Influence of chylomicron remnants on human monocyte activation in vitro

Background and aims

Atherosclerosis is known to be an inflammatory disease and there is increasing evidence that chylomicron remnants (CMR), the lipoproteins which carry dietary fats in the blood, cause macrophage foam cell formation and inflammation. In early atherosclerosis the frequency of activated monocytes in the peripheral circulation is increased, and clearance of CMR from blood may be delayed, however, whether CMR contribute directly to monocyte activation and subsequent egress into the arterial wall has not been established. Here, the contribution of CMR to activation of monocyte pro-inflammatory pathways was assessed using an in vitro model.

Methods and results

Primary human monocytes and CMR-like particles (CRLP) were used to measure several endpoints of monocyte activation. Treatment with CRLP caused rapid and prolonged generation of reactive oxygen species by monocytes. The pro-inflammatory chemokines MCP-1 and IL-8 were secreted in nanogram quantities by the cells in the absence of CRLP. IL-8 secretion was transiently increased after CRLP treatment, and CRLP maintained secretion in the presence of pharmacological inhibitors of IL-8 production. In contrast, exposure to CRLP significantly reduced MCP-1 secretion. Chemotaxis towards MCP-1 was increased in monocytes pre-exposed to CRLP and was reversed by addition of exogenous MCP-1.

Conclusion

Our findings indicate that CRLP activate human monocytes and augment their migration in vitro by reducing cellular MCP-1 expression. Our data support the current hypothesis that CMR contribute to the inflammatory milieu of the arterial wall in early atherosclerosis, and suggest that this may reflect direct interaction with circulating blood monocytes.     

Monocyte activation in patients with Wegener's granulomatosis

OBJECTIVE: Wegener's granulomatosis (WG) is an inflammatory disorder characterised by granulomatous inflammation, vasculitis, and necrotising vasculitis and is strongly associated with anti-neutrophil cytoplasmic antibodies (ANCA). Activated monocytes/macrophages are present in renal biopsy specimens and participate in granuloma formation by synthesising and secreting a variety of chemoattractants, growth factors, and cytokines. In view of these findings, in vivo monocyte activation was evaluated in patients with WG and the findings related to parameters of clinical disease activity. METHODS: Monocyte activation was analysed by measuring plasma concentrations of soluble products of monocyte activation, that is neopterin and interleukin 6 (IL6), by ELISA, and by quantitating the surface expression of activation markers on circulating monocytes by flow cytometry. RESULTS: Twenty-four patients with active WG were included in this study. Ten of these patients were also analysed at the time of remission. Twelve patients with sepsis served as positive controls, and 10 healthy volunteers as negative controls for monocyte activation. Patients with active disease had increased monocyte activation compared with healthy controls as shown by increased concentrations of neopterin (p < 0.0001) and increased surface expression of CD11b (p < 0.05) and CD64 (p < 0.05). In those patients with increased concentrations of IL6 during active disease plasma concentrations of IL6 decreased during follow up when patients went into remission (p < 0.0001). In addition, neopterin (r = 0.37, r = 0.44), IL6 (r = 0.37, r = 0.60) and CD63 expression (r = 0.39, r = 0.45) correlated significantly with disease activity as measured by the Birmingham Vasculitis Activity Score and C reactive protein values, respectively. Compared with patients with sepsis, all markers of monocyte activation in patients with vasculitis were lower. CONCLUSION: It is concluded that disease activity in WG correlates with the extent of activation of monocytes, compatible with their role in the pathophysiology of this disease.     

丹参多酚酸盐对急性心肌梗死PCI术后CD14~+单核细胞活性氧和Toll样受体的影响

目的:观察丹参多酚酸盐对急性心肌梗死经皮冠状动脉介入术(PCI)术后,外周血单核细胞脂质过氧化损伤、单核细胞趋化因子-1及Toll样受体的影响,以进一步探讨丹参多酚在拮抗急性心肌梗死单核细胞氧化损伤中的作用。方法:选取我院2011年2月至2012年8月,因急性ST段抬高性心肌梗死入院患者320例,所有患者均行急诊PCI治疗。将320例患者随机分为常规治疗组(155例)和丹参多酚治疗组(165例)。常规治疗组予以阿司匹林、氯吡格雷及阿托伐他汀等药物口服。而丹参多酚治疗组除上述治疗外,予以丹参多酚注射液400mg静点,此后每日1次,持续7d。入院时及治疗7d末取静脉血6 mL分离单核细胞,通过流式细胞仪、ELISA等方法检测单核细胞活性氧ROS、血浆单核细胞趋化因子-1(MCP-1)以及单核细胞Toll受体4(TLR4)表达的变化。另取30例自愿者为对照组。结果:治疗7d末,两组患者单核细胞ROS、血浆MCP-1及TLR4水平均较入院前明显降低(P<0.01),而与常规治疗组比较,治疗7d末丹参多酚治疗组单核细胞ROS、血浆MCP-1及TLR4水平显著降低(P<0.05)。结论:注射用丹参多酚能通过抑制CD14+单核细胞TLR4信号转导途径,抑制单核细胞过氧化损伤,降低急性心肌梗死PCI术后患者单核细胞引起的炎性损伤。     

Increased subpopulations of CD16(+) and CD56(+) blood monocytes in patients with active Crohn's disease

BACKGROUND: Circulating monocytes may be subdivided according to the presence or absence of the Fcgamma receptor CD16 and the neural cell adhesion molecule CD56. Monocytes classified into these subpopulations are characterized by distinct phenotypic and functional features. We hypothesized that patients with active Crohn's disease differ in their peripheral monocyte subpopulations. METHODS: Using flow cytometry we investigated the expression of CD16 and CD56 on circulating monocytes in 11 patients with active Crohn's disease and 11 controls. These monocyte subpopulations were then analyzed for expression of the chemokine receptor fractalkine, CX(3)CR1, and the monocyte chemoattractant protein-1, CCR2. RESULTS: We found a median 3.7-fold increase in the number of CD16(+) monocytes related to the population with high expression of the pattern recognition receptor CD14 compared to that in the controls (P < 0.001). By studying the percentage of monocytes expressing CX(3)CR1, and their relative fluorescence intensity (RFI), we found significant differences, with both the highest percentage and the highest RFI in the CD14(low)CD16(+) subpopulation, whereas the CD14(high)CD16(+) subgroup represented an intermediate population. Inversely, CCR2 expression was highest in the populations with high expression of CD14, whereas the CD14(low)CD16(+) subpopulation showed the lowest percentage and the lowest RFI for CCR2. We found the percentage of CD14(+)CD56(+) monocytes in patients with active Crohn's disease to be increased 2.7 times compared to the controls (P = 0.011). CONCLUSIONS: These results show that subsets of peripheral monocytes with a more mature phenotype are expanded in patients with active Crohn's disease.     

单核细胞株THP-1分化为泡沫细胞过程中清道夫受体A变化及阿托伐他汀的干预作用

目的 研究单核细胞株THP-1分化为巨噬细胞、泡沫细胞过程中细胞清道夫受体A（SRA）表达、单核细胞趋化蛋白-1（MCP-1）分泌及应用阿托伐他汀干预的情况。方法 佛波醇酯诱导THP-1细胞分化为巨噬细胞,将其分为对照组、OX-LDL组（泡沫细胞组）、OX-LDL＋阿托伐他汀组（再分为低、中、高浓度3组）。用ELISA法,测定细胞培养液中MCP-1浓度。将SRA特异性结合配体DiI-Ac-LDL与细胞孵育,应用荧光显微镜观察各组细胞SRA蛋白表达及活性情况。并将MCP-1浓度与SRA蛋白活性进行相关性分析。结果400倍光镜下观察到细胞株THP-1在佛波醇酯诱导下转变为泡沫细胞。与对照组相比,OX-LDL组MCP-1表达升高,6h后升高明显（P〈0．05）,12h达高峰（P〈0．01）,24h后逐渐下降（P〈0．01）。阿托伐他汀药物干预,呈剂量依赖性降低MCP．1水平。OX-LDL组SRA蛋白活性水平明显高于对照组（P〈0．01）。阿托伐他汀干预,呈剂量依赖性下调SRA蛋白活性水平。各组细胞12hMCP．1浓度与SRA蛋白活性水平呈明显正相关（r=0．683,P〈0．01）。结论SRA、炎症因子MCP-1在THP-1细胞分化为泡沫细胞过程中发挥重要作用,阿托伐他汀抑制MCP-1与SRA表达,可能是其抗动脉粥样硬化形成的重要机制。     

Oxidized LDL specifically promotes the initiation of monocyte invasion during transendothelial migration with upregulated PECAM-1 and downregulated VE-cadherin on endothelial junctions

It is poorly understood how oxidized LDL (oxLDL) promotes monocyte dynamics in transendothelial migration (TEM) in atherogenesis. We developed an in vitro 3D-live-single cell TEM assay system with subendothelial oxLDL embedded in ultra-thin collagen gels, mimicking subendothelial oxLDL accumulation in vivo. With dividing monocyte dynamics into three stages (1: adhesion on endothelium, 2: invasion and 3: complete transmigration below endothelium), we analyzed the stage transition dynamics of individual living human monocytes. OxLDL did not enhance initial monocyte adhesion to endothelium (stage 1), but it specifically primed adherent monocytes to start invasion (stage 1-->2). Once invasion started, it had no effect thereafter on monocyte stage transition (stage 2-->3). OxLDL upregulated PECAM-1 and downregulated VE-cadherin on endothelial junctions without monocyte addition, both of which could promote monocyte entry by enhanced homophilic binding to monocyte PECAM-1, and by disrupted junctional barrier, respectively. Meanwhile, monocyte speed at neither locomotion on endothelium (stage 1) nor subendothelial migration (stage 3) was altered by oxLDL. These data indicate that before monocyte adhesion, endothelial junctions changed their conformation to more monocyte-acceptable state in response to oxLDL, resulting the stage-specific promotion of monocyte TEM (stage 1-->2; initiation of invasion) with no enhancement of its initial adhesion or migration speed.     

Interrelations between interleukin-6, interleukin-1 beta, plasma C-reactive protein values, and in vitro C-reactive protein generation in patients with inflammatory bowel disease.

Acute phase proteins are released from the liver in response to cytokines, and measurement of serum concentrations offers a valuable means of assessing inflammatory bowel disease. C-reactive protein (CRP) is a participating prominent component of the acute phase response in active Crohn's disease. This study aimed at determining the comparative role of the cytokines interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6), in driving CRP production in inflammatory bowel disease, and to test the hypothesis that there is a difference in the profile of cytokines generated in these two conditions. Serum CRP, the release of the cytokines IL-1 beta and IL-6 from monocytes, and the ability of monocyte conditioned medium to stimulate CRP synthesis by hepatocytes in an in vitro system was measured in patients with ulcerative colitis and Crohn's disease. Monocytes from patients with Crohn's disease produced more 1L beta-1 than monocytes from patients with ulcerative colitis or normal controls. There was no increased tendency for monocytes from Crohn's disease patients to produce more 1L-6, so the greater circulating values of IL-6 reported by a number of authors in Crohn's disease may reflect the participation of a larger number of cells of the monocyte-macrophage series, or production of IL-6 by other cell types. Correlation of cytokine production by monocytes with in vitro CRP release from cultured hepatocytes in response to monocyte conditioned medium showed that, in that system, IL-1 beta was the stronger stimulus to CRP production. Some of the differences in the inflammatory processes of ulcerative colitis and Crohn's disease may reflect differences in the amount of IL-1beta and IL-6 generated from macrophages and monocytes.     

Increased expression of CD23 (Fc(epsilon) receptor II) by peripheral blood monocytes of aids patients

Monocytes expressing the Fcepsilon receptor II (CD23) play important roles in inflammatory and allergic immune responses. We found that peripheral blood monocytes of AIDS patients express increased levels of CD23, compared with monocytes of healthy HIV-1-seronegative individuals (controls) (p < 0.05). We compared expression of monocyte CD23 with expression of monocyte Fcgamma receptors (CD16, CD32, CD64), plasma/serum levels of IgE (also IgM, IgG, IgA), and Th1 (IFN-gamma) and Th2 (IL-4, IL-10) cytokines. We found that monocyte CD23 expression directly correlated with monocyte CD16 expression (p < 0.01, R = 0.58), which was also increased in AIDS patients; there was no correlation with CD32 or CD64 or with soluble factors in plasma/serum (i.e., IgE, IL-4, IL-10, and IFN-gamma). Interestingly, despite the known ability of IL-10 to downregulate monocyte CD23 expression, plasma IL-10 levels were increased in these AIDS patients compared with controls (p < 0.05). We thus evaluated the effect of AIDS and control plasma or rhIL-10 to regulate CD23 expression by monocytes in cultures (24 hr) of healthy human cells +/- treatment with anti-IL-10R blocking antibody. We found that anti-IL-10R blocking antibody treatment had no effect on monocyte CD23 expression in cultures containing AIDS plasma, but increased monocyte CD23 expression in cultures containing control plasma (p < 0.05) or rhIL-10. In conclusion, the identification of increased monocyte CD23 expression in AIDS patients may further characterize the aberrant activated phenotype of monocytes during the immunopathogenesis of HIV-1 disease. Further, monocyte CD23 expression does not appear to be suppressed by the IL-10-enriched environment in AIDS.     

阿托伐他汀对冠心病患者血中趋化因子水平的影响

目的观察冠心病患者血浆中趋化因子的变化,并进一步探讨阿托伐汀对趋化因子的影响。方法健康对照组25例,血脂正常的冠心病70例,随机将冠心病患者分为常规治疗组和阿托伐汀组,检测血浆中单核细胞趋化蛋白-1(monocyte chemoattract protein-1,MCP-1)、T细胞表达和分泌活化蛋白(regulated upon activation normal T cell expressed and secreted,RANTES)及白介素-8(interleukin-8,IL-8)水平。结果与健康对照组相比,冠心病患者血浆中MCP-1、RANTES、IL-8水平明显升高(P<0.01)。常规治疗组和阿托伐汀组治疗4周后MCP-1、RANTES、IL-8水平均有下降(P<0.05,P<0.01),阿托伐汀治疗组较常规治疗组降趋化因子的作用更明显(P<0.05)。结论阿托伐汀降低冠心病患者血浆中趋化因子水平,可能起到稳定粥样斑块的作用。     

Rapid recruitment of inflammatory monocytes is independent of neutrophil migration

Early neutrophil entry into an inflammatory site is thought to mediate a chemokine switch, inducing subsequent monocyte recruitment through the regulation of monocyte chemoattractant protein-1 (MCP-1) release. As the murine monocyte is poorly characterized and difficult to identify, there has been little examination of either its early recruitment in inflammatory models or of the factors that influence its early migration. The phenotyping of rapidly recruited inflammatory leukocytes with 7/4 and Gr-1 monoclonal antibodies (mAbs) identifies 2 distinct populations, which we characterize as murine monocytes and neutrophils. Monocytes migrate in the first 2 hours of inflammation making use of alpha4beta1 but not of Mac-1 or lymphocyte function-associated antigen-1 (LFA-1) integrins. Early migration is dependent on MCP-1, but neither MCP-1 release nor monocyte recruitment is affected by the reduced neutrophil migration seen in LFA-1-/- mice. Endogenous peritoneal macrophages and mesothelial cells lining the peritoneum contain MCP-1, which is released following thioglycollate stimulation. The murine monocyte therefore responds rapidly to chemokines produced in situ by tissue cells at the site of inflammation with no requirement for prior influx of neutrophils.     

9-cis retinoic acid induces monocyte chemoattractant protein-1 secretion in human monocytic THP-1 cells.

Monocyte migration and activation are regulated by monocyte chemoattractant protein-1 (MCP-1). Prior studies have shown MCP-1 expression is modulated by a variety of ligands that act through extracellular receptors. In the current study, we show 9-cis retinoic acid (RA), a ligand for the nuclear hormone receptor retinoid X receptor (RXR) and retinoic acid receptor (RAR), markedly induces the expression of MCP-1. In human THP-1 monocytic leukemia cells cultured with RA (0.05 to 500 nmol/L), MCP-1 expression was induced rapidly, significantly, and dose-dependently by as much as 165-fold. MCP-1 RNA level was also increased in RA-treated cells. Expression of PPARgamma, a heterodimer partner of RXR, is also markedly induced by RA in THP-1 cells. However, BRL49653, a PPARgamma ligand, failed to induce MCP-1 secretion either alone or to modify the expression level induced by RA. In contrast, BRL49653 significantly increased MCP-1 (biotinylated MCP-1) binding to THP-1 cells, whereas RA had no effect. Other peroxisome proliferator activated receptor (PPAR) ligands, 15d-PGJ(2) and troglitazone (PPARgamma), Wy14,643 (PPARalpha), and PD195599 (PPARbeta) inhibited the induction of MCP-1 by RA. RA's effect on MCP-1 expression in human elutriated monocytes were similar to that observed in the THP-1 cells. These studies identify RA as a nuclear signal for MCP-1 induction in undifferentiated human monocytic cells. These studies also suggest monocyte MCP-1 expression induced through RA may modulate cell migration.     

Redox regulation of MAPK phosphatase 1 controls monocyte migration and macrophage recruitment

Monocytic adhesion and chemotaxis are regulated by MAPK pathways, which in turn are controlled by redox-sensitive MAPK phosphatases (MKPs). We recently reported that metabolic disorders prime monocytes for enhanced recruitment into vascular lesions by increasing monocytes’ responsiveness to chemoattractants. However, the molecular details of this proatherogenic mechanism were not known. Here we show that monocyte priming results in the S-glutathionylation and subsequent inactivation and degradation of MKP-1. Chronic exposure of human THP-1 monocytes to diabetic conditions resulted in the loss of MKP-1 protein levels, the hyperactivation of ERK and p38 in response to monocyte chemoattractant protein-1 (MCP-1), and increased monocyte adhesion and chemotaxis. Knockdown of MKP-1 mimicked the priming effects of metabolic stress, whereas MKP-1 overexpression blunted both MAPK activation and monocyte adhesion and migration induced by MCP-1. Metabolic stress promoted the S-glutathionylation of MKP-1, targeting MKP-1 for proteasomal degradation. Preventing MKP-1 S-glutathionylation in metabolically stressed monocytes by overexpressing glutaredoxin 1 protected MKP-1 from degradation and normalized monocyte adhesion and chemotaxis in response to MCP-1. Blood monocytes isolated from diabetic mice showed a 55% reduction in MKP-1 activity compared with nondiabetic mice. Hematopoietic MKP-1 deficiency in atherosclerosis-prone mice mimicked monocyte priming and dysfunction associated with metabolic disorders, increased monocyte chemotaxis in vivo, and accelerated atherosclerotic lesion formation. In conclusion, we identified MKP-1 as a central redox-sensitive regulator of monocyte adhesion and migration and showed that the loss of MKP-1 activity is a critical step in monocyte priming and the metabolic stress-induced conversion of blood monocytes into a proatherogenic phenotype.     

Mean platelet volume: a useful marker of inflammatory bowel disease activity

OBJECTIVES: We investigated whether the mean platelet volume would be a useful marker in the evaluation of inflammatory bowel disease activity. METHODS: Complete blood count, C-reactive protein, erythrocyte sedimentation rate, serum thrombopoietin and erythropoietin, plasma beta-thromboglobulin, and platelet factor 4 were measured in 93 patients with ulcerative colitis, 66 patients with Crohn's disease, and 38 healthy blood donors. Disease activity was assessed by the Clinical Colitis Activity Index in patients with ulcerative colitis and by the Crohn's Disease Activity Index in patients with Crohn's disease. RESULTS: Mean platelet count was increased in patients with active compared to inactive ulcerative colitis (p < 0.05), and in patients with active compared to inactive Crohn's disease (p = 0.0002) or healthy controls (p < 0.0001). On the other hand, mean platelet volume was significantly decreased in patients with active compared to inactive ulcerative colitis (p = 0.02) or healthy controls (p < 0.0001), and in patients with active compared to inactive Crohn's disease (p = 0.0005) or healthy controls (p < 0.0001). Mean platelet volume was inversely correlated with the white blood cell count (r = -0.17, p = 0.02), C-reactive protein (r = -0.46, p = 0.009) and erythrocyte sedimentation rate (r = -0.28, p = 0.008). No significant correlations were found between mean platelet volume and serum thrombopoietin or erythropoietin levels; however, a strong negative correlation between mean platelet volume and beta-thromboglobulin (r = -0.34, p < 0.0001) and platelet factor 4 (r = -0.30, p = 0.0002) was observed. CONCLUSIONS: Mean platelet volume is significantly reduced in active inflammatory bowel disease and is negatively correlated with the known inflammatory bowel disease activity markers and the platelet activation products. We propose that mean platelet volume provides a useful marker of activity in inflammatory bowel disease.     

Circulating human leucocyte elastase in patients with inflammatory bowel disease.

The plasma concentration of human leucocyte elastase (HLE), measured by enzyme immunoassay as the complex with alpha 1-proteinase inhibitor, was determined in 94 patients with active and inactive inflammatory bowel disease. In Crohn's disease and in ulcerative colitis human leucocyte elastase levels were raised significantly above normal when the disease was active, and fell on remission. The mean human leucocyte elastase level in 31 cases of active Crohn's disease was significantly greater than the mean human leucocyte elastase level in 23 patients with active ulcerative colitis (p = 0.013). The values of human leucocyte elastase correlated significantly with Crohn's disease activity index scores (p = 0.05) and with the circulating concentration of C-reactive protein (p less than 0.05 and p less than 0.01 for ulcerative colitis and Crohn's disease respectively), but not with the erythrocyte sedimentation rate. These results indicate that the concentration of human leucocyte elastase in the plasma of patients with inflammatory bowel disease reflects the activity of their intestinal disease and suggest that serial measurements of human leucocyte elastase may be useful in the assessment and clinical management of these conditions.     

CD14 expression on monocytes and soluble CD14 plasma levels in correlation to the promotor polymorphism of the endotoxin receptor CD14 gene in patients with inactive Crohn's disease

BACKGROUND/AIMS: The association of the single nucleotide polymorphism in the promotor of the lipopolysaccharide receptor CD14 gene (T/C at position -159) with Crohn's disease has recently been demonstrated. This CD14 polymorphism is a potential predisposition factor responsible for inter-individual differing inflammatory reactions involving the CD14 receptor. We studied the correlation between the CD14 genotype (CC, CT, TT) and the membrane-bound CD14 monocyte expression and soluble CD14 in patients with inactive Crohn's disease. METHODOLOGY: In 23 patients and 29 healthy volunteers the membrane-bound CD14 density on unstimulated monocytes and soluble CD14 plasma levels were examined using quantitative flow cytometry and enzyme-linked immunosorbent assay. RESULTS: In normal controls membrane-bound CD14 monocyte density did not differ significantly between the genotypes CC, CT, or TT. In contrast, patients with inactive Crohn's disease and genotype TT showed a significantly lower membrane-bound CD14 density on monocytes compared to patients with genotype CC. Soluble CD14 plasma levels were significantly higher in patients with inactive Crohn's disease compared to the same genotype of healthy controls, but there was no significant difference between the genotypes CC, CT, and TT. CONCLUSIONS: Our data show that the membrane-bound CD14 monocyte expression and the soluble CD14 plasma levels in patients with inactive Crohn's disease completely differ from that in healthy individuals. In order to develop individualized therapy strategies further studies should be carried out to evaluate whether the TT genotype is associated with differences in the clinical course of Crohn's disease and in the response to antibacterial treatment.     

The impact of early administration of low-dose atorvastatin treatment on inflammatory process, in patients with unstable angina and low cholesterol level

BACKGROUND: Lipid-lowering agents are known to reduce long-term mortality in patients with stable angina or multiple risk factors. However, the effects of lipid-lowering treatment on inflammatory process during and immediately after the acute phase of unstable angina remain unclear. In this study we assessed the effects of low-dose atorvastatin treatment, on inflammatory process in patients admitted for unstable angina with low cholesterol level. METHODS: Forty-seven normocholesterolemic patients with unstable angina were randomized into two groups, and received atorvastatin 10 mg/day (n = 24) or no statin (n = 23) for 6 weeks. Circulating levels of inteleukin 6 (IL-6), monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor alpha (TNF-alpha) and soluble vascular cell adhesion molecule (sVCAM-1) were measured by their admission, and at the 1st and 6th week of the study. RESULTS: Serum levels of MCP-1 and sVCAM-1 were significantly increased in the control group (p < 0.05) while remained unaffected in the atorvastatin-treated group six weeks after admission. However, IL-6 and TNF-alpha levels were similarly decreased in both atorvastatin-treated and control groups. CONCLUSION: Low-dose atorvastatin treatment modifies inflammatory process in patients with unstable angina and low cholesterol level, an effect seen at 6 weeks but not 1 week after admission.     

Alterations in the function of circulating mononuclear cells derived from patients with Crohn＇s disease treated with mastic

AIM： To assess the effects of mastic administration on cytokine production of circulating mononuclear cells of patients with active Crohn＇s disease （CD）. METHODS： The study was conducted in patients with established mildly to moderately active CD, attending the outpatient clinics of the hospital, and in healthy controls. Recruited to a 4 wk treatment with mastic caps （6 caps/d, 0.37 g/cap） were 10 patients and 8 controls, all of who successfully completed the protocol. Interleukin-6 （IL-6）, tumor necrosis factor-alpha （TNF-α）, monocyte chemotactic protein-1 （MCP-1）, macrophage migration inhibitory factor （MIF） and intracellular antioxidant glutathione （GSH） were evaluated in peripheral blood mononuclear cells （PBMC） before and after treatment. RESULTS： Treating CD patients with mastic resulted in the reduction of TNF-α secretion （2.1 ± 0.9 ng/mL vs 0.5 ± 0.4 ng/mL, P = 0.028）. MIF release was significantly increased （1.2±0.4 ng/mL vs 2.5 ± 0.7 ng/mL, P = 0.026） meaning that random migration and chemotaxis of monocytes/macrophages was inhibited. No significant changes were observed in IL-6, MCP-1 and GSH concentrations. CONCLUSION： This study shows that mastic acts as an immunomodulator on PBIC, acting as a TNF-α inhibitor and a MIF stimulator. Although further double-blind, placebo-controlled studies in a large number of patients is required to clarify the role of this natural product, this finding provides strong evidence that mastic might be an important regulator of immunity in CD.     

Protein S-100ß: A Biochemical Marker for Increased Intracranial Pressure in Pigs with Acute Hepatic Failure

Background: Inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) synthesis are increased in epithelial cells and in tissue macrophages of the inflamed mucosa from patients with inflammatory bowel disease (IBD). Since tissue macrophages are derived from circulating monocytes, we studied iNOS expression in circulating monocytes and related this expression to disease activity. In view of the possible role of NO in monocyte function, we also studied iNOS expression in relation to markers of monocyte activation. Methods: The expression of iNOS in circulating monocytes from 15 patients with active IBD, 6 patients who went into remission and 18 healthy controls was quantified by flow cytometry and correlated with surface markers (CD63, CD11b, HLA-DR) for monocyte activation. In addition, iNOS expression in circulating monocytes was assessed by Western blotting, immunocytochemistry and measurement of the NO metabolites nitrite and nitrate in plasma. Results: The expression of iNOS in circulating monocytes and the percentage of iNOS-positive monocytes were increased in patients with active IBD compared to healthy controls (fluorescence index: 1.3 (0.1-6.3) versus 0.8 (0.0- 1.8); P < 0.05; percentage of iNOS positive monocytes: 37.3 (1.0-77.0)% versus 5.3 (0.0-43.3)%; P < 0.01). The six patients who went into remission all had a marked reduction of iNOS expression. iNOS expression was confirmed by Western blotting and immunocytochemistry. Plasma nitrite and nitrate levels were elevated in three patients with active IBD. The surface markers for monocyte activation, CD63 and CD11b, were not elevated. HLA-DR expression was decreased on circulating monocytes from patients with active ulcerative colitis. Conclusions: iNOS is increased in circulating monocytes from patients with active IBD and this increased expression correlates with disease activity. Considering the decreased HLA-DR expression and absence of monocyte activation markers, NO produced by iNOS may have a function in suppressing systemic monocyte activation.     

Activation of blood coagulation in Crohn's disease. Increased plasma fibrinopeptide A levels and enhanced generation of monocyte tissue factor activity

We have examined the relationships among activation of blood coagulation, generation of monocyte procoagulant activity, and clinical activity in patients with Crohn's disease. Subclinical activation of blood coagulation was measured using a radioimmunoassay for fibrinopeptide A. Fibrinopeptide A levels were strongly correlated with the level of disease activity as measured by the Crohn's disease activity index. Patients with active disease who were successfully treated either medically or surgically demonstrated a reduction of fibrinopeptide A levels. Failure of fibrinopeptide A to return to the normal range predicted an early relapse. Monocyte tissue factor generation was assessed in both unstimulated and lipopolysaccharide-stimulated mononuclear cell cultures obtained from the peripheral blood of patients with Crohn's disease. A strong correlation (r = 0.89) was observed between plasma fibrinopeptide A levels and monocyte tissue factor generation. These results suggest that monocyte procoagulant generation may contribute to the activation of blood coagulation in this inflammatory bowel disease. Moreover, fibrinopeptide A levels in Crohn's disease may provide a useful quantitative measure of inflammatory activity.