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

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

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

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

Histidine-Rich Glycoprotein Can Prevent Development of Mouse Experimental Glioblastoma

Extensive angiogenesis, formation of new capillaries from pre-existing blood vessels, is an important feature of malignant glioma. Several antiangiogenic drugs targeting vascular endothelial growth factor (VEGF) or its receptors are currently in clinical trials as therapy for high-grade glioma and bevacizumab was recently approved by the FDA for treatment of recurrent glioblastoma. However, the modest efficacy of these drugs and emerging problems with anti-VEGF treatment resistance welcome the development of alternative antiangiogenic therapies. One potential candidate is histidine-rich glycoprotein (HRG), a plasma protein with antiangiogenic properties that can inhibit endothelial cell adhesion and migration. We have used the RCAS/TV-A mouse model for gliomas to investigate the effect of HRG on brain tumor development. Tumors were induced with platelet-derived growth factor-B (PDGF-B), in the presence or absence of HRG. We found that HRG had little effect on tumor incidence but could significantly inhibit the development of malignant glioma and completely prevent the occurrence of grade IV tumors (glioblastoma)

The Kunitz-Like Modulatory Protein Haemangin Is Vital for Hard Tick Blood-Feeding Success

Ticks are serious haematophagus arthropod pests and are only second to mosquitoes as vectors of diseases of humans and animals. The salivary glands of the slower feeding hard ticks such as Haemaphysalis longicornis are a rich source of bioactive molecules and are critical to their biologic success, yet distinct molecules that help prolong parasitism on robust mammalian hosts and achieve blood-meals remain unidentified. Here, we report on the molecular and biochemical features and precise functions of a novel Kunitz inhibitor from H. longicornis salivary glands, termed Haemangin, in the modulation of angiogenesis and in persistent blood-feeding. Haemangin was shown to disrupt angiogenesis and wound healing via inhibition of vascular endothelial cell proliferation and induction of apoptosis. Further, this compound potently inactivated trypsin, chymotrypsin, and plasmin, indicating its antiproteolytic potential on angiogenic cascades. Analysis of Haemangin-specific gene expression kinetics at different blood-feeding stages of adult ticks revealed a dramatic up-regulation prior to complete feeding, which appears to be functionally linked to the acquisition of blood-meals. Notably, disruption of Haemangin-specific mRNA by a reverse genetic tool significantly diminished engorgement of adult H. longicornis, while the knock-down ticks failed to impair angiogenesis in vivo. To our knowledge, we have provided the first insights into transcriptional responses of human microvascular endothelial cells to Haemangin. DNA microarray data revealed that Haemangin altered the expression of 3,267 genes, including those of angiogenic significance, further substantiating the antiangiogenic function of Haemangin. We establish the vital roles of Haemangin in the hard tick blood-feeding process. Moreover, our results provide novel insights into the blood-feeding strategies that enable hard ticks to persistently feed and ensure full blood-meals through the modulation of angiogenesis and wound healing processes

Myeloid Heme Oxygenase-1 Haploinsufficiency Reduces High Fat Diet-Induced Insulin Resistance by Affecting Adipose Macrophage Infiltration in Mice

Increased adipose tissue macrophages contribute to obesity-induced metabolic syndrome. Heme oxygenase-1 (HO-1) is a stress-inducible enzyme with potent anti-inflammatory and proangiogenic activities in macrophages. However, the role of macrophage HO-1 on obesity-induced adipose inflammation and metabolic syndrome remains unclear. Here we show that high-fat diet (HFD) feeding in C57BL/6J mice induced HO-1 expression in the visceral adipose tissue, particularly the stromal vascular fraction. When the irradiated C57BL/6J mice reconstituted with wild-type or HO-1+/− bone marrow were fed with HFD for over 24 weeks, the HO-1+/− chimeras were protected from HFD-induced insulin resistance and this was associated with reduced adipose macrophage infiltration and angiogenesis, suggesting that HO-1 affects myeloid cell migration toward adipose tissue during obesity. In vivo and in vitro migration assays revealed that HO-1+/− macrophages exhibited an impaired migration response. Chemoattractant-induced phosphorylation of p38 and focal adhesion kinase (FAK) declined faster in HO-1+/− macrophages. Further experiments demonstrated that carbon monoxide and bilirubin, the byproducts derived from heme degradation by HO-1, enhanced macrophage migration by increasing phosphorylation of p38 and FAK, respectively. These data disclose a novel role of hematopoietic cell HO-1 in promoting adipose macrophage infiltration and the development of insulin resistance during obesity

Absence of Host Plasminogen Activator Inhibitor 1 Prevents Cancer Invasion and Vascularization

Acquisition of invasive/metastatic potential through protease expression is an essential event in tumor progression. High levels of components of the plasminogen activation system, including urokinase, but paradoxically also its inhibitor, plasminogen activator inhibitor 1 (PAI1), have been correlated with a poor prognosis for some cancers. We report here that deficient PAI1 expression in host mice prevented local invasion and tumor vascularization of transplanted malignant keratinocytes. When this PAI1 deficiency was circumvented by intravenous injection of a replication-defective adenoviral vector expressing human PAI1, invasion and associated angiogenesis were restored. This experimental evidence demonstrates that host-produced PAI is essential for cancer cell invasion and angiogenesis

Thrombospondins in the heart: potential functions in cardiac remodeling

Cardiac remodeling after myocardial injury involves inflammation, angiogenesis, left ventricular hypertrophy and matrix remodeling. Thrombospondins (TSPs) belong to the group of matricellular proteins, which are non-structural extracellular matrix proteins that modulate cell–matrix interactions and cell function in injured tissues or tumors. They interact with different matrix and membrane-bound proteins due to their diverse functional domains. That the expression of TSPs strongly increases during cardiac stress or injury indicates an important role for them during cardiac remodeling. Recently, the protective properties of TSP expression against heart failure have been acknowledged. The current review will focus on the biological role of TSPs in the ischemic and hypertensive heart, and will describe the functional consequences of TSP polymorphisms in cardiac disease

Assessment of endogenous fibrinolysis in clinical using novel tests - Ready for clinical roll-out?

© The Author(s) 2021. This article is licensed under a Creative Commons Attribution 4.0 International License. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.The occurrence of thrombotic complications, which can result in excess mortality and morbidity, represent an imbalance between the pro-thrombotic and fibrinolytic equilibrium.The mainstay treatment of these complications involves the use of antithrombotic agents but despite advances in pharmacotherapy, there remains a significant proportion of patients who continue to remain at risk.Endogenous fibrinolysis is a physiological counter-measure against lasting thrombosis and may be measured using several techniques to identify higher risk patients who may benefit from more aggressive pharmacotherapy. However, the assessment of the fibrinolytic systemis not yet accepted into routine clinical practice.In this review, we will revisit the different methods of assessing endogenous fibrinolysis (factorial assays, turbidimetric lysis assays, viscoelastic and the global thrombosis tests), including the strengths, limitations, correlation to clinical outcomes of each method and howwe might integrate the assessment of endogenous fibrinolysis into clinical practice in the future.Peer reviewedFinal Published versio