Inhibition Of ICAM-1 Expression Research Articles

Study on the Antioxidant and Anti-Inflammatory Effects of Scutellaria baicalensis Extract and Fractions Based on Indicator Compound Content

The skin, as a barrier against environmental stressors, is highly affected by rapid industrialization and pollution, leading to oxidative stress, aging, and inflammation. Growing interest in natural antioxidants, especially in functional cosmetics, has highlighted the need for safe, effective alternatives to synthetic ingredients. <i>Scutellaria baicalensis</i> is traditionally known for its antioxidant and anti-inflammatory properties, but research on its specific fractions remains limited. This study explores both the overall and fraction-specific efficacy of <i>Scutellaria baicalensis</i> extract for potential use in skincare. This study summarizes the evaluation of various biological activities of <i>Scutellaria baicalensis</i> extract and its fractions. In the first stage, DPPH and ABTS radical scavenging activities were investigated, identifying fractions with higher scavenging activity than the extract itself. In the second stage, cell viability was tested to determine concentrations that do not induce cytotoxicity. In RAW 264.7 macrophages, inflammation was induced using LPS, and the suppression of Nitrite production demonstrated the anti-inflammatory effects. In HaCaT cells, skin inflammation was induced with TNF-α and IFN-γ, and the reduction in interleukin (IL)-6 production was observed. Additionally, specific fractions exhibited a significant ability to regulate key chemokines like IL-8, RANTES, and TARC. The study also showed that the inhibition of ICAM-1 expression helped control cellular adhesion in the early stages of inflammation. Moreover, HPLC analysis confirmed that specific fractions contained higher concentrations of active ingredients compared to the extract, reinforcing their potential as natural cosmetic ingredients. <i>Scutellaria baicalensis</i> fractions showed exceptional antioxidant and anti-inflammatory effects, even in small amounts, indicating their potential use in functional cosmetics and anti-aging products. Future studies are expected to explore the development of natural cosmetic ingredients from these fractions, contributing to the growth of the functional cosmetics and anti-aging market.

Thrombopoietin mimetic reduces mouse lung inflammation and fibrosis after radiation by attenuating activated endothelial phenotypes.

Radiation-induced lung injury (RILI) initiates radiation pneumonitis and progresses to fibrosis as the main side effect experienced by patients with lung cancer treated with radiotherapy. There is no effective drug for RILI. Sustained vascular activation is a major contributor to the establishment of chronic disease. Here, using a whole thoracic irradiation (WTI) mouse model, we investigated the mechanisms and effectiveness of thrombopoietin mimetic (TPOm) for preventing RILI. We demonstrated that administering TPOm 24 hours before irradiation decreased histologic lung injury score, apoptosis, vascular permeability, expression of proinflammatory cytokines, and neutrophil infiltration in the lungs of mice 2 weeks after WTI. We described the expression of c-MPL, a TPO receptor, in mouse primary pulmonary microvascular endothelial cells, showing that TPOm reduced endothelial cell-neutrophil adhesion by inhibiting ICAM-1 expression. Seven months after WTI, TPOm-treated lung exhibited less collagen deposition and expression of MMP-9, TIMP-1, IL-6, TGF-β, and p21. Moreover, TPOm improved lung vascular structure, lung density, and respiration rate, leading to a prolonged survival time after WTI. Single-cell RNA sequencing analysis of lungs 2 weeks after WTI revealed that TPOm shifted populations of capillary endothelial cells toward a less activated and more homeostatic phenotype. Taken together, TPOm is protective for RILI by inhibiting endothelial cell activation.

Mechanisms underlying neutrophils adhesion to triple-negative breast cancer cells via CD11b-ICAM1 in promoting breast cancer progression

BackgroundTriple-negative breast cancer (TNBC) is recognized as the most aggressive and immunologically infiltrated subtype of breast cancer. A high circulating neutrophil-to-lymphocyte ratio (NLR) is strongly linked to a poor prognosis among patients with breast cancer, emphasizing the critical role of neutrophils. Although the involvement of neutrophils in tumor metastasis is well documented, their interactions with primary tumors and tumor cells are not yet fully understood.MethodsClinical data were analyzed to investigate the role of neutrophils in breast cancer. In vivo mouse model and in vitro co-culture system were used for mechanism researches. Blocking experiments were further performed to identify therapeutic agents against TNBC.ResultsTNBC cells secreted GM-CSF to sustain the survival of mature neutrophils and upregulated CD11b expression. Through CD11b, neutrophils specifically binded to ICAM1 on TNBC cells, facilitating adhesion. Transcriptomic sequencing combined with human and murine functional experiments revealed that neutrophils, through direct CD11b-ICAM1 interactions, activated the MAPK signaling pathway in TNBC cells, thereby enhancing tumor cell invasion and migration. Atorvastatin effectively inhibited ICAM1 expression in tumor cells, and tumor cells with ICAM1 knockout or treated with atorvastatin were unresponsive to neutrophil activation. The MAPK pathway and MMP9 expression were significantly inhibited in the tumor tissues of TNBC patients treated with atorvastatin.ConclusionsTargeting CD11b-ICAM1 with atorvastatin represented a potential clinical approach to reduce the malignant characteristics of TNBC.Graphical

Discovery of common molecular signatures and drug repurposing for COVID-19/Asthma comorbidity: ACE2 and multi-partite networks

ABSTRACT Angiotensin-converting enzyme 2 (ACE2) is identified as the functional receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of the ongoing global coronavirus disease-2019 (COVID-19) pandemic. This study aimed to elucidate potential therapeutic avenues by scrutinizing approved drugs through the identification of the genetic signature associated with SARS-CoV-2 infection in individuals with asthma. This exploration was conducted through an integrated analysis, encompassing interaction networks between the ACE2 receptor and common host (co-host) factors implicated in COVID-19/asthma comorbidity. The comprehensive analysis involved the identification of common differentially expressed genes (cDEGs) and hub-cDEGs, functional annotations, interaction networks, gene set variation analysis (GSVA), gene set enrichment analysis (GSEA), and module construction. Interaction networks were used to identify overlapping disease modules and potential drug targets. Computational biology and molecular docking analyzes were utilized to discern functional drug modules. Subsequently, the impact of the identified drugs on the expression of hub-cDEGs was experimentally validated using a mouse model. A total of 153 cDEGs or co-host factors associated with ACE2 were identified in the COVID-19 and asthma comorbidity. Among these, seven significant cDEGs and proteins – namely, HRAS, IFNG, JUN, CDH1, TLR4, ICAM1, and SCD—were recognized as pivotal host factors linked to ACE2. Regulatory network analysis of hub-cDEGs revealed eight top-ranked transcription factors (TFs) proteins and nine microRNAs as key regulatory factors operating at the transcriptional and post-transcriptional levels, respectively. Molecular docking simulations led to the proposal of 10 top-ranked repurposable drug molecules (Rapamycin, Ivermectin, Everolimus, Quercetin, Estradiol, Entrectinib, Nilotinib, Conivaptan, Radotinib, and Venetoclax) as potential treatment options for COVID-19 in individuals with comorbid asthma. Validation analysis demonstrated that Rapamycin effectively inhibited ICAM1 expression in the HDM-stimulated mice group (p < 0.01). This study unveils the common pathogenesis and genetic signature underlying asthma and SARS-CoV-2 infection, delineated by the interaction networks of ACE2-related host factors. These findings provide valuable insights for the design and discovery of drugs aimed at more effective therapeutics within the context of lung disease comorbidities.

MiR-146a encapsulated liposomes reduce vascular inflammatory responses through decrease of ICAM-1 expression, macrophage activation, and foam cell formation.

Vascular insults can create an inflammatory cascade involving endothelial cell, smooth muscle cell, and macrophage activation which can eventually lead to vascular disease such as atherosclerosis. Several studies have identified microRNA 146a's (miR-146a) anti-inflammatory potential based on its role in regulating the nuclear factor kappa beta (NF-κβ) pathway. Therefore, in this study, we introduced exogenous miR-146a encapsulated by liposomes to lipopolysaccharide (LPS) stimulated vascular cells and macrophages to reduce inflammatory responses. First, the miR-146a encapsulated liposomes showed uniform size (radius 96.4 ± 4.22 nm) and round shape, long term stability (at least two months), high encapsulation efficiency (69.73 ± 0.07%), and were well transfected to human aortic endothelial cells (HAECs), human aortic smooth muscle cells (SMCs), and human differentiated monocytes (U937 cells). In addition, we demonstrated that miR-146a encapsulated liposomes reduced vascular inflammation responses in HAECs and SMCs through inhibition of ICAM-1 expression and decreased monocyte adhesion. In macrophages, miR-146a liposome treatment demonstrated decreased production of proinflammatory cytokines, tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β), as well as reduced oxidized low-density lipoprotein (ox-LDL) uptake and foam cell formation. Thus, based on these results, miR-146a encapsulated liposomes may be promising for reducing vascular inflammation by targeting its multiple associated mediators.

Mesenchymal stem cell exosome-derived miR-223 alleviates acute graft-versus-host disease via reducing the migration of donor T cells

BackgroundMesenchymal stem cells (MSCs) have been utilized in treating acute graft-versus-host disease (aGvHD) as they show strong immunosuppressive capacity through the release of various mediators, including immunosuppressive molecules, growth factors, chemokines, and exosomes. MicroRNAs (miRNAs) derived from MSC exosomes (MSCs-Exo) play a critical role in the regulation of immune responses. However, the function of miRNAs in treating aGvHD remains unknown. Here, we performed expression profiling of exosome-miRNAs from human umbilical cord MSCs (huc-MSCs) and murine compact bone MSCs (mb-MSCs) to investigate their immunoregulation effects in aGvHD.MethodsHuc-MSCs-Exo and mb-MSCs-Exo were isolated and constructed MSCs-Exo-derived miRNA expression profiling using high-throughput sequencing. High expression of miR-223 was identified in both kinds of MSCs-Exo by bioinformatics analysis and quantitative real-time PCR (qPCR). In vitro cell crawling assay, transmigration assay and adhesion assay were subsequently applied to investigate the regulation of miR-223 on T cells. MiR-223 target gene was analyzed by western blot, luciferase analysis, and qPCR. Moreover, murine aGvHD model was established by infusing splenocytes and bone marrow nuclear cells from C57BL/6j mice (H-2Kb) into BALB/c recipient mice (H-2Kd). For therapeutic effect, MSCs or miR-223 Agomir were injected via tail vein. The general conditions of the mice in each group were monitored. Hematoxylin-eosin (H&E) staining was used to detect pathological changes of mice spleen, liver, and intestine. Mechanistically, immunofluorescence and flow cytometry were used to evaluate donor T cell migration, and enzyme-linked immunosorbent assay (ELISA) was used to detect the expression of serum inflammatory cytokines IFN-γ, TNF-α, and IL-17.ResultsHigh-throughput sequencing revealed high expression of miR-223 in huc-MSCs-Exo and mb-MSCs-Exo. MiR-223 could restrain adhesion and migration of T cells by inhibiting ICAM-1 expression in mouse lymphatic endothelial cells. MiR-223Agomir infusion attenuated aGvHD clinical symptoms, reduced the donor T cell infiltration into the spleen, liver, and intestine, and decreased inflammatory cytokines IFN-γ, TNF-α, and IL-17.ConclusionMSCs-Exo-derived miR-223 could attenuate aGvHD in mice through decreasing donor T cell migration. Our results unveil a new role of MSCs-Exo containing miR-223 in the treatment of aGvHD.

Tissue Integrity and COVID-19

Tissue integrity depends on biological tissue stiffness. Tissue integrity can protect both against age-related diseases and against severity of COVID-19. The disruption of tight junctions and increase of tissue permeability with advancing age can be related with age-related diseases as well as with age-dependent COVID-19. Release of tightly bound water from collagen fibrils leads to the increase of extracellular matrix stiffness and to the associated with matrix stiffness increased tissue permeability. The link between arterial stiffness and oxidative stress has been reported and is expected to be studied in more detail in the future. Trehalose can be suggested for retardation of tightly bound water release and subsequent extracellular matrix crosslinking by advanced glycation end products. Increase in tissue permeability can be blocked by polyphenols that inhibit ICAM-1 expression and mitigate cytoskeleton reorganization. NF-κB activation as a result of increased stiffness and cytoskeleton reorganization can cause both cardiovascular pathologies and COVID-19. Increased cholesterol content in cell membrane leads to increased virus entry into cell and increase of cholesterol is linked with cardiovascular diseases. Statins and chitosan are known as cholesterol-lowering substances. Nrf2 inhibits NF-κB activation and NF-κB inhibits Nrf2 pathway.

P3475Dabigatran inhibits the activation of endothelial progenitor cells induced by thrombin

Abstract Introduction Thrombin is a coagulation serine protease, which also activates various cell types, including endothelial cells. Late-outgrowth endothelial cells (OECs) are a type of endothelial progenitor cells that contribute to endothelial regeneration and angiogenesis. Dabigatran is a direct oral anticoagulant that inhibits thrombin's action and is widely used in everyday clinical practice. Purpose We investigated the effect of dabigatran on thrombin-induced activation of OECs, using the membrane expression of adhesion molecule ICAM-1 as activation marker. Control experiments were conducted in mature human umbilical vein endothelial cells (HUVECs). Methods CD34+ cells were isolated from cord blood mononuclear cells, using human CD34 Microbead Kit and appropriately cultured for OEC formation. HUVECs were purchased from Lonza. Confluent OECs and HUVECs were incubated with 10 or 20 μM dabigatran for 10 min, before activation with 8 U/mL thrombin, for 24 h. The effect of dabigatran on ICAM-1 expression (anti-CD54-PE) was evaluated in both cell types as % gated CD31+/CD54+ cells, using flow cytometry. Results Thrombin induced ICAM-1 membrane expression by 33±15% on OECs and by 123±22% on HUVECs, compared with respective untreated cells (p&lt;0.05 for both comparisons, from 6 different experiments). Dabigatran at 10 μM significantly inhibited ICAM-1 expression by 36±7% on OECs and 57±8% on HUVECs, as well as at 20 μM by 51±8% on OECs and 79±7% on HUVECs (p&lt;0.05 for all comparisons, from 4 different experiments). Conclusions We show for the first time that dabigatran at concentrations relevant to those existing in vivo after oral administration, inhibits the thrombin-induced membrane expression of the adhesion molecule ICAM-1 on OECs, a phenomenon that is also observed on mature endothelial cells. The significance of this effect regarding the pathophysiological role of OECs and mature endothelial cells at the clinical level remains to be established. Acknowledgement/Funding Operational Program “Human Resources Development, Education and Lifelong Learning”. Co-financed by the European Union and Greek national funds

Effect of Quercetin on the Cell Cycle and Adhesion Molecules of NOD/SCID Mice with Acute B Lymphocytic Leukemia

To investigate the effect of Quercetin on cell cycle and adhesive molecules of NOD.SCID mice with acule B lymphocytic leuaemia(B-ALL). 5×106 Nalm-6(B-ALL cell line) cells were injected into the tail vein of 48 NOD/SCID mice to establish the NOD/SCID mice with B-ALL. After 15 day, the NOD/SCID mice with B-ALL were randomly divided into 3 groups: salive group as control (injection with saline of 0.2 ml/mouse), cyclophos-phamid group (injection with cyclophosphamide of 100µg/kg) and quercetin group(injection with quercetin of 3 mg/kg). After treatment for 21 d, the perecntage of Nalm-6 cells in G1, G2, M and S phases was detected by flow cytonetry; the B lymphocytes Nalm-6 cells, neutrophils and WBC in while blood were counted before and after treatment; the expression of intercellalar. Adhesion molecole-1(FCAIU-1), vascular cell adhesion molecule-1(VCAM-1) and P-selectin was detected by double autibody soundwich method. Compared with level before treatment, the expression of ICAM-1, VCAM-1 and P-selectin decreased after treatment with guercetin, The hemogram showed that the peripheral blood nentrophil level obviously increased, while the levels of B lymphocytes, Nalm-6 cells and WBC count decreased obviously after treatment with guercetin. The cell proliferatim rario in G0/G1 phase decreased, yet the cell proliferation ratio in S and G2/M phases increased after treatment with guercetin. The guercetin can decrease the intercellular adhesion through inhibition of ICAM-1 expression, and arrests Nalm-6 cells in S and G2/M phases. The guercetin has obviously inhibitory effect on B-ALL cells.

P02.12 Moringa oleifera, Lam extract as anti tumor through inhibition of ICAM-1 expression in brain tissue of white rat with metabolic syndrome

P02.12 Moringa oleifera, Lam extract as anti tumor through inhibition of ICAM-1 expression in brain tissue of white rat with metabolic syndrome

Angiopoietin-Like Protein 3 Directly Regulates HDL Metabolism and Function

Background: Angiopoietin-like protein 3 (ANGPTL3) is produced by hepatocytes. Once secreted, it inhibits endothelial lipase activity and regulates angiogenesis. We previously reported the positive association between serum ANGPTL3 and HDL function in nondiabetic participants. Hereby, we investigated whether ANGPTL3 directly modulates HDL function. Methods: HDLs were isolated from plasma. ANGPTL3 in HDLs was studied by western blot and ELISA. Cholesterol efflux to HDLs was analyzed in macrophages preloaded with fluorescent cholesterol. Recombinant human ANGPTL3 was subjected to endothelial cells treated with TNF-α and ICAM-1 expression was analyzed by FACS. Similar experiments were repeated in db mice injected with AAV encoding human ANGPTL3 cDNA or shRNA. Results: ANGPTL3 was present in human HDLs, which were lower in T2DM patients than controls (p=0.04). ANGPLT3 in HDLs was positively associated with cholesterol efflux in nondiabetic controls (r=0.33, p=0.044) but not in T2DM patients (r=0.05, p=0.77). Recombinant ANGPTL3 reduced ICAM-1 expression in endothelial cells exposed to TNF-α (p=0.0001). Similarly, ANGPTL3 levels in HDLs were lower in db mice at 18 weeks old than 8 weeks old, which was accompanied with reduced cholesterol reflux to HDLs and rendered inhibition of ICAM-1 expression in endothelial cells (p&amp;lt;0.for all). Following AAV-mediated ANGPTL3 cDNA transfer, ANGTL3 levels were increased in HDLs of db mice, which increased cholesterol efflux and decreased ICAM-1 expression compared with db mice with vector transfer. Vice versa, decreased ANGPTL3 levels in HDLs by AAV-mediated shRNA transfer abrogated HDL function mentioned above (p&amp;lt;0.for both). When placed on high fat diet for 10 weeks, atherosclerotic plaque was larger in db mice with ANGPTL3 shRNA transfer than other gene transferred groups (p&amp;lt;0.05). Conclusion: ANGPTL3 is a HDL component and direct regulates HDL function. Decrease of ANGPTL3 expression may reduce HDL function and accelerate atherosclerosis in T2DM. Disclosure L. Yang: None. Y. Wang: None. Y. Fu: None. J. Lang: None. D. Zhao: None. Y. Feng: None.

Abstract 029: Interleukin-35 Suppresses Endothelial Activation by Inhibiting Mitochondrial Reactive Oxygen Species Mediated Site-specific Acetylation of Histone 3 Lysine 14

Recently we reported that IL-35 is an inflammation-induced ant-inflammatory cytokine; thus, function as a HAMP (homeostasis associated molecular patterns). However, whether IL-35 affects endothelial cell activation and subsequent development of atherosclerosis is not known. Hence, we studied the expression of IL-35 during early atherosclerosis; and its roles and mechanisms in suppressing EC activation. Initially, we analyzed whether endogenous IL-35 levels and its receptor expression was increased in early atherosclerosis. Our data revealed that IL-35 level was significantly increased in the plasma of hyperlipidemic patients and apolipoprotein E (ApoE-/-) deficient mice. Further, the expression of IL-35 receptor components were increased in ApoE-/- mouse aortas. IL-35 inhibited monocyte recruitment on to human aortic endothelial cells (EC) activated by pro-atherogenic lysophosphatidylcholine (LPC). Furthermore, our RNA-Seq analysis showed that IL-35 selectively inhibited ICAM-1 expression that mediates EC activation. Further analysis by flowcytometry and electron spin resonance analysis revealed that IL-35 blocked LPC-mediated mitochondrial-reactive oxygen species (mtROS) production. The subsequent mass spectrometry, ChIP-Seq analysis and EMSA showed that LPC-mediated mtROS promotes acetylation of histone 3 lysine 14 (H3K14). Acetylation of histones facilitate the transcription of EC activation-related genes. We found that acetylated H3K14 increased the binding of pro-inflammatory transcription factor Ap-1 on to ICAM-1 promoter region and induction of ICAM-1 gene transcription. IL-35 mediated Inhibition of mtROS attenuated H3K14 acetylation, inhibited AP-1 binding to ICAM-1 promoter and suppressed ICAM-1 transcription. Finally, IL-35 cytokine treatment inhibited the progression and development of atherosclerosis in ApoE-/- mice. Our study concluded that IL-35 is induced during atherosclerosis and plays an anti-atherogenic role by suppressing EC activation through mtROS and H3K14 acetylation dependent mechanism.

AMA0428, A Potent Rock Inhibitor, Attenuates Early and Late Experimental Diabetic Retinopathy

ABSTRACTPurpose: Diabetic retinopathy (DR) is characterized by an early stage of inflammation and vessel leakage, and an advanced vasoproliferative stage. Also, neurodegeneration might play an important role in disease pathogenesis. The aim of this study was to investigate the effect of the Rho kinase (ROCK) inhibitor, AMA0428, on these processes.Methods: The response to ROCK inhibition by AMA0428 (1 µg) was studied in vivo using the murine model for streptozotocin (STZ)-induced diabetes, focusing on early non-proliferative DR features and the oxygen-induced retinopathy (OIR) model to investigate proliferative DR. Intravitreal (IVT) administration of AMA0428 was compared with murine anti-VEGF-R2 antibody (DC101, 6.2 µg) and placebo (H2O/PEG; 1C8). Outcome was assessed by analyzing leukostasis using fluorescein isothiocyanate coupled concanavalin A (FITC-ConA) and vessel leakage (bovine serum albumin conjugated with fluorescein isothiocyanate; FITC-BSA)/neovascularization and neurodegeneration by immunohistological approaches (hematoxylin and eosin (H&E), terminal deoxynucleotidyl transferase–mediated biotinylated UTP nick end labeling (TUNEL), Brn3a). ELISA and Western blotting were employed to unravel the consequences of ROCK inhibition (1 µM AMA0428) on myosin phosphatase target protein (MYPT)-1 phosphorylation, endothelial nitric oxide synthase (eNOS) phosphorylation, and vascular endothelial growth factor (VEGF) levels in retinas of diabetic mice, on NF-κβ activity and ICAM-1 expression in endothelial cells (ECs).Results: In vivo, AMA0428 significantly reduced vessel leakage and neovascularization, respectively, in the STZ and OIR model, comparable to DC101 therapy. Additionally, the ROCK inhibitor decreased neurodegeneration in both models and inhibited leukostasis by 30% (p < 0.05) in the STZ model (p < 0.05), while DC101 had no positive effect on the outcome of these latter processes. ROCK activity was upregulated in the diabetic retina and AMA0428 administration resulted in decreased phospho-MYPT-1, enhanced phospho-eNOS, and reduced VEGF levels. In vitro, AMA0428 interfered with NF-κβ activity, thereby inhibiting ICAM-1 expression in ECs.Conclusions: Targeting ROCK with AMA0428 effectively attenuated outcome in an early DR model (STZ) and a late vasoproliferative retinopathy model (OIR). These findings make AMA0428 a promising candidate with an additional anti-inflammatory and neuroprotective benefit for DR patients, as compared with anti-VEGF treatment.

Lycopene inhibits ICAM-1 expression and NF-κB activation by Nrf2-regulated cell redox state in human retinal pigment epithelial cells

AimsAge-related macular degeneration (AMD) is one of the most common diseases leading to blindness in elderly people. The progression of AMD may be prevented through anti-inflammation and antioxidation in retinal pigment epithelium (RPE) cells. Lycopene, a carotenoid, has been shown to possess both antioxidative and anti-inflammatory properties. This research was conducted to detail the mechanisms of these effects of lycopene-treated RPE cells. Main methodsWe exposed ARPE-19 cells to TNFα after pretreatment with lycopene, and measured monocyte adhesion, ICAM-1 expression, NF-κB nuclear translocation, and transcriptional activity. Cell viability was assayed with Alamar Blue. The cell redox state was tested by glutathione (GSH) and reactive oxygen species (ROS) levels. The importance of the Nrf2 pathway was tested in nuclear translocation, promoter reporter assay, and siRNA. Key findingsLycopene could reduce TNF-α-induced monocyte adhesion and H2O2– induced cell damage in RPE cells. Furthermore, lycopene inhibits ICAM-1 expression and abolishes NF-κB activation for up to 12h in TNFα-treated RPE cells. Lycopene upregulates Nrf2 levels in nuclear extracts and increases the transactivity of antioxidant response elements. The use of Nrf2 siRNA blocks the inhibitory effect of lycopene in TNF-α-induced ICAM-1 expression and NF-κB activation. Glutamate-cysteine ligase (GCL) is the rate-limiting enzyme in the de novo synthesis of GSH. We found that lycopene increases intracellular GSH levels and GCL expression. Following lycopene treatment, TNF-α-induced ROS production was abolished. SignificanceThe Nrf2-regulated antioxidant property plays a pivotal role in the anti-inflammatory mechanism underlying the inhibition of NF-κB activation in lycopene-treated ARPE-19 cells.

Abstract 357: Antioxidant and Anti-inflammatory Strategies Prevent Endothelial Dysfunction in Chronic Kidney Disease

Accelerated atherosclerosis in chronic kidney disease (CKD) is preceded by the development of endothelial dysfunction (ED), with development of a proinflammatory and prothrombotic phenotype and enhanced oxidative stress. The effect of anti-inflammatory and antioxidant strategies on the endothelium has been evaluated in an in vitro model of ED in uremia. Endothelial cells (ECs) were pretreated with the antioxidant enzyme mimetics ebselen, EUK-134 and EUK-118; the flavonoids apigenin, genistein and quercetin, with both antioxidant and anti-inflammatory potential; and two commercially available compounds: N-acetylcysteine (NAC) and defibrotide (DF). There is increasing evidence demonstrating that both NAC and DF exhibit both properties. ECs were exposed to medium containing serum from patients on dialysis (n=10) or from healthy donors (n=15). Changes in the expression of the adhesion receptor ICAM-1 and the production of intracellular reactive oxygen species (ROS) were assessed. Activation of inflammation-related proteins p38 MAPK and NFkappaB (NFκB) were also evaluated. Exposure of ECs to uremic media resulted in a significantly increased expression of ICAM-1, overproduction of ROS and activation of p38MAPK and NFκB compared to control ECs (p&lt;0.05). Ebselen, EUK 134, and EUK118 inhibited ICAM-1 expression and ROS generation in the uremic condition (p&lt;0.01). Regarding flavonoids, only quercetin showed a moderated but significant inhibitory effect on both parameters (p&lt;0.05). NAC and DF exhibited a protective effect on ECs exposed to the uremic insult (p&lt;0.05 for ICAM-1 expression and ROS generation). All the compounds reduced p38MAPK activation (p&lt;0.05). The antioxidant-enzyme mimetics and NAC were able to inhibit the activation of NFκB induced by the uremic media (p&lt;0.05). Endothelial dysfunction associated with CKD is considered to be the first step in the progression of atherosclerosis. Our results indicate that the antioxidant enzyme mimetics, NAC and DF exhibit not only antioxidant but also anti-inflammatory effects on the endothelium. Therefore, further research on the protective effects of these compounds may provide new strategies for the prevention of the cardiovascular complications in uremia.

A novel class of copper(II)- and zinc(II)-bound non-steroidal anti-inflammatory drugs that inhibits acute inflammation in vivo.

BackgroundThe ability of Zn(II) and Cu(II) metal complexes of non-steroidal anti-inflammatory drugs (NSAIDs) to inhibit acute arterial inflammation in vivo has been studied. ResultsWhen acute vascular inflammation was induced in normocholesterolemic New Zealand White rabbits by inserting a non-occlusive silastic collar around the common carotid artery, a single oral dose of Cu(II)-indomethacin (Cu(II)Indo, 3 mg/kg) administered by laparotomy achieved a 67 % (8.2 ± 1.7 vs. 2.7 ± 0.4 image units, p < 0.05) reduction in endothelial expression of vascular cell adhesion molecule-1 (VCAM-1) but did not inhibit endothelial intercellular adhesion molecule (ICAM-1) expression significantly. Treatment with Cu(II)-acemetacin (Cu(II)ACM, 3 mg/kg) led to a profound 88 % (8.2 ± 1.7 vs. 1.0 ± 0.5 image units, p < 0.01) reduction in endothelial VCAM-1 expression but did not inhibit ICAM-1 expression, while treatment with Zn(II)-acemetacin (Zn(II)ACM, 3 mg/kg) led to an 84 % (19.3 ± 1.0 vs. 3.1 ± 1.2 image units, p < 0.01) reduction in endothelial ICAM-1 expression and did not inhibit VCAM-1 expression. No adverse gastric, hepatic or renal effects were observed in treated animals. ConclusionThese findings provide the “proof of concept” that this novel class of drug, where there is complexation of NSAIDs with metal ions, has substantial anti-inflammatory effects in an animal model of acute vascular inflammation with the possibility of low rates of adverse effects.

Hypochlorite-Modified Albumin Upregulates ICAM-1 Expression via a MAPK-NF-κB Signaling Cascade: Protective Effects of Apocynin.

Hypochlorite-modified albumin (HOCl-alb) has been linked to endothelial dysfunction, which plays an important role in the development of hypertension, diabetes, and chronic kidney disease. However, whether HOCl-alb induces endothelial dysfunction via vascular inflammation and whether a signaling pathway is involved are unknown and have not been investigated. HOCl-alb was found to upregulate ICAM-1 expression in human umbilical vein endothelial cells (HUVECs) in a time- and dose-dependent manner. HOCl-alb time-dependently phosphorylated ERK1/2 and p38MAPK. HOCl-alb also activated NF-κB. ICAM-1 expression was dose-dependently inhibited by U0126 (a specific inhibitor of MEK1/2, a signal upstream from ERK1/2), SB203580 (a specific inhibitor of p38MAPK), and SN50 (a specific inhibitor of NF-κB). U0126 and SB203580 both counteracted the activation of NF-κB, whereas the phosphorylation of ERK1/2 and p38MAPK was not blocked by SN50. ERK1/2 phosphorylation was blocked by U0126 but not by SB203580, and p38MAPK activity was reduced by SB203580 but not by U0126. Apocynin, a specific NADPH oxidase (NOX) inhibitor, inhibited ICAM-1 expression and the activity of ERK1/2, p38MAPK, and NF-κB. These results indicate that HOCl-alb-induced ICAM-1 expression is caused by the activation of a redox-sensitive intracellular signal cascade involving ERK1/2 and p38MAPK, culminating in the activation of NF-κB and involving NOXs among the upstream signals.

MicroRNA-141 regulates the expression level of ICAM-1 on endothelium to decrease myocardial ischemia-reperfusion injury.

A growing number of studies have suggested microRNAs (miRNAs) are involved in the modulation of myocardial ischemia-reperfusion (MI/R) injury; however, the role of endogenous miRNAs targeting endothelial cells (ECs) and its interaction with ICAM-1 in the setting of MI/R remain poorly understood. Our microarray results showed that miR-146a, miR-146b-5p, miR-155*, miR-155, miR-497, and miR-451 were significantly upregulated, whereas, miR-141 and miR-564 were significantly downregulated in the ECs challenged with TNF-α for 6 h. Real-time PCR analyses additionally validated that the expression levels of miR-146a, miR-155*, and miR-141 were consistent with the microarray results. Then, ICAM-1 was identified as a novel target of miR-141 by Target Scan software and the reporter gene system. Further functional experiments showed that elevated levels of miR-141 inhibited ICAM-1 expression and diminished leukocytes adhesion to ECs in vitro. In an in vivo murine model of MI/R injury, pretreatment with miR-141 mimics through the tail vein downregulated the expression level of ICAM-1 in heart and attenuated MI/R injury as evidenced by decreased infarct size and decline of serum cardial troponin I (cTnI) and lactate dehydrogenase (LDH) concentration. The cardioprotective effects of miR-141 mimics may be attributed to the decreased infiltration of CD11b(+) cells and F4/80(+) macrophages into ischemic myocardium tissue. In conclusion, our results demonstrate that miR-141, as a novel repressor of ICAM-1, is involved in the attenuation of MI/R injury via antithetical regulation of ICAM-1 and inflammatory cells infiltration. Thus miR-141 may constitute a new therapeutic target in the setting of ischemic heart disease.

Lipoxin A4 attenuates endothelial dysfunction during experimental cerebral malaria.

A breakdown of the brain-blood barrier (BBB) due to endothelial dysfunction is a primary feature of cerebral malaria (CM). Lipoxins (LX) are specialized pro-resolving mediators that attenuate endothelial dysfunction in different vascular beds. It has already been shown that LXA4 prolonged Plasmodium berghei-infected mice survival by a mechanism that depends on inhibiting IL-12 production and CD8(+)IFN-γ(+) T cells in brain tissue; however, the effects of this treatment on endothelial dysfunction induced during experimental cerebral malaria (ECM) remains to be elucidated. Herein, we investigate the role of LXA4 on endothelial dysfunction during ECM. The treatment of P. berghei-infected mice with LXA4 prevented BBB breakdown and ameliorated behavioral symptoms but did not modulate TNF-α production. In addition, microcirculation analysis showed that treatment with LXA4 significantly increased functional capillary density in brains of P. berghei-infected C57BL/6 mice. Furthermore, histological analyses of brain sections demonstrated that exogenous LXA4 reduced capillary congestion that was accompanied by reduced ICAM-1 expression in the brain tissue. In agreement, LXA4 treatment of endothelial cells stimulated by Plasmodium berghei (Pb)- or Plasmodium falciparum (Pf)-parasitized red blood cells (RBCs) inhibited ICAM-1 expression. Additionally, LXA4 treatment restored the expression of HO-1 that is reduced during ECM. As well, LXA4 treatment inhibits PbRBC and PfRBC adhesion to endothelial cells that was reversed by the use of an HO-1 inhibitor (ZnPPIX). Our results demonstrate for the first time that LXA4 ameliorates endothelial dysfunction during ECM by modulating ICAM-1 and HO-1 expression in brain tissue.

Tang-Tong-Fang Confers Protection against Experimental Diabetic Peripheral Neuropathy by Reducing Inflammation.

Tang-tong-fang (TTF) is a Chinese herbal formula that has been shown to be beneficial in diabetic peripheral neuropathy (DPN), a common complication secondary to diabetic microvascular injury. However, the underlying mechanism of protection in nerve ischemia provided by TTF is still unclear. We hypothesized that TTF alleviates DPN via inhibition of ICAM-1 expression. Therefore, we tested the effect of TTF in a previously established DPN model, in which nerve injury was induced by ischemia/reperfusion in streptozotocin-induced diabetic rats. We found that the conduction velocity and amplitude of action potentials of sciatic nerve conduction were reduced in the DPN model group but were rescued by TTF treatment. In addition, TTF treatment also attenuated the effect of DPN on other parameters including histology and ultrastructural changes, expression of ICAM-1, MPO, and TNF-α in rat sciatic nerves, and plasma sICAM-1 and MPO levels. Together, our data suggest that TTF treatment may alleviate DPN via ICAM-1 inhibition.