CD100 and plexins B2 and B1 mediate monocyte-endothelial cell adhesion and might take part in atherogenesis

Leukocyte migration is essential for the function of the immune system. Their recruitment from the vessels to the tissues involves sequential molecular interactions between leukocytes and endothelial cells (ECs). Many adhesion molecules involved in this process have already been described. However, additional molecules may be important in this interaction, and here we explore the potential role for CD100 and plexins in monocyte-EC binding.CD100 was shown to be involved in platelet-endothelial cell interaction, an important step in atherogenesis and thrombus formation. In a recent work we have described CD100 expression in monocytes and in macrophages and foam cells of human atherosclerotic plaques. In the present work, we have identified plexin B2 as a putative CD100 receptor in these cells. We have detected CD100 expression in the endothelium as well as in in vitro cultured endothelial cells. Blocking of CD100, plexin B1 and/or B2 in adhesion experiments have shown that both CD100 and plexins act as adhesion molecules involved in monocyte-endothelial cell binding. This effect may be mediated by CD100 expressed in both cell types, probably coupled to the receptors endothelial plexin B1 and monocytic plexin B2. These results can bring new insights about a possible biological activity of CD100 in monocyte adhesion and atherosclerosis, as well as a future candidate for targeting therapeutics.     

LAG-3 (CD223) reduces macrophage and dendritic cell differentiation from monocyte precursors

Major histocompatibility complex (MHC) class II molecules expressed on monocytes may play a role in the control of differentiation of antigen-presenting cells. A soluble LAG-3 (CD223) molecule (sLAG-3) is a natural, high-affinity ligand for MHC class II. It is known to induce maturation of monocyte-derived dendritic cells in vitro and is used as a vaccine adjuvant to induce CD4 T helper type 1 responses and CD8 T-cell responses in vivo. Here, we demonstrate that sLAG-3 (but not an MHC class II-specific monoclonal antibody) reduces the differentiation of monocytes into macrophages in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF) as well as their differentiation into dendritic cells in the presence of GM-CSF and interleukin-4, as shown by a decrease in CD14 and CD1a expression, respectively. Dendritic cells derived from monocytes in the presence of sLAG-3 showed impaired antigen-presentation function, as assessed by the reduced capability to induce proliferation of T cells. Our results suggest that activated LAG-3(+) lymphocytes present at sites of inflammation may reduce the differentiation of monocytes into macrophages or fully competent antigen-presenting dendritic cells, thus limiting the magnitude of the ongoing T-cell immune responses.     

Monocyte-derived macrophages matured under prolonged hypoxia transcriptionally up-regulate HIF-1α mRNA

This study tested the hypothesis that prolonged severe hypoxia during monocyte to macrophage differentiation results in macrophages with a pattern of gene expression and phenotype distinct from those maturing in normal oxygen levels. Macrophages accumulate in hypoxic and anoxic areas within pathological sites such as tumours, wounds, and arthritic joints, and have been proposed as vehicles for gene therapy delivery to such tissues. Several non-pathological tissues are also hypoxic. We therefore argue that differentiation from monocyte to macrophage in hypoxic conditions is a common occurrence. However, the effect of long term severe hypoxia on monocyte to macrophage differentiation has not been studied. Here, using primary human peripheral blood monocytes, we show that maturation for 5 days in 0.2% oxygen results in decreased phagocytosis, and decreased CD40 and CD206 expression. Chronic hypoxia induced much higher mRNA levels of the pro-angiogenic cytokine, VEGF, in adherence-purified macrophages (27-fold), CD14-magnetic bead purified monocytes (90-fold), and PBMC (104-fold) compared to acute (24 h) hypoxia (11, 17 and 9-fold, respectively). This suggests that macrophages may play an even greater role in angiogenesis than previously appreciated. Furthermore, chronic hypoxia resulted in up-regulation of HIF-1α mRNA, in all monocyte-derived macrophage types studied. Actinomycin D experiments indicate that the increases in HIF-1α mRNA were not due to increased mRNA stability. To our knowledge this is the first study demonstrating up-regulation of HIF-1α mRNA by hypoxia in macrophages. Taken together, the data support the hypothesis that hypoxia affects monocyte to macrophage maturation, resulting in a distinct gene expression pattern and phenotype.     

Differential expression of the chemokine receptors CX3CR1 and CCR1 by microglia and macrophages in myelin-oligodendrocyte-glycoprotein-induced experimental autoimmune encephalomyelitis

Chemokines are important for the recruitment of immune cells into sites of inflammation. To better understand their functional roles during inflammation we have here studied the in vivo expression of receptors for the chemokines CCL3/CCL5/CCL7 (MIP-1alpha/RANTES/MCP-3) and CX3CL1 (fractalkine), CCR1 and CX3CR1, respectively, in rat myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis. Combined in situ hybridization and immunohistochemistry demonstrated intensely upregulated CCR1 mRNA expression in early, actively demyelinating plaques, whereas CX3CR1 displayed a more generalized expression pattern. CX3CR1 mRNA expressing cells were identified as microglia on the basis of their cellular morphology and positive GSA/B4 lectin staining. In contrast, CCR1 mRNA was preferentially expressed by ED1+ GSA/B4+ macrophages. The notion of differential chemokine receptor expression in microglia and monocyte-derived macrophages was corroborated at the protein level by extraction and flow cytometric sorting of cells infiltrating the spinal cord using gating for the surface markers CD45, ED-2 and CD11b. These observations suggest a differential receptor expression between microglia and monocyte-derived macrophages and that mainly the latter cell type is responsible for active demyelination. This has great relevance for the possibility of therapeutic intervention in demyelinating diseases such as multiple sclerosis, for example by targeting signaling events leading to monocyte recruitment.     

Migratory fate and differentiation of blood monocyte subsets

Monocytes are established circulating precursors for tissue macrophages and dendritic cells (DCs). Monocyte-derived macrophages and DCs fulfill critical roles in innate and adaptive immunity during inflammation, and it is believed that monocytes also maintain these populations in peripheral tissues during homeostasis. However, the continuous replenishment of any DC pool by blood monocytes in the steady state remains to be established, and some macrophage populations may be self-renewing in the steady state. Recent identification of mouse monocyte subsets that closely resemble human monocyte subsets has inspired a variety of techniques wherein monocytes can be readily traced in vivo to address these critical questions. There are two major monocyte subsets that vary in chemokine receptor (CCR) and adhesion molecule expression, and in migratory and differentiation properties. In humans, 'classical' CD14+ CD16- monocytes express CCR2, CD64, CD62L, whereas 'non-classical' CD14low CD16+ monocytes lack CCR2. Their counterparts in mice are CCR2+ Gr-1hi and CCR2- Gr-1low monocytes, respectively. Gr-1hi (Ly6Chi) monocytes are recruited to inflammatory sites, e.g. inflamed skin or acutely inflamed peritoneum and give rise to macrophages and DCs in inflammatory or infectious disease models and to epidermal Langerhans cells after skin inflammation. Gr-1low monocytes have been proposed as precursors for steady state DCs, but experimental evidence is as of yet limited. Fortunately, the rate of progress in the study of monocyte fate is rapidly picking up pace, giving rise to the expectation that we will soon know much more about the biology of monocytes in the steady state and inflammation.     

棕榈酸通过上调脂肪酸转位酶诱导THP-1细胞的炎症反应

目的:探讨脂肪酸转位酶(fatty acid translocase,FAT/CD36)在棕榈酸诱导的人源单核巨噬细胞THP-1炎症反应中的作用。方法:给予不同浓度棕榈酸(0 mmol/L、0.1 mmol/L和0.2 mmol/L)处理THP-1细胞24h。Transwell迁移实验检测细胞迁移能力;real-time PCR检测CD36、肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)及单核细胞趋化蛋白1(MCP-1)的m RNA表达水平;ELISA和Western blot法检测靶蛋白的蛋白含量。利用RNA干扰技术构建低表达CD36(si CD36)的THP-1细胞模型,观察抑制CD36表达对细胞迁移、炎症及趋化因子表达的影响。结果:棕榈酸促进了THP-1细胞CD36的m RNA和蛋白表达,且增强了THP-1细胞炎症/趋化因子的m RNA和蛋白表达,差异有统计学意义(P0.05)。棕榈酸处理组THP-1细胞的迁移能力明显强于对照组。与阴性对照组细胞相比,si CD36组炎症因子的m RNA和蛋白表达明显降低(P0.05),THP-1细胞迁移水平也明显降低(P0.05)。结论:棕榈酸通过上调THP-1巨噬细胞中CD36表达,促进了巨噬细胞的迁移,促使细胞产生大量炎症/趋化因子,导致巨噬细胞炎症反应。     

Molecular mechanisms regulating CD13‐mediated adhesion

CD13/Aminopeptidase N is a transmembrane metalloproteinase that is expressed in many tissues where it regulates various cellular functions. In inflammation, CD13 is expressed on myeloid cells, is up‐regulated on endothelial cells at sites of inflammation and mediates monocyte/endothelial adhesion by homotypic interactions. In animal models the lack of CD13 alters the profiles of infiltrating inflammatory cells at sites of ischaemic injury. Here, we found that CD13 expression is enriched specifically on the pro‐inflammatory subset of monocytes, suggesting that CD13 may regulate trafficking and function of specific subsets of immune cells. To further dissect the mechanisms regulating CD13‐dependent trafficking we used the murine model of thioglycollate‐induced sterile peritonitis. Peritoneal monocytes, macrophages and dendritic cells were significantly decreased in inflammatory exudates from global CD13^KO animals when compared with wild‐type controls. Furthermore, adoptive transfer of wild‐type and CD13^KO primary myeloid cells, or wild‐type myeloid cells pre‐treated with CD13‐blocking antibodies into thioglycollate‐challenged wild‐type recipients demonstrated fewer CD13^KO or treated cells in the lavage, suggesting that CD13 expression confers a competitive advantage in trafficking. Similarly, both wild‐type and CD13^KO cells were reduced in infiltrates in CD13^KO recipients, confirming that both monocytic and endothelial CD13 contribute to trafficking. Finally, murine monocyte cell lines expressing mouse/human chimeric CD13 molecules demonstrated that the C‐terminal domain of the protein mediates CD13 adhesion. Therefore, this work verifies that the altered inflammatory trafficking in CD13^KO mice is the result of aberrant myeloid cell subset trafficking and further defines the molecular mechanisms underlying this regulation.     

Reprint of: Alterations of TRIM21-mRNA expression during monocyte maturation

Tripartite motif-containing protein 21 (TRIM21) play a dual role in the cytoplasm of the cells where it facilitates destruction of some antibody-coated viruses and some bacteria, and initiates synthesis of proinflammatory cytokines. Macrophages and CD16⁺ monocyte subset can particularly participate in a proinflammatory response caused by viral infection, however, the molecular mechanisms underlying these processes are not fully understood. The aim of this study was to determine the level of TRIM21-mRNA expression in monocyte subsets including: classical (CD14⁺⁺CD16⁻), intermediate (CD14⁺⁺CD16⁺) and non-classical (CD14⁺CD16⁺⁺) monocytes, as well as during in vitro differentiation of the isolated monocytes towards dendritic cells or macrophages.Our results revealed that the level of TRIM21 mRNA expression was significantly lower in CD16- monocytes, when compared to CD16⁺ cells and the whole monocyte population, yet no significant differences were observed when CD16⁺ population was divided into intermediate and non-classical subsets. More pronounced differences were observed in the case of monocyte-derived macrophages (MDM) and dendritic cells (DCs). TRIM21-mRNA expression level was app. 6-fold higher in DCs, and app. 16-fold higher in MDM (p < 0,01), when compared to freshly isolated monocytes.Our results may suggest the new mechanism of increased proinflammatory cytokine production by CD16⁺ (intermediate and non-classical) monocytes and macrophages, at least in patients with acute or chronic infections, caused by enveloped viruses. We suggest that TRIM21 may be one of the factors associated with the “switching on” the proinflammatory programme in CD16⁺ monocytes or monocyte-derived macrophages.     

Alterations of TRIM21-mRNA expression during monocyte maturation

Tripartite motif-containing protein 21 (TRIM21) play a dual role in the cytoplasm of the cells where it facilitates destruction of some antibody-coated viruses and some bacteria, and initiates synthesis of proinflammatory cytokines. Macrophages and CD16⁺ monocyte subset can particularly participate in a proinflammatory response caused by viral infection, however, the molecular mechanisms underlying these processes are not fully understood. The aim of this study was to determine the level of TRIM21-mRNA expression in monocyte subsets including: classical (CD14⁺⁺CD16⁻), intermediate (CD14⁺⁺CD16⁺) and non-classical (CD14⁺CD16⁺⁺) monocytes, as well as during in vitro differentiation of the isolated monocytes towards dendritic cells or macrophages.Our results revealed that the level of TRIM21 mRNA expression was significantly lower in CD16- monocytes, when compared to CD16⁺ cells and the whole monocyte population, yet no significant differences were observed when CD16⁺ population was divided into intermediate and non-classical subsets. More pronounced differences were observed in the case of monocyte-derived macrophages (MDM) and dendritic cells (DCs). TRIM21-mRNA expression level was app. 6-fold higher in DCs, and app. 16-fold higher in MDM (p < 0,01), when compared to freshly isolated monocytes.Our results may suggest the new mechanism of increased proinflammatory cytokine production by CD16⁺ (intermediate and non-classical) monocytes and macrophages, at least in patients with acute or chronic infections, caused by enveloped viruses. We suggest that TRIM21 may be one of the factors associated with the “switching on” the proinflammatory programme in CD16⁺ monocytes or monocyte-derived macrophages.     

L-Selectin Expression on the Surface of Peripheral Blood Leucocytes from Rheumatoid Arthritis Patients is Linked to Disease Activity

Recruitment of mononuclear cells from the circulation to sites of inflammation relies on migration across vessel endothelium. T and B cells, macrophages and neutrophils infiltrate synovial tissue of rheumatoid arthritis (RA) patients. The authors have analysed the numbers of circulating CD3⁺, CD19⁺ lymphocytes, monocytes, and granulocytes expressing adhesion molecules (L-selectin, CD44 and CD11a), together with levels of expression in RA patients compared to healthy individuals. Numbers of leucocytes expressing the adhesion molecules detected were similar in RA and control groups. Lower levels of expression of L-selectin on all cells were found in RA patients compared to controls. Expression of L-selectin on T and B cells was found to correlate with disease activity in RA. The authors have observed a characteristic pattern of adhesion molecule expression in RA patients, particularly when analysing the relationships between cells. The close regulation of these molecules between RA patients and healthy individuals is discussed.     

Up-regulation of ICAM-1, CD11a/CD18 and CD11c/CD18 on human THP-1 monocytes stimulated by Streptococcus suis serotype 2

Streptococcus suis serotype 2 is known to be a major pathogen of swine, causing mainly meningitis. It is also a zoonotic agent leading predominantly to meningitis in humans working in close contact with pigs. In this study, we investigated the ability of S. suis to up-regulate the expression of adhesion molecules involved in inflammation, using an enzyme-linked immunosorbent assay. S. suis serotype 2 stimulated the up-regulation of the expression of intercellular adhesion molecule-1 (ICAM-1, CD54), CD11a/CD18 and CD11c/CD18 on human THP-1 monocytes, but did not change that of ICAM-1, vascular cell adhesion molecule-1 (VCAM-1, CD106) and E-selectin (CD62E) on human endothelial cells. The up-regulation of adhesion molecules was time- and bacterial concentration-dependent, and cell wall components were largely responsible for such stimulation. To a lesser extent, purified haemolysin of S. suis also stimulated adhesion molecule expression. Stimulation of monocytes with strains of different origin showed that there was no clear tendency for human strains to induce a higher expression of adhesion molecules than strains from diseased pigs. Finally, monocytes stimulated with S. suis also showed an increase in adherence to endothelial cells. Hence, S. suis is capable of up-regulating important adhesion molecules involved in inflammation, which may result in an increased leucocyte recruitment into sites of infection, thus providing a possible mechanism for some of the inflammatory features of meningitis caused by this pathogen.     

Atherosclerotic lesions grow through recruitment and proliferation of circulating monocytes in a murine model

Macrophage-derived foam cells in developing atherosclerotic lesions may potentially originate either from recruitment of circulating monocytes or from migration of resident tissue macrophages. In this study, we have determined the source of intimal macrophages in the apoE-knockout mouse flow-cessation/hypercholesterolemia model of atherosclerosis using a bone marrow transplantation approach. We also examined the time course and spatial distribution of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 expression to assess whether endothelial adhesion molecules were involved in recruitment of either circulating monocytes or resident macrophages. We used allelic variants of the mouse common leukocyte antigen (CD45) to distinguish host-derived and donor-derived white blood cells (WBCs) both in blood and in macrophage-rich carotid lesions. We found that the distribution of CD45 isoforms in lesions is similar to that of circulating WBCs, whereas the host-type CD45 isoform is more prevalent in resident adventitial macrophages. These data indicate that macrophage-derived foam cells in the lesion derive mainly from circulating precursors rather than from resident macrophages. The corresponding time course of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 expression suggests that recruitment of circulating WBCs by endothelial adhesion molecules is likely to be more important during lesion initiation than during the later phase of rapid lesion growth.     

The chemokine CCL5 induces selective migration of bovine classical monocytes and drives their differentiation into LPS-hyporesponsive macrophages in vitro

Human and mouse studies indicate distinct roles of selected chemokines for monocyte subset attraction. We therefore analyzed the still unknown sensitivity and response of bovine monocyte subsets toward two monocyte-attracting chemokines (CCL2, CCL5). Only CCL5 induced a significant Ca²⁺influx and migration response in bovine monocytes, with classical and intermediate monocytes being significantly stimulated and attracted compared to nonclassical monocytes. The presence of CCL5 during in vitro macrophage differentiation did not alter their capacity to phagocytize or to generate reactive oxygen species upon stimulation with E. coli. However, macrophages differentiated in the presence of CCL5 displayed an altered phenotype with significantly less expressed CD14 and MHC class II molecules, whereas CD16 was upregulated. Moreover, CCL5-differentiated macrophages displayed a reduced upregulation of CXCL8, ARG1, IL6 and IL10 mRNA. Taken together, CCL5 but not CCL2 mainly attract bovine classical monocytes and promote their differentiation into LPS-hypo-responsive macrophages.     

Human β defensin‐3 induces chemokines from monocytes and macrophages: diminished activity in cells from HIV‐infected persons

Human β defensin‐3 (hBD‐3) is an antimicrobial peptide with diverse functionality. We investigated the capacity of hBD‐3 and, for comparison, Pam3CSK4 and LL‐37 to induce co‐stimulatory molecules and chemokine expression in monocytes. These stimuli differentially induced CD80 and CD86 on the surface of monocytes and each stimulant induced a variety of chemokines including monocyte chemoattractant protein 1 (MCP‐1), Gro‐α, macrophage‐derived chemokine (MDC) and macrophage inflammatory protein 1β (MIP1β), while only hBD‐3 and Pam3CSK4 significantly induced the angiogenesis factor, vascular endothelial growth factor (VEGF). Human BD‐3 induced similar chemokines in monocyte‐derived macrophages and additionally induced expression of Regulated upon activation normal T‐cell expressed and presumably secreted (RANTES) in these cells. Comparison of monocytes from HIV⁺ and HIV^– donors indicated that monocytes from HIV⁺ donors were more likely to spontaneously express certain chemokines (MIP‐1α, MIP‐1β and MCP‐1) and less able to increase expression of other molecules in response to hBD‐3 (MDC, Gro‐α and VEGF). Chemokine receptor expression (CCR5, CCR2 and CXCR2) was relatively normal in monocytes from HIV⁺ donors compared with cells from HIV^– donors with the exception of diminished expression of the receptor for MDC, CCR4, which was reduced in the patrolling monocyte subset (CD14⁺ CD16⁺⁺) of HIV⁺ donors. These observations implicate chemokine induction by hBD‐3 as a potentially important mechanism for orchestrating cell migration into inflamed tissues. Alterations in chemokine production or their receptors in monocytes of HIV‐infected persons could influence cell migration and modify the effects of hBD‐3 at sites of inflammation.     

Down-regulation by tumor necrosis factor-alpha of neutrophil cell surface expression of the sialophorin CD43 and the hyaluronate receptor CD44 through a proteolytic mechanism.

Adhesion of human neutrophils to endothelial cells is a crucial step during migration to the extravascular sites of inflammation. A large number of molecules, including the CD44 and LAM-1 antigens, have been described to participate in this process. We have investigated the regulation by human recombinant tumor necrosis factor-alpha (TNF-alpha) of human neutrophil plasma membrane expression of both CD44 and LAM-1 adhesion molecules, as well as that of CD43 sialophorin, which has been involved in adhesion and activation of leukocytes. The expression of these three antigens was down-regulated in neutrophils upon TNF-alpha treatment, as determined by immunofluorescence and immunoprecipitation experiments. However, the expression of other cell surface molecules, such as CD45 or CD11b, was up-regulated. Similar regulatory effects were also observed upon neutrophil treatment with other activating agents such as the chemoattractant peptide formyl-Met-Leu-Phe, the calcium ionophore A23187, or the phorbol ester phorbol 12-myristate 13-acetate. Protease inhibitors virtually abrogated the TNF-alpha-induced down-regulation of CD43 and CD44 expression, but not that of LAM-1, suggesting the involvement of a protease activity in this process. These results underline the role of TNF-alpha on the differential regulation of cell surface expression of neutrophil adhesion molecules, thus implying modifications in the neutrophil adhesive properties.     

PECAM-1 and leukosialin (CD43) expression correlate with heightened inflammation in rat adjuvant-induced arthritis.

A hallmark of both adjuvant-induced arthritis (AIA) and rheumatoid arthritis is chronic joint inflammation characterized by ingress of leukocytes into the inflamed synovial tissue. The timing of expression of adhesion molecules, which govern the ingress of leukocytes, is important in the orchestration of an inflammatory response. We examined the expression of vascular cell adhesion molecule-1 (VCAM-1), sialo adhesin, platelet and endothelial cell adhesion molecule-1 (PECAM-1), and leukosialin (CD43) in AIA, starting at adjuvant injection (day 0), through the peak of inflammation (day 18 postadjuvant injection), until day 54. VCAM-1 is constitutively expressed on the lining layer and ECs and its expression levels do not change throughout the progression of AIA. Sialoadhesin synovial tissue lining cell expression is decreased after adjuvant injection. In contrast, PECAM-1 expression is increased on synovial tissue lining cells on day 7 and is elevated through day 54 (peaking on day 54 with six-fold more cells expressing PECAM-1). PECAM-1 expression on endothelial cells peaks on day 7 with three-fold more cells expressing it, while on macrophages expression maximizes on day 25 with six-fold more cells expressing PECAM-1. CD43 expression is increased on synovial tissue lining cells, macrophages, neutrophils, and lymphocytes on days 18 and 25, before going back to basal levels. The increased expression of PECAM-1 and CD43 on leukocytes at the height of inflammation in AIA suggests important roles for these adhesion molecules in potentially binding their EC ligands resulting in leukocyte ingress into the synovial tissue.     

Defective up-regulation of CD49d in final maturation of NOD mouse macrophages

Macrophages are potent regulators of both innate and adaptive immunity. They play a central role in the development of autoimmune diabetes and are among the first cells to appear in peri-islet infiltrates of NOD mice that spontaneously develop diabetes. Since efficient adhesion and migration are crucial for proper macrophage trafficking, we examined the migration and fibronectin (FN) adhesion capacity of NOD macrophages, as well as the regulation and expression of the FN receptors alpha4beta1 and alpha5beta1. When compared to macrophages from control strains, resident NOD macrophages showed a reduced ability to adhere to and migrate on FN, a delayed clearance following peritoneal inflammation, and substantially lower expression levels of the alpha4beta1 integrin alpha chain, CD49d. NOD bone marrow-derived macrophages were specifically defective in the LPS-induced increase in CD49d expression. Moreover, the mitogen-activated protein kinase extracellular signal-regulated kinase-1/2 negatively regulated macrophage CD49d expression and strongly suppressed its expression in NOD macrophages. The data presented herein indicate that the LPS-activated signaling cascade plays a critical role in CD49d expression of macrophages. Mature NOD macrophages are characterized by decreased CD49d expression and show defective CD49d-mediated adhesion to FN.     

Differential regulation of CD11b on gammadelta T cells and monocytes in response to unripe apple polyphenols

Leukocyte adhesion and migration are mediated partially by CD11b/CD18 (membrane-activated complex-1, CR3). Earlier studies have demonstrated a role for green tea polyphenols in down-regulating CD11b on CD8(+) T cells and monocytes. We have shown recently a stimulatory effect of unripe apple polyphenols (APP) on gammadelta T cells. Thus, we compared the effect of APP on bovine gammadelta T cell and monocyte CD11b expression. Purified bovine monocytes and monocyte-depleted PBLs were cultured with APP. CD11b levels decreased on monocytes in response to APP. In contrast, a gammadelta T cell subset responded to APP by up-regulating CD11b. The CD11b regulation was not seen on gammadelta T cells or monocytes treated with APP fractions depleted of tannins. The APP-induced down-regulation of CD11b on monocytes was inhibited by an anti-CD11b mAb, consistent with previous studies showing that polyphenols bind CD11b. As expected, the anti-CD11b mAb had no effect on the APP response in resting gammadelta T cells, as these cells lacked CD11b. Consistent with the changes in surface CD11b expression, APP-treated gammadelta T cells showed increased adherence to plastic, whereas monocyte adhesion was reduced. APP also induced cytokine gene expression in gammadelta T cells. Some polyphenols are thought of as anti-inflammatory agents; however, these data, as well as other ongoing studies, indicate they have a proinflammatory effect on gammadelta T cells. In vivo, plant polyphenols may enhance gammadelta T cell migration and function at sites of inflammation, where they could induce rapid, immune-regulatory and innate-like immune responses.     

In vivo transmigrated monocytes from patients with stable coronary artery disease have a reduced expression of CD11b

Coronary artery disease (CAD) is characterized by infiltration of monocyte derived cells in the intima of the vessel wall. We hypothesized that accumulation of these cells is caused partly by an altered monocyte transmigration process in CAD. To gain insight into this issue we applied the skin blister method that allows collection of in vivo transmigrated cells at sites of local inflammation. Nineteen patients with stable CAD and 19 matched controls were enrolled. Markers of inflammation and gradients of chemokines, as well as adhesion molecule expression and up-regulation capacity, were studied. The expression of inflammatory markers, such as C-reactive protein, interleukin (IL)-6, tumour necrosis factor-alpha and IL-10, was similar in patients and controls, indicating that patients were in a stable phase of the disease. Expression of adhesion molecules, CD11b and very late activation antigen-4, on peripheral monocytes did not differ between patients and controls. However, following in vivo transmigration, monocytes in patients with CAD had a significantly reduced expression and mobilization of CD11b. The effect on CD11b could not be reproduced by in vitro stimulation with blister fluid, representing a local inflammatory milieu, or in an in vitro system of transmigration. These findings point towards differences in monocyte CD11b expression and availability at an inflammatory site between patients with CAD and healthy controls.     

Differential detergent fractionation for non-electrophoretic bovine peripheral blood monocyte proteomics reveals proteins involved in professional antigen presentation

Professional antigen presenting cells (APC), dendritic cells (DC) and their myeloid progenitors, monocytes/macrophages are critical controllers of innate and adaptive immunity. Here we show that differential detergent fractionation (DDF) analysis of bovine monocytes reveals proteins related to antigen pattern recognition, uptake and presentation to immunocompetent lymphocytes. We identify 53 bovine proteins involved in immune function of professional APC. In particular, 13 adhesion molecules, three toll-like receptors (TLR1, 6 and 8), three antigen uptake-related proteins (including mannose receptor [MR] precursor), and eight actin-like proteins involved in active endocytosis were identified. In addition, MHC class I and II-related proteins, cytokines, active substances and growth factors have been identified. We conclude that the DDF approach can provide interpretable and meaningful functional information concerning protein expression profiles associated with monocyte activation, transformation into macrophages and/or immature DC, and maturation of monocyte-derived DC in the presence of multiple bovine pathogens.