Monocyte Chemotactic Protein-1 Gene Expression Research Articles

Triterpenoid CDDO-EA inhibits lipopolysaccharide-induced inflammatory responses in skeletal muscle cells through suppression of NF-κB.

Chronic inflammation is a major contributor to the development of obesity-induced insulin resistance, which then can lead to the development of type 2 diabetes (T2D). Skeletal muscle plays a pivotal role in insulin-stimulated whole-body glucose disposal. Therefore, dysregulation of glucose metabolism by inflammation in skeletal muscle can adversely affect skeletal muscle insulin sensitivity and contribute to the pathogenesis of T2D. The mechanism underlying insulin resistance is not well known; however, macrophages are important initiators in the development of the chronic inflammatory state leading to insulin resistance. Skeletal muscle consists of resident macrophages which can be activated by lipopolysaccharide (LPS). These activated macrophages affect myocytes via a paracrine action of pro-inflammatory mediators resulting in secretion of myokines that contribute to inflammation and ultimately skeletal muscle insulin resistance. Therefore, knowing that synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acids (CDDOs) can attenuate macrophage pro-inflammatory responses in chronic disorders, such as cancer and obesity, and that macrophage pro-inflammatory responses can modulate skeletal muscle inflammation, we first examined whether CDDO-ethyl amide (CDDO-EA) inhibited chemokine and cytokine production in macrophages since this had not been reported for CDDO-EA. CDDO-EA blocked LPS-induced tumor necrosis factor-alpha (TNF-α), monocyte chemotactic protein-1 (MCP-1), interleukine-1beta (IL-1β), and interleukine-6 (IL-6) production in RAW 264.7 mouse and THP-1 human macrophages. Although many studies show that CDDOs have anti-inflammatory properties in several tissues and cells, little is known about the anti-inflammatory effects of CDDOs on skeletal muscle. We hypothesized that CDDO-EA protects skeletal muscle from LPS-induced inflammation by blocking nuclear factor kappa B (NF-κB) signaling. Our studies demonstrate that CDDO-EA prevented LPS-induced TNF-α and MCP-1 gene expression by inhibiting the NF-κB signaling pathway in L6-GLUT4myc rat myotubes. Our findings suggest that CDDO-EA suppresses LPS-induced inflammation in macrophages and myocytes and that CDDO-EA is a promising compound as a therapeutic agent for protecting skeletal muscle from inflammation.

Changes in MCP-1, HGF, and IGF-1 expression in endometrial stromal cells, PBMCs, and PFMCs of endometriotic women following 1,25(OH)2D3 treatment.

1,25(OH)2D3 has anti-inflammatory and growth inhibitory effects. Our study explored the effect of 1,25(OH)2D3 treatment on the expression of monocyte chemotactic protein-1 (MCP-1), hepatocyte growth factor (HGF), and insulin-like growth factor-1 (IGF-1) by peripheral blood mononuclear cells (PBMCs), peritoneal fluid mononuclear cells (PFMCs), endometrial stromal cells (ESCs), and its effect on the proliferation of PBMCs and PFMCs of patients with endometriosis compared with controls. PBMCs, PFMCs, and ESCs were obtained from 10 endometriosis patients and 10 non-endometriotic individuals. After treating cells with 0.1μM of 1,25(OH)2D3 for 6, 24, and 48 h, the gene and protein expression of mentioned factors were evaluated by real-time PCR and ELISA methods, respectively. 1,25(OH)2D3 treatment significantly reduced the protein expression of MCP-1, HGF, and IGF-1 in PBMCs and PFMCs of endometriotic patients at 48 h (p< 0.05-<0.01). Also, this treatment significantly reduced MCP-1, HGF, and IGF-1 gene and/or protein expression in EESCs and EuESCs at 24 and 48 h (p< 0.05-<0.01). 1,25(OH)2D3 treatment also reduced the proliferation of PBMCs and PFMCs of endometriotic patients compared with controls (p < 0.01). 1,25(OH)2D3 can be considered as a potentially effective agent in the prevention and treatment of endometriosis along with other therapies.

Resveratrol treatment reduces expression of MCP-1, IL-6, IL-8 and RANTES in endometriotic stromal cells.

Endometriosis is an inflammatory disease affecting reproductive‐aged women. Immunologic disturbance, as well as inflammation, have crucial roles in the pathogenesis of endometriosis. In this study, we evaluated the effects of resveratrol treatment on expression of monocyte chemotactic protein‐1 (MCP‐1), interleukin‐6 (IL‐6), IL‐8, and regulated upon activation, normal T cell expressed and secreted (RANTES) in endometrial stromal cells from patients with endometriosis compared with non‐endometriotic controls. Thirteen eutopic (EuESCs) and nine ectopic (EESCs) endometrial stromal cells from endometriotic patients as well as eleven endometrial stromal cells from non‐endometriotic controls (CESCs) were treated with resveratrol (100 μmol/L) or ethanol, and gene and/or protein expression of MCP‐1, IL‐6, IL‐8 and RANTES was examined at 6, 24 and 48 hours following treatment in the cells from all origins. Resveratrol treatment significantly reduced gene and protein expression of MCP‐1, IL‐6, and IL‐8 in EuESCs and EESCs compared with CESCs (P < .05‐.001, P < .05‐.001 and P < .05‐<.01, respectively), and this reduction was more noticeable in EESCs than EuESCs (P < .05‐<.001). Besides, resveratrol treatment significantly reduced RANTES protein expression in EESCs in all time intervals (P < .05). Resveratrol treatment significantly reduced the expression of MCP‐1, IL‐6, IL‐8 and RANTES in EESCs.

λ-Carrageenan initiates inflammation via activation of heterodimers TLR2/6 and TLR4/6

Abstract λ-Carrageenan (λCGN) is frequently used to model local inflammation, however, the mechanism by which λCGN functions has not been fully examined. Swine whole blood was stimulated with λCGN and biomarkers were evaluated. Monocyte Chemotactic Protein-1(MCP-1) and Interleukin-8 protein levels were increased, while Serum Amyloid A (SAA) and Tumor Necrosis Factor-α (TNFα) protein levels were unchanged. Gene expression of MCP-1, Phosphatidylinositol 3-Kinase, and Alveolar Macrophage Chemotactic–II were increased; gene expression of SAA, Interleukin-6, and TNFα were unchanged; and gene expression of Thrombospondin-2 Precursor were decreased. Assessment of which TLRs (Toll-Like Receptors) were involved in the λCGN activation pathway, HEK-293 cells transfected with one plasmid containing either TLR2 or TLR4, along with a second plasmid to release SEAP (secreted embryonic alkaline phosphatase) upon NFκB activation were stimulated with λCGN. TLR2 and TLR4 both showed increased SEAP release after λCGN stimulation. The lack of TNFα production after λCGN stimulation suggested that it was not working through a classic TLR4 homodimer. To determine if a common mechanism could explain these results, RNA silencing studies were initiated. TLR6 was silenced in SEAP HEK-293 cells containing a plasmid for either TLR2 or TLR4. In both cell lines where TLR6 was silenced, λCGN was unable to induce a SEAP response, while these cells maintained responsiveness to a TLR1/2 ligand and a TLR 4 ligand, respectively. These results suggest that λCGN initiates inflammation via TLR2/6 and TLR4/6 heterodimers.

Characterization of the inflammatory effects of λ-carrageenan at the genomic and proteomic levels

Abstract λ-Carrageenan (λCGN) is often used to model local inflammation, but the mechanism by which this compound functions at the genomic and proteomic level has not been delineated. Whole blood from swine was stimulated with λCGN to determine inflammatory biomarkers. We also utilized a HEK-293 (human embryonic kidney) cell line transfected with a SEAP (secreted embryonic alkaline phosphatase)-release plasmid to determine which TLR (Toll-Like Receptor; TLR) signaling cascade was stimulated. Results indicated that Monocyte Chemotactic Protein-1 gene expression and protein were increased by λCGN stimulation while Serum Amyloid A gene expression and protein remained unchanged. λCGN stimulation increased mRNA levels of Phosphatidylinositol 3-Kinase and Alveolar Macrophage Chemotactic –II, decreased mRNA levels of Thrombospondin-2 Precursor, and had no effect on Interleukin-6 mRNA levels. λCGN stimulation increased protein levels of Interleukin-8 and had no effect on protein levels of Tumor Necrosis Factor-α. SEAP release from HEK-293 cells stimulated with λCGN indicated that both TLR2 and TLR4 signaling cascades were initiated. λCGN stimulation did not activate TLR3, TLR5, TLR7, TLR8, TLR9, NOD1, NOD2, Dectin-1a, or Dectin-1b. Experiments silencing either TLR1 or TLR6 to determine which TLR2 heterodimer is activated by λCGN are currently underway. In conclusion, while λCGN stimulation appears to signal through both the TLR2 and TLR4 pathways, significant differences exist between the genes and proteins stimulated by λCGN and pure TLR4 agonists such as E. coli-derived lipopolysaccharide.

Effects of Shock Waves on Expression of IL-6, IL-8, MCP-1, and TNF-α Expression by Human Periodontal Ligament Fibroblasts: An In Vitro Study.

BackgroundExtracorporeal shock wave therapy (ESWT) can modulate cell behavior through mechanical information transduction. Human periodontal ligament fibroblasts (hPDLF) are sensible to mechanical stimulus and can express pro-inflammatory molecules in response. The aim of this study was to evaluate the impacts of shock waves on interleukin-6 (IL-6), interleukin-8 (IL-8), monocyte chemotactic protein 1 (MCP-1), and tumor necrosis factor-alpha (TNF-α) expression by hPDLF.Material/MethodsAfter being treated by shock waves with different parameters (100–500 times, 0.05–0.19 mJ/mm2), cell viability was tested using CCK-8. IL-6, IL-8, MCP-1, and TNF-α gene expression was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and IL-6 and IL-8 protein was measured by enzyme-linked immunosorbent assay (ELISA) at different time points.ResultsShock waves with the parameters used in this study had no significant effects on the viability of hPDLF. A statistical inhibition of IL-6, IL-8, MCP-1, and TNF-α expression during the first few hours was observed (P<0.05). Expression of IL-8 was significantly elevated in the group receiving the most pulses of shock wave (500 times) after 4 h (P<0.05). At 8 h and 24 h, all treated groups demonstrated significantly enhanced IL-6 expression (P<0.05). TNF-α expression in the groups receiving more shock pulses (300, 500 times) or the highest energy shock treatment (0.19 mJ/mm2) was statistically decreased (P<0.05) at 24 h.ConclusionsUnder the condition of this study, a shock wave with energy density no higher than 0.19 mJ/mm2 and pulses no more than 500 times elicited no negative effects on cell viability of hPDLF. After a uniform initial inhibition impact on expression of inflammatory mediators, a shock wave could cause dose-related up-regulation of IL-6 and IL-8 and down-regulation of TNF-α.

Monocyte Chemotactic Protein-1 Gene Expression in Blood and Ascitic Fluid of Cirrhotic Patients with Spontaneous Bacterial Peritonitis

Background and study aim: Cirrhotic patients with ascites show a higher susceptibility to bacterial infections, monocyte chemotactic protein-1 (MCP-1) secretion is up-regulated during chronic hepatitis and correlates with the severity of hepatic inflammation. The aim of this work is to determine the level of expression of MCP-1 gene in blood and ascitic fluid in cirrhotic patients with and without spontaneous bacterial peritonitis (SBP) to evaluate its role in pathogenesis of SBP and its role in diagnosis. Patients and Methods: This study included 15 healthy subjects served as control group in addition to 35 cirrhotic patients due to HCV infection with ascites; classified into two groups, cirrhosis without SBP (15 patients) and cirrhosis with SBP (20 patients). All groups were subjected to quantitative estimation of MCP-1 gene expression in blood by real time PCR. In SBP and non SBP groups the gene expression were assessed in ascitic fluid also at diagnosis and reassessed in SBP group after treatment. Results: Blood and ascitic fluid expression of MCP-1 gene were significant higher in SBP group than non SBP group and control group. SBP group showed a significant decrease in level of MCP-1 gene expression in blood and ascitic fluid after resolution of infection by appropriate treatment of SBP. Conclusion: MCP-1 gene expression in both blood and ascitic fluid may be related to pathophysiology and course of SBP and can be used as a marker for diagnosis.

Chlorella Protein Hydrolysate Attenuates Glucose Metabolic Disorder and Fatty Liver in High-fat Diet-induced Obese Mice.

Chlorella (Parachlorella beijerinckii) powder is reported to show a preventive effect against metabolic syndromes such as arteriosclerosis, hyperlipidemia, and hypertension. Approximately 60% of the chlorella content is protein. In order to understand the role of chlorella protein, we prepared a chlorella protein hydrolysate (CPH) by protease treatment. Male C57BL/6 mice were divided into three groups: a normal diet group, high-fat diet (HFD) group, and high-fat diet supplemented with CPH (HFD+CPH) group. The CPH administration improved glucose intolerance, insulin sensitivity, and adipose tissue hypertrophy in the high-fat diet-fed mice. In addition, the HFD+CPH group had significantly decreased liver total cholesterol and triglyceride levels compared with those in the HFD group. Furthermore, the HFD+CPH group had a decreased level of monocyte chemotactic protein-1 (MCP-1) in serum and a lower MCP-1 mRNA expression level in adipose tissue compared with the HFD group. The present study suggests that chlorella protein hydrolysate can prevent a high-fat diet-induced glucose disorder and fatty liver by inhibiting adipocyte hypertrophy and reducing the MCP-1 protein and gene expression.

Cachexia-associated adipose tissue morphological rearrangement in gastrointestinal cancer patients.

Background and aimsCachexia is a syndrome characterized by marked involuntary loss of body weight. Recently, adipose tissue (AT) wasting has been shown to occur before the appearance of other classical cachexia markers. We investigated the composition and rearrangement of the extracellular matrix, adipocyte morphology and inflammation in the subcutaneous AT (scAT) pad of gastrointestinal cancer patients.MethodsSurgical biopsies for scAT were obtained from gastrointestinal cancer patients, who were signed up into the following groups: cancer cachexia (CC, n = 11), weight‐stable cancer (WSC, n = 9) and weight‐stable control (non‐cancer) (control, n = 7). The stable weight groups were considered as those with no important weight change during the last year and body mass index <25 kg/m2. Subcutaneous AT fibrosis was quantified and characterized by quantitative PCR, histological analysis and immunohistochemistry.ResultsThe degree of fibrosis and the distribution and collagen types (I and III) were different in WSC and CC patients. CC patients showed more pronounced fibrosis in comparison with WSC. Infiltrating macrophages surrounding adipocytes and CD3 Ly were found in the fibrotic areas of scAT. Subcutaneous AT fibrotic areas demonstrated increased monocyte chemotactic protein 1 (MCP‐1) and Cluster of Differentiation (CD)68 gene expression in cancer patients.ConclusionsOur data indicate architectural modification consisting of fibrosis and inflammatory cell infiltration in scAT as induced by cachexia in gastrointestinal cancer patients. The latter was characterized by the presence of macrophages and lymphocytes, more evident in the fibrotic areas. In addition, increased MCP‐1 and CD68 gene expression in scAT from cancer patients may indicate an important role of these markers in the early phases of cancer.

Endocannabinoids and inflammatory response in periodontal ligament cells.

Endocannabinoids are associated with multiple regulatory functions in several tissues. The main endocannabinoids, anandamide (AEA) and 2-arachidonylglycerol (2-AG), have been detected in the gingival crevicular fluid of periodontitis patients, but the association between periodontal disease or human periodontal ligament cells (hPdLCs) and endocannabinoids still remain unclear. The aim of the present study was to examine the effects of AEA and 2-AG on the proliferation/viability and cytokine/chemokine production of hPdLCs in the presence/absence of Porphyromonas gingivalis lipopolysaccharide (P. gingivalis LPS). The proliferation/viability of hPdLCs was measured using 3,4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide (MTT)-assay. Interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemotactic protein-1 (MCP-1) levels were examined at gene expression and protein level by real-time PCR and ELISA, respectively. AEA and 2-AG did not reveal any significant effects on proliferation/viability of hPdLCs in the absence of P. gingivalis LPS. However, hPdLCs viability was significantly increased by 10–20 µM AEA in the presence of P. gingivalis LPS (1 µg/ml). In the absence of P. gingivalis LPS, AEA and 2-AG did not exhibit any significant effect on the expression of IL-8 and MCP-1 expression in hPdLCs, whereas IL-6 expression was slightly enhanced by 10 µM 2-AG and not affected by AEA. In P.gingivalis LPS stimulated hPdLCs, 10 µM AEA down-regulated gene-expression and protein production of IL-6, IL-8, and MCP-1. In contrast, 10 µM 2-AG had an opposite effect and induced a significant up-regulation of gene and protein expression of IL-6 and IL-8 (P<0.05) as well as gene-expression of MCP-1 in P. gingivalis LPS stimulated hPdLCs. Our data suggest that AEA appears to have an anti-inflammatory and immune suppressive effect on hPdLCs’ host response to P.gingivalis LPS, whereas 2-AG appears to promote detrimental inflammatory processes. In conclusion, AEA and 2-AG might play an important role in the modulation of periodontal inflammation.

Molecular detection of monocyte chemotactic protein-1 polymorphism in spontaneous bacterial peritonitis patients.

To investigate the association of the functional monocyte chemotactic protein-1 (MCP-1) promoter polymorphism (A-2518G) with spontaneous bacterial peritonitis (SBP). Fifty patients with post-hepatitis C liver cirrhosis and ascites were categorized into two groups; group I included 25 patients with SBP and group II included 25 patients free from SBP. In addition, a group of 20 healthy volunteers were included. We assessed the MCP-1 gene polymorphism and gene expression as well as interleukin (IL)-10 levels in both blood and ascitic fluid. A significant MCP-1 gene polymorphism was detected in groups I and II (P = 0.001 and 0.02 respectively). Group I was associated with a significantly higher frequency of AG genotype [control 8 (40%) vs SBP 19 (76.0%), P < 0.001], and group II was associated with a significantly higher frequency of GG genotype when compared to healthy volunteers [control 1 (5%) vs cirrhotic 16 (64%), P < 0.001]. Accordingly, the frequency of G allele was significantly higher in both groups (I and II) [control 10 (25%) vs SBP 27 (54%), P < 0.001 and vs cirrhotic 37 (74.0%), P < 0.001, respectively]. The total blood and ascetic fluid levels of IL-10 and MCP-1 gene expression were significantly higher in group I than in group II. Group I showed significant reductions in the levels of MCP-1 gene expression and IL-10 in the whole blood and ascetic fluid after therapy. MCP-1 GG genotype and G allele may predispose HCV infected patients to a more progressive disease course, while AG genotype may increase the susceptibility to SBP. Patients carrying these genotypes should be under supervision to prevent or restrict further complications.

Monocyte chemotactic protein-1 (MCP-1) is the central mediator of inflammation in the ovarian follicle of obese women

To investigate serum, follicular fluid (FF) and granulosa cell (GC) inflammatory marker levels in normal, overweight and obese women undergoing ART. Prospective controlled study. 84 women (age:37±5) who underwent COH followed by oocyte retrieval were included. Serum was collected in early follicular phase of the IVF cycle. FF was collected only from the first follicle. Cumulus GCs from large follicles >14mm were isolated from oocytes. FF inflammatory marker levels (MCP-1, IL-1α, IL-1β, IL-2, Il-4, IL-6, IL-8, IL-10, EGF, GM-CSF, TGFα, TNFα, TNFβ, VEGF, Eotaxin, FGF2) were measured using Milliplex assay. Serum and FF levels of CRP, OxLDL,Catalase and IL-18 were measured using ELISA. mRNA was isolated from GCs, and gene expression of MCP-1, CCR2 (MCP-1 receptor), SRB, ATF, GPx and Lox1 was quantified using qRT-PCR. Statistical analyses: Pearson and Spearman rank correlation, Student t and Mann-Whitney U tests with signifance at 0.05. In FF only MCP-1 was significantly correlated with BMI (r=0.45, p=0.003), while in serum, Catalase and CRP were significantly correlated with BMI (r=0.4, p=0.04 and r=0.6 p=0.02). Obese women (BMI>=30; n=13) had higher MCP-1 levels in their FF than lean women (BMI<25, n=15) (821±403 vs 444±210 pg/ml, p=0.003). MCP-1 was also expressed in the GCs whereas CCR2 was not. While FF MCP-1 correlated with BMI, GC MCP-1 did not (p=0.85). Other inflammatory markers: SRB, ATF, GPx and Lox1 were also detected in GCs, but their levels did not correlate to BMI. MCP-1 is a marker of inflammation and chemotactic for macrophages. Our study shows MCP-1 to be the only inflammatory marker that is significantly increased in FF of obese women, suggesting a role for MCP-1 in obesity-induced ovarian dysfunction. However CCR2 was not detected in the GCs. These data suggest that GCs are not the target of MCP-1 but may contribute to MCP-1 production. Hence in the obese patient, MCP-1 may attract monocytes and lead to inflammation in the ovarian follicle.

The anti-inflammatory effect of kaempferol on early atherosclerosis in high cholesterol fed rabbits.

BackgroundAtherosclerosis has been widely accepted as an inflammatory disease of vascular, adhesion molecules play an important role in the early progression of it. The aim of the present study was to evaluate the effect of kaempferol on the inflammatory molecules such as E-selectin (E-sel), intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesionmolecule-1 (VCAM-1) and monocyte chemotactic protein-1 (MCP-1) in high cholesterol induced atherosclerosis rabbit models.MethodsThirty male New Zealand white (NZW) rabbits were randomly divided into five groups, control group, model group, fenofibrate (12mg/kg) group and kaempferol groups (150 mg/kg and 30 mg/kg). The rabbits were fed with a normal diet or a high cholesterol diet for 10 weeks. Levels of blood lipids, serum tumour-necrosis factor-alpha (TNF-α) and serum interleukin-1beta (IL-1β) were detected at the end of the sixth and tenth week. Malonaldehyde (MDA) level and superoxide dismutase (SOD) activity in serum were also determined. Lesion areas of the aorta were measured with morphometry analysis after ten weeks. Gene expression of E-sel, ICAM-1, VCAM-1 and MCP-1 in aortas was determined by RT-PCR (reverse transcription-polymerase chain reaction). Immunohistochemical staining was employed to measure protein expression of E-sel, ICAM-1, VCAM-1 and MCP-1.ResultsModel rabbits fed with ten weeks of high-cholesterol diet developed significant progression of atherosclerosis. Compared with the control, levels of blood lipids, TNF-α, IL-1β and MDA increased markedly in serum of model rabbits, while SOD levels decreased. Gene and protein expressions of E-sel, ICAM-1, VCAM-1 and MCP-1 in atherosclerotic aortas increased remarkably in model group. However, comparing to the model rabbits, levels of TNF-α, IL-1β and MDA decreased significantly and serum SOD activity increased, gene and protein expressions of E-sel, ICAM-1, VCAM-1 and MCP-1 in aortas decreased significantly with the treatment of kaempferol.ConclusionKaempferol shows anti-atherosclerotic effect by modulating the gene and protein expression of inflammatory molecules.

Sevoflurane suppresses tumour necrosis factor-α-induced inflammatory responses in small airway epithelial cells after anoxia/reoxygenation

Lung ischaemia-reperfusion (I/R) injury is correlated with poor clinical outcome. The inflammatory cytokines interleukin (IL)-6, IL-8, and monocyte chemotactic protein-1 (MCP-1) are produced by pulmonary epithelial cells during lung transplantation and are considered to be involved in I/R injury. The volatile anaesthetic sevoflurane has been shown to exert a protective effect on I/R injury in various organs. We investigated the effect of sevoflurane on the inflammatory functions of pulmonary epithelial cells in vitro. Human normal small airway epithelial cells (SAEC) were incubated under anoxic conditions for 24 h with or without sevoflurane and then stimulated with tumour necrosis factor (TNF)-α under hyperoxic conditions for 5 h with or without sevoflurane. After incubation, IL-6, IL-8, and MCP-1 mRNA expression was analysed by quantitative real-time RT-PCR. The production of IL-6, IL-8, and MCP-1 was assayed by enzyme-linked immunosorbent assay, the effects of sevoflurane on inflammatory gene expression were examined by DNA microarray analysis, and the effects of sevoflurane on NF-κB-mediated inflammatory cytokine production were examined by immunoblotting. Sevoflurane suppressed TNF-α-induced IL-6, IL-8, and MCP-1 gene expression and the production of IL-6 and IL-8 in SAEC under anoxia/reoxygenation conditions. DNA microarray analysis indicated that sevoflurane modulated NF-κB-related gene expression. Sevoflurane significantly inhibited TNF-α-induced translocation of p65 NF-κB into the nucleus. Sevoflurane enhanced TNF-α-induced gene expression of inhibitor κB (IκB) but not of NF-κB. Sevoflurane suppressed the NF-κB-mediated production of pulmonary epithelial cell-derived inflammatory cytokines, including IL-6 and IL-8, which are capable of causing I/R injury.

MCP-1 Expression Is Specifically Regulated During Activation of Skeletal Repair and Remodeling

Monocyte chemotactic protein-1 (MCP-1) belongs to the CC chemokine superfamily and plays a critical role in the recruitment and activation of leukocytes during acute inflammation. We hypothesize that MCP-1 is also an important chemokine that regulates the recruitment and activation of bone cells required for skeletal repair and remodeling. We used the ulnar stress fracture (SFx) model, which allows investigation of focal remodeling with a known time course and precise anatomical location. SFx were created in the right ulna of female Wistar rats using cyclic end loading. Unloaded animals were used as a control. Rats were killed 4h and 1, 4, 7, and 14days after loading (n=10/group); RNA was extracted and converted to cDNA for quantitative PCR analysis using TaqMan gene expression assays. Four hours after loading, MCP-1 gene expression was increased ~30-fold (P<0.001), remained elevated at 24h (~12-fold, P<0.001), then declined by day 14. Relative to the contralateral limb, expression of the receptors CCR1 and CCR2 increased over the 14days, being significant by 4days for CCR1 and 14days for CCR2 (P<0.05). Other inflammation-related chemokines (RANTES, MIP1a) were not increased at these early time points. Using in situ hybridization and immunohistochemistry in separate animal groups (n=5/group, control, days 1, 4, 7), MCP-1 mRNA and protein were localized in periosteal osteoblasts associated with woven bone formation at the fracture exit point but not in osteocytes adjacent to the SFx. These data support an important role for MCP-1 in the early phase of SFx repair and activated remodeling.

Resistin mediates the hepatic stellate cell phenotype

To describe the role of resistin in liver fibrosis. For the in vivo animal study, Sprague Dawley rats were subjected to bile duct ligation (BDL) for 4 wk. Rat liver, adipose tissue (epididymal fat) and serum were analyzed for resistin expression. For the in vitro experiment, rat primary hepatic stellate cells (HSCs) and Kupffer cells (KCs) were used. HSCs were exposed to recombinant resistin, and collagen I, transforming growth factor β1, α smooth muscle actin, tissue inhibitor of metalloproteinase 1 and connective tissue growth factor expression were analyzed. Resistin gene and protein expression was quantified as was the expression of pro-inflammatory cytokines including tumor necrosis factor α (TNFα), interleukin (IL)-1, IL-6, IL-8 and monocyte chemotactic protein-1 (MCP-1). The effects of resistin on HSC proliferation, migration and apoptosis were determined. The effects of resistin on KCs were also investigated. Following BDL, rat epididymal fat and serum rather than liver showed higher resistin expression compared to control rats. In liver, resistin was expressed in quiescent HSCs and KCs. Resistin treatment resulted in enhancement of TNFα, IL-6, IL-8 and MCP-1 gene expression and increased IL-6 and MCP-1 protein in HSCs. Resistin activated HSC phospho-MAPK/p38, and p38 inhibition diminished IL-6 and MCP-1 expression. Furthermore, resistin facilitated HSC proliferation and migration, but decreased apoptosis which was via an IL-6 and MCP-1 mechanism. Finally, resistin-induced transforming growth factor β1 from KCs enhanced HSC collagen I expression. Resistin directly and indirectly modulates HSC behavior towards a more pro-fibrogenic phenotype.

Alterations in Pulse Pressure Affect Artery Function

Pulse pressure changes in response to cardiovascular diseases and interventions, but its effect on vascular wall structure and function is poorly understood. We examined the effect of increased or decreased pulse pressure on artery function, cellular function, and extracellular matrix remodeling. Porcine carotid arteries were cultured under non-pulsatile (100 mmHg), pulsatile (70-130 mmHg), or hyper-pulsatile pressure (50-150 mmHg) for 1 to 3 days. Vasomotor response, wall permeability, cell proliferation, apoptosis, extracellular matrix remodeling, and proteins involved in atherogenesis were examined. Our results showed that hyper-pulsatile pressure decreased the artery response to sodium nitroprusside, basal tone, and wall permeability after three days. Non-pulsatile pressure increased cell proliferation. Neither hyper-pulsatile nor non-pulsatile pressure caused a change in the extracellular matrix or in the expression of matrix metalloproteinase-2 (MMP-2), MMP-9, caveolin-1, or α-actin. Hyper-pulsatile pressure increased monocyte chemotactic protein-1 gene expression. Taken together, these changes indicate that pulse pressure has a limited effect on the artery immediately after its application. Specifically an increase in pulse pressure alters the artery tone and wall permeability while a decrease in pulse pressure alters cell proliferation. Overall these results provide insight into how the artery initially responds to changes in pulse pressure.

Induction of brain natriuretic peptide and monocyte chemotactic protein-1 gene expression by oxidized low-density lipoprotein: relevance to ischemic heart failure

Brain natriuretic peptide (BNP) and monocyte chemotactic protein-1 (MCP-1) are biomarkers of heart failure (HF). The aim of the present study was to determine the role of oxidized low-density lipoprotein (Ox-LDL) in the induction of these biomarkers and the signaling pathways involved in vitro. Incubation of HL-1 cardiomyocytes and human myocytes with Ox-LDL induced the expression of BNP and MCP-1 genes, while native LDL had no effect. When peroxides associated with Ox-LDL were reduced to hydroxides, the ability to induce BNP and MCP-1 gene expression was abolished. Furthermore, exposure of HL-1 cells to ischemic conditions alone had no effect on BNP gene expression, while ischemia followed by reperfusion resulted in increased expression of BNP gene. Inhibitors of ERK and JNK inhibited the induction of BNP. Signaling array results suggested that the induction of both MAPK and NF-κB pathways is involved in the induction of BNP by Ox-LDL. These results suggest that Ox-LDL or peroxidized lipids formed in oxidatively stressed myocytes during ischemia-reperfusion injury may play a role in the induction of BNP and MCP-1.

Sulforaphane inhibits de novo synthesis of IL-8 and MCP-1 in human epithelial cells generated by cigarette smoke extract

Chronic obstructive pulmonary disease (COPD) is currently the fifth leading cause of death worldwide. Exposure to cigarette smoke (CS) is the primary factor associated with the COPD development. CS activates epithelial cells to secrete chemokines such as interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) that recruit neutrophils and macrophages to the lung. These inflammatory cells then release additional chemokines and cytokines leading to chronic inflammation that initiates apoptosis in epithelial and endothelial cells and destruction of alveolar structure. Pulmonary epithelium responds to oxidative stress mediated by CS through activating NRF2-dependent pathways, leading to an increased expression of antioxidant and cytoprotective enzymes thereby providing a protective response against CS-induced lung injury. We hypothesized that activating NRF2-dependent cytoprotective gene expression with sulforaphane (SFN) affords protection against CS-induced lung damage by inhibiting chemokine production. Results indicate that in the human BEAS-2B epithelial cell line, 5 μM SFN activated NRF2-dependent gene expression by triggering the translocation of NRF2 to the nucleus and significantly increased the expression of NRF2-dependent genes such as NADPH quinone oxidoreductase-1, heme oxygenase-1, and glutamate cysteine ligase modulatory subunit. Cigarette smoke extract (CSE) exposure of BEAS-2B cells significantly increased production of both IL-8 and MCP-1. Production of both chemokines was significantly reduced with SFN given prior to CSE; SFN inhibited IL-8 and MCP-1 gene expression at the transcription level. Our results indicate that activating NRF2 pathways with SFN inhibits CSE-induced chemokine production in human epithelial cells. However, the mechanism by which the production of chemokines is inhibited through SFN still remains to be elucidated. SFN may enhance NRF2 transcriptional activity resulting in the inhibition of proinflammatory pathways such as NF-κB.

The ethanolamide metabolite of DHA, docosahexaenoylethanolamine, shows immunomodulating effects in mouse peritoneal and RAW264.7 macrophages: evidence for a new link between fish oil and inflammation

Several mechanisms have been proposed for the positive health effects associated with dietary consumption of long-chain n-3 PUFA (n-3 LC-PUFA) including DHA (22:6n-3) and EPA (20:5n-3). After dietary intake, LC-PUFA are incorporated into membranes and can be converted to their corresponding N-acylethanolamines (NAE). However, little is known on the biological role of these metabolites. In the present study, we tested a series of unsaturated NAE on the lipopolysaccharide (LPS)-induced NO production in RAW264.7 macrophages. Among the compounds tested, docosahexaenoylethanolamine (DHEA), the ethanolamide of DHA, was found to be the most potent inhibitor, inducing a dose-dependent inhibition of NO release. Immune-modulating properties of DHEA were further studied in the same cell line, demonstrating that DHEA significantly suppressed the production of monocyte chemotactic protein-1 (MCP-1), a cytokine playing a pivotal role in chronic inflammation. In LPS-stimulated mouse peritoneal macrophages, DHEA also reduced MCP-1 and NO production. Furthermore, inhibition was also found to take place at a transcriptional level, as gene expression of MCP-1 and inducible NO synthase was inhibited by DHEA. To summarise, in the present study, we showed that DHEA, a DHA-derived NAE metabolite, modulates inflammation by reducing MCP-1 and NO production and expression. These results provide new leads in molecular mechanisms by which DHA can modulate inflammatory processes.