the cardiac response post-MI in WT mice bred heterozygous in-house versus commercially purchased WT mice revealed differences in cardiac rupture. CONCLUSION: These data demonstrate a redundant role for WARP in the wound healing process after MI but demonstrate that the continental/breeding/housing origin of mice of the same C57Bl6-J strain is critical in determining the susceptibility to cardiac rupture and stress the importance of using the correct littermate controls.

The microRNA-221/-222 cluster balances the antiviral and inflammatory response in viral myocarditis.

AIMS: Viral myocarditis (VM) is an important cause of heart failure and sudden cardiac death in young healthy adults; it is also an aetiological precursor of dilated cardiomyopathy. We explored the role of the miR-221/-222 family that is up-regulated in VM. METHODS AND RESULTS: Here, we show that microRNA-221 (miR-221) and miR-222 levels are significantly elevated during acute VM caused by Coxsackievirus B3 (CVB3). Both miRs are expressed by different cardiac cells and by infiltrating inflammatory cells, but their up-regulation upon myocarditis is mostly exclusive for the cardiomyocyte. Systemic inhibition of miR-221/-222 in mice increased cardiac viral load, prolonged the viraemic state, and strongly aggravated cardiac injury and inflammation. Similarly, in vitro, overexpression of miR-221 and miR-222 inhibited enteroviral replication, whereas knockdown of this miR-cluster augmented viral replication. We identified and confirmed a number of miR-221/-222 targets that co-orchestrate the increased viral replication and inflammation, including ETS1/2, IRF2, BCL2L11, TOX, BMF, and CXCL12. In vitro inhibition of IRF2, TOX, or CXCL12 in cardiomyocytes significantly dampened their inflammatory response to CVB3 infection, confirming the functionality of these targets in VM and highlighting the importance of miR-221/-222 as regulators of the cardiac response to VM. CONCLUSIONS: The miR-221/-222 cluster orchestrates the antiviral and inflammatory immune response to viral infection of the heart. Its inhibition increases viral load, inflammation, and overall cardiac injury upon VM.