Prevention of myocardial reperfusion injury in rats by an antibody against monocyte chemotactic and activating factor/monocyte chemoattractant protein-1

MCAF (monocyte chemotactic and activating factor)/MCP-1 (monocyte chemoattractant protein-1) is an important mediator of monocyte recruitment to inflammatory sites. However, its pathophysiologic role in myocardial reperfusion injury remains unknown. Male Wistar rats were anesthetized, and the left anterior descending coronary artery was ligated for an hour, after which the ligature was released. Northern blotting analysis revealed that MCAF/MCP-1 mRNA expression increased 16-fold in the reperfused region at 12 hours after reperfusion. MCAF/MCP-1 concentration in plasma and the heart was already elevated after hour of ischemia in this model. Goat polyclonal antibodies were prepared by repeated immunization of animals with purified, recombinant rat MCAF/MCP-1, and the neutralizing activities of this antibody were confirmed by monocyte chemotaxis assay and administration to rats with crescentic glomerulonephritis. Intravenous injection of anti-MCAF/MCP-1 antibody significantly reduced the infarct size at 24 hours after reperfusion compared with the injection of control IgG (33.9 +/- 5.1% vs 49.4 +/- 2.7% of ischemic area, mean +/- SEM). Administration of this antibody markedly decreased the intercellular adhesion molecule-1 mRNA expression and infiltration of macrophages, which suggested the pathophysiologic role of MCAF/MCP-1. Neutralization of MCAF/MCP-1 is beneficial by preventing reperfusion injury in a rat model of myocardial ischemia and reperfusion.     

Increased expression of monocyte chemotactic protein-1 during active hepatic fibrogenesis: correlation with monocyte infiltration.

Monocyte chemotactic protein (MCP)-1 is a chemoattractant and activator for circulating monocytes and T lymphocytes. We investigated MCP-1 protein and gene expression during chronic liver disease at different stages, using immunohistochemistry and in situ hybridization, respectively. In normal liver, a modest expression of MCP-1 was confined to few peri-sinusoidal cells and to bile duct epithelial cells. During chronic hepatitis, MCP-1 immunostaining and gene expression were evident in the inflammatory infiltrate of the portal tract. In tissue from patients with active cirrhosis, MCP-1 expression was clearly up-regulated and was present in the portal tract, in the epithelial cells of regenerating bile ducts, and in the active septa surrounding regenerating nodules. A combination of in situ hybridization for MCP-1 and immunohistochemistry showed that activated stellate cells and monocyte/macrophages contribute to MCP-1 expression in vivo together with bile duct epithelial cells. Comparison of serial sections of liver biopsies from patients with various degrees of necro-inflammatory activity showed that infiltration of the portal tracts with monocytes/macrophages is directly correlated with the expression of MCP-1. These data expand previous in vitro studies showing that secretion of MCP-1 may contribute to the formation and maintenance of the inflammatory infiltrate observed during chronic liver disease.     

Expression of monocyte chemoattractant protein 1 in human inflamed gingival tissues.

Gingival inflammation is initiated by bacterial colonization on the tooth surface. It is characterized by infiltration of mononuclear cells, a common feature of many forms of chronic inflammation. Monocyte chemoattractant protein 1 (MCP-1) is the predominant monocyte chemoattractant secreted by a variety of different cells in vitro. For this report, we examined MCP-1 expression in bacterially induced gingival inflammation by immunohistochemistry and in situ hybridization. The cell types expressing MCP-1 are identified as vascular endothelial cells and monocytes/macrophages. Correlation analysis shows that the number of cells expressing MCP-1 is related to the degree of inflammation. Our finding that MCP-1 is expressed in inflamed gingival tissue suggests that MCP-1 plays an important role in the recruitment of monocytes and amplification of inflammatory signals in bacterially induced inflammation.     

Evidence for increased expression of eotaxin and monocyte chemotactic protein-4 in atopic dermatitis

BACKGROUND: Atopic dermatitis (AD) is a chronic inflammatory skin disease associated with tissue eosinophilia and the activation of T lymphocytes. The novel eosinophil chemoattractants, eotaxin and monocyte chemotactic protein (MCP)-4, are up-regulated at sites of allergic inflammation, yet their contribution to the pathophysiologic mechanisms of AD remains to be determined. OBJECTIVE: We sought to investigate the expression of eotaxin and MCP-4 in acute and chronic lesions from patients with AD and to determine their relationship to the numbers of resident inflammatory cells. METHODS: With use of in situ hybridization, the expression of eotaxin and MCP-4 messenger RNA (mRNA) in skin biopsy specimens from patients with acute and chronic AD skin lesions was compared with that of uninvolved skin from these patients and skin from healthy volunteers. RESULTS: There was a constitutive expression of eotaxin and MCP-4 mRNA in skin biopsy specimens from healthy subjects. Positive signal for chemokine mRNA was observed both within the epidermis and inflammatory cells (macrophages, eosinophils, and T cells) of the subepidermis in AD skin lesions. Within the subepithelium acute and chronic skin lesions exhibited a significant increase in the numbers of eotaxin and MCP-4 mRNA-positive cells compared with uninvolved skin (P <.01), whereas the numbers of eotaxin and MCP-4 mRNA-positive cells were significantly higher in chronic AD compared with acute AD skin lesions (P <.005, P <.001, respectively). Correlations were observed between the expression of eotaxin and MCP-4 mRNA and the presence of eosinophils and macrophages, respectively, in AD lesions (r(2) = 0.84, r(2) = 0.94). CONCLUSION: There is an increased expression of eotaxin and MCP-4 in acute and chronic lesions, suggesting that these chemotactic factors play a major role in the pathophysiologic mechanisms of AD.     

Spindle cell ductal carcinoma in situ. An unusual variant of ductal intra-epithelial neoplasia that simulates ductal hyperplasia or a myoepithelial proliferation

It is well known that the number of peritoneal macrophages is increased in patients with pelvic endometriosis. We measured the concentration of monocyte chemoattractant protein-1 (MCP-1) using an enzyme-linked immunosorbent assay (ELISA) in the peritoneal fluid of women with and without endometriosis. The expression of MCP-1 in pelvic endometriotic lesions obtained from the peritoneum was also examined using immunohistochemistry and nonradioactive in situ hybridization. The mean concentration of MCP-1 in the peritoneal fluid was significantly higher in the patients with endometriosis (P<0.05). The most significant elevation, compared with non-endometriosis patients, was found in stage I of the disease (P<0.05). However, no statistically significant difference was found among endometriosis stages I, II, III, and IV. Immunohistochemical staining revealed that MCP-1-positive cells were localized in the glandular epithelium of the endometriotic lesions and in the stromal macrophages distributed in those lesions, but normal peritoneal cells were negative. The in situ hybridization method demonstrated expression of MCP-1 mRNA on the endometriotic glandular epithelium and stromal macrophages. These findings suggest that MCP-1 may be involved in the histogenesis and early development of peritoneal endometriosis.     

Modulation of JE/MCP-1 expression in dermal wound repair.

The tissue macrophage plays a prominent role in wound repair, yet the parameters that influence macrophage migration into the wound bed are not well understood. To better understand the process of macrophage recruitment, the production of JE, the murine homologue of monocyte chemoattractant protein 1(JE/MCP-1), was examined in a murine model of dermal wound repair. High levels of JE/MCP-1 mRNA were found in dermal punch wounds at 12 hours and 1 day (24 hours) after wounding; mRNA levels slowly decreased to undetectable by day 21. In situ hybridization analysis of wounds revealed that JE/MCP-1 was predominantly expressed by monocytic and macrophage-like cells, as well as by occasional fibroblasts and other interstitial cells. To correlate JE/MCP-1 production with macrophage migration, macrophage infiltration into the wound bed was quantitated. The number of macrophages within the wound increased to a maximum at day 3 (11.3 +/- 4.5 macrophages per high power field), began to decrease at day 5 (4.8 +/- 1.9 macrophages per high power field), and reached near base line at day 10 (3.0 +/- 1.1 macrophages per high power field). The results demonstrate that JE/MCP-1 production within wounds is closely linked to the time course and distribution of macrophage infiltration, with maximal JE/MCP-1 mRNA levels occurring 1 to 2 days before maximal macrophage infiltration. The results support a role for JE/MCP-1 in the recruitment of wound macrophages and suggest that macrophages, through the production of JE/MCP-1, may sustain the recruitment of additional monocytes and macrophages into sites of injury.     

Increased expression of monocyte chemotactic protein-1 in the endometrium of women with endometriosis.

The pathogenesis of endometriosis, a disease widely believed to arise from an aberrant growth of endometrial tissue outside the uterus, is still unclear. We have previously observed that cytokine-stimulated endometrial cells of women with endometriosis secrete in vitro increased amounts of monocyte chemotactic protein-1 (MCP-1). This factor may be important in the recruitment and activation of peritoneal macrophages observed in endometriosis patients. The present study reports that, in the presence of the disease, such an up-regulation of MCP-1 expression arises in vivo and can be encountered in situ in the intrauterine endometrium. In women with endometriosis, MCP-1 expression was elevated in endometrial glands, both at the level of the protein (immunohistochemistry) and the mRNA (in situ hybridization). This was observed throughout the menstrual cycle and varied according to the stage of the disease. These findings strongly argue in favor of the presence of pathophysiological changes in the eutopic endometrium of patients with endometriosis and make plausible MCP-1 as a key effector cell mediator involved in the pathogenesis of the disease.     

Effects of monocyte chemoattractant protein 1 on blood-borne cell recruitment after transient focal cerebral ischemia in mice

Monocyte chemoattractant protein 1 (MCP-1) plays an important role in inflammatory reactions following cerebral ischemia. It is known that MCP-1 overexpression leads to increased infarct volume and elevated hematogenous cell recruitment, while MCP-1-deficient mice develop smaller infarcts. It was supposed that MCP-1 dependent macrophage recruitment might be the underlying mechanism of ischemic brain damage but a precise distinction of local microglia and invading macrophages was not performed. In this study we investigated the differential role of MCP-1 on inflammatory cells in MCP-1-deficient mice, using green fluorescent protein (GFP) transgenic bone marrow chimeras. After 30-min of focal cerebral ischemia microglia was rapidly activated and was not different between MCP-1-deficient mice and wild type controls. Activated microglia outnumbered GFP-positive macrophages over the study period. Furthermore, macrophage infiltration was significantly reduced at day 7 in MCP-1-deficient animals (31.2±20.1 cells/mm²) compared to MCP-1 wild type mice (131.5±66.7 cells/mm², P<0.001). Neutrophils were also significantly reduced in MCP-1-deficient mice (62% on day 4% and 87% on day 7; P<0.001). This is the first investigation in cerebral ischemia showing that MCP-1 is necessary for recruiting blood-borne cells to the injury site whereas it does not affect the microglia activation and migration. However, the remarkable predominance of activated microglia and the additional attenuation of invading macrophages suggest that different mechanisms than macrophage recruitment are responsible for the MCP-1-mediated neuroprotective effects after experimental stroke.     

复方丹参对脑缺血再灌注后大鼠海马和齿状回神经细胞凋亡及Bcl-2 mRNA表达的影响

目的:探讨中药复方丹参对大鼠脑缺血再灌注后海马和齿状回神经细胞凋亡及Bcl-2 mRNA表达的影响。方法:采用大脑中动脉内栓线法建立大鼠大脑中动脉缺血再灌注模型,应用原位细胞凋亡检测和原位杂交技术检测大鼠海马和齿状回神经细胞凋亡和Bcl-2 mRNA的表达并做图像分析。结果:与假手术对照组比较,缺血再灌注组凋亡神经细胞主要位于缺血侧海马CA1、CA3区,齿状回凋亡细胞较少。3个区神经细胞Bcl-2mRNA的表达在缺血再灌注2 h后升高,随时间的延长逐渐增强。复方丹参组神经细胞Bcl-2 mRNA的表达明显强于缺血再灌组,而凋亡神经细胞数明显较低。结论:复方丹参可通过上调神经细胞Bcl-2 mRNA的表达,抑制神经细胞凋亡,从而减轻缺血再灌注对大鼠海马和齿状回的损伤。     

Expression of interleukin-6 and monocyte chemoattractant protein-1 by peritoneal sub-mesothelial cells during abdominal operations

Cytokines and chemokines including interleukin-6 (IL-6) and monocyte chemoattractant protein-1 (MCP-1) are secreted in response to major abdominal operations. The aim of this study was to identify the peritoneal cells that produce IL-6 and MCP-1. Samples of peritoneal tissue were taken from patients at the beginning and end of major abdominal operations. The samples were incubated in culture medium on microtitre plates for 5 h. The concentrations of IL-6 and MCP-1 were measured in culture supernatants by enzyme-linked immunosorbent assay (ELISA). In paraffin sections, cells that expressed IL-6 or MCP-1 were identified by combined in situ hybridization and immunohistochemistry. Antibodies against CD68, CD34, actin, and calretinin were included in these experiments. The median production of IL-6 increased significantly from 6256 pg/ml at the start of the operation to 20,000 pg/ml at the end. Production of MCP-1 rose from 7700 pg/ml to 11,820 pg/ml. IL-6 mRNA was mainly confined to endothelial cells. MCP-1 was expressed by a broader range of cells, consisting of actin-positive smooth muscle cells and endothelial cells, fibroblast-like cells, as well as occasional macrophages and mesothelial cells. Peritoneal endothelial cells contribute to the transient increase in concentrations of IL-6 in the circulation after surgical trauma. Recruitment of monocytes to the site of the trauma seems to be mainly effected by actin-positive smooth muscle cells and endothelial cells.     

七氟烷对心肌缺血再灌注内皮细胞细胞间黏附分子-1、血管细胞黏附分子-1和E-选择素表达的影响

目的： 探讨七氟烷对心肌缺血再灌注内皮细胞促炎作用的细胞间黏附分子-1（ ICAM-1）、血管细胞黏附 分子-1（ VCAM-1）和E- 选择素表达的影响。方法 ：在大鼠心肌缺血再灌注模型的基础上,将大鼠随机分为假 手术组（ 对照组）、缺血再灌注损伤组（ I/R 组）和七氟烷组;观察各组大鼠手术前、缺血15 min 和再灌注4 h 的 心率、平均动脉压和心率-收缩压乘积（ RPP ）;免疫组织化学法检测心肌组织中CD68+ 巨噬细胞数目、内皮细胞 ICAM-1、VCAM-1 和E- 选择素的表达;TUNEL 染色法检测凋亡细胞的比例。结果：缺血15 min 时,I/R 组和 七氟烷组平均动脉压和RPP 均显著下降;再灌注4 h 时,七氟烷组平均动脉压和RPP 均有所上升,相对于I/R 组, 差异具有统计学意义;与对照组比较,I/R 组内皮细胞ICAM-1、VCAM-1 和E- 选择素的表达均显著升高,七氟 烷则能够有效抑制I/R 引起的内皮细胞促炎分子的表达;对照组CD68+ 巨噬细胞为5.83 个/ 高倍镜视野（ HPF）, I/R 组数目为55.67 个/HPF,两组间差异具有统计学意义;七氟烷能够显著减少心组织内巨噬细胞的浸润,与I/R 组比较,降低了66.46%;TUNEL 染色结果显示对照组心肌细胞凋亡率2.20%,I/R 组为28.63%,两组间差异明显; 七氟烷能够显著降低心肌细胞的凋亡,相对于I/R 组,降低了51.76%。结论：七氟烷可降低缺血再灌注损伤后内 皮细胞表面促炎分子的表达,减少心肌组织巨噬细胞浸润和细胞凋亡。     

Enhanced production of monocyte chemoattractant protein-1 in the dorsal root ganglia in a rat model of neuropathic pain: possible involvement in the development of neuropathic pain

Chemokines are a family of peptides originally identified as the factors regulating the migration of leukocytes in inflammatory and immune responses. Recently, they have been shown to be produced in the central and peripheral nervous systems under various pathological conditions and act on neuronal and glial cells. In this study, we examined the production of monocyte chemoattractant protein-1 (MCP-1), a well-characterized chemokine, in dorsal root ganglia (DRG) in a rat model of neuropathic pain. Partial ligation of the sciatic nerve induced mechanical allodynia in the ipsilateral hindpaw with weaker allodynia in the contralateral one. Immunohistochemical analyses revealed that the number of MCP-1 immunoreactivity (ir)-positive cells was increased in the ipsilateral DRG. The increase started by 4h after the ligation, peaked at 24h and continued to at least 48 h. The weaker but significant increase was observed in the contralateral DRG. Double immunofluorescent staining demonstrated that almost all of the MCP-1ir-positive cells were neuronal cells. In situ hybridization histochemistry showed that MCP-1 mRNA expression was markedly upregulated in the ipsilateral DRG with weaker increase in the contralateral one at 24 h after the ligation, indicating that the elevation in MCP-1ir detected by immunohistochemistry was due to an upregulation of MCP-1 production by the DRG neurons themselves. Furthermore, intrathecal administration of MCP-1 induced mechanical allodynia. These results suggest that MCP-1 produced in the DRG neurons is involved in the development of mechanical allodynia induced by nerve injury.     

Inducible and endothelial nitric oxide synthase expression during development of transplant arteriosclerosis in rat aortic grafts.

In the vascular system, distinct isoforms of nitric oxide synthase (NOS) generate nitric oxide (NO), which acts as a biological messenger. Its role in the development of transplant arteriosclerosis (TA) is still unclear. To investigate whether NO is involved in TA, we studied the expression of NOS isoforms, inducible NOS (iNOS) and endothelial NOS (eNOS), by immunohistochemistry and in situ hybridization during the first two post-transplantation months and their relation with cold ischemia (1 to 24 hours) and reperfusion injury using an aortic transplantation model in the rat. We found an increased iNOS expression in the intima and adventitia and a decreased expression in the media, whereas eNOS expression was not significantly altered during the development of TA. Co-localization studies suggested that iNOS-positive cells were vascular smooth muscle cells, monocyte-derived macrophages, and endothelial cells. Prolonged ischemic storage time resulted in an increase in eNOS expression in the neointima. In situ hybridization showed iNOS mRNA expression by vascular cells in the neointima and media. NO produced by iNOS and eNOS may be involved, at least in part, in the pathogenesis of TA in aortic grafts. Additional studies are needed to confirm the modulatory mechanism of NO during the development of TA.     

Pioglitazone, a peroxisome proliferator-activated receptor-gamma agonist, attenuates myocardial ischemia/reperfusion injury in a rat model

Thiazolidinediones are insulin-sensitizing drugs, ligands for peroxisome proliferator-activated receptor-gamma (PPAR-gamma), which play an important role in the modulation of inflammatory responses. Myocardial ischemia/reperfusion (MI/R) injury is associated with inflammation, in which various cells, particularly monocytes and macrophages, are involved. This study examined the effects of the thiazolidinedione peroxisome proliferator-activated receptor-gamma ligand, pioglitazone, in a rat model of MI/R injury. Pioglitazone at 3 mg/kg/day or the vehicle was administered for 7 days before rats were subjected to 30 minutes of coronary ligation followed by 24 hours of reperfusion. The mRNA expression [monocyte chemoattractant protein-1 (MCP-1) and intercellular adhesion molecule-1] in the ischemic region, the number of infiltrating macrophages in the ischemic region, and the myocardial infarct size were examined. The inhibitory effects of pioglitazone on activated macrophages were studied in vitro. Phorbol 12-myristate 13-acetate-induced MCP-1 production, in the absence or presence of pioglitazone, were assayed in cultured macrophages. Compared with the control group, (1). mRNA levels of MCP-1 and intercellular adhesion molecule-1 and the number of infiltrating macrophages in the ischemic region were significantly lower in the pioglitazone-treated group; and (2). myocardial infarct size was significantly smaller in the pioglitazone-treated group. Phorbol 12-myristate 13-acetate-stimulated cultured macrophages in the presence of pioglitazone produced significantly lower levels of MCP-1 than the stimulated control in the absence of pioglitazone. These observations demonstrate that pioglitazone has anti-inflammatory effects in MI/R injury that are independent of its insulin-sensitizing effect.     

Expression of monocyte chemotactic protein-1 in human melanoma in vivo.

A common feature of human melanoma is infiltration by monocytes at early stages of tumorigenesis. This infiltration may be highly significant since macrophages have the capacity to alter the behavior of tumor cells. The authors previously demonstrated that the predominant monocyte chemoattractant produced by tumor cells in vitro was monocyte chemotactic protein-1 (MCP-1). The authors identify the expression of MCP-1 in pathologic specimens of both primary and metastatic human melanoma but not in normal skin. The finding that MCP-1 is produced by malignant melanoma suggests that specific genes are expressed in tumor cells that can induce the recruitment of monocytes in vivo.     

Neutralization of macrophage inflammatory protein 2 (MIP-2) and MIP-1alpha attenuates neutrophil recruitment in the central nervous system during experimental bacterial meningitis

Chemokines are low-molecular-weight chemotactic cytokines that have been shown to play a central role in the perivascular transmigration and accumulation of specific subsets of leukocytes at sites of tissue damage. Using in situ hybridization (ISH), we investigated the mRNA induction of macrophage inflammatory protein 2 (MIP-2), MIP-1alpha, monocyte chemoattractant protein 1 (MCP-1), and RANTES. Challenge of infant rats' brains with Haemophilus influenzae type b intraperitoneally resulted in the time-dependent expression of MIP-2, MIP-1alpha, MCP-1, and RANTES, which was maximal 24 to 48 h postinoculation. Immunohistochemistry showed significant increases in neutrophils and macrophages infiltrating the meninges, the ventricular system, and the periventricular area. The kinetics of MIP-2, MIP-1alpha, MCP-1, and RANTES mRNA expression paralleled those of the recruitment of inflammatory cells and disease severity. Administration of anti-MIP-2 or anti-MIP-1alpha antibodies (Abs) resulted in significant reduction of neutrophils. Administration of anti-MCP-1 Abs significantly decreased macrophage infiltration. Combined studies of ISH and immunohistochemistry showed that MIP-2- and MIP-1alpha-positive cells were neutrophils and macrophages. MCP-1-positive cells were neutrophils, macrophages, and astrocytes. Expression of RANTES was localized predominantly to resident astrocytes and microglia. The present study indicates that blocking of MIP-2 or MIP-1alpha bioactivity in vivo results in decreased neutrophil influx. These data are also the first demonstration that the C-C chemokine MIP-1alpha is involved in neutrophil recruitment in vivo.     

Role of oxygen in postischemic myocardial injury

Myocardial function is dependent on a constant supply of oxygen from the coronary circulation. A reduction of oxygen supply due to coronary obstruction results in myocardial ischemia, which leads to cardiac dysfunction. Reperfusion of the ischemic myocardium is required for tissue survival. Thrombolytic therapy, coronary artery bypass surgery and coronary angioplasty are some of the treatments available for the restoration of blood flow to the ischemic myocardium. However, the restoration of blood flow may also lead to reperfusion injury, resulting in myocyte death. Thus, any imbalance between oxygen supply and metabolic demand leads to functional, metabolic, morphologic, and electrophysiologic alterations, causing cell death. Myocardial ischemia reperfusion (IR) injury is a multifactorial process that is mediated by oxygen free radicals, neutrophil activation and infiltration, calcium overload, and apoptosis. Controlled reperfusion of the ischemic myocardium has been advocated to prevent the IR injury. Studies have shown that reperfusion injury and postischemic cardiac function are related to the quantity and delivery of oxygen during reperfusion. Substantial evidence suggests that controlled reoxygenation may ameliorate postischemic organ dysfunction. In this review, we discuss the role of oxygenation during reperfusion and subsequent biochemical and pathologic alterations in reperfused myocardium and recovery of heart function.     

Potential role for monocyte chemotactic protein-4 (MCP-4) in monocyte/macrophage recruitment in acute renal inflammation

The CC chemokine, monocyte chemoattractant protein-4 (MCP-4), is an important chemoattractant for monocytes and T cells. Recent data indicate a role in renal inflammation. This study has used in situ hybridization and immunohistochemical analysis of cryostat sections of biopsy material taken from patients with acute renal allograft rejection and vasculitic glomerulonephritis to demonstrate renal expression of MCP-4, both at message and protein level. MCP-4 was primarily expressed at peritubular, periglomerular, and perivascular sites, irrespective of the inflammatory condition, and was associated with infiltrating CD3-positive lymphocytes and CD68-positive monocyte/macrophages. In addition, proximal tubular epithelial cells grown in culture from cortical fragments of human kidney showed low levels of constitutive MCP-4 expression, detectable by western blotting; this expression of MCP-4 was up-regulated in response to the pro-inflammatory cytokines, tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma). CCR3-, CCR5- and CCR2-expressing leukocyte populations were identified at sites of MCP-4 expression. Double-staining techniques revealed that CC chemokine receptor-expressing cells were primarily CD68-positive. These studies suggest an important role for MCP-4 in the recruitment and retention of monocytes/macrophages in renal inflammation.     

IL-9 and c-Kit+ mast cells in allergic rhinitis during seasonal allergen exposure: effect of immunotherapy

Background IL-9 is an important stimulus for tissue infiltration by mast cells, a feature requiring concomitant activation of c-Kit. Objectives We assessed IL-9 expression and c-Kit + mast cells in the nasal mucosa of patients with allergic rhinitis during seasonal pollen exposure and observed the effects of allergen immunotherapy. Methods We studied 44 patients with seasonal rhinitis and asthma before and 2 years after a double-blind trial of grass pollen immunotherapy. Nasal mucosal IL-9 + cells and c-Kit + mast cells were assessed by means of immunochemistry. Cell types expressing IL-9 protein were determined by means of dual immunofluorescence. IL-9 mRNA-positive cells were assessed by means of in situ hybridization, and their phenotype was determined by using sequential immunohistochemistry and in situ hybridization. Results Nasal mucosal c-Kit + mast cells were increased during the pollen season ( P = .0001). IL-9 mRNA-positive cells also tended to increase ( P = .1) and correlated with nasal EG2 + eosinophils ( r = 0.47, P = .05) and IL-5 mRNA-positive cells ( r = 0.54, P = .02). The cell sources of IL-9 included T cells, eosinophils, neutrophils, and mast cells. When compared with placebo, successful pollen immunotherapy markedly inhibited seasonal increases in nasal mucosal c-Kit + mast cells ( P = .001) and the seasonal expression of IL-9 mRNA-positive cells ( P = .06). Immunotherapy also inhibited IL-9 protein expression from nonendothelial cell sources ( P = .0007). Conclusion IL-9 is upregulated in the nasal mucosa during the pollen season and correlates with tissue infiltration by eosinophils. Successful pollen immunotherapy is associated with inhibition of seasonal increases in both nasal c-Kit + mast cells and eosinophils. This effect might be explained, at least in part, by the reduced local expression of IL-9.     

Toll-like receptor 2 deficiency leads to delayed exacerbation of ischemic injury

ABSTRACT: BACKGROUND: : Using live imaging approach we have previously shown that microglia activation after stroke is characterized by marked and long-term induction of the Toll like Receptor 2 (TLR2) biophotonic signals. However, the role of TLR2 (and potentially other TLRs), beyond the acute innate immune response and an early neuroprotection against ischemic injury is not well understood. METHOD: S: The TLR2 -/- mice were subjected to transient middle cerebral artery occlusion (MCAO) followed by different reperfusion times. Analyses assessing microglial activation profile/innate immune response were performed using in situ hybridization, immunohistochemistry analysis, flow cytometry and inflammatory cytokine array. The effects of the TLR2 deficiency on the evolution of ischemic brain injury were analyzed using a cresyl violet staining of brain sections with appropriate lesion size estimation. RESULTS: : Here we report that TLR2 deficiency markedly affects post-stroke immune response resulting in delayed exacerbation of the ischemic injury. The temporal analysis of the microglia/macrophage activation profiles in TLR2 -/- mice and age-matched controls revealed reduced microglia/macrophage activation after stroke, reduced capacity of resident microglia to proliferate as well as decreased levels of MCP-1 and consequently lower levels of CD45high/CD11b+ expressing cells as shown by flow cytometry analysis. Importantly, although acute ischemic lesions (24-72hrs) were smaller in TLR2 -/- mice, the observed alterations in innate immune response were more pronounces at later time-points (at day 7) after initial stroke, which finally resulted in delayed exacerbation of ischemic lesion leading to larger chronic infarctions as compared to WT mice. Moreover, our results revealed that TLR2 deficiency is associated with significant decrease in the levels of neurotrophic/antiapoptotic factor IGF-1, expressed by microglia in the areas in- and around ischemic lesion. CONCLUSION: Altogether our results clearly suggest that optimal and timely microglial activation/innate immune response is needed to limit neuronal damage after stroke.