Monocyte Chemoattractant Protein-1 Expression In Macrophages Research Articles

Cell-specific and athero-protective roles for RIPK3 in a murine model of atherosclerosis.

ABSTRACTReceptor-interacting protein kinase 3 (RIPK3) was recently implicated in promoting atherosclerosis progression through a proposed role in macrophage necroptosis. However, RIPK3 has been connected to numerous other cellular pathways, which raises questions about its actual role in atherosclerosis. Furthermore, RIPK3 is expressed in a multitude of cell types, suggesting that it may be physiologically relevant to more than just macrophages in atherosclerosis. In this study, Ripk3 was deleted in macrophages, endothelial cells, vascular smooth muscle cells or globally on the Apoe−/− background using Cre-lox technology. To induce atherosclerosis progression, male and female mice were fed a Western diet for three months before tissue collection and analysis. Surprisingly, necroptosis markers were nearly undetectable in atherosclerotic aortas. Furthermore, en face lesion area was increased in macrophage- and endothelial-specific deletions of Ripk3 in the descending and abdominal regions of the aorta. Analysis of bone-marrow-derived macrophages and cultured endothelial cells revealed that Ripk3 deletion promotes expression of monocyte chemoattractant protein 1 (MCP-1) and E-selectin in these cell types, respectively. Western blot analysis showed upregulation of MCP-1 in aortas with Ripk3-deficient macrophages. Altogether, these data suggest that RIPK3 in macrophages and endothelial cells protects against atherosclerosis through a mechanism that likely does not involve necroptosis. This protection may be due to RIPK3-mediated suppression of pro-inflammatory MCP-1 expression in macrophages and E-selectin expression in endothelial cells. These findings suggest a novel and unexpected cell-type specific and athero-protective function for RIPK3.This article has an associated First Person interview with the first author of the paper.

Inhibitory Effect of 3-(4-Hydroxyphenyl)-1-(thiophen-2-yl) prop-2-en-1-one, a Chalcone Derivative on MCP-1 Expression in Macrophages via Inhibition of ROS and Akt Signaling.

Chalcones (1,3-diaryl-2-propen-1-ones), a subfamily of flavonoid, are widely known to possess potent anti-inflammatory and anti-oxidant properties. In this study, we investigated the effect of 3-(4-Hydroxyphenyl)-1-(thio3-(4-Hydroxyphenyl phen-2-yl)prop-2-en-1-one (TI-I-175), a synthetic chalcone derivative, on endotoxin-induced expression of monocyte chemoattractant protein-1 (MCP-1), one of the key chemokines that regulates migration and infiltration of immune cells, and its potential mechanisms. TII-175 potently inhibited MCP-1 mRNA expression stimulated by lipopolysaccharide (LPS) in RAW 264.7 macrophages without significant effect on cell viability. Treatment of cells with TI-I-175 markedly prevented LPS-induced transcriptional activation of activator protein-1 (AP-1) as measured by luciferase reporter assay, while nuclear factor-κB (NF-κB) activity was not inhibited by TI-I-175, implying that TI-I-175 suppressed MCP-1 expression probably via regulation of AP-1. In addition, TI-I-175 treatment significantly inhibited LPS-induced Akt phosphorylation and led to a significant decrease in reactive oxygen species (ROS) production by LPS, which act as up-stream signaling events required for AP-1 activation in RAW 264.7 macrophages. Taken together, these results indicate that TI-I-175 suppresses MCP-1 gene expression in LPS-stimulated RAW 264.7 macrophages via suppression of ROS production and Akt activation.

Abstract 18763: Characteristics and Anti-Atherosclerotic Properties of (-)-Epigallocatechin Gallate Loaded Lipid Nanoparticles in vitro

BACKGROUND: Atherosclerotic cardiovascular disease is the leading cause of death and disability in the United States and other developed countries. Macrophages are major cells responsible for atherosclerotic lesion development. (-)-Epigallocatechin gallate (EGCG), a natural compound found in green tea, is valuable to prevent atherosclerosis. But EGCG’s low level of stability, bioavailability and targeting specificity in the body makes administering it in therapeutic doses unrealistic. METHODS AND RESULTS: We have successfully synthesized EGCG loaded lipid nanoparticles (ENP), which are composed of biocompatible and biodegradable lipids, surfactants and EGCG. The size of nanoparticles was less than 100 nm in diameter measured using a dynamic light scattering method and a transmission electron microscope. The encapsulation efficiency was around 90%. As compared to native EGCG, ENP significantly increased EGCG stability and macrophage EGCG content at 37°C. Remarkably, ENP significantly decreased human macrophage cholesteryl ester content measured using a high performance liquid chromatography (HPLC) system. ENP also significantly lowered both gene expression and protein secretion of monocyte chemoattractant protein 1 (MCP-1) in/from human macrophages measured using real-time PCR and ELISA assays, respectively. We also incorporated target ligands on the surface of ENP. The target ligands further increased the binding and uptake of those nanoparticles in human macrophages. CONCLUSIONS: ENP significantly enhanced EGCG stability, increased its cellular bioavailability, decreased cholesteryl ester content and MCP-1 expression in macrophages. Target ligands increased the target specificity of those nanoparticles to human macrophages. ENP with target ligands have a potential for preventing atherosclerotic lesion development through enhancing EGCG uptake by macrophages and decreasing macrophage cholesterol accumulation and inflammatory responses. Funding support: The project described was supported by Grant Number R15AT007013 from NIH NCCAM.

Genistein Inhibits ox-LDL-induced VCAM-1, ICAM-1 and MCP-1 Expression of HUVECs Through Heme Oxygenase-1

Genistein, a principal component of soybean isoflavones, plays an important role in the prevention of atherosclerosis. However, the detailed mechanisms have not been fully investigated. The aims of this study were to evaluate the anti-atherosclerotic effect and investigate potential pharmacological mechanism of genistein. A model of oxidized low-density lipoprotein (ox-LDL)-induced injury in on human umbilical vein endothelial cells (HUVECs) was established to evaluate the protective role of genistein. Macrophage/monocyte chemoattractant protein-1 (MCP-1), vascular cellular adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1) secretion and their messenger RNA transcription were observed via enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase PCR (RT-PCR). Meanwhile, the study investigated the role of Nrf2/HO-1 pathway during the process. Pretreatment with genistein markedly reduced ox-LDL-induced MCP-1, VCAM-1 and ICAM-1 secretion and mRNA transcription, which was further decreased by the inducer of HO and reversed by the inhibitor of HO; additionally, the effects were accompanied with upregulating HO-1 mRNA and protein expression and markedly abolished with Nrf2 siRNA. Anti-inflammatory effect of genistein on endothelial cells may be associated with the activation of Nrf2/HO-1 pathway.

Reciprocal Interaction between Macrophages and T cells Stimulates IFN-γ and MCP-1 Production in Ang II-induced Cardiac Inflammation and Fibrosis

BackgroundThe inflammatory response plays a critical role in hypertension-induced cardiac remodeling. We aimed to study how interaction among inflammatory cells causes inflammatory responses in the process of hypertensive cardiac fibrosis.Methodology/Principal FindingsInfusion of angiotensin II (Ang II, 1500 ng/kg/min) in mice rapidly induced the expression of interferon γ (IFN-γ) and leukocytes infiltration into the heart. To determine the role of IFN-γ on cardiac inflammation and remodeling, both wild-type (WT) and IFN-γ-knockout (KO) mice were infused Ang II for 7 days, and were found an equal blood pressure increase. However, knockout of IFN-γ prevented Ang II-induced: 1) infiltration of macrophages and T cells into cardiac tissue; 2) expression of tumor necrosis factor α and monocyte chemoattractant protein 1 (MCP-1), and 3) cardiac fibrosis, including the expression of α-smooth muscle actin and collagen I (all p<0.05). Cultured T cells or macrophages alone expressed very low level of IFN-γ, however, co-culture of T cells and macrophages increased IFN-γ expression by 19.8±0.95 folds (vs. WT macrophage, p<0.001) and 20.9 ± 2.09 folds (vs. WT T cells, p<0.001). In vitro co-culture studies using T cells and macrophages from WT or IFN-γ KO mice demonstrated that T cells were primary source for IFN-γ production. Co-culture of WT macrophages with WT T cells, but not with IFN-γ-knockout T cells, increased IFN-γ production (p<0.01). Moreover, IFN-γ produced by T cells amplified MCP-1 expression in macrophages and stimulated macrophage migration.Conclusions/SignificanceReciprocal interaction between macrophages and T cells in heart stimulates IFN-γ expression, leading to increased MCP-1 expression in macrophages, which results a forward-feed recruitment of macrophages, thus contributing to Ang II-induced cardiac inflammation and fibrosis.

Immunohistochemical study of monocyte chemoattractant protein-1 in the pancreas of NOD mice following cyclophosphamide administration and during spontaneous diabetes

In type 1 diabetes mellitus (T1DM), the processes which control the recruitment of immune cells into pancreatic islets are poorly defined. Complex interactions involving adhesion molecules, chemokines and chemokine receptors may facilitate this process. The chemokine, monocyte chemoattractant protein-1 (MCP-1), previously shown to be important in leukocyte trafficking in other disease systems, may be a key participant in the early influx of blood-borne immune cells into islets during T1DM. In the non-obese diabetic (NOD) mouse, the expression of MCP-1 protein has not been demonstrated. We employed dual-label immunohistochemistry to examine the intra-islet expression, distribution and cellular source of MCP-1 in the NOD mouse following cyclophosphamide administration. NOD mice were treated with cyclophosphamide at day 72-73 and MCP-1 expression studied at days 0, 4, 7, 11 and 14 after treatment and comparisons were made between age-matched NOD mice treated with diluent and non-diabetes-prone CD-1 mice. Pancreatic expression of MCP-1 was also examined in NOD mice at various stages of spontaneous diabetes. In the cyclophosphamide group at day 0, MCP-1 immunolabelling was present in selective peri-islet macrophages but declined at day 4. It increased slightly at day 7 but was more marked from day 11, irrespective of diabetes development. The pattern of MCP-1 expression in macrophages was different over time in both the cyclophosphamide and control groups. In the cyclophosphamide group, there was a change over time with an increase at day 11. In the control group, there was little evidence of change over time. There was no significant difference in the mean percentage of MCP-1 positive macrophages between the cyclophosphamide-treated diabetic and non-diabetic mice. During spontaneous diabetes in the NOD mouse, only a few peri-islet MCP-1 cells appeared at day 45. These became more numerous from day 65 but were absent at diabetes onset. We speculate that a proportion of early islet-infiltrating macrophages which express MCP-1 may attract additional lymphocytes and macrophages into the early inflamed islets and intensify the process of insulitis.

Homocysteine induces monocyte chemoattractant protein-1 expression by activating NF-κB in THP-1 macrophages

Homocysteinemia is an independent risk factor for cardiovascular disorders. The recruitment of monocytes is an important event in atherogenesis. Monocyte chemoattractant protein-1 (MCP-1) is a potent chemokine that stimulates monocyte migration into the intima of arterial walls. The objective of the present study was to investigate the effect of homocysteine on MCP-1 expression in macrophages and the underlying mechanism of such effect. Human monocytic cell (THP-1)-derived macrophages were incubated with homocysteine. By nuclease protection assay and ELISA, homocysteine (0.05-0.2 mM) was shown to significantly enhance the expression of MCP-1 mRNA (up to 2.6-fold) and protein (up to 4.8-fold) in these cells. Homocysteine-induced MCP-1 expression resulted in increased monocyte chemotaxis. The increase in MCP-1 expression was associated with activation of nuclear factor (NF)-kappaB due to increased phosphorylation of the inhibitory protein (IkappaB-alpha) as well as reduced expression of IkappaB-alpha mRNA in homocysteine-treated cells. In conclusion, our results demonstrate that homocysteine, at pathological concentration, stimulates MCP-1 expression in THP-1 macrophages via NF-kappaB activation.

Oxidized low density lipoprotein and very low density lipoprotein enhance expression of monocyte chemoattractant protein-1 in rabbit peritoneal exudate macrophages

The migration of monocytes into arterial subendothelial space is one of the earliest events in atherogenesis. Monocyte chemoattractant protein 1 (MCP-1) is a potent monocyte chemoattractant. The purpose of this work was to examine whether oxidized low density lipoprotein (OX-LDL) and very low density lipoprotein (OX-VLDL) have any effect on the expression of MCP-1 mRNA and protein in rabbit peritoneal exudate macrophages. The total RNA was extracted from the macrophages after 24 h exposure to LDL, VLDL, OX-LDL, and OX-VLDL, respectively, and the media (LDL-CM, VLDL-CM, OX-LDL-CM and OX-VLDL-CM) conditioned by the macrophages exposed to the above-mentioned lipoproteins were collected. The MCP-1 mRNA expression in macrophages was examined by Northern blot analysis. Meanwhile, MCP-1 protein in the conditioned media was determined by sandwich ELISA. The chemotactic activity of the conditioned media for monocytes was determined by micropore filter assay. The results revealed that the macrophages can express MCP-1, and 24 h exposure to OX-LDL and OX-VLDL induced a 3.2-fold and a 3.4-fold increase in MCP-1 mRNA expression in macrophages and a 2.2-fold and a 2.5-fold increase in the level of MCP-1 protein in the conditioned media, respectively. However, 24 h exposure to LDL and VLDL only induced a slight increase in the expression of MCP-1 mRNA and protein in macrophages. Furthermore, the migration distance of monocyte induced by OX-LDL-CM and OX-VLDL-CM was longer than that induced by LDL-CM and VLDL-CM, as well as by CM. We conclude that the macrophages can express MCP-1, and OX-LDL and OX- VLDL induce stronger MCP-1 expression. It suggests that macrophages may amplify the recruitment of monocytes into subendothelial space, and OX-LDL and OX-VLDL may play an important role in the pathogenesis of atherosclerosis through enhancing the MCP-1 expression in macrophages.