Chemoattractant MCP-1 Research Articles

Role of Graft-Derived CCL2/MCP-1 in Antibody-Mediated Rejection of Cardiac Allografts.

The role of monocytes/macrophages in antibody-mediated rejection (AMR) remains unclear. We have reported that murine CCR5-/-/CD8-/- recipients produce high titers of donor-specific antibody and reject MHC-mismatched heart allografts. The current studies tested the magnitude of injury and alloimmune responses to grafts deficient in expression of the monocyte chemoattractant MCP-1/CCL2 during AMR. B6.CCR5-/-/CD8-/- recipients rejected MHC-mismatched wild-type A/J allografts with high donor-reactive antibody titers and diffuse C4d deposition in the large vessels and myocardial capillaries, features consistent with AMR. Donor-specific CD4 T cells producing IFN-γ in the recipient spleen were significantly decreased in response to A/J.CCL2-/- vs. A/J allografts and this resulted in decreased donor-reactive antibody production and C4d deposition in allografts in B6. CCR5-/-/CD8-/- recipients at day 7 post-transplant. Consequently, A/J.CCL2-/- allograft survival was slightly but significantly longer than that of A/J allografts (MST: wild-type A/J, day 8 vs. A/J.CCL2-/-, day 10, P < 0.01). Allograft CCL2 deficiency reduced classically activated (M1) macrophage and other inflammatory leukocyte infiltration into the allografts. Strikingly, A/J.CCL2-/- allograft survival was much longer than wild-type A/J allografts in recipients treated with 0.25 mg anti-CD40L mAb at the time of transplant (MST: wild-type A/J, day 21.5 vs. A/J.CCL2-/-, day 58, P < 0.01) with lower titers of donor-reactive antibody and decreased leukocyte infiltration. CCL2 deficiency also reduced pro-fibrotic gene expression and delayed the development of interstitial fibrosis when compared to wild-type A/J allografts. The results indicate that allograft-derived CCL2/MCP-1 plays an important role in directing M1 macrophages and other inflammatory cell infiltration into allografts and in the development of donor-reactive antibody and vascular rejection and suggest CCL2/MCP-1 as a therapeutic target for antibody-mediated vascular rejection and chronic graft injury.

Exercise effects on polyp burden and immune markers in the ApcMin/+ mouse model of intestinal tumorigenesis.

Many observational epidemiologic studies suggest an association between exercise and colon cancer risk. The mechanisms contributing to a preventative effect of exercise on colon cancer are complex and multifaceted. Altered immune system function is one possible mechanism that has been largely unexplored. Therefore, the purpose of this study was to examine the effects of exercise on markers associated with macrophages and select T cell populations in a mouse model of intestinal tumorigenesis and to relate this to polyp characteristics. Male ApcMin/+ mice were randomly assigned to either sedentary (Sed) or exercise (Ex) treatment (n=6–9/group). The exercise treatment consisted of treadmill running for 1 h/day and 6 days a week at 15 m/min from 4 until 16 weeks of age. Intestinal polyps were counted and categorized by size. Mucosal tissue was analyzed for mRNA expression of overall macrophages (F4/80), for genes associated with M1 (IL-12, IL-23 and Nos2) and M2 (CD206, IL-10, IL-4, CCL17, CCL22 and Arg-1) macrophages and the macrophage chemoattractants MCP-1, fetuin A and CXCL14. Markers for cytotoxic T cells (CTLs) and regulatory T cells were also examined by measuring mRNA expression of CD8 and Foxp3, respectively. While there was no significant difference in overall polyp number between the groups (Sed, 23.3±4.3; and Ex, 16.5±4.3), Ex did have a reduction in the number of large polyps (Sed, 6.1±1.1; and Ex, 3.0±0.6) (P<0.05). This was consistent with a decrease in spleen weight (P<0.05). Similarly, Ex reduced mRNA expression of overall macrophages (F4/80) as well as markers associated with both M1 (IL-12) and M2 (CD206, CCL22 and Arg-1) subtypes (P<0.05) but there was no significant decrease in macrophage chemoattractants. CD8 expression was increased while Foxp3 expression was decreased with Ex (P<0.05). Overall the data provide important new information on immune regulation as a possible mechanism for the documented benefits of exercise training on reducing colon cancer progression.

Migration of Human Monocytes is stimulated by Jagged-1 and DLL-4 via the Notch pathway

The Notch signaling pathway is a well-conserved signaling pathway. As known so far Notch signaling is engaged in physiological processes in pulmonary hypertension (PAH). As shown recently, a significant increase of perivascular numbers of macrophages (CD68(+)) and monocytes (CD14(+)) can be observed in PAH. The aim of this study was to elucidate the role of the Notch signaling pathway in the migration of human monocytes. Human monocytes were isolated from venous blood, taken from healthy donors. For the chemotactic assays modified 48-well microchemotaxis chambers were used. Cells migrated through a 5 µm pore sized cellulose membrane filter for 45 min in a humidified atmosphere against a gradient of the substances tested (fMLP, Jagged-1, DLL-4). Migration depth of the cells was quantified microscopically by measuring the distance [µm] from the surface of the filters to the leading front of the cells. The Notch ligands DLL-4 and Jagged-1 significantly stimulated direct and indirect migration in a dose dependent manner revealing a maximal effect at 100ng/ml. The effect was comparable with the maximal stimulation elicited by the well-known chemoattractants fMLP and MCP-1 (10 -8 M). Via checkerboard assay was shown, that monocytes migrate in a chemotactic and chemokinetic manner. Interestingly, at least 30 min of preincubation was necessary to induce chemokinesis. Inhibition of migration was reached by preincubation of cells with ADAM 17. Jagged-1 and DLL-4 stimulated migration of human monocytes in vitro. Thus, it can be expected, that Jagged-1 and DLL-4 are involved in innate and adaptive immunological responses in PAH. Consequently, the Notch pathway may serve as a new therapeutic target in future.

Ursolic acid protects monocytes against metabolic stress-induced priming and dysfunction by preventing the induction of Nox4

AimsDietary supplementation with ursolic acid (UA) prevents monocyte dysfunction in diabetic mice and protects mice against atherosclerosis and loss of renal function. The goal of this study was to determine the molecular mechanism by which UA prevents monocyte dysfunction induced by metabolic stress. Methods and resultsMetabolic stress sensitizes or “primes” human THP-1 monocytes and murine peritoneal macrophages to the chemoattractant MCP-1, converting these cells into a hyper-chemotactic phenotype. UA protected THP-1 monocytes and peritoneal macrophages against metabolic priming and prevented their hyper-reactivity to MCP-1. UA blocked the metabolic stress-induced increase in global protein-S-glutathionylation, a measure of cellular thiol oxidative stress, and normalized actin-S-glutathionylation. UA also restored MAPK phosphatase-1 (MKP1) protein expression and phosphatase activity, decreased by metabolic priming, and normalized p38 MAPK activation. Neither metabolic stress nor UA supplementation altered mRNA or protein levels of glutaredoxin-1, the principal enzyme responsible for the reduction of mixed disulfides between glutathione and protein thiols in these cells. However, the induction of Nox4 by metabolic stress, required for metabolic priming, was inhibited by UA in both THP-1 monocytes and peritoneal macrophages. ConclusionUA protects THP-1 monocytes against dysfunction by suppressing metabolic stress-induced Nox4 expression, thereby preventing the Nox4-dependent dysregulation of redox-sensitive processes, including actin turnover and MAPK-signaling, two key processes that control monocyte migration and adhesion. This study provides a novel mechanism for the anti-inflammatory and athero- and renoprotective properties of UA and suggests that dysfunctional blood monocytes may be primary targets of UA and related compounds.

Obesity and Breast Cancer: A Multipartite Connection

Obesity and Breast Cancer: A Multipartite Connection

Aluminium, carbonyls and cytokines in human nipple aspirate fluids: Possible relationship between inflammation, oxidative stress and breast cancer microenvironment

The human breast is likely exposed to Al (aluminium) from many sources including diet and personal care products. Underarm applications of aluminium salt-based antiperspirant provide a possible long-term source of exposure, especially after underarm applications to shaved and abraded skin. Al research in breast fluids likely reflects the intraductal microenvironment. We found increased levels of aluminium in noninvasively collected nipple aspirate fluids (NAF) from 19 breast cancer patients compared with 16 healthy control subjects (268 vs 131μg/l, respectively; p<0.0001). In the same NAF samples we found significantly increased levels of protein oxidative carbonyls in cancer patients compared to healthy women (2.35 vs 0.41nmol/mg prot, respectively; p<0.0001). Aluminium content and carbonyl levels showed a significant positive linear correlation (r2 0.6628, p<0.0001). In cancer NAF samples (containing higher amounts of aluminium salts) we also found a significantly increased levels of pro-inflammatory cytokines (IL-1β, IL-6, IL-12 p70, and TNF-α) and chemoattractant CC and CXC chemokines (IL-8, MIP-1α and MCP-1). In 12 invasive cancer NAF samples we found a significant positive linear correlation among aluminium, carbonyls and pro-inflammatory IL-6 cytokine (Y=64.79x−39.63, r2 0.8192, p<0.0005), as well as pro-inflammatory monocyte chemoattractant MCP-1 cytokine (Y=2026x−866, r2 0.9495, p<0.0001). In addition to emerging evidence, our results support the possible involvement of aluminium ions in oxidative and inflammatory status perturbations of breast cancer microenvironment, suggesting aluminium accumulation in breast microenvironment as a possible risk factor for oxidative/inflammatory phenotype of breast cells.

Critical role of aldehydes in cigarette smoke-induced acute airway inflammation.

BackgroundCigarette smoking (CS) is the most important risk factor for COPD, which is associated with neutrophilic airway inflammation. We hypothesize, that highly reactive aldehydes are critical for CS-induced neutrophilic airway inflammation.MethodsBALB/c mice were exposed to CS, water filtered CS (WF-CS) or air for 5 days. Levels of total particulate matter (TPM) and aldehydes in CS and WF-CS were measured. Six hours after the last exposure, inflammatory cells and cytokine levels were measured in lung tissue and bronchoalveolar lavage fluid (BALF). Furthermore, Beas-2b bronchial epithelial cells were exposed to CS extract (CSE) or WF-CS extract (WF-CSE) in the absence or presence of the aldehyde acrolein and IL-8 production was measured after 24 hrs.ResultsCompared to CS, in WF-CS strongly decreased (CS; 271.1 ± 41.5 μM, WF-CS; 58.5 ± 8.2 μM) levels of aldehydes were present whereas levels of TPM were only slightly reduced (CS; 20.78 ± 0.59 mg, WF-CS; 16.38 ± 0.36 mg). The numbers of mononuclear cells in BALF (p<0.01) and lung tissue (p<0.01) were significantly increased in the CS- and WF-CS-exposed mice compared to air control mice. Interestingly, the numbers of neutrophils (p<0.001) in BALF and neutrophils and eosinophils (p<0.05) in lung tissue were significantly increased in the CS-exposed but not in WF-CS-exposed mice as compared to air control mice. Levels of the neutrophil and eosinophil chemoattractants KC, MCP-1, MIP-1α and IL-5 were all significantly increased in lung tissue from CS-exposed mice compared to both WF-CS-exposed and air control mice. Interestingly, depletion of aldehydes in WF-CS extract significantly reduced IL-8 production in Beas-2b as compared to CSE, which could be restored by the aldehyde acrolein.ConclusionAldehydes present in CS play a critical role in inflammatory cytokine production and neutrophilic- but not mononuclear airway inflammation.

P38 MAPK in Myeloma Cells Regulates Osteoclast and Osteoblast Activity and Induces Bone Destruction

p38 mitogen-activated protein kinase (MAPK), which is constitutively activated in human myeloma, has been implicated in bone destruction by this cancer, but the processes it recruits are obscure. In this study, we show that p38 activity in myeloma inhibits osteoblast differentiation and bone formation, but also enhances osteoclast maturation and bone resorption. p38 regulated the expression and secretion of the Wnt pathway antagonist DKK-1 and the monocyte chemoattractant MCP-1. Attenuating p38, DKK-1, or MCP-1 were each sufficient to reduce bone lesions in vivo. Although it is well known that DKK-1 inhibits osteoblast differentiation, we found that together with MCP-1, it could also promote osteoclast differentiation and bone resorption. The latter effects were mediated by enhancing expression of RANK in osteoclast progenitor cells and by upregulating secretion of its ligand RANKL from stromal cells and mature osteoblasts. In summary, our study defined the mechanisms by which p38 signaling in myeloma cells regulates osteoblastogenesis, osteoclastogenesis, and bone destruction. Our findings, which may have implications for bone invasion by other cancers where p38 is elevated, strongly suggests that targeting p38 for inhibition may offer an effective therapeutic approach to treat osteolytic bone lesions in patients with myeloma.

Effects of acute and chronic low density lipoprotein exposure on neutrophil function

Mounting evidence suggests that obesity and the metabolic syndrome have significant but often divergent effects on the innate immune system. These effects have been best established in monocytes and macrophages, particularly as a consequence of the hypercholesterolemic state. We have recently described defects in neutrophil function in the setting of both obesity and hypercholesterolemia, and hypothesized that exposure to elevated levels of lipoproteins, particularly LDL its oxidized forms, contributed to these defects. As a model of chronic cholesterol exposure, we examined functional responses of bone marrow neutrophils isolated from non-obese mice with diet-induced hypercholesterolemia compared to normal cholesterol controls. Chemotaxis, calcium flux, CD11b display, and F-actin polymerization were assayed in response to several chemoattractants, while neutrophil cytokine transcriptional response was determined to LPS. Following this, the acute effects of isolated LDL and its oxidized forms on normal neutrophils were assayed using the same functional assays. We found that neutrophils from non-obese hypercholesterolemic mice had blunted chemotaxis, altered calcium flux, and normal to augmented CD11b display with prolonged actin polymerization in response to stimuli. In response to acute exposure to lipoproteins, neutrophils showed chemotaxis to LDL which increased with the degree of LDL oxidation. Paradoxically, LDL oxidation yielded the opposite effect on LDL-induced CD11b display and actin polymerization, and both native and oxidized LDL were found to induce neutrophil transcription of the monocyte chemoattractant MCP-1. Together these findings suggest that chronic hypercholesterolemia impairs neutrophil functional responses, and these defects may be in part due to protracted signaling responses to LDL and its oxidized forms.

Macrophages promote cyst growth in polycystic kidney disease.

Polycystic kidney disease (PKD) exhibits an inflammatory component, but the contribution of inflammation to cyst progression is unknown. Macrophages promote the proliferation of tubular cells following ischemic injury, suggesting that they may have a role in cystogenesis. Furthermore, cultured Pkd1-deficient cells express the macrophage chemoattractants Mcp1 and Cxcl16 and stimulate macrophage migration. Here, in orthologous models of both PKD1 and PKD2, abnormally large numbers of alternatively activated macrophages surrounded the cysts. To determine whether pericystic macrophages contribute to the proliferation of cyst-lining cells, we depleted phagocytic cells from Pkd1(fl/fl);Pkhd1-Cre mice by treating with liposomal clodronate from postnatal day 10 until day 24. Compared with vehicle-treated controls, macrophage-depleted mice had a significantly lower cystic index, reduced proliferation of cyst-lining cells, better-preserved renal parenchyma, and improved renal function. In conclusion, these data suggest that macrophages home to cystic areas and contribute to cyst growth. Interruption of these homing and proliferative signals could have therapeutic potential for PKD.

Interleukin-17 Promotes Early Allograft Inflammation

Acute cellular rejection of organ transplants is executed by donor-reactive T cells, which are dominated by interferon-gamma-producing cells. As interferon-gamma is dispensable for graft destruction, we evaluated the contribution of interleukin-17A (IL-17) to intragraft inflammation in major histocompatibility complex-mismatched heart transplants. A/J (H-2(a)) cardiac allografts placed into wild-type BALB/c (H-2(d)) mice induced intragraft IL-17 production on day 2 after transplant. Allografts placed into BALB/c IL-17(-/-) recipients demonstrated diminished production of the chemokines CXCL1 and CXCL2 and delayed neutrophil and T cell recruitment. However, by day 7 after transplant, allografts from IL-17(-/-) and wild-type recipients had comparable levels of cellular infiltration. The priming of donor-specific T cells was not affected by the absence of IL-17, and the kinetics of cardiac allograft rejection were similar in wild-type and IL-17(-/-) recipients. In contrast, IL-17(-/-) mice depleted of CD8 T cells rejected A/J allografts in a delayed fashion compared with CD8-depleted wild-type recipients. Although donor-reactive CD4 T cells were efficiently activated in both groups, the infiltration of effector T cells into allografts was impaired in IL-17(-/-) recipients. Our data indicate that locally produced IL-17 amplifies intragraft inflammation early after transplantation and promotes tissue injury by facilitating T cell recruitment into the graft. Targeting the IL-17 signaling network in conjunction with other graft-prolonging therapies may decrease this injury and improve the survival of transplanted organs.

Abstract 2339: Stimulation of sialyl Lewis X carbohydrate expression in lung carcinoma cells by neutrophils: Implications for metastasis

Abstract Sialyl Lewis X (sLeX) is a carbohydrate epitope expressed on leukocytes and carcinoma cells that promotes binding of both cell types to vascular endothelium by selectin mediated processes. These binding events are crucial for leukocyte trafficking to lymphoid tissues and sites of inflammation and metastasis of cancer cells. Increased sLeX expression is correlated with advanced disease and a poor prognosis for several types of carcinomas. Synthesis of sLeX is dependent on activity of the enzymes α-(1,3)-fucosyltransferase (FucT-III) and α2,3-sialyltransferase. Exposure to tumor necrosis factor alpha (TNF-α) has been described to upregulate FucT-III resulting in increased sLeX in the airways of patients with respiratory disease, however, little is known about the molecular mechanisms involved in regulation of sLeX expression in the inflammatory tumor microenvironment. We stably transfected lung carcinoma cells with FucT-III to investigate the role of TNF-α in regulation of sLeX expression, selectin binding ability, and metastatic potential of lung carcinoma cells. Stimulation of FucT-III transfected but not mock-transfected H1299 and A549 lung carcinoma cells with recombinant TNF-α resulted in a concentration dependent upregulation of sLeX expression, increased binding of carcinoma cells to E-selectin, and increased migration of transfectants through Matrigel. Lung carcinoma cells transfected with FucT-III demonstrated high secretion of the neutrophil chemoattractant cytokines IL-8, GRO-α and MCP-1. Co-culture of lung carcinoma cells with neutrophil conditioned media and neutrophils isolated from blood at an optimal ratio of 1:5 resulted in significantly increased sLeX expression (p = 0.02 and p = 0.002 respectively). Exposure of transfected carcinoma cells to neutrophil conditioned media and neutrophils resulted in morphological changes consistent with increased invasive potential and a TNF-α dependent increased cellular migration through Matrigel. In patient lung carcinoma specimens but not in normal lung tissues, we observed areas of high sLeX expression that correlated with increased neutrophil infiltration in 80% of tumors. We conclude that FucT-III activity and sLeX expression in lung carcinomas may recruit neutrophils in the tumor microenvironment that enhance the metastatic potential of lung tumor cells. This work was supported in part by funds from the National Institutes of Health/National Cancer Institute grant K08 CA111829-04. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2339.

Chronic morphine treatment inhibits LPS-induced angiogenesis: Implications in wound healing

Delayed wound healing is a chronic problem in opioid drug abusers. We investigated the role chronic morphine plays on later stages of wound healing events using an angiogenesis model. Our results show that morphine treatment resulted in a significant decrease in inflammation induced angiogenesis. To delineate the mechanisms involved we investigate the role of hypoxia inducible factor 1 alpha (HIF-1 alpha), a potent inducer of angiogenic growth factor. Morphine treatment resulted in a significant decrease in the expression and nuclear translocation of HIF-1 alpha with a concurrent suppression in vascular endothelial growth factor (VEGF) synthesis. Cells of the innate immune system play a dominant role in the angiogenic process. Morphine treatment inhibited early recruitment of both neutrophils and monocytes towards an inflammatory signal with a significant decrease in the monocyte chemoattractant MCP-1. Taken together, our studies show that morphine regulates the wound repair process on multiple levels. Morphine acts both directly and indirectly in suppressing angiogenesis.

An adventitial IL-6/MCP1 amplification loop accelerates macrophage-mediated vascular inflammation leading to aortic dissection in mice

Vascular inflammation contributes to cardiovascular diseases such as aortic aneurysm and dissection. However, the precise inflammatory pathways involved have not been clearly defined. We have shown here that subcutaneous infusion of Ang II, a vasopressor known to promote vascular inflammation, into older C57BL/6J mice induced aortic production of the proinflammatory cytokine IL-6 and the monocyte chemoattractant MCP-1. Production of these factors occurred predominantly in the tunica adventitia, along with macrophage recruitment, adventitial expansion, and development of thoracic and suprarenal aortic dissections. In contrast, a reduced incidence of dissections was observed after Ang II infusion into mice lacking either IL-6 or the MCP-1 receptor CCR2. Further analysis revealed that Ang II induced CCR2+CD14hiCD11bhiF4/80- macrophage accumulation selectively in aortic dissections and not in aortas from Il6-/- mice. Adoptive transfer of Ccr2+/+ monocytes into Ccr2-/- mice resulted in selective monocyte uptake into the ascending and suprarenal aorta in regions of enhanced ROS stress, with restoration of IL-6 secretion and increased incidence of dissection. In vitro, coculture of monocytes and aortic adventitial fibroblasts produced MCP-1- and IL-6-enriched conditioned medium that promoted differentiation of monocytes into macrophages, induced CD14 and CD11b upregulation, and induced MCP-1 and MMP-9 expression. These results suggest that leukocyte-fibroblast interactions in the aortic adventitia potentiate IL-6 production, inducing local monocyte recruitment and activation, thereby promoting MCP-1 secretion, vascular inflammation, ECM remodeling, and aortic destabilization.

Macrophages express a TCRαβ-based variable immunoreceptor (134.34)

Abstract Immunological dogma holds that variable immunoreceptors are restricted to lymphocytes. However, recent studies demonstrate that a subpopuation of human neutrophils expresses the TCRαβ (PNAS 103:14441, 2006) and mouse thymic granulocytes exhibit rearrangement of their TCR loci (Nature 452:764, 2008). Here, we provide evidence that a subpopulation of circulating human monocytes, monocyte-derived macrophages and tissue macrophages constitutively express a TCRαβ-based variable immunoreceptor. TCRαβ + monocyte/macrophage subpopulations were assessed using with laser scanning cytometry and repertoire diversities were determined by CDR3 spectratyping. We find that human macrophages express individual-specific TCR Vαβ repertoires. Th1 or Th2 cytokine dependent in vitro differentiation triggers expression of distinct Vαβ repertoires in macrophages. Moreover, Th1 polarized macrophages show enhanced secretion of the major monocyte chemoattractant MCP-1 in response to costimulation with αCD3/CD28 . The presence of a variable immunoreceptor in macrophages together with our previous identification of the TCR in a subset of neutrophils challenge the traditional dichotomy of the vertebrate immune system into non-specific/myeloid and antigen-specific/lymphoid immunity. They unravel a much closer kinship between both lineages than commonly assumed. Supported by DFG grants KA 1078/4-1, BE 1768/5-1

Monocyte chemoattractant proteins in the pathogenesis of systemic sclerosis

Activation of the immune system and increased synthesis of extracellular matrix proteins by fibroblasts are hallmarks in the pathogenesis of SSc. The molecular mechanisms underlying the infiltration of inflammatory cells into the skin and the subsequent activation of fibroblasts are still largely unknown. Chemokines are a family of small molecules that are classified according to the position of the NH(2)-terminal cysteine motif. Recent data indicate that chemokines and in particular two members of the subfamily of monocyte chemoattractant proteins, MCP-1 (CCL-2) and MCP-3 (CCL-7), might be involved in the pathogenesis of SSc. MCP-1 and -3 are overexpressed by SSc fibroblasts and in skin lesions from SSc patients compared to healthy controls. MCP-1 and -3 are chemotactic for inflammatory cells and stimulate their migration into the skin. In addition to their pro-inflammatory effects, MCP-1 and -3 contribute to tissue fibrosis by activating the synthesis of extracellular matrix proteins in SSc fibroblasts. Therapeutic strategies targeting MCP-1 have revealed promising results in several animal models of SSc. Antagonists against the receptor CCR2 are currently tested in clinical trials of a variety of diseases and also represent interesting candidates for target-directed therapy in SSc.

Disruption of CXC Motif Chemokine Ligand-14 in Mice Ameliorates Obesity-induced Insulin Resistance

In obese individuals, white adipose tissue (WAT) is infiltrated by large numbers of macrophages, resulting in enhanced inflammatory responses that contribute to insulin resistance. Here we show that expression of the CXC motif chemokine ligand-14 (CXCL14), which targets tissue macrophages, is elevated in WAT of obese mice fed a high fat diet (HFD) compared with lean mice fed a regular diet. We found that HFD-fed CXCL14-deficient mice have impaired WAT macrophage mobilization and improved insulin responsiveness. Insulin-stimulated phosphorylation of Akt kinase in skeletal muscle was severely attenuated in HFD-fed CXCL14+/- mice but not in HFD-fed CXCL14-/- mice. The insulin-sensitive phenotype of CXCL14-/- mice after HFD feeding was prominent in female mice but not in male mice. HFD-fed CXCL14-/- mice were protected from hyperglycemia, hyperinsulinemia, and hypoadiponectinemia and did not exhibit increased levels of circulating retinol-binding protein-4 and increased expression of interleukin-6 in WAT. Transgenic overexpression of CXCL14 in skeletal muscle restored obesity-induced insulin resistance in CXCL14-/- mice. CXCL14 attenuated insulin-stimulated glucose uptake in cultured myocytes and to a lesser extent in cultured adipocytes. These results demonstrate that CXCL14 is a critical chemoattractant of WAT macrophages and a novel regulator of glucose metabolism that functions mainly in skeletal muscle.

Possible involvement of muscularis resident macrophages in impairment of interstitial cells of Cajal and myenteric nerve systems in rat models of TNBS-induced colitis

Resident macrophages are distributed in the network of interstitial cells of Cajal (ICC) and the myenteric nerve within the myenteric plexus. We evaluated changes in chemoattractant protein mRNA expression in macrophages and neutrophils, the ICC, nerve and macrophages in the myenteric plexus of model rats with TNBS-induced colitis. Chemoattractant proteins, MCP-1, GRO, MIP-2 and CINC-2alpha were upregulated in the colonic muscle layer after inflammation. Leukocyte infiltration and MPO activity were increased in the muscle layer. Electron microscopy indicated an irregular contour of the myenteric ganglia into which numerous macrophages had penetrated. Macrophages were also distributed near the ICC in the inflamed myenteric plexus. Immunohistochemistry showed that the ICC network and myenteric nerve system had disappeared from the inflamed region, whereas the number of resident macrophages was increased. TTX-insensitive, possibly ICC-mediated, rhythmic contractions of circular smooth muscle strips and enteric neuron-mediated TTX-sensitive peristalsis in the whole proximal colon tissue were significantly inhibited in the inflamed colon, indicating that the ICC-myenteric nerve system was dysfunctional in the inflamed muscle layer. Their accumulation around the myenteric nerve plexus and the ICC network suggests that macrophages play an important role in inducing intestinal dysmotility in gut inflammation.

Role of chemokines, neuronal projections, and the blood-brain barrier in the enhancement of cerebral EAE following focal brain damage.

The role of focal brain damage as a trigger for autoimmune inflammation in multiple sclerosis (MS) is unclear. In this study we examine mechanisms by which experimental autoimmune encephalomyelitis (EAE) is enhanced by focal brain damage. EAE was produced in Lewis rats by footpad inoculation; focal brain damage, in the form of a cortical cryolesion (cryolesion-EAE), was induced 8 days post-inoculation (d.p.i.). The distribution of inflammation and chemokine production in cryolesion-EAE and EAE-only were compared. Inflammation in the brain, measured by immunocytochemistry for T lymphocytes (W3/13) and microglial activation (MHC class II -OX6), was significantly enhanced in cryolesion-EAE 11-15 d.p.i. (p < 0.01-0.05) but by 20-40 d.p.i., equated with EAE-only. Inflammation in cryolesion-EAE related to breakdown of the blood-brain barrier (BBB) at the site of the cryolesion and also to the corticospinal tracts and thalamus, reflecting the afferent and efferent neuronal connections with the cryolesioned cortex. Semiquantitative RT/PCR dot-blot hybridization assay showed a 6-fold increase in mRNA for specific chemokines in the brain in cryolesion-EAE at 9 d.p.i. (MCP-1) and 11 d.p.i. (MCP-1 and MCP-5) with no significant increase in RANTES, GRO-alpha, or MIP-1alpha. By 14 d.p.i., the levels of MCP-1 and MCP-5 mRNA equated with EAE-only animals. These results suggest that enhancement and location of autoimmune inflammation in the brain following focal cortical injury initially involve chemokines such as the macrophage chemoattractants MCP-1 and MCP-5, and the activities of afferent and efferent neuronal connections with the site of damage. By analogy, similar factors may modulate or reactivate autoimmune inflammation in MS.

The human astrocytoma cell line U373MG produces monocyte chemotactic protein (MCP)-1 upon stimulation with β-amyloid protein

Astrocytes associated with β-amyloid (Aβ) accumulate in senile plaques of Alzheimer's disease (AD). To investigate the biological effects of Aβ/astrocyte interaction, we examined chemokine production by the human astrocytoma cell line U373MG stimulated with Aβ peptides. Northern blot analysis and specific immunoassays demonstrate that Aβ [1–42] and Aβ [25–35] induce mRNA expression and release of monocyte chemotactic protein (MCP)-1 but not of γ-interferon inducible protein (IP)-10 by U373MG cells. The observation that Aβ induces astrocyte production of the potent microglia chemoattractant MCP-1 contributes to understanding mechanism of damage exerted by Aβ in AD senile plaques.