Anti-VCAM1 monoclonal antibody, clone 5F9

Inflammation is central to heart failure progression. Innate immune signaling increases expression of the transmembrane proteoglycan syndecan-4 in cardiac myocytes and fibroblasts, followed by shedding of its ectodomain. Circulating shed syndecan-4 is increased in heart failure patients, however the pathophysiological and molecular consequences associated with syndecan-4 shedding remain poorly understood. Here we used lipopolysaccharide (LPS) challenge to investigate the effects of syndecan-4 shedding in the heart. Wild-type mice (10 mg/kg, 9 h) and cultured neonatal rat cardiomyocytes and fibroblasts were subjected to LPS challenge. LPS increased cardiac syndecan-4 mRNA without altering full-length protein. Elevated levels of shedding fragments in the myocardium and blood from the heart confirmed syndecan-4 shedding in vivo. A parallel upregulation of ADAMTS1, ADAMTS4 and MMP9 mRNA suggested these shedding enzymes to be involved. Echocardiography revealed reduced ejection fraction, diastolic tissue velocity and prolonged QRS duration in mice unable to shed syndecan-4 (syndecan-4 KO) after LPS challenge. In line with syndecan-4 shedding promoting immune cell recruitment, expression of immune cell markers (CD8, CD11a, F4/80) and adhesion receptors (Icam1, Vcam1) were attenuated in syndecan-4 KO hearts after LPS. Cardiomyocytes and fibroblasts exposed to shed heparan sulfate-substituted syndecan-4 ectodomains showed increased Icam1, Vcam1, TNF alpha and IL-1 beta expression and NF-kappa B-activation, suggesting direct regulation of immune cell recruitment pathways. In cardiac fibroblasts, shed ectodomains regulated expression of extracellular matrix constituents associated with collagen synthesis, cross-linking and tumover. Higher syndecan-4 levels in the coronary sinus vs. the radial artery of open heart surgery patients suggested that syndecan-4 is shed from the human heart. Our data demonstrate that shedding of syndecan-4 ectodomains is part of the cardiac innate immune response, promoting immune cell recruitment, extracellular matrix remodeling and mitigating cardiac dysfunction in response to LPS. (C) 2015 Elsevier Ltd. All rights reserved.

Microvascular dysfunction leads to multi-organ failure and mortality in sepsis. Our previous studies demonstrated that administration of exogenous endothelial progenitor cells (EPCs) confers protection in sepsis as evidenced by reduced vascular leakage, improved organ function, and increased survival. We hypothesize that EPCs protect the microvasculature through the exosomes-mediated transfer of microRNAs (miRNAs). Mice were rendered septic by cecal ligation and puncture (CLP), and EPC exosomes were administered intravenously at 4 hr after CLP. EPC exosomes treatment improved survival, suppressing lung and renal vascular leakage, and reducing liver and kidney dysfunction in septic mice. EPC exosomes attenuated sepsis-induced increases in plasma levels of cytokines and chemokine. Moreover, we determined miRNA contents of EPC exosomes with next-generation sequencing and found abundant miR-126-3p and 5p. We demonstrated that exosomal miR-126-5p and 3p suppressed LPS-induced high mobility group box 1 (HMGB1) and vascular cell adhesion molecule 1 (VCAM1) levels, respectively, in human microvascular endothelial cells (HMVECs). Inhibition of microRNA-126-5p and 3p through transfection with microRNA-126-5p and 3p inhibitors abrogated the beneficial effect of EPC exosomes. The inhibition of exosomal microRNA-126 failed to block LPS-induced increase in HMGB1 and VCAM1 protein levels in HMVECs and negated the protective effect of exosomes on sepsis survival. Thus, EPC exosomes prevent microvascular dysfunction and improve sepsis outcomes potentially through the delivery of miR-126.