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CXC chemokine ligand 8 (CXCL8) is highly expressed in many human tumors including colorectal cancer, and it can promote the malignant progression of tumors. It was reported that M2 macrophages were abundant in colorectal cancer microenvironment, but whether CXCL8 affects the infiltration of M2 macrophages and its potential mechanism are not yet clear. The study aimed to investigate the effect of CXCL8 on M2 macrophage infiltration and chemotaxis in the colorectal cancer. Firstly, we analyzed the CXCL8 expression and immune cell infiltration in human colorectal cancer tissues from TCGA RNA-seq data. The expression of CXCL8 was verified by immunohistochemistry in tissues obtained from Shanxi Provincial Cancer Hospital. Then, Western blot and qRT-PCR were employed to detect CXCL8 expression in five colorectal cancer cell lines. THP-1 cells were allowed to differentiate into M2 macrophages via the phorbol myristate acetate (PMA) and IL-4 treatment, followed by detection of the chemotaxis of M2 macrophages towards HCT116, SW480 and CXCL8-HCT116, CXCL8-SW480 cell lines. HCT116 and SW480 cells were treated with interleukin 1β (IL-1β) to detect the expression of CXCL8, and co-cultured with M2 macrophages to analyze the chemotactic activity. The results revealed that the expression of CXCL8 was increased in pairs of CRC tissues versus normal adjacent tissues, and there were more M2 macrophage infiltration in cancer tissues with high expression of CXCL8. The mRNA and protein expression of CXCL8 in HCT116 and SW480 were increased after the IL-1β treatment (P < 0. 05). We confirmed that CXCL8 is a chemotactic factor for M2 macrophages by transwell assays (P<0. 05). In conclusion, CXCL8 in colorectal cancer cells can be induced by IL-1β in colorectal cancer cells and the upregulation of CXCL8 can promote the chemotaxis of M2 macrophages. The massive infiltration of M2 macrophages in colorectal cancer microenvironment may be related with the increased expression of CXCL8.
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Objective To investigate the effect of IFN-γ on the proliferation and migration of esophageal squamous cell carcinoma cell line Eca9706 and related mechanism. Methods Cells were cultured in vitro and treated with interferon-γ. Cell morphology changes were observed under microscope, cell proliferation ability was detected by CCK-8 experiment, and cell migration ability was detected by cell scratch experiment and Transwell experiment. Real-time PCR method was used to detect the expression efficiency of chemokine CXCL8 (interleukin 8), and the ELISA experiment was used to detect the change of CXCL8 secretion. Results Compared with the blank control group, Eca9706 cells treated with different concentrations of interferon-γ did not change significantly in cell morphology. CCK8 experiment confirmed that the proliferation ability of Eca9706 cells after IFN-γ treatment was significantly reduced (P < 0.01). Cell scratch experiment found that IFN-γ significantly decreased the migration ability of Eca9706 cells (P < 0.01). Transwell experiment showed that after IFN-γ treatment, the migration ability of Eca9706 cells was significantly inhibited (P < 0.01). CXCL8 gene expression level in Eca9706 cells treated with interferon-γ was significantly down-regulated (P < 0.01), and the amount of CXCL8 secretion significantly reduced (P < 0.05). Conclusion Interferon-γ can inhibit the proliferation and migration of esophageal cancer cell line Eca9706, which may be related to its inhibition of the expression and secretion of CXCL8.
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Objective:To investigate the efficacy of dexmedetomidine combined with transversus abdominis plane block in laparoscopic radical resection of colorectal cancer and its effect on serum CXCL8 level.Methods:A total of 72 patients who planned to undergo laparoscopic radical resection of colorectal cancer in Changzhou Hospital of Traditional Chinese Medicine from March 2017 to March 2021 were selected as the research subjects, and they were divided into transversus abdominis plane block anesthesia group (group A) and dexmedetomidine combined with transversus abdominis plane block anesthesia group (group B) by random number table method with 36 cases in each group. The operation time, intraoperative blood loss, fluid supplementation, visual analogue score (VAS) and Ramsay sedation score at different times after operation, and serum CXCL8 level after operation were compared between the two groups of patients. The adverse reactions of the two groups of patients were compared.Results:There was no significant difference in operation time and fluid supplementation between the two groups (both P > 0.05). The VAS scores of patients in group B at 4, 8 and 24 hours after operation were lower than those in group A [(2.8±0.6) points vs. (4.2±1.2) points, (2.1±1.0) points vs. (3.4±1.1) points, (1.8±0.4) points vs. (2.5±0.7) points, all P < 0.05], and the Ramsay sedation scores of patients in group B at 4, 8 and 24 hours after operation were higher than those in group A [(4.3±1.2) points vs. (2.7±0.7) points, (3.5±1.1) points vs. (2.2±1.0) points, (2.4±0.9) points vs. (1.6±0.6) points, all P<0.05]. Serum CXCL8 levels of patients in group B at 2, 24 and 48 hours after operation were lower than those in group A [(78±16) ng/ml vs. (87±19) ng/ml, (68±14) ng/ml vs. (75±15) ng/ml, (52±10) ng/ml vs. (61±13) ng/ml, all P<0.05]. The incidence rates of adverse reactions in group A and group B were 8.3% (3/36) and 13.9% (5/36), and the difference was not statistically significant ( P > 0.05). Conclusions:Dexmedetomidine infusion during laparoscopic radical resection of colorectal cancer under general anesthesia combined with transversus abdominis plane block can help reduce postoperative pain, increase sedative effect, and reduce serum CXCL8 level.
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ObjectiveTo study the effects of Chinese herbal compound Youguiwan on angiogenesis of rats with ovarian dysfunction caused by natural aging and its relationship with chemokine interleukin 8 （CXCL8）/CXC chemokine receptor 1/2 （CXCR1/2） signaling pathway， angiopoietin 1 （Ang-1）， and angiopoietin 2 （Ang-2）， so as to explore its mechanism in improving the ovarian function. MethodFifty six female SD rats were randomly divided into the young control group （n=8） and modeling group （n=48， ovarian dysfunction caused by natural aging）. Rats in both the young control and modeling groups were routinely fed， during which the ones in the modeling group underwent exfoliative cytology of vaginal smears for five to seven days. The ones presented with prolonged estrous cycle， followed by continuous estrus and repeated pseudopregnancy revealed by vaginal cytology during four consecutive estrous cycles indicated early aging， and the young rats with keratinocyte proliferation index higher than 50% for 10 consecutive days were classified into the young control group. The successfully modeled rats were randomly divided into the early-aged group， estrogen （65 μg·kg-1·d-1） group， Zuoguiwan （33 g·kg-1·d-1） group， as well as the low-， medium-， and high-dose （1.2， 2.4， 4.8 g·kg-1·d-1） Youguiwan groups. Rats in the young control group and the early-aged group were gavaged with the same volume of normal saline for 30 days. After the experiment， the morphological changes in rat ovary were observed by hematoxylin-eosin （HE） staining. The protein expression levels of chemokines CXCL8， CXCR1， CXCR2， Ang-1， and Ang-2 in rat ovary were detected by Western blotting and immunohistochemistry， and the mRNA expression levels of CXCL8， CXCR1， CXCR2， Ang-1， and Ang-2 by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultCompared with the young control group， the early-aged group exhibited reduced number of growing follicles， corpus luteum， and blood vessels at all levels， elevated atretic follicles （P<0.01）， up-regulated protein and mRNA expression of CXCL8， CXCR1， and CXCR2 in the ovarian tissue （P<0.01）， and down-regulated Ang-1 and Ang-2 protein and mRNA expression （P<0.05）. Compared with the early-aged group， each medication remarkably increased the number of growing follicles， corpus luteum， and blood vessels （P<0.05）， lowered the number of atretic follicles （P<0.05）， down-regulated the protein and mRNA expression levels of CXCL8， CXCR1， and CXCR2 in the ovarian tissue （P<0.05）， and up-regulated the protein and mRNA expression levels of Ang-1 and Ang-2 （P<0.05）. ConclusionYouguiwan down-regulates the levels of CXCL8， CXCR1， and CXCR2 in rat ovary and up-regulates the levels of Ang-1 and Ang-2 to promote ovarian angiogenesis and improve rat ovarian function.
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Objective To explore sensitive indicators for the initiation,development,and metastasis of gastric cancer and to provide objective evidence for the early diagnosis,treatment,and progression monitoring of gastric cancer.Methods A total of 108 patients with gastric cancer were enrolled in this study.The expression of interleukin receptor 1 (CXCR1)in samples from gastric cancer and adjacent tissues was detected by immunohistochemistry and patient clinical data were collected for correlation analysis.Logistic regression analysis of the 5-year survival rate of patients was conducted.Results The positive CXCR1 expression rate in gastric neoplasm tissues was significantly higher than that in adjacent tissues.Nevertheless,CXCR1 was correlated with tumor differentiation (P =0.017),TNM staging (P =0.006),and the existence of lymphatic metastasis (P =0.035).The overall survival rate (P =0.043) and recurrence-free survival rate (P=0.029) of patients with positive CXCR1 were lower than those of patients with negative CXCR1.Conclusions CXCR1 expression levels increase in gastric neoplasm tissues and are associated with tumor differentiation,TNM staging,and lymphatic metastasis.Positive CXCR1 is correlated with poor prognosis and has the potential to serve as one of clinical prognostic indicators.
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Objective To detect the expression of CXCL8 in colorectal tumor tissues and marginal tissues,and analyze the correlation between CXCL8 and clinicopathological parameters. Methods Sixty colorectal cancer specimens and the mar-ginal tissues were collected from the colorectal cancer patients who were surgically removed in gastrointestinal surgery in Zhengzhou Central Hospital Affiliated to Zhengzhou University from November 2014 to November 2016 and were all confirmed by pathology. The expression of CXCL8 protein and mRNA in colorectal tumor tissues and marginal tissues was detected by im-munohistochemistry and real time fluorescent quantitation polymerase chain reaction respectively. Results The expression of CXCL8 mRNA in colorectal tumor tissues and marginal tissues was 157. 6 ± 26. 2 and 42. 7 ± 9. 6. The expression of CXCL8 mRNA in colorectal tumor tissues was higher than that in the marginal tissues(P < 0. 05). The positive expression rate of CXCL8 protein in the colorectal tumor tissue and marginal tissue was 83. 3％(50 / 60)and 25. 0％(15 / 60),respectively. The positive expression rate of CXCL8 in the colorectal tumor tissues was higher than that in the marginal tissues(χ2 = 41. 10,P <0. 01). The expression of CXCL8 in colorectal tumor tissues was correlated with lymph node metastasis and tumor staging(P <0. 01). The expression of CXCL8 in colorectal tumor tissues had not correlation with age,gender and differentiated degree colo-rectal tumor(P > 0. 05). Conclusion CXCL8 has high expression in colorectal cancer tissues,and it is closely related to the condition of lymph node metastasis and tumor staging in patients with colorectal cancer.
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Objective To investigate the expression and clinical significance of CXC chemokine receptors 1 and 2 (CXCR1 and CXCR2) and CXCL8 in peripheral blood mononuclear cells (PBMCs) and liver biopsy tissues from patients with primary hepatocellular carcinoma (PHC).Methods Serum specimens were collected from 36 patients with PHC, 30 patients with liver cirrhosis and 28 healthy subjects.Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to measure the expression of CXCR1, CXCR2 and CXCL8 at mRNA level in PBMCs.Streptavidin-perosidase (SP) immunohistochemistry was used to detect the expression of CXCR1, CXCR2 and CXCL8 at protein level in liver biopsy tissues.Levels of C-reactive protein (CRP), alpha-fetoprotein (AFP) and ferritin (FER) in the serum specimens were detected by chemiluminescence immunoassay.Then the correlations between these markers were analyzed.Results All of the results showed that the expression of CXCR1, CXCR2 and CXCL8 at mRNA level in PBMCs from the PHC group were higher than those of the healthy control group (P<0.01) as well as those of the liver cirrhosis group (P<0.05).Up-regulated expression of CXCR1, CXCR2 and CXCL8 in patients with PHC were associated with the depth of tumor invasion, lymph node or distant metastasis, clinical stage and levels of CRP, AFP and FER in serum (P<0.05).The expression of CXCR1, CXCR2 and CXCL8 at protein level in liver biopsy tissues were also significantly increased in the PHC group in comparison with those of the healthy control group as indicated by the result of SP immunohistochemistry (P<0.05).Conclusion Levels of CXCR1, CXCR2 and CXCL8 in the patients with PHC are significantly increased and positively correlated with the levels of AFP, FER and CRP in serum, suggesting that the signal transduction process mediated by CXCR1, CXCR2 and their common ligand CXCL8 may play a key role in the pathological process of PHC.This study may provide a potential new strategy for immune intervention in hepatocellular cancer.
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Objective:To investigate the expression levels of CXCR1,CXCR2 and their common ligand CXCL8 in peripheral blood mononuclear cells (PBMCs) and liver biopsy from the patients with hepatitis B related hepatocellular carcinoma and their clinical significances.Methods:Quantitative real time polymerase chain reaction (qRT-PCR) was used to detect the mRNA levels of CXCR1,CXCR2,CXCL8 in the peripheral blood mononuclear cells of thirty-six hepatitis B related hepatocellular carcinoma and the protein levels of CXCR1 and CXCR2 and CXCL8 in liver biopsy were detected by SP immunohistochemical method.The level of C-reactive protein in serum was determined by chemiluminescence immunoassay respectively.Then,the correlations between CRP and the mRNA of CXCR1,CXCR2 and CXCL8 were analyzed.Results:The mRNA levels of CXCR1 (0.952 7±0.197 2),CXCR2 (0.896 9±0.173 0),CXCL8 (1.771 9±0.248 9) in the PBMCs of hepatitis B related hepatocellular carcinoma were significantly higher than those in controls (P＜0.01).And the protein levels of CXCR1,CXCR2 and CXCL8 were also obviously increased in liver biopsy of hepatitis B related hepatocellular carcinoma (P＜0.05).In addition,there was positive correlations between the level of serum C-reactive protein and the mRNA expression of CXCR1 (r =0.54,P＜0.01),CXCR2 (r =0.49,P＜0.01),CXCL8 (r =0.63,P＜0.01).Conclusion:The levels of CXCR1,CXCR2 and CXCL8 significantly increased in hepatitis B related hepatocellular carcinoma patients and positively correlated with serum CRP,suggesting that CXCR1,CXCR2 and their common ligand CXCL8 signal transduction process may play a key role in the pathological process of hepatitis B related hepatocellular carcinoma,which may provide a new direction for the immune intervention therapy of hepatocellular carcinoma.
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ABSTRACT An increasing body of evidence suggests that the use of probiotic bacteria is a promising intervention approach for the treatment of inflammatory diseases with a polymicrobial etiology. P. gingivalis has been noted to have a different way of interacting with the innate immune response of the host compared to other pathogenic bacteria, which is a recognized feature that inhibits CXCL8 expression. Objective The aim of the study was to determine if P. gingivalis infection modulates the inflammatory response of gingival stromal stem cells (G-MSSCs), including the release of CXCL8, and the expression of TLRs and if immunomodulatory L. rhamnosus ATCC9595 could prevent CXCL8 inhibition in experimental inflammation. Material and Methods G-MSSCs were pretreated with L. rhamnosus ATCC9595 and then stimulated with P. gingivalis ATCC33277. CXCL8 and IL-10 levels were investigated with ELISA and the TLR-4 and 2 were determined through flow cytometer analysis. Results CXCL8 was suppressed by P. gingivalis and L. rhamnosus ATCC9595, whereas incubation with both strains did not abolish CXCL8. L. rhamnosus ATCC9595 scaled down the expression of TLR4 and induced TLR2 expression when exposed to P. gingivalis stimulation (p<0.01). Conclusions These findings provide evidence that L. rhamnosus ATCC9595 can modulate the inflammatory signals and could introduce P. gingivalis to immune systems by inducing CXCL8 secretion.
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Humains , Jeune adulte , Interleukine-8/analyse , Porphyromonas gingivalis/immunologie , Probiotiques/pharmacologie , Lacticaseibacillus rhamnosus/physiologie , Cellules souches mésenchymateuses/microbiologie , Parodontite/microbiologie , Adhérence bactérienne/immunologie , Test ELISA , Cellules cultivées , Interleukine-8/immunologie , Interféron gamma/analyse , Interféron gamma/immunologie , Interleukine-10 , Statistique non paramétrique , Récepteur de type Toll-4/analyse , Récepteur de type Toll-4/immunologie , Cytométrie en flux , Immunité innéeRésumé
Objective:To study the expression of CXCL 8 in the serum and CXCL8 mRNA in the peripheral blood mononuclear cells(PBMCs) of the children with Mycoplasma pneumoniae pneumonia (MPP) and its clinical significance.Methods: Forty-eight children(severe cases 12,light cases 36) with MPP were recruited from October 2013 to March 2015 in the Maternal and Child Health-Care Hospital of Huainan.The concentration of the CXCL8 in serum and the level of CXCL8 mRNA in the PBMCs were measured by enzyme linked immunosorbent assay ( ELISA) and polymerase chain reaction ( PCR).Taking GAPDH as the internal reference ,the ratio of lgcDNA/lgGAPDH was regarded as the extreme level of CXCL 8 mRNA.Results: The serum level of CXCL8 and expression of CXCL8 mRNA in PBMCs in the children with MPP were ( 298.917 ±51.860 ) pg/ml and ( 1.848 ±0.525 ) lgcDNA/lgGAPDH.Compared with the normal control ,there were significant differences between the two groups ( P0.05).However, the expression of CXCL8 mRNA in peripheral blood of the children with severe illness was significantly higher than those in light cases (P<0.05).Intravenous infusion of Erythromycin was provided in the acute phase for seven to ten days ,so that the children′s condition could be significantly controlled , and the symptoms of pulmonary inflammation were also relieved .Followed by the use of sequential therapy of Azithromycin for about two to three weeks ,the children′s condition were gradually from acute stage to recovery stage .At this time,the CXCL8 and its mRNA levels in peripheral blood of the sick children were all significantly decreased comparing with those in the acute stage(P<0.05).Conclusion: The expression of CXCL8 and its mRNA were increased in the peripheral blood of the sick children with Mycoplasma pneumonia ,and also correlated with the severity of the disease .CXCL8 can participate in the pathogenesis of Mycoplasma pneumonia ,and has a certain cue effect on the severity and prognosis of the disease .Azithromycin can reduce the content of CXCL8 in serum of the sick children via the pathway of inhibiting the proliferation of Mycoplasma pneumoniae ,and down regulate the expression of mRNA ,so that the immune injury mediated by Mycoplasma pneumoniae may be gradually inhibited .
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Peptidoglycan (PG), the gram positive bacterial pathogen-associated molecular patterns (PAMP), is detected in a high proportion in macrophage-rich atheromatous regions, and expression of chemokine CXCL8, which triggers monocyte arrest on early atherosclerotic endothelium, is elevated in monocytes/macrophages in human atherosclerotic lesion. The aim of this study was to investigate whether PG induced CXCL8 expression in the cell type and to determine cellular signaling pathways involved in that process. Exposure of THP-1 cell, human monocyte/macrophage cell line, to PG not only enhanced CXCL8 release but also profoundly induced il8 gene transcription. PG-induced release of CXCL8 and induction of il8 gene transcription were blocked by OxPAPC, an inhibitor of TLR-2/4 and TLR4, but not by polymyxin B, an inhibitor of LPS. PG-mediated CXCL8 release was significantly attenuated by inhibitors of PI3K-Akt-mTOR pathways. PKC inhibitors, MAPK inhibitors, and ROS quenchers also significantly attenuated expression of CXCL8. The present study proposes that PG contributes to inflammatory reaction and progression of atherosclerosis by inducing CXCL8 expression in monocytes/macrophages, and that TLR-2, PI3K-Akt-mTOR, PKC, ROS, and MAPK are actively involved in the process.
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Humains , Athérosclérose , Lignée cellulaire , Endothélium , Interleukine-8 , Monocytes , Peptidoglycane , Polymyxine BRésumé
Objective:To study on the levels of CXCL8 and its receptors (CXCR1 and CXCR2) in peripheral blood neutrophils of the patients with chronic hepatitis B.Methods:The neutrophils were isolated and purified by neutrophil isolation medium,and the loads of HBV-DNA in neutrophils were detected by PCR,and the levels of HBeAg in serum were measured by ELISA.The patients were divided into different groups according to the detective results so that the expressions of CXCL8 and its receptors ( CXCR1,CXCR2) in neutrophils were detected by the methods of streptavidin-biotin complex ( SABC ) immunocytochemistry stain.Results:The data of SABC immunocytochemical stain showed that the positive color of CXCL8 was mainly located in the cytoplasm of PMNs.However,the most positive color of CXCR1and CXCR2 was mainly expressed in the cytoplasm and cell membrane.Interestingly,the deeper immune coloring of CXCL8 and CXCR1, and relatively shallow immune coloring of CXCR2 were explored in the group with positive of HBeAg.The similar detective results also had been found in the cases with positive of HBV DNA in neutrophils.Compared with the normal control group,the levels of CXCL8 and CXCR1 in the patients were significantly increased ( P0.05).Conclusion:After neutrophils occult infected by HBV,not only the secretion of CXCL8 can be promoted, but also the expression of CXCR1 will be further increased.The data of immunohistochemical staining have been shown that the color degree of CXCL8 and its receptors ( CXCR1, CXCR2 ) are positive correlation to the level of HBeAg and the loads of HBV DNA.More PMNs can be chemotactic attraction to lesion so as to participate in the local inflammatory injury and tissue repair via the interactive pathway of the high expression of CXCR1 on surface of neutrophils with CXCL8.
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Objective To investigate the level of chemotactic factors(CXCL5 and CXCL8)in hepatic fibrosis and cirrhosis tissue.Methods Hepatic tissues were obtained from 9 patients with hepatic hemangioma (hepatic hemangioma group),10 patients with liver fibrosis(liver fibrosis group)and 11 patients with liver cirrhosis(1iver cirrhosis group)at Nanfang Hospital from May 2008 to May 2009.The contents of CXCL5 and CXCL8 in hepatic tissue were assayed by ELISA.All data were analyzed by one-way ANOVA,Pearson rank correlation or Spearman correlation.Results The contents of CXCL5 and CXCL8 were(0.8±0.7)ng/g and(6.2±3.7)ng/g in hepatic hemangioma group,(2.0±2.0)ng/g and(11.6±3.5)ng/g in liver fibrosis group and (17.1±4.8)ng/g and(12.3±3.9)ng/g in liver cirrhosis group,with significant difference among the 3 groups (F=60.050,7.690,P＜0.05).The expression of CXCL5 was correlated with the content of alanine aminotransferase(ALT),aspartate aminotransferase(AST)and prothrombin time(PT)(r=0.502,0.468,0.523,P＜0.05):the expression of CXCL8 was correlated with the content of ALT,AST.total bilirubin and PT(r=0.477,0.504,0.537,0.431,P＜0.05).Conclusions With the aggravation of hepatic fibrosis,the contents of CXCL5 and CXCL8 are increased with different patterns.The changes of CXCL5 and CXCL8 are related with the injury of liver,but the changes of CXCL5 and CXCL8 do not correspond with the degree of the injury of liver.
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BACKGROUND: 15d-PGJ2 has been known to act as an anti-inflammatory agent and has anti-hypertensive effects. As a result of these properties, we examined the effect of 15d-PGJ2 on the LPS-induced IL-8/CXCL8 mRNA expression in VSMCs from SHR. METHODS: Effect and action mechanism of 15d-PGJ2 on the expression of LPS-induced IL-8/CXCL8 mRNA in VSMCs from SHR and WKY were examined by using real-time polymerase chain reaction, electrophoretic mobility shift assay for NF-kappaB avtivity, Western blotting analysis for ERK and p38 phosphorylation and flow cytometry for NAD(P)H oxidase activity. RESULTS: 15d-PGJ2 decreased the expression of LPS-induced IL-8/CXCL8 mRNA in WKY VSMCs, but increased the expression of LPS-induced IL-8/CXCL8 mRNA in SHR VSMCs. The upregulatory effect of 15d-PGJ2 in SHR VSMCs was mediated through PPAR gamma, and dependent on NF-kappaB activation and ERK phosphorylation. However, inhibition of the p38 signaling pathway augmented the upregulatory effect of 15d-PGJ2 on LPS-induced IL-8/CXCL8 mRNA. A NAD(P)H oxidase inhibitor inhibited the upregulatory effect of 15d-PGJ2 on LPS-induced IL-8/CXCL8 mRNA expression in SHR VSMCs, and an increase in NAD(P)H oxidase activity was detected in SHR VSMCs treated with 15d-PGJ2/LPS. CONCLUSION: Our results indicate that the upregulatory effect of 15d-PGJ2 on LPS-induced IL-8/CXCL8 expression in SHR VSMCs is mediated through the PPAR gamma and ERK pathway, and may be related to NAD(P)H oxidase activity. However, p38 inactivation may also play an important role in 15d-PGJ2/LPS-induced IL-8/CXCL8 expression in SHR VSMCs.
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Technique de Western , Test de retard de migration électrophorétique , Cytométrie en flux , Système de signalisation des MAP kinases , Muscles lisses vasculaires , NADPH oxidase , Facteur de transcription NF-kappa B , Phosphorylation , Récepteur PPAR gamma , Prostaglandine D2 , Rats de lignée SHR , Réaction de polymérisation en chaine en temps réel , ARN messagerRésumé
Upon contact with airway epithelial cells, mycobacteria activate several signal transduction events that are required for induction of inflammatory cytokines/chemokines. In this study, we found that Mycobacterium tuberculosis (Mtb)induced reactive oxygen species (ROS) production is essential for the expression of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and CXC-chemokine ligand (CXCL) 8 through the activation of mitogen-activated protein kinases [MAPKs; extracellular signal-regulated kinase (ERK) 1/2 and p38 MAPK] in A549 cells representing alveolar epithelial cells. We observed that Mtb rapidly enhanced ROS production after stimulation in a toll-like receptor (TLR) 2-dependent manner. In addition, Mtb triggered ERK1/2 and p38 MAPK signaling pathways which were dependent on ROS generation in A549 cells. Moreover, Mtb stimulation significantly increased the secretion of TNF-alpha, IL-6, and CXCL8 over that in untreated controls. Pretreatment of A549 cells with the antioxidant, N-acetylcysteine and the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor, diphenylene iodonium, substantially inhibited Mtb-induced production of TNF-alpha, IL-6, and CXCL8. Studies using inhibitors selective for ERK1/2 and p38 MAPK pathways showed that both pathways play an essential role in the induction of TNF-alpha, IL-6, and CXCL8 at transcriptional levels in A549 cells. Collectively, our findings indicate the critical role of TLR2-dependent ROS in the Mtb-induced inflammatory cytokine/chemokine production in alveolar epithelial cells through MAPK-dependent signaling pathways.
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Acétylcystéine , Cellules épithéliales , Interleukine-6 , Interleukines , Mitogen-Activated Protein Kinases , Mycobacterium , Mycobacterium tuberculosis , NADP , Composés onium , Oxidoreductases , Oxygène , p38 Mitogen-Activated Protein Kinases , Phosphotransferases , Protein kinases , Espèces réactives de l'oxygène , Transduction du signal , Récepteurs de type Toll , Facteur de nécrose tumorale alphaRésumé
BACKGROUND: We previously demonstrated remarkable differences in the expression of IL-8/CXCL8 in aortic tissues and vascular smooth muscle cells (VSMC) from spontaneously hypertensive rats (SHR) compared to VSMC from normotensive Wistar-Kyoto rats (WKY). In the present study, we investigated the direct effect of IL-8/CXCL8 on expression of 12-lipoxygenase (LO), a hypertensive modulator, in SHR VSMC. METHODS: Cultured aortic VSMC from SHR and WKY were used. Expression of 12-LO mRNA was determined by real-time polymerase chain reaction. Phosphorlyation of ERK1/2 and production of 12-LO and angiotensin II subtype 1 (AT1) receptor were assessed by Western blots. IL-8/CXCL8-stimulated DNA synthesis was determined by measuring incorporation of [3H]-thymidine. And effect of IL-8/CXCL8 on vascular tone was determined by phenylephrine-induced contraction of thoracic aortic rings. RESULTS: Treatment with IL-8/CXCL8 greatly increased 12-LO mRNA expression and protein production compared to treatment with angiotensin II. IL-8/CXCL8 also increased the expression of the AT1 receptor. The increase in 12-LO induced by IL-8/CXCL8 was inhibited by treatment with an AT1 receptor antagonist. The induction of 12-LO mRNA production and the proliferation of SHR VSMC by IL-8/CXCL8 was mediated by the ERK pathway. The proliferation of SHR VSMC and the vascular contraction in the thoracic aortic ring, both of which were induced by IL-8/CXCL8, were inhibited by baicalein, a 12-LO inhibitor. CONCLUSION: These results suggest that the potential role of IL-8/CXCL8 in hypertensive processes is likely mediated through the 12-LO pathway.
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Animaux , Rats , Angiotensine-II , Arachidonate 12-lipoxygenase , Technique de Western , Contrats , ADN , Flavanones , Système de signalisation des MAP kinases , Muscles lisses vasculaires , Rats de lignée SHR , Réaction de polymérisation en chaine en temps réel , ARN messagerRésumé
Endothelin-1 (ET-1) has been characterized as a potent vasoconstrictor secreted by the endothelium, and play a major role in the regulation of vascular tone. It has been also known to participate in inflammatory reactions. The production of ET-1 by macrophages during infection and inflammation is related to tissue perfusion and leukocyte extravasation. The aim of this study is to investigate the role of IL-8/CXCL8, as a major inflammatory chemokine, for ET-1 expression in macrophges. Expression of ET-1 mRNA in mouse peritoneal macrophages (PeM phi) was weaker than that in vascular smooth muscle cells (VSMCs) from spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY). However, expression of IL-8/CXCL8-induced ET-1 mRNA in PeM phi was much more stronger than that in SHR and WKY VSMCs. Maximum expression of ET-1 mRNA was observed at 50 ng/ml dose of IL-8/CXCL8 and occurred at 2 h after addition of IL-8/CXCL8. Expression of ET-1 by IL-8/CXCL8 was dependent on NF-kappaB activation and ERK1/2 phosphorylation. Baicalein, a 12-lipoxygenase (LO) inhibitor, inhibited the expression of IL-8/CXCL8-induced ET-1 mRNA. This inhibitory action of baicalein was mediated via ERK1/2 inactivation. Induction of 12-LO mRNA by IL-8/CXCL8 and expression of ET-1 mRNA by 12-LO metabolite, 12(S)-HETE were also detected. The expression of IL-8/CXCL8-induced ET-1 mRNA was not detected in PeM phi transfected with 12-LO siRNA. These results suggest that IL-8/CXCL8 can act as one of main inducers of ET-1 in vascular inflammatory reactions, and ET-1 expression by IL-8/CXCL8 is related to 12-LO pathway in PeM phi.