Inhibition Of Adhesion Molecule Expression Research Articles

Meteorin-like protein (METRNL)/IL-41 improves LPS-induced inflammatory responses via AMPK or PPARδ–mediated signaling pathways

PurposeMeteorin-like protein (METRNL) (also known as IL-41), recently identified as a myokine, is released in response to muscle contraction. It improves the skeletal muscle insulin sensitivity through exerting a beneficial anti-inflammatory effect. However, no independent studies have been published to verify the effects of METRNL on human umbilical vein endothelial cells (HUVECs) and THP-1 human monocytes. Materials and methodsThe levels of NFκB and IκB phosphorylation as well as the expression of adhesion molecules were assessed by Western blotting analysis. Cell adhesion assay demonstrated the interactions between HUVEC and THP-1 ​cells. We used enzyme-linked immunosorbent assay (ELISA) to measure the levels of TNFα and MCP-1 in culture medium. ResultsTreatment with METRNL suppressed the secretion of TNFα and MCP-1 as well as NFκB and IκB phosphorylation and inflammatory markers in lipopolysaccharide (LPS)-treated HUVECs and THP-1 ​cells. Furthermore, treatment with METRNL ameliorated LPS-induced attachment of THP-1 monocytes to HUVECs via inhibition of adhesion molecule expression and apoptosis. Treatment of HUVEC and THP-1 ​cells with METRNL enhanced AMPK phosphorylation and PPARδ expression in a dose-dependent manner. Small interference (si) RNA-mediated suppression of AMPK or PPARδ restored all these changes. ConclusionsIt has therefore been shown that METRNL ameliorates inflammatory responses through AMPK and PPARδ-dependent pathways in LPS-treated HUVEC. In sum, the current study may suggest the suppressive potential of METRNL against endothelial inflammation.

Endothelial HSPA12B Exerts Protection Against Sepsis-Induced Severe Cardiomyopathy via Suppression of Adhesion Molecule Expression by miR-126.

Heat shock protein A12B (HSPA12B) is predominately expressed in endothelial cells (ECs) and has been reported to protect against cardiac dysfunction from endotoxemia or myocardial infarction. This study investigated the mechanisms by which endothelial HSPA12B protects polymicrobial sepsis–induced cardiomyopathy. Wild-type (WT) and endothelial HSPA12B knockout (HSPA12B–/–) mice were subjected to polymicrobial sepsis induced by cecal ligation and puncture (CLP). Cecal ligation and puncture sepsis accelerated mortality and caused severe cardiac dysfunction in HSPA12B–/– mice compared with WT septic mice. The levels of adhesion molecules and the infiltrated immune cells in the myocardium of HSPA12B–/– septic mice were markedly greater than in WT septic mice. The levels of microRNA-126 (miR-126), which targets adhesion molecules, in serum exosomes from HSPA12B–/– septic mice were significantly lower than in WT septic mice. Transfection of ECs with adenovirus expressing HSPA12B significantly increased miR-126 levels. Increased miR-126 levels in ECs prevented LPS-stimulated expression of adhesion molecules. In vivo delivery of miR-126 carried by exosomes into the myocardium of HSPA12B–/– mice significantly attenuated CLP sepsis increased levels of adhesion molecules, and improved CLP sepsis–induced cardiac dysfunction. The data suggest that HSPA12B protects against sepsis-induced severe cardiomyopathy via regulating miR-126 expression which targets adhesion molecules, thus decreasing the accumulation of immune cells in the myocardium.

MicroRNA-181a-5p and microRNA-181a-3p cooperatively restrict vascular inflammation and atherosclerosis

MicroRNAs have emerged as important post-transcriptional regulators of gene expression and are involved in diverse diseases and cellular process. Decreased expression of miR-181a has been observed in the patients with coronary artery disease, but its function and mechanism in atherogenesis is not clear. This study was designed to determine the roles of miR-181a-5p, as well as its passenger strand, miR-181a-3p, in vascular inflammation and atherogenesis. We found that the levels of both miR-181a-5p and miR-181a-3p are decreased in the aorta plaque and plasma of apoE−/− mice in response to hyperlipidemia and in the plasma of patients with coronary artery disease. Rescue of miR-181a-5p and miR-181a-3p significantly retards atherosclerotic plaque formation in apoE−/− mice. MiR-181a-5p and miR-181a-3p have no effect on lipid metabolism but decrease proinflammatory gene expression and the infiltration of macrophage, leukocyte and T cell into the lesions. In addition, gain-of-function and loss-of-function experiments show that miR-181a-5p and miR-181a-3p inhibit adhesion molecule expression in HUVECs and monocytes-endothelial cell interaction. MiR-181a-5p and miR-181a-3p cooperatively receded endothelium inflammation compared with single miRNA strand. Mechanistically, miR-181a-5p and miR-181a-3p prevent endothelial cell activation through blockade of NF-κB signaling pathway by targeting TAB2 and NEMO, respectively. In conclusion, these findings suggest that miR-181a-5p and miR-181a-3p are both antiatherogenic miRNAs. MiR-181a-5p and miR-181a-3p mimetics retard atherosclerosis progression through blocking NF-κB activation and vascular inflammation by targeting TAB2 and NEMO, respectively. Therefore, restoration of miR-181a-5p and miR-181a-3p may represent a novel therapeutic approach to manage atherosclerosis.

Isobavachalcone attenuates Sephadex-induced lung injury via activation of A20 and NRF2/HO-1 in rats

The aim of this study is to investigate the protective effect and underlying molecular mechanisms of isobavachalcone on Sephadex-induced lung injury in rats. The result showed isobavachalcone inhibited massive granulomas, decreased inflammatory cells infiltration and oxidative stress markers level, but it can increase antioxidant enzymes level. The ELISA assay exhibited isobavachalcone decreased TNF-α production in BALF and lung tissue. Western blotting analysis showed isobavachalcone can inhibit NF-κB pathway that may be mediated by upregulation of A20. Furthermore, we also found isobavachalcone can activate NRF2/HO-1 pathway and inhibit adhesion molecule expression. Taken together, the present results suggested that isobavachalcone can attenuate Sephadex-induced lung injury that may be related to inhibition of NF-κB mediated by upregulation of A20 and activation of NRF2/HO-1 signaling pathway.

The proteasome and proteasome inhibitors in multiple myeloma.

Proteasome inhibitors are one of the most important classes of agents to have emerged for the treatment of multiple myeloma in the past two decades, and now form one of the backbones of treatment. Three agents in this class have been approved by the United States Food and Drug Administration-the first-in-class compound bortezomib, the second-generation agent carfilzomib, and the first oral proteasome inhibitor, ixazomib. The success of this class of agents is due to the exquisite sensitivity of myeloma cells to the inhibition of the 26S proteasome, which plays a critical role in the pathogenesis and proliferation of the disease. Proteasome inhibition results in multiple downstream effects, including the inhibition of NF-κB signaling, the accumulation of misfolded and unfolded proteins, resulting in endoplasmic reticulum stress and leading to the unfolded protein response, the downregulation of growth factor receptors, suppression of adhesion molecule expression, and inhibition of angiogenesis; resistance to proteasome inhibition may arise through cellular responses mediating these downstream effects. These multiple biologic consequences of proteasome inhibition result in synergistic or additive activity with other chemotherapeutic and targeted agents for myeloma, and proteasome inhibitor-based combination regimens have become established as a cornerstone of therapy throughout the myeloma treatment algorithm, incorporating agents from the other key classes of antimyeloma agents, including the immunomodulatory drugs, monoclonal antibodies, and histone deacetylase inhibitors. This review gives an overview of the critical role of the proteasome in myeloma and the characteristics of the different proteasome inhibitors and provides a comprehensive summary of key clinical efficacy and safety data with the currently approved proteasome inhibitors.

Cordycepin inhibits vascular adhesion molecule expression in TNF-α-stimulated vascular muscle cells.

Atherosclerosis is a chronic inflammatory disease, which is associated with the increased expression of adhesion molecules in vascular smooth muscle cells (VSMCs). Cordycepin is one of the major bioactive components of Ophiocordyceps sinensis that has been demonstrated to exert anti-atherogenic activity; however, its molecular mechanisms are poorly understood. The aim of the present study was to examine the in vitro effects of cordycepin on the tumor necrosis factor (TNF)-α-induced suppression of adhesion molecule expression. The results of the present study demonstrated that cordycepin markedly inhibited the expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) in TNF-α-stimulated human aortic vascular smooth muscle cells (HA-VSMCs). Cordycepin significantly inhibited the TNF-α-induced mitogen-activated protein kinase (MAPK) and protein kinase B (Akt) activation (P<0.05), markedly inhibited the TNF-α-induced expression level of nuclear factor (NF)-κB p65 and markedly prevented the TNF-α-associated degradation of IκBα in HA-VSMCs. The results of the present study suggest that cordycepin inhibits the expression of VCAM-1 and ICAM-1 in TNF-α-stimulated HA-VSMCs via downregulating the MAPK/Akt/NF-κB signaling pathway. Therefore, cordycepin may have a potential therapeutic application for preventing the advancement of atherosclerotic lesions.

Montelukast inhibits oxidized low-density lipoproteins (ox-LDL) induced vascular endothelial attachment: An implication for the treatment of atherosclerosis

Recruitment of monocytes to endothelial cells is important during early stages of atherosclerosis development, which is activated in response to a number of inflammatory stimuli, including oxidized low-density lipoprotein (ox-LDL). Montelukast is a licensed drug approved by the Food and Drug Administration (FDA) and clinically used for the treatment of asthma by reducing the eosinophilic inflammation in the airway. Little information regarding the effects of Montelukast on endothelial inflammation has been reported before. In the current study, we found that Montelukast markedly reduced ox-LDL-induced monocyte adhesion to human umbilical vein endothelial cells. In addition, the inhibitory mechanism of Montelukast was associated with suppression of adhesion molecule expression, including VCAM-1 and E-selectin. Mechanistically, ERK5 mediated expression of the transcriptional factor KLF2 was found to be involved in the anti-inflammation effects of Montelukast against ox-LDL induced endothelial inflammation. Results indicate that Montelukast plays a protective role in the early stages of atherosclerosis.

The Nutraceutical Dehydrozingerone and Its Dimer Counteract Inflammation- and Oxidative Stress-Induced Dysfunction of In Vitro Cultured Human Endothelial Cells: A Novel Perspective for the Prevention and Therapy of Atherosclerosis.

Atherosclerosis is characterized by endothelial dysfunction, mainly induced by inflammation and oxidative stress. Increased reactive oxygen species (ROS) production together with increased adhesion molecules and thrombogenic tissue factor (TF) expression on endothelial cells has a key role in proatherogenic mechanisms. Therefore downmodulation of these molecules could be useful for reducing the severity of inflammation and atherosclerosis progression. Dehydrozingerone (DHZ) is a nutraceutical compound with anti-inflammatory and antioxidant activities. In this study we evaluated the ability of DHZ and its symmetric dimer to modulate hydrogen peroxide- (H2O2-) induced ROS production in human umbilical vein endothelial cells (HUVEC). We also evaluated intercellular adhesion molecule- (ICAM-) 1, vascular cell adhesion molecule- (VCAM-) 1, and TF expression in HUVEC activated by tumor necrosis factor- (TNF-) α. HUVEC pretreatment with DHZ and DHZ dimer reduced H2O2-induced ROS production and inhibited adhesion molecule expression and secretion. Of note, only DHZ dimer was able to reduce TF expression. DHZ effects were in part mediated by the inhibition of the nuclear factor- (NF-) κB activation. Overall, our findings demonstrate that the DHZ dimer exerts a potent anti-inflammatory, antioxidant, and antithrombotic activity on endothelial cells and suggest potential usefulness of this compound to contrast the pathogenic mechanisms involved in atherosclerosis progression.

Goniolactone C, a styryl lactone derivative, inhibits PDGF-BB-induced vascular smooth muscle cell migration and proliferation via PDGFR/ERK signaling.

Platelet-derived growth factor-BB (PDGF-BB) and its downstream effector, extracellular signal-regulated kinase 1/2 (ERK1/2) MAP kinase, initiate a multitude of biological effects, including vascular smooth muscle cell (VSMC) proliferation and migration, which are critical events in the initiation and development of restenosis following percutaneous transluminal coronary angioplasty (PTCA). Styryl lactones are natural products that have been demonstrated to possess anti-proliferative activities. Goniolactone C is a styryl lactone derivative that was first extracted from Goniothalamus cheliensis Hu. In the present study, we investigated the effects of goniolactone C on VSMC migration and proliferation. We found that goniolactone C preferentially interacted with cellular systems that rely on PDGF signaling but not those that rely on epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) signaling. Goniolactone C strongly inhibited PDGF-BB-induced VSMC migration and proliferation. goniolactone C-mediated inhibition of VSMC proliferation was associated with cell cycle arrest, while goniolactone C-mediated inhibition of VSMC migration was associated with the suppression of adhesion molecule expression. In addition, goniolactone C directly inhibited PDGFR-β kinase activity, thereby blocking the downstream effector of PDGF-BB. Thus, the results of the present study suggest a novel adjunctive pharmacological strategy that may be used to prevent angioplasty-related restenosis.

Antiangiogenic Activity of Hypoxylonol C

Hypoxylonol C (1), isolated from the inedible mushroom Hypoxylon truncatum, exhibited inhibitory activities against the migration and tube formation of HUVECs. A cDNA microarray analysis was performed to investigate the target of hypoxylonol C (1) in HUVECs, and it was found that the genes related to cell cycle and adhesion were down-regulated. The down-regulation of mRNA levels of cell cycle and adhesion genes was confirmed by real-time RT-PCR. Cell cycle arrest and suppression of adhesion molecule expression might be plausible mechanisms of actions for the antiangiogenic activity of hypoxylonol C (1).

EP01 A stinky remedy for atherosclerosis

Being the last key enzyme of the reverse trans-sulfuration pathway, cystathionine gamma-lyase (CSE) catalyzes the conversion of cystathionine into cysteine, α-ketobutyrate, and ammonia. It further catalyzes cysteine into a gasotransmitter H2S [1] . CSE-mediated H2S production affects lipid metabolism and cardiovascular health, which is exemplarily manifested in the pathogenesis of atherosclerosis [2] . In order to identify the role of endogenous H2S in the development of atherosclerosis, we tried to feed CSE deficient (CSE-KO) mice with a high fat diet as an atherosclerosis model. The control diet for this treatment is the cysteine-limited casein based rodent diet. With this control diet, CSE-KO mice showed growth retardation, decreased cysteine, H2S, and glutathione levels, and increased plasma homocysteine level. None of the CSE-KO mice survived with the cysteine-limited diet. Supplementation of cysteine, but not NaHS, to the animals sustained their lives [3] . The linkage between cysteine biosynthesis and CSE deficiency for life sustainability is thus established [3] , [4] . In the following studies, cysteine had been added to control chow or atherogenic paigen-type diet to feed CSE-KO and WT mice for 12 weeks. While WT mice and CSE-KO mice did not develop any atherosclerosis with the control chow, CSE-KO mice fed with atherogenic diet developed early fatty streak lesions in the aortic root. We further show that increased atherosclerotic lesion formation in CSE-KO mice was associated with elevated plasma cholesterol and LDL-cholesterol levels, enhanced aortic cell proliferation and oxidative stress as well as increased adhesion molecule expression. Supplementation of NaHS to the animals significantly reversed the atherosclerosis development. The mice with double knockout of CSE and apolipoprotein E (apoE) gene expression exhibited more severe atherosclerosis than CSE or apoE knockout alone [5] . Our results demonstrate that a physiological level of endogenous H2S limits the development of atherosclerosis by reducing vessel intimal proliferation and inhibiting adhesion molecule expression, and that the down-regulation of CSE/H2S system predisposes vascular tissues to vascular remodelling and early development of atherosclerosis. We may now claim that H2S is a stinky remedy for atherosclerosis [2] .

Decreased Endogenous Production of Hydrogen Sulfide Accelerates Atherosclerosis

Cystathionine γ-lyase (CSE) produces hydrogen sulfide (H2S) in the cardiovascular system. The deficiency of CSE in mice leads to a decreased endogenous H2S level, an age-dependent increase in blood pressure, and impaired endothelium-dependent vasorelaxation. To date, there is no direct evidence for a causative role of altered metabolism of endogenous H2S in atherosclerosis development. Six-week-old CSE gene knockout and wild-type mice were fed with either a control chow or atherogenic paigen-type diet for 12 weeks. Plasma lipid profile and homocysteine levels, blood pressure, oxidative stress, atherosclerotic lesion size in the aortic roots, cell proliferation, and adhesion molecule expression were then analyzed. CSE-knockout mice fed with atherogenic diet developed early fatty streak lesions in the aortic root, elevated plasma levels of cholesterol and low-density lipoprotein cholesterol, hyperhomocysteinemia, increased lesional oxidative stress and adhesion molecule expression, and enhanced aortic intimal proliferation. Treatment of CSE-knockout mice with NaHS, but not N-acetylcysteine or ezetimibe, inhibited the accelerated atherosclerosis development. Double knockout of CSE and apolipoprotein E gene expression in mice exacerbated atherosclerosis development more than that in the mice with only apolipoprotein E or CSE knockout. Endogenously synthesized H2S protects vascular tissues from atherogenic damage by reducing vessel intimal proliferation and inhibiting adhesion molecule expression. Decreased endogenous H2S production predisposes the animals to vascular remodeling and early development of atherosclerosis. The CSE/H2S pathway is an important therapeutic target for protection against atherosclerosis.

Berberine-attenuated monocyte adhesion to endothelial cells induced by oxidized low-density lipoprotein via inhibition of adhesion molecule expression

Recruitment of monocytes to endothelial cells is important during early stages of atherosclerosis development. This process is predominantly mediated by cellular adhesion molecules, including vascular cell adhesion molecule‑1 (VCAM‑1) and intercellular adhesion molecule‑1 (ICAM‑1), which are expressed by activated endothelial cells in response to a number of inflammatory stimuli, including oxidized low‑density lipoprotein (oxLDL). Previous studies have demonstrated that berberine, a natural extract from Rhizoma coptidis, prevents oxLDL‑induced endothelial cellular apoptosis. However, its effect on the adhesion of monocytes to endothelial cells and the mechanism associated with this process remains unclear. In the present study, berberine was revealed to markedly reduce oxLDL‑induced monocyte adhesion to human umbilical vein endothelial cells. In addition, the inhibitory mechanism of berberine was associated with suppression of adhesion molecule expression, including VCAM‑1 and ICAM‑1. Results indicate that berberine plays a protective role in the early stages of atherosclerosis.

Effect of Tea Polyphenols on the Adhesion of Highly Metastatic Human Lung Carcinoma Cell Lines to Endothelial Cells in Vitro

Tea polyphenols are known to play roles in critical steps of human lung carcinoma cell metastasis. For understanding the mechanisms whereby they inhibit tumor metastasis, the present study was conducted to investigate their effects on the adhesion of highly metastatic lung carcinoma cell lines (PG cells) to endothelial cells (EC cells) and adhesion molecule expression in vitro. The expression of CD44 or CD54 in the PG cells was detected by flow cytometry and adhesion of PG cells to EC cells was assessed by confocal microscopy double fluorescence staining. The results showed that tea polyphenols: (1) inhibited the expression of CD44 and CD54, two important adhesion molecules in the PG cells in a dose-dependent manner; (2) significantly blocked the adhesion of PG cells to EC cells not only in a state of rest but also when active; and (3) influenced CD44 and CD54 expression during the adhesion process of PG cells to EC cells. The data indicated that the blocking role of tea polyphenols in the adhesion of PG cells to EC cells is related to CD44 and CD54. The mechanism of tea polyphenol prevention of human lung carcinoma metastasis might be through inhibiting adhesion molecule expression to block cancer cell adhesion.

A Pentapeptide Monocyte Locomotion Inhibitory Factor Protects Brain Ischemia Injury by Targeting the eEF1A1/Endothelial Nitric Oxide Synthase Pathway

Ischemic stroke is a major cause of death worldwide but lacks viable treatment or treatment targets. Monocyte locomotion inhibitory factor (MLIF) is a small heat-stable pentapeptide produced by Entamoeba histolytica in axenic culture, which is supposed to protect the brain from ischemic injury; the mechanism, however, remains unknown. In this study, we further investigated the mechanism underlying the protective role of MLIF in brain ischemia. A middle cerebral artery occlusion model in rats was used for detecting the effect of MLIF in the brain ischemia in vivo. To identify targets of MLIF in brain endothelial cells, we performed immunoprecipitation of biotin-conjugated MLIF and mass spectrometry. MLIF can protect the brain from ischemic injury in vivo, yielding decreased ischemic volume, prolonged survival, and improved neurological outcome. In vitro studies showed that MLIF displayed protective effects through inhibition of expression of pathological inflammatory adhesion molecules and enhancing endothelial nitric oxide synthase expression and nitric oxide release in the cerebrovascular endothelium. The target screening experiments demonstrated binding of MLIF to the ribosomal protein translation elongation factor eEF1A1. MLIF enhanced endothelial nitric oxide synthase expression through stabilization of endothelial nitric oxide synthase mRNA, and eEF1A1 was shown to be necessary for this enhanced expression. Knockdown of eEF1A1 or inhibition of endothelial nitric oxide synthase attenuated MLIF-mediated inhibition of adhesion molecule expression. In this study, we identified a new potential pharmacologically targetable mechanism underlying MLIF's protective effects in brain ischemia through the eEF1A1/endothelial nitric oxide synthase pathway.

Suppression of adhesion molecule expression by phenanthrene-containing extract of bulbils of Chinese Yam in vascular smooth muscle cells through inhibition of MAPK, Akt and NF-κB

Atherosclerosis is a chronic inflammatory disease associated with increased expression of adhesion molecules in vascular smooth muscle cells (VSMCs). The objective of this study was to examine the in vitro effects of extract from aerial Bulbil of Dioscorea batatas Decne (Db-Ex) on the ability to suppress the expression of adhesion molecules induced by TNF-α. We also identified bioactive components from a methanol extract. VSMCs pre-exposed to Db-Ex (10–100μg/ml) were stimulated with TNF-α (10ng/ml). Preincubation of VSMCs for 2h with Db-Ex dose-dependently inhibited TNF-α-induced adhesion of THP-1 monocytic cells and mRNA and protein expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1). Db-Ex treatment decreased ROS production and the amount of phosphorylated form of p38, ERK, JNK and Akt in TNF-α-stimulated cells, suggesting that Db-Ex inhibits adhesion molecule expression possibly through MAPK and Akt regulation. Db-Ex also suppressed TNF-α-activation NK-κB. This effect was mediated through degradation of IκBα and nuclear translocation of the p65 subunit of NF-κB. These results suggest that Db-Ex inhibits monocyte adhesion and the TNF-α-mediated induction of adhesion molecules in VSMC by downregulating the MAPK/Akt/NF-κB signaling pathway, which may explain the ability of Db-Ex to suppress inflammation within the atherosclerotic lesion.

Transfection of Short-Interfering RNA Silences Adhesion Molecule Expression on Cardiac Microvascular Cells

Acute rejection reactions and the development of graft arterial disease are serious limitations after transplantation. Both are connected to the expression of adhesion molecules on the activated microvascular endothelium of the allograft. siRNA-mediated silencing of ESELE, ICAM-1, and VCAM-1 on human cardiac microvascular cells (HCMEC) was investigated in order to inhibit leukocyte-endothelial interactions. HCMEC were investigated for the time-dependent expression of ESELE, ICAM-1, and VCAM-1 after TNF-α stimulation and for siRNA mediated suppression using a nonviral transfecting approach. Furthermore, the effects of siRNA transfection on leukocyte binding to the endothelium were analyzed. Transfection with siRNA induced a significant suppression of adhesion molecule expression, regardless of whether there had been a prior single or cocktail transfection of the sequences ( P < 0.05). The quantity of attaching leukocytes was significantly reduced after an equal silencing of adhesion molecules ( P < 0.05). This investigation demonstrates that liposomal transfection of HCMEC with specific siRNA sequences is capable of both repressing adhesion molecule expression and of reducing subsequent leukocyte-endothelial actions.

Metabolic conversion of dietary flavonoids alters their anti-inflammatory and antioxidant properties

The notion that dietary flavonoids exert beneficial health effects in humans is often based on in vitro studies using the glycoside or aglycone forms of these flavonoids. However, flavonoids are extensively metabolized in humans, resulting in the formation of glucuronide, methyl, and sulfate derivatives, which may have different properties than their parent compounds. The goal of this study was to investigate whether different chemical modifications of the same flavonoid molecule affect its biological and antioxidant activities. Hence, we studied the anti-inflammatory effects of several major human metabolites of quercetin and (−)-epigallocatechin-3-O-gallate (EGCG) by assessing their inhibitory effects on tumor necrosis factor α (TNFα)-induced protein expression of cellular adhesion molecules in human aortic endothelial cells (HAEC). HAEC were incubated with 1–30μM quercetin, 3′- or 4′-O-methyl-quercetin, quercetin-3-O-glucuronide, and quercetin-3′-O-sulfate or 20–100μM EGCG, 4′′-O-methyl-EGCG, and 4′,4′′-di-O-methyl-EGCG, prior to coincubation with 100 U/ml of TNFα. 3′-O-Methyl-quercetin, 4′-O-methyl-quercetin, and their parent aglycone compound, quercetin, all effectively inhibited expression of intercellular adhesion molecule-1 (ICAM-1) with IC50 values (concentration required for 50% inhibition) of 8.0, 5.0, and 4.4μM, respectively; E-selectin expression was suppressed to a somewhat lesser but still significant degree by all three compounds, whereas vascular cell adhesion molecule-1 (VCAM-1) was not affected. In contrast, quercetin-3-O-glucuronide (20–100μM), quercetin-3′-O-sulfate (10–30μM), and phenolic acid metabolites of quercetin (20–100μM) did not inhibit adhesion molecule expression. 4′,4′′-Di-O-methyl-EGCG selectively inhibited ICAM-1 expression with an IC50 value of 94μM, whereas EGCG (20–60μM) and 4′′-O-methyl-EGCG (20–100μM) had no effect. The inhibitory effects of 3′-O-methyl-quercetin and 4′,4′′-di-O-methyl-EGCG on adhesion molecule expression were not related either to inhibition of NF-κB activation or to their antioxidant reducing capacity. Our data indicate that flavonoid metabolites have different biological and antioxidant properties than their parent compounds, and suggest that data from in vitro studies using nonmetabolites of flavonoids are of limited relevance in vivo.

Aurintricarboxylic acid inhibits the nuclear factor-κB-dependent expression of intercellular cell adhesion molecule-1 and endothelial cell selectin on activated human endothelial cells

Activation of the vascular endothelium and increased adhesion of circulating leukocytes to the activated endothelium are important events in inflammation and coagulation. Aurintricarboxylic acid (ATA), a triphenylmethyl dye compound, is known to inhibit platelet adhesion by interfering with the binding of von Willebrand factor to platelet glycoprotein Ib. However, the effect of ATA on the inflammatory response of endothelial cells has not yet been investigated. Here, we investigated the functional role and molecular mechanism of ATA on the activation of human endothelial cells. ATA inhibited the expression of intercellular adhesion molecule-1 (ICAM-1), and endothelial cell selectin (E-selectin) was upregulated on human umbilical vein endothelial cells (HUVECs) in response to tumor necrosis factor-α or lipopolysaccharide (LPS). We also observed the inhibitory effect of ATA on LPS-induced mRNA expression of ICAM-1 and E-selectin. Furthermore, ATA inhibited the binding of leukocytes to activated HUVECs. ATA significantly inhibited the nuclear translocation of nuclear factor-κB (NF-κB) and degradation of IκB on activated HUVECs, suggesting that ATA inhibits NF-κB signaling. Finally, three NF-κB inhibitors effectively inhibited the expressions of ICAM-1 and E-selectin on activated endothelial cells. The present data suggest that ATA exerts beneficial effect in various inflammation conditions through inhibition of adhesion molecule expression in activated endothelial cells and the resulting inhibition of leukocytes tissue accumulation.

抗GBM腎炎ラットの糸球体における接着分子発現に対する柴苓湯(TJ-114)の効果

In order to clarify the antinephritic mechanisms of Sairei-to (TJ-114), the effects of TJ-114 on the expression of adhesion molecules were evaluated in rats with anti-GBM nephritis. TJ-114 was administered orally once a day from the day (1st day) after anti-GBM serum injection throughout the experiment. TJ-114 inhibited crescent formation in glomeruli at the 10th day compared to control rats with anti-GBM nephritis. The ICAM-1- or VCAM-1-positive area in the glomeruli was increased in the nephritic control rats at the 10th day. In contrast, TJ-114 prevented increase in the ICAM-1- or VCAM-1-positive area in the glomeruli of nephritic rats. TJ-114 inhibited increase in the number of LFA-1- or VLA-4-positive cells in the glomeruli. One of the constituent components of TJ-114, Syo-saiko-to (TJ-9) at the dose of 750 mg/kg, also significantly inhibited increase in the ICAM-1- or VCAM-1-positive area in the glomeruli. Gorei-san (TJ-17), another component of TJ-114, inhibited increase in the ELAM-1-positive area and increase in the number of VLA-4-positive cells in the glomeruli. Prednisolone markedly inhibited increase in the positive area of those adhesion molecules and increase in the number of ligand-molecule-positive cells in the glomeruli. These results indicate that the antinephritic action of TJ-114 may be partially due to inhibition of adhesion molecule expression in the glomeruli.