GAS6 receptor antagonism impairs adipocyte differentiation and adipose tissue development in mice (HRL, VC and IS)

ABSTRACT A low Mr receptor tyrosine kinase inhibitor (R428) with high affinity and selectivity for the GAS6 (growth arrest specific protein 6) receptor Axl was used to study a potential role of GAS6 signaling in adiposity. In vitro, R428 caused a concentration-dependent inhibition of preadipocyte differentiation into mature adipocytes, as evidenced by reduced lipid uptake. Inhibition of Axl mediated signaling was confirmed by reduced levels of phospho-Akt activity. In vivo, oral administration of R428 during 5 weeks to mice kept on high fat diet resulted in significantly reduced weight gain and subcutaneous (SC) and gonadal (GON) fat mass. This was associated with marked adipocyte hypotrophy, enhanced macrophage infiltration and apoptosis. Thus, affecting GAS6 signaling through receptor antagonism using a low Mr Axl antagonist impairs adipocyte differentiation and reduces adipose tissue development in a murine model of nutritionally induced obesity. JPET # 178046 -4

Effect of VPAC1 Blockade on Adipose Tissue Formation and Composition in Mouse Models of Nutritionally Induced Obesity

Background. The pituitary adenylate cyclase activating polypeptide (PACAP) may affect adipogenesis and adipose tissue formation through interaction with its G-protein-coupled receptor VPAC1. Methods. We have used a monoclonal antibody (MAb 23A11) blocking VPAC1 in mouse models of nutritionally induced obesity. Results. Administration of MAb 23A11 (25 mg/kg body weight i.p. twice weekly) to 5-week old male C57Bl/6 mice kept on a high-fat diet for 15 weeks had no significant effect on weight gain, nor on subcutaneous (SC) or gonadal (GON) adipose tissue mass, as compared to the control MAb 1C8. However, adipocyte hypertrophy was observed in SC adipose tissue of MAb 23A11 treated mice. In a second study, 24 weeks old obese mice were treated for 5 weeks with MAb 23A11, without effect on body weight or fat mass, as compared to treatment with MAb 1C8. In addition, MAb 23A11 had no significant effect on glucose tolerance or insulin resistance in lean or obese C57Bl/6 mice. Conclusion. Blocking VPAC1 does not significantly affect adipose tissue formation in mouse models of diet-induced obesity, although it may be associated with mild adipocyte hypertrophy

Exogenously added GPI-anchored tissue inhibitor of matrix metal loproteinase-1 (TIMP-1) displays enhanced and novel biological activities

The family of tissue inhibitors of metalloproteinases (TIMPs) exhibits diverse physiological/biological functions including the inhibition of active matrix metalloproteinases, regulation of proMMP activation, cell growth, and the modulation of angiogenesis. TIMP-1 is a secreted protein that can be detected on the cell surface through its interaction with surface proteins. The diverse biological functions of TIMP-1 are thought to lie, in part, in the kinetics of TIMP-1/MMP/surface protein interactions. Proteins anchored by glycoinositol phospholipids (GPIs), when purified and added to cells in vitro, are incorporated into their surface membranes. A GPI anchor was fused to TIMP-1 to generate a reagent that could be added directly to cell membranes and thus focus defined concentrations of TIMP-1 protein on any cell surface independent of protein-protein interaction. Unlike native TIMP-1, exogenously added GPI-anchored TIMP-1 protein effectively blocked release of MMP-2 and MMP-9 from osteosarcoma cells. TIMP-1-GP1 was a more effective modulator of migration and proliferation than TIMP-1. While control hTIMP-1 protein did not significantly affect migration of primary microvascular endothelial cells at the concentrations tested, the GPI-anchored TIMP-1 protein showed a pronounced suppression of endothelial cell migration in response to bFGF. In addition, TIMP-1-GPI was more effective at inducing microvascular endothelial proliferation. In contrast, fibroblast proliferation was suppressed by the agent. Reagents based on this method should assist in the dissection of the protease cascades and activities involved in TIMP biology. Membrane-fixed TIMP-1 may represent a more effective version of the protein for use in therapeutic expression

Adipocyte extracellular matrix composition, dynamics and role in obesity

The central role of the adipose tissue in lipid metabolism places specific demands on the cell structure of adipocytes. The protein composition and dynamics of the extracellular matrix (ECM) is of crucial importance for the functioning of those cells. Adipogenesis is a bi-phasic process in which the ECM develops from a fibrillar to a laminar structure as cells move from the commitment phase to the growth phase characterized by storage of vast amounts of triglycerides. Mature adipocytes appear to spend a lot of energy on the maintenance of the ECM. ECM remodeling is mediated by a balanced complement of constructive and destructive enzymes together with their enhancers and inhibitors. ECM remodeling is an energy costing process regulated by insulin, by the energy metabolism, and by mechanical forces. In the obese, overgrowth of adipocytes may lead to instability of the ECM, possibly mediated by hypoxia

Macrophage-Stimulated Cardiac Fibroblast Production of IL-6 Is Essential for TGF β/Smad Activation and Cardiac Fibrosis Induced by Angiotensin II

Interleukin-6 (IL-6) is an important cytokine participating in multiple biologic activities in immune regulation and inflammation. IL-6 has been associated with cardiovascular remodeling. However, the mechanism of IL-6 in hypertensive cardiac fibrosis is still unclear. Angiotensin II (Ang II) infusion in mice increased IL-6 expression in the heart. IL-6 knockout (IL-6-/-) reduced Ang II-induced cardiac fibrosis: 1) Masson trichrome staining showed that Ang II infusion significantly increased fibrotic areas of the wild-type mouse heart, which was greatly suppressed in IL-6-/- mice and 2) immunohistochemistry staining showed decreased expression of α-smooth muscle actin (α-SMA), transforming growth factor β1 (TGF-β1) and collagen I in IL-6-/- mouse heart. The baseline mRNA expression of IL-6 in cardiac fibroblasts was low and was absent in cardiomyocytes or macrophages; however, co-culture of cardiac fibroblasts with macrophages significantly increased IL-6 production and expression of α-SMA and collagen I in fibroblasts. Moreover, TGF-β1 expression and phosphorylation of TGF-β downstream signal Smad3 was stimulated by co-culture of macrophages with cardiac fibroblasts, while IL-6 neutralizing antibody decreased TGF-β1 expression and Smad3 phosphorylation in co-culture of macrophage and fibroblast. Taken together, our results indicate that macrophages stimulate cardiac fibroblasts to produce IL-6, which leads to TGF-β1 production and Smad3 phosphorylation in cardiac fibroblasts and thus stimulates cardiac fibrosis

Plasmin Plays an Essential Role in Amplification of Psoriasiform Skin Inflammation in Mice

BACKGROUND: Although increased levels of plasminogen activators have been found in psoriatic lesions, the role of plasmin converted from plasminogen by plasminogen activators in pathogenesis of psoriasis has not been investigated. METHODOLOGY/PRINCIPAL FINDINGS: Here we examined the contribution of plasmin to amplification of inflammation in patients with psoriasis. We found that plasminogen was diminished, but that the amount and activity of its converted product plasmin were markedly increased in psoriasis. Moreover, annexin II, a receptor for plasmin was dramatically increased in both dermis and epidermis in psoriasis. Plasmin at sites of inflammation was pro-inflammatory, eliciting production of inflammatory factors, including CC chemokine ligand 20 (CCL20) and interleukin-23 (IL-23), that was mediated by the nuclear factor-kappaB (NF-κB) signaling pathway and that had an essential role in the recruitment and activation of pathogenic C-C chemokine receptor type 6 (CCR6)+ T cells. Moreover, intradermal injection of plasmin or plasmin together with recombinant monocyte/macrophage chemotactic protein-1 (MCP-1) resulted in induction of psoriasiform skin inflammation around the injection sites with several aspects of human psoriasis in mice. CONCLUSIONS/SIGNIFICANCE: Plasmin converted from plasminogen by plasminogen activators plays an essential role in amplification of psoriasiform skin inflammation in mice, and targeting plasmin receptor--annexin II--may harbor therapeutic potential for the treatment of human psoriasis

Determinants of serum zinc in a random population sample of four Belgian towns with different degrees of environmental exposure to cadmium

This report investigated the distribution of serum zinc and the factors determining serum zinc concentration in a large random population sample. The 1977 participants (959 men and 1018 women), 20–80 years old, constituted a stratified random sample of the population of four Belgian districts, representing two areas with low and two with high environmental exposure to cadmium. For each exposure level, a rural and an urban area were selected. The serum concentration of zinc, frequently used as an index for zinc status in human subjects, was higher in men (13.1 μmole/L, range 6.5–23.0 μmole/L) than in women (12.6 μmole/L, range 6.3–23.2 μmole/L). In men, 20% of the variance of serum zinc was explained by age (linear and squared term, R = 0.29), diurnal variation (r = 0.29), and total cholesterol (r = 0.16). After adjustment for these covariates, a negative relationship was observed between serum zinc and both blood (r = −0.10) and urinary cadmium (r = −0.14). In women, 11% of the variance could be explained by age (linear and squared term, R = 0.15), diurnal variation in serum zinc (r = 0.27), creatinine clearance (r = −0.11), log γ-glutamyltranspeptidase (r = 0.08), cholesterol (r = 0.07), contraceptive pill intake (r = −0.07), and log serum ferritin (r = 0.06). Before and after adjustment for significant covariates, serum zinc was, on average, lowest in the two districts where the body burden of cadmium, as assessed by urinary cadmium excretion, was highest. These results were not altered when subjects exposed to heavy metals at work were excluded from analysis