Human Monocyte Chemoattractant Protein-1 Research Articles

Transgenic human C-reactive protein affects oxidative stress but not inflammation biomarkers in the aorta of spontaneously hypertensive rats

BackgroundC-reactive protein (CRP) is an acute inflammatory protein detected in obese patients with metabolic syndrome. Moreover, increased CRP levels have been linked with atherosclerotic disease, congestive heart failure, and ischemic heart disease, suggesting that it is not only a biomarker but also plays an active role in the pathophysiology of cardiovascular diseases. Since endothelial dysfunction plays an essential role in various cardiovascular pathologies and is characterized by increased expression of cell adhesion molecules and inflammatory markers, we aimed to detect specific markers of endothelial dysfunction, inflammation, and oxidative stress in spontaneously hypertensive rats (SHR) expressing human CRP. This model is genetically predisposed to the development of the metabolic syndrome.MethodsTransgenic SHR male rats (SHR-CRP) and non-transgenic SHR (SHR) at the age of 8 months were used. Metabolic profile (including serum and tissue triglyceride (TAG), serum insulin concentrations, insulin-stimulated incorporation of glucose, and serum non-esterified fatty acids (NEFA) levels) was measured. In addition, human serum CRP, MCP-1 (monocyte chemoattractant protein-1), and adiponectin were evaluated by means of ELISA, histological analysis was used to study morphological changes in the aorta, and western blot analysis of aortic tissue was performed to detect expression of endothelial, inflammatory, and oxidative stress markers.ResultsThe presence of human CRP was associated with significantly decreased insulin-stimulated glycogenesis in skeletal muscle, increased muscle and hepatic accumulation of TAG and decreased plasmatic cGMP concentrations, reduced adiponectin levels, and increased monocyte chemoattractant protein-1 (MCP-1) levels in the blood, suggesting pro-inflammatory and presence of multiple features of metabolic syndrome in SHR-CRP animals. Histological analysis of aortic sections did not reveal any visible morphological changes in animals from both SHR and SHR-CRP rats. Western blot analysis of the expression of proteins related to the proper function of endothelium demonstrated significant differences in the expression of p-eNOS/eNOS in the aorta, although endoglin (ENG) protein expression remained unaffected. In addition, the presence of human CRP in SHR in this study did not affect the expression of inflammatory markers, namely p-NFkB, P-selectin, and COX2 in the aorta. On the other hand, biomarkers related to oxidative stress, such as HO-1 and SOD3, were significantly changed, indicating the induction of oxidative stress.ConclusionsOur findings demonstrate that CRP alone cannot fully induce the expression of endothelial dysfunction biomarkers, suggesting other risk factors of cardiovascular disorders are necessary to be involved to induce endothelial dysfunction with CRP.

MCP-5 suppresses osteoclast differentiation through Ccr5 upregulation.

Human monocyte chemoattractant protein-1 (MCP-1) in mice has two orthologs, MCP-1 and MCP-5. MCP-1, which is highly expressed in osteoclasts rather than in osteoclast precursor cells, is an important factor in osteoclast differentiation. However, the roles of MCP-5 in osteoclasts are completely unknown. In this study, contrary to MCP-1, MCP-5 was downregulated during receptor activator of nuclear factor kappa B ligand(RANKL)-induced osteoclast differentiation and was considered an inhibitory factor in osteoclast differentiation. The inhibitory role of MCP-5 in osteoclast differentiation was closely related to the increase in Ccr5 expression and the inhibition of IκB degradation by RANKL. Transgenic mice expressing MCP-5 controlled by Mx-1 promoter exhibited an increased bone mass because of a decrease in osteoclasts. This result strongly supported that MCP-5 negatively regulated osteoclast differentiation. MCP-5 also prevented severe bone loss caused by RANKL.

Serum levels of monocyte chemoattractant protein-1 has no correlation with gender and age in polytrauma

<p class="abstract"><strong>Background:</strong> Trauma is one of the commonest causes of death among young people with significant increase in morbidity and mortality. Following injuries to tissues, the body responds in an attempt to cause repair of the damaged tissue. Although its significance depends on the interplay of various factors involving neurohormonal and immune responses. Monocyte chemoattractant protein-1 (MCP-1) is a pro-inflammatory chemoattractant produced by the monocyte and causes the further release of chemokines and cytokines needed at the site of tissue injury. It therefore plays a critical role in tissue healing.</p><p class="abstract"><strong>Methods:</strong> This was a prospective hospital-based study carried out at University College Hospital (UCH), Ibadan. Polytrauma patients admitted through the accident and emergency department that met a defined criteria were recruited and had their blood samples taken into an endotoxin free test tube at 48±2 hours after trauma. MCP-1 levels in the serum were estimated though the human MCP-1 enzyme-linked immunosorbent assay (ELISA) kit. This process was carried out using the ELISA technique based on the producer’s guide.<strong></strong></p><p class="abstract"><strong>Results:</strong> 110 patients were recruited for the study. There was no significant difference in gender variations in MCP-1 level (t=-0.935, p=0.351). For the male and female variations in MCP-1 levels, it ranges from 10 to 2841 and 22 to 2687 respectively. Likewise, there was no significant relationship in age variations in MCP-1 values (f=0.959, p>0.05).</p><p class="abstract"><strong>Conclusions:</strong> This study demonstrated no correlation between serum level of monocyte chemoattractant protein-1 with age and gender in polytraumatized patient according to age and gender.</p>

Cross-sectional associations of adipokines and abdominal fat distribution with aging in men

Objective To investigate the relationship of adipokines and abdominal fat distribution with aging in men. Methods In a cross-sectional study, a total of 218 participants aged 40–79 years were recruited as a subset of the Prospective Urban Rural Epidemiology (PURE) China Action on Spine and Hip status (CASH) study population. Analysis of variance (ANOVA) and multivariable regression were used to estimate the associations of interest. Results With the increasing of age, waist circumference, waist-to-hip ratio, waist-to-height ratio, total adipose tissue (TAT), visceral adipose tissue (VAT), VAT/subcutaneous adipose tissue (SAT), leptin, adiponectin-to-leptin ratio, and human monocyte chemo-attractant protein-1 (MCP-1) increased significantly (p < 0.05), while adiponectin decreased significantly (p < 0.05). Adiponectin, adiponectin/leptin, and adiponectin/resistin varied inversely with the VAT quartiles (p < 0.05). There was a significant negative correlation among adiponectin, adiponectin-to-leptin ratio, adiponectin-to-resistin ratio, and all the body fat distribution parameters. VAT was inversely and significantly associated with adiponectin, adiponectin-to-leptin ratio, and adiponectin-to-resistin ratio (p < 0.05). Conclusions It showed that aging, abdominal fat distribution, and adipokines were related with each other, which support the hypothesis that regulation of VAT and adipokines is closely linked to aging.

Optimization of the Solid Phase Synthesis of the Ingramon Peptide Antagonist of the Human Monocyte Chemoattractant Protein 1 (MCP-1)

The solid phase synthesis (SPS) of the ingramon peptide antagonist of the human monocyte chemoattractant protein-1 (H-Asp-His-Leu-Asp-Lys-Gln-Thr-Gln-Thr-Pro-Lys-Thr-OH), which exhibited the anti-inflammatory activity, was optimized. Side products of the Fmoc-SPS of this peptide were studied. Their structures were determined by the 1H NMR spectroscopy and confirmed by the synthesis. Methodological ways to minimize of a formation of impurities in the course of the ingramon SPS were found. The reproducible SPS procedure of the ingramon preparation that resulted in the formation of the minimum amount of such side products as [Asi4]-, [D-Asp4]-, and [βAsp4]-peptides was elaborated.

Preliminary evaluation and mechanism of adipose-derived stem cell transplantation from allogenic diabetic rats in the treatment of diabetic rat wounds

Preliminary evaluation and mechanism of adipose-derived stem cell transplantation from allogenic diabetic rats in the treatment of diabetic rat wounds

Liraglutide Attenuates Preestablished Atherosclerosis in Apolipoprotein E-Deficient Mice via Regulation of Immune Cell Phenotypes and Proinflammatory Mediators.

We have shown that the glucagon-like peptide-1 receptor agonist (GLP-1RA) liraglutide (Lir) inhibits development of early atherosclerosis in vivo by modulating immune cell function. We hypothesized that Lir could attenuate pre-established disease by modulating monocyte or macrophage phenotype to induce atheroprotective responses. Human atherosclerotic plaques obtained postendarterectomy and human peripheral blood macrophages were treated ex vivo with Lir. In parallel, apolipoprotein E-deficient (ApoE-/-) mice received a high-fat, high-cholesterol diet to induce atherosclerosis for 8 weeks, after which ApoE-/- mice received 300 μg/kg of Lir daily or vehicle control for a further 4 weeks to investigate the attenuation of atherosclerosis. Lir inhibited proinflammatory monocyte chemoattractant protein-1 secretion from human endarterectomy samples and monocyte chemoattractant protein-1, tumor necrosis factor-α, and interleukin (IL)-1β secretion from human macrophages after ex vivo treatment. An increase in CD206 mRNA and IL-10 secretion was also detected, which implies resolution of inflammation. Importantly, Lir significantly attenuated pre-established atherosclerosis in ApoE-/- mice in the whole aorta and aortic root. Proteomic analysis of ApoE-/- bone marrow cells showed that Lir upregulated the proinflammatory cathepsin protein family, which was abolished in differentiated macrophages. In addition, flow cytometry analysis of bone marrow cells induced a shift toward reduced proinflammatory and increased anti-inflammatory macrophages. We concluded that Lir attenuates pre-established atherosclerosis in vivo by altering proinflammatory mediators. This is the first study to describe a mechanism through which Lir attenuates atherosclerosis by increasing bone marrow proinflammatory protein expression, which is lost in differentiated bone marrow-derived macrophages. This study contributes to our understanding of the anti-inflammatory and cardioprotective role of GLP-1RAs. SIGNIFICANCE STATEMENT: It is critical to understand the mechanisms through which liraglutide (Lir) mediates a cardioprotective effect as many type 2 diabetic medications increase the risk of myocardial infarction and stroke. We have identified that Lir reduces proinflammatory immune cell populations and mediators from plaque-burdened murine aortas in vivo and augments proresolving bone marrow-derived macrophages in attenuation of atherosclerotic disease, which provides further insight into the atheroprotective effect of Lir.

Evaluation and extension of the two-site, two-step model for binding and activation of the chemokine receptor CCR1

Interactions between secreted immune proteins called chemokines and their cognate G protein-coupled receptors regulate the trafficking of leukocytes in inflammatory responses. The two-site, two-step model describes these interactions. It involves initial binding of the chemokine N-loop/β3 region to the receptor's N-terminal region and subsequent insertion of the chemokine N-terminal region into the transmembrane helical bundle of the receptor concurrent with receptor activation. Here, we test aspects of this model with C-C motif chemokine receptor 1 (CCR1) and several chemokine ligands. First, we compared the chemokine-binding affinities of CCR1 with those of peptides corresponding to the CCR1 N-terminal region. Relatively low affinities of the peptides and poor correlations between CCR1 and peptide affinities indicated that other regions of the receptor may contribute to binding affinity. Second, we evaluated the contributions of the two CCR1-interacting regions of the cognate chemokine ligand CCL7 (formerly monocyte chemoattractant protein-3 (MCP-3)) using chimeras between CCL7 and the non-cognate ligand CCL2 (formerly MCP-1). The results revealed that the chemokine N-terminal region contributes significantly to binding affinity but that differences in binding affinity do not completely account for differences in receptor activation. On the basis of these observations, we propose an elaboration of the two-site, two-step model-the "three-step" model-in which initial interactions of the first site result in low-affinity, nonspecific binding; rate-limiting engagement of the second site enables high-affinity, specific binding; and subsequent conformational rearrangement gives rise to receptor activation.

AN AMYLOID-BETA-DERIVED PEPTIDE TRANSFORMS RAT GLIAL PROGENITOR CELLS INTO MICROGLIA

Septapeptides (“septas”), previously identified as meningitis-specific antigens, defined by a rubella virus monoclonal antibody, were found in human Monocyte Chemoattractant Protein (hMCP-1) and on the surface of meningitis-causing bacteria, viruses and spirochetes.1-3 Some bacterial septas were tested for Ca+2 mobilization through receptor-associated heterotrimeric G-protein binding on THP-1 cells, progenitor cells of circulating peripheral macrophages. Certain of the free septas acted on their own as mild agonists of Ca+2 mobilization. Their signal transduction activity may be mediated through a single (or a single class) of receptor, but the data do not link this with the MCP-1 receptor on THP-1 cells. These data support the notion that (1) infectious organisms may conserve and employ these sequences in order to facilitate their transport through the BBB to infect the CNS and (2) the hypothesis that the septas represent muteins of the MCP-1 active site for monocyte stem cell activation to macrophages. Other MCP muteins have since been identified in the AD-associated agents, amyloid beta and prions, as well as in viruses like HIV, known to establish chronic infections in the CNS. Rat glial progenitor cells in tissue culture 4-6 were used to test for hMCP-1 activity in septas derived from amyloid beta. Nanomolar concentrations of the amyloid beta septa 13HHQKLVF19 were found to transform more than 60% of rat glial progenitor cells into mature microglia in tissue culture7.8. These data support the hypothesis that an amyloid beta-derived septa may serve to activate stem cells to increase the number of microglia available to combat chronic infection in the CNS. 1VanAlstyne & Sharma. April 23, 1996. US Patent No. 5,510,264. 2VanAlstyne & Sharma. Sept. 17, 1996. US Patent No. 5,556,757. 3Van Alstyne & Sharma. Dec. 6, 2011. US Patent No. 8,071,102. 4Singh and Van Alstyne. 1978.Brain Res. 155:418-421. 5Pope and Van Alstyne. 1981.Virology 113(2):776-780. 6Van Alstyne and Paty. 1983.Virology 124:173-180 7Van Alstyne, Jan. 4, 2018. US Patent Application 62/613,621. 8Van Alstyne. June 20, 2018 Poster(3035) presentation at Keystone Symposia Conference “Advances in Neurodegenerative Disease Research and Therapy”. Session Z3.

Abstract B55: Tumor-derived MCP-1 regulates invasiveness in triple-negative breast cancer via the MAP kinase pathway

Abstract Background: Triple-negative breast cancer (TNBC) is the most aggressive type of breast cancer compared to other breast cancer subtypes. The frequency of TNBC expression is highest in young African-American women, leading to significant cancer health disparity in this population. Furthermore, TNBC is difficult to treat due to lack of known receptor targets at protein or gene level. Hence, it is imperative to identify novel therapeutic strategies for treatment of TNBC. Here we aim to show that the Monocyte Chemoattractant Protein -1 (MCP-1) is a reliable biochemical marker to assess TNBC progression. Experimental Design: We employed ELISA method to measure secreted MCP-1 in cell conditioned media, and real-time PCR to determine the mRNA status of MCP-1 in different TNBC cell lines. Boyden chamber assay was used to determine the effect of recombinant MCP-1 on cellular invasiveness. Immunohistochemistry staining was utilized for detecting protein of interest in tissue samples from breast cancer patients. MCP-1 knockdown was performed using lentiviral vector with shRNA targeting MCP-1 coding regions. RNAseq was performed with recombinant human MCP-1. Results: Our data show that MCP-1 is upregulated in TNBC cell lines, both transcriptionally and in secreted protein levels compared to ER-positive cell line, MCF-7. Breast cancer patients also showed high expression of MCP-1. MCP-1 treatment induced MDA-MB-231 and MCF-7 cell invasion, without affecting cell proliferation. Small-molecule antagonists against chemokine receptor 2 (CCR2), cognate receptor for MCP-1, and the MAP kinase pathway inhibitor U0106 negatively affected MDA-MB-231 cell invasion as measured by the Boyden chamber assay. This suggests that MCP-1-CCR2 axis may regulate invasiveness via the MAP kinase pathway. Knocking down MCP-1 by shRNA decreased cell invasion in TNBC cell line, BT-549, along with downregulation of key epithelial-to-mesenchymal transition markers, N-cadherin and Vimentin. Recombinant MCP-1 treatment in TNBC MDA-MB-231 cells upregulated genes associated with cytokine signaling. Conclusion: Our study suggests that a high MCP-1 level in TNBC cells may be responsible for increase in cell invasion via the MAP kinase pathway. Thus, MCP-1-mediated pathways could be potential therapeutic targets for the treatment of TNBC and reduce cancer health disparities. Citation Format: Pranabananda Dutta, Kimberly Paico, Yanyuan Wu, Marianna Sarkissyan, Jaydutt Vadgama. Tumor-derived MCP-1 regulates invasiveness in triple-negative breast cancer via the MAP kinase pathway [abstract]. In: Proceedings of the Tenth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2017 Sep 25-28; Atlanta, GA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2018;27(7 Suppl):Abstract nr B55.

Display of human and rabbit monocyte chemoattractant protein-1 on human embryonic kidney 293T cell surface.

Monocyte chemoattractant protein-1 (MCP-1/CCL2) is a protein that is secreted immediately upon endothelial injury, and thereby it plays a key role in inflammation via recruitment of leucocytes to the site of inflammation at the beginning and throughout the inflammatory processes. Aim of this study was to develop two separate cell lines displaying either human MCP-1 (HMCP-1) or rabbit MCP-1 (RMCP-1) on their surface. A DNA fragment containing HMCP-1- or RMCP-1-encoding sequence was inserted into a pcDNA plasmid. Escherichia coli cells strain TOP 10F’ was separately transformed with the pcDNA/RMCP-1 or /HMCP-1 ligation mixture. Following the cloning and construct verification, human embryonic kidney cell line (HEK 293T) was transfected with either of the linearized plasmids. Plasmid integration into the genomic DNA of HEK 293T cells was verified by polymerase chain reaction (PCR). HMCP-1 and RMCP-1 expression was evaluated at RNA and protein levels by real-time PCR and flow cytometry, respectively. PCR products of the expected sizes were amplified from the chromosomal DNA of transfected HEK 293T cells, i.e. 644 bp for H-MCP1 and 737 bp for RMCP-1 constructs. Real-time PCR revealed that the copy numbers of RMCP1 and HMCP1 mRNA per cell were 294 and 500, respectively. Flow cytometry analysis indicated 85% for RMCP-1 and 87% for HMCP-1 expression levels on the surface of transfected cells, when compared with an isotype control. The experiments thus confirmed that the MCP-1 genes were integrated into the HEK 293T genomic DNA and the encoded proteins were stably expressed on the cell surface.

Role of Organic Anion Transporters in the Uptake of Protein-Bound Uremic Toxins by Human Endothelial Cells and Monocyte Chemoattractant Protein-1 Expression

Organic anion transporters (OATs) are involved in the uptake of uremic toxins such as p-cresyl sulfate (PCS) and indoxyl sulfate (IS), which play a role in endothelial dysfunction in patients with chronic kidney diseases (CKD). In this study, we investigated the role of OAT1 and OAT3 in the uptake of PCS and IS into human endothelial cells. PCS was synthesized via p-cresol sulfation and characterized using analytical methods. The cells were treated with PCS and IS in the absence and presence of probenecid (Pb), an OAT inhibitor. Cell viability was assessed using the MTT assay. The absorbed toxins were analyzed using chromatography, OAT expression using immunocytochemistry and western blot, and monocyte chemoattractant protein-1 (MCP-1) expression using enzyme-linked immunosorbent assay. Cell viability decreased after toxin treatment in a dose-dependent manner. PCS and IS showed significant internalization after 60 min treatment, while no internalization was observed in the presence of Pb, suggesting that OATs are involved in the transport of both toxins. Immunocytochemistry and western blot demonstrated OAT1 and OAT3 expression in endothelial cells. MCP-1 expression increased after toxins treatment but decreased after Pb treatment. PCS and IS uptake were mediated by OATs, and OAT blockage could serve as a therapeutic strategy to inhibit MCP-1 expression.

Chondroitin Sulfate Inhibits Monocyte Chemoattractant Protein-1 Release From 3T3-L1 Adipocytes: A New Treatment Opportunity for Obesity-Related Inflammation?

Monocyte chemoattractant protein-1 (MCP-1) overproduction from inflamed adipose tissue is a major contributor to obesity-related metabolic syndromes. 3T3-L1 embryonic fibroblasts were cultured and differentiated into adipocytes using an established protocol. Adipocytes were treated with lipopolysaccharide (LPS) to induce inflammation and thus MCP-1 release. At the same time, varying concentrations of chondroitin sulfate (CS) were added in a physiologically relevant range (10-200 µg/mL) to determine its impact on MCP-1 release. Chondroitin sulfate, a natural glycosaminoglycan of connective tissue including the cartilage extracellular matrix, was chosen on the basis of our previous studies demonstrating its anti-inflammatory effect on macrophages. Because the main action of MCP-1 is to induce monocyte migration, cultured THP-1 monocytes were used to test whether CS at the highest physiologically relevant concentration could inhibit cell migration induced by human recombinant MCP-1. Chondroitin sulfate (100-200 µg/mL) inhibited MCP-1 release from inflamed adipocytes in a dose-dependent manner (P < .01, 95% confidence interval [CI]: −5.89 to −3.858 at 100 µg/mL and P < .001, 95% CI: −6.028 to −3.996 at 200 µg/mL) but had no effect on MCP-1–driven chemotaxis of THP-1 monocytes. In summary, CS could be expected to reduce macrophage infiltration into adipose tissue by reduction in adipocyte expression and release of MCP-1 and as such might reduce adipose tissue inflammation in response to pro-inflammatory stimuli such as LPS, now increasingly recognized to be relevant in vivo.

Protection of kinsenoside against AGEs-induced endothelial dysfunction in human umbilical vein endothelial cells

AimsKinsenoside is the major ingredient of Anoectochilus roxburghii which is a traditional Chinese herb using for the treatment of diabetes. The present study investigated the safety and vascular protection of kinsenoside related to advanced glycation end products (AGEs) in human umbilical vein endothelial cells (HUVECs) and the underlying mechanisms. Materials and methodsHUVECs were pre-incubated with AGEs (200μg/mL) for 1h, and then co-treated with different concentrations of kinsenoside (10–30μg/mL) for another 48h. After the supernatant was collected, the contents of nitric oxide (NO), the levels of reactive oxygen species (ROS) and inflammatory cytokines, and the expressions of AGEs receptor (RAGE) and nuclear factor kappa B (NF-κB) were measured. Key findingsNo significant changes in cell viability were found in kinsenoside-treated cells at the range of 10–70μg/mL. Pretreatment with kinsenoside induced a significant increase in NO production in AGEs-induced cells. In addition, kinsenoside not only inhibited the expression of RAGE but also decreased intracellular ROS generation induced by AGEs. Furthermore, kinsenoside suppressed the protein and gene expression of NF-κB, and reduced the release of intercellular adhesion molecule-1 (ICAM-1) and human monocyte chemoattractant protein-1 (MCP-1) in a dose-dependent manner remarkably. SignificanceThese results indicated that kinsenoside might attenuate AGEs-induced endothelial dysfunction via AGEs-RAGE-NF-κB pathway. Considering the relatively low toxicity of kinsenoside, it might be a promising agent for treatment of vasculopathy in diabetic patients.

Suppression of methylmercury-induced IL-6 and MCP-1 expressions by N-acetylcysteine in U-87MG human astrocytoma cells.

The aim of this study was to clarify the involvement of oxidative stress in methylmercury (MeHg)-induced pro-inflammatory cytokine expressions and the suppressive effects of N-acetylcysteine (NAC) in MeHg-induced cytokine expression. Using U-87-MG human astrocytoma cell line, interleukin (IL)-6 and monocyte chemoattractant protein (MCP)-1 expressions induced by 4 μM MeHg were measured at mRNA and protein levels. Hydrogen peroxide (H2O2) and superoxide anion (O2(-)) were quantified by flow-cytometry analysis. To examine the suppressive effects of NAC on the cytokine expressions among different timing of NAC treatment, cells were treated with 0.5 or 5mM NAC before, simultaneously, or after MeHg administration. MeHg exposure at 4 μM, a non-cytotoxic concentration, significantly induced MCP-1 and IL-6 expressions at both mRNA and protein levels. A significant increase of H2O2 production but not O2(-) was observed. MeHg-induced expression of MCP-1 and IL-6 mRNA was reduced by 10-20% in the presence of 5mM NAC (co-treatment experiment) compared to cells treated with MeHg only. Pre-treatment of cells with 0.5 or 5mM NAC at 0.5 or 1h and its subsequent washout before MeHg addition suppressed MCP-1 and IL-6 cytokine expressions. Post-treatment of cells with NAC after MeHg addition also suppressed the cytokine induction, but the magnitude of suppression was evidently lower than in co-treated cells even though the H2O2 generation was almost completely suppressed by NAC. NAC may effectively suppress the MeHg-induced cytokine production through both, inhibition of reactive oxygen species as well as extracellular chelation of MeHg in astrocytes.

Clinical significance of expression of human macrophage metalloelastase and monocyte chemoattractant protein-1 in gastric cancer

目的：研究人巨噬细胞金属弹力酶（human macrophage metalloelastase,HME）和单核细胞趋化蛋白-1（monocyte chemoattractant protein-1,MCP-1）在胃癌组织中的表达及临床意义.方法：应用免疫组织化学SP法检测胃癌和癌旁组织石蜡包埋组织标本30例中的HME和MCP-1蛋白的表达情况,并分析HME与MCP-1的相关性及两者与临床病理的关系.结果：（1）HME和MCP-1在胃癌组织中表达的阳性率为70.0%（21/30）和80.0%（24/30）,而在癌旁组织中表达的阳性率为26.7%（8/30）和16.7%（5/30）,二者在胃癌组织中的表达显著高于胃正常组织,差异有统计学意义（P〈0.05）;（2）HME在胃癌组织中的阳性表达与胃癌的分化程度,血管浸润有关（P〈0.05）,与其他临床病理参数无明显相关;MCP-1的表达与患者的肿瘤的浸润深度、肿瘤大小、肿瘤分期、淋巴转移、远处转移有关（P〈0.05）,与其他临床病理参数无明显相关;（3）HME与MCP-1在胃癌组织中的阳性表达无相关性（列联分析r=0.115,P〉0.05）.结论：胃癌组织中HME、MCP-1的高表达与胃癌的血管生成、浸润、转移有关,参与人胃癌的发生和发展,有助于探讨胃癌的发生机制.

MCP-1 Stimulates MMP-9 Expression via ERK 1/2 and p38 MAPK Signaling Pathways in Human Aortic Smooth Muscle Cells

Objective: We investigated the molecular mechanism underlying the role of monocyte chemoattractant protein-1 (MCP-1) in the formation and development of human abdominal aortic aneurysm (AAA). Methods: We examined protein expression profiles using a protein array and found that MCP-1 was the most highly expressed protein in AAA tissues compared with normal aortas. To investigate the potential mechanism of MCP-1 involvement in the pathogenesis of AAA, we treated human aortic smooth muscle cells (HASMCs) with human recombinant MCP-1. Results: MCP-1 was the most highly expressed protein in AAA tissues compared with normal aorta; matrix metalloproteinase-9 (MMP-9) expression was also significantly increased. Treatment with MCP-1 significantly increased the expression and activation of MMP-9 and activated the three major mitogen activated protein kinases (MAPKs) extracellular signal regulated kinase (ERK), c-Jun amino terminal kinase (JNK1/2) and p38 MAPK. Furthermore, MCP-1-induced secretion of MMP-9 was inhibited by U0126 (inhibitor of the ERK 1/2 pathway) and SB203580 (inhibitor of the p38 MAPK pathway), but not SP600125 (inhibitor of the JNK1/2 pathway). Conclusion: These data demonstrate that MCP-1 stimulates secretion of MMP-9 directly through the ERK1/2 and p38 MAPK mediated pathways in HASMCs. Thus, inhibition of this molecular mechanism might be a potential therapeutic target in the non-surgical treatment of AAA.

MCP-1 binds to oxidized LDL and is carried by lipoprotein(a) in human plasma

Lipoprotein oxidation plays an important role in pathogenesis of atherosclerosis. Oxidized low density lipoprotein (OxLDL) induces profound inflammatory responses in vascular cells, such as production of monocyte chemoattractant protein-1 (MCP-1) [chemokine (C-C motif) ligand 2], a key chemokine in the initiation and progression of vascular inflammation. Here we demonstrate that OxLDL also binds MCP-1 and that the OxLDL-bound MCP-1 retains its ability to recruit monocytes. A human MCP-1 mutant in which basic amino acids Arg-18 and Lys-19 were replaced with Ala did not bind to OxLDL. The MCP-1 binding to OxLDL was inhibited by the monoclonal antibody E06, which binds oxidized phospholipids (OxPLs) in OxLDL. Because OxPLs are carried by lipoprotein(a) [Lp(a)] in human plasma, we tested to determine whether Lp(a) binds MCP-1. Recombinant wild-type but not mutant MCP-1 added to human plasma bound to Lp(a), and its binding was inhibited by E06. Lp(a) captured from human plasma contained MCP-1 and the Lp(a)-associated endogenous MCP-1 induced monocyte migration. These results demonstrate that OxLDL and Lp(a) bind MCP-1 in vitro and in vivo and that OxPLs are major determinants of the MCP-1 binding. The association of MCP-1 with OxLDL and Lp(a) may play a role in modulating monocyte trafficking during atherogenesis.

Platelet-derived growth factor (PDGF)-BB-mediated induction of monocyte chemoattractant protein 1 in human astrocytes: implications for HIV-associated neuroinflammation

Chemokine (C-C motif) ligand 2, also known as monocyte chemoattractant protein 1 (MCP-1) is an important factor for the pathogenesis of HIV-associated neurocognitive disorders (HAND). The mechanisms of MCP-1-mediated neuropathogenesis, in part, revolve around its neuroinflammatory role and the recruitment of monocytes into the central nervous system (CNS) via the disrupted blood-brain barrier (BBB). We have previously demonstrated that HIV-1/HIV-1 Tat upregulate platelet-derived growth factor (PDGF)-BB, a known cerebrovascular permeant; subsequently, the present study was aimed at exploring the regulation of MCP-1 by PDGF-BB in astrocytes with implications in HAND. Specifically, the data herein demonstrate that exposure of human astrocytes to HIV-1 LAI elevated PDGF-B and MCP-1 levels. Furthermore, treating astrocytes with the human recombinant PDGF-BB protein significantly increased the production and release of MCP-1 at both the RNA and protein levels. MCP-1 induction was regulated by activation of extracellular-signal-regulated kinase (ERK)1/2, c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein (MAP) kinases and phosphatidylinositol 3-kinase (PI3K)/Akt pathways and the downstream transcription factor, nuclear factor κB (NFκB). Chromatin immunoprecipitation (ChIP) assays demonstrated increased binding of NFκB to the human MCP-1 promoter following PDGF-BB exposure. Conditioned media from PDGF-BB-treated astrocytes increased monocyte transmigration through human brain microvascular endothelial cells (HBMECs), an effect that was blocked by STI-571, a tyrosine kinase inhibitor (PDGF receptor (PDGF-R) blocker). PDGF-BB-mediated release of MCP-1 was critical for increased permeability in an in vitro BBB model as evidenced by blocking antibody assays. Since MCP-1 is linked to disease severity, understanding its modulation by PDGF-BB could aid in understanding the proinflammatory responses in HAND. These results suggest that astrocyte activation by PDGF-BB exaggerates monocyte recruitment into the brain via MCP-1 and underscores the critical role astrocytes play in HAND.

15-deoxy-Δ¹²,¹⁴ -prostaglandin J₂ inhibits human immunodeficiency virus-1 tat-induced monocyte chemoattractant protein-1/CCL2 production by blocking the extracellular signal-regulated kinase-1/2 signaling pathway independently of peroxisome proliferator-activated receptor-γ and heme oxygenase-1 in rat hippocampal slices.

Human immunodeficiency virus (HIV)-induced inflammation, and its consequences within the central nervous system (CNS), must be countered by multiple pharmacologic agents, and 15-deoxy-Δ(12,14) -prostaglandin J(2) (15d-PGJ2) may hold promise in the treatment of pathologies associated with this inflammatory response. 15d-PGJ2 can repress the inflammatory response by means of peroxisome proliferator-activated receptor-γ (PPARγ)-dependent and -independent mechanisms. However, its precise role and antiinflammatory mechanism in the hippocampus remain poorly understood. In the present study, rat hippocampal slices were stimulated with full-length HIV-1 Tat protein to investigate the role of 15d-PGJ2 8in the hippocampal inflammatory response. Pretreatment of slices with 15d-PGJ2 markedly reduced Tat-induced monocyte chemoattractant protein-1 (MCP-1/CCL2) production. Interestingly, the PPARγ antagonist GW9662 did not inhibit action of 15d-PGJ2, confirming the latter's PPARγ-independent mechanism of mediating antiinflammatory effects. Despite 15d-PGJ2's increasing the expression of heme oxygenase-1 (HO-1), its action was not abrogated by the HO-1 inhibitor zinc protoporphyrin IX (ZnPPIX), nor was it recapitulated by HO-1 inducers such as cobalt protoporphyrin (CoPP). Moreover, short interfering RNA (siRNA)-directed knockdown of HO-1 did not abolish the antiinflammatory action of 15d-PGJ2 against Tat-induced MCP-1 production in human microglia-like THP-1 cells. Conversely, 15d-PGJ2 suppressed Tat-induced ERK1/2 activation, decreasing MCP-1 production upon Tat stimulation. The NADPH oxidase inhibitors DPI and apocynin also abrogated Tat-stimulated ERK1/2 activation, reducing MCP-1 production. Collectively, these data demonstrate that the antiinflammatory effects of 15d-PGJ2 on the hippocampus are exerted through inhibition of Tat-mediated ERK1/2 activation, coupled with that of a redox-sensitive pathway, independent of PPARγ and HO-1.