Functional characterization of histamine receptor subtypes in a human bronchial epithelial cell line

Histamine is a well-known mediator eliciting a broad range of responses in different cell types. Four different subtypes of G protein-coupled histamine receptors (H1-H4) have been cloned and pharmacologically characterized. However, involvement of the different histamine receptor subtypes in immunomodulatory functions of bronchial epithelium has only been investigated marginally. The expression and function of histamine receptor subtypes on the human bronchial epithelial cell line BEAS-2B was analyzed by PCR, intracellular Ca++ -measurements and ELISA. We show mRNA expression of the histamine receptor subtypes H1, H2, and H3, but not H4 in the human bronchial epithelial cell line BEAS-2B. Using intracellular Ca++ -measurements, we demonstrated functional expression of the H1 and H3 receptors. To characterize the biological properties of histamine in airway epithelial biology, we also investigated its effects on cytokine secretion by BEAS-2B cells. Thereby, we were able to show up-regulation of the proinflammatory mediators IL-6 and CXCL8/ IL-8 via activation of the H1, H2 and H3 receptor subtypes. The Th1 cytokines CXCL9/MIG and CXCL10/IP-10 and the chemokine CCL5/RANTES were regulated in a distinct manner: Whereas histamine inhibited the IFN-gamma/TNF-alpha-induced secretion of MIG via the histamine receptor subtypes H1, H2, and H3, the histamine-induced suppression of RANTES was due to activation of the H2 and H3 receptors, while reduction of cytokine-triggered IP-10 secretion was mediated only by triggering the H2 receptor. In summary our data provide evidence that histamine released during allergic lung diseases exerts regulatory influence on airway epithelial cells.     

Interleukin-5 enhances eosinophil adhesion to bronchial epithelial cells

BACKGROUND: Eosinophil-bronchial epithelial cell interactions are thought to be central to the pathogenesis of asthma, both in terms of the epithelium as a source of pro-inflammatory mediators and as a target for eosinophil-mediated damage. We have therefore investigated adhesion interactions between these two cell types. OBJECTIVES: To determine the role of eosinophil and epithelial activation on eosinophil adhesion to bronchial epithelium and to characterize the adhesion receptors mediating eosinophil adhesion. METHODS: Eosinophils were purified from human peripheral blood by immunomagnetic selection and adhesion to confluent cultures of the airway epithelial cell lines A549 and BEAS-2B was studied. RESULTS: Stimulation of A549 cells with TNFalpha, IFNgamma or a combination of 50 ng/mL of TNFalpha, IFNgamma and IL-1 (cytomix) did not effect eosinophil binding despite an increase in ICAM-1 expression. Similarly stimulation of eosinophils with PAF or IL-5 had no effect on eosinophil binding to medium- or cytokine-treated A549 cells. In contrast stimulation of BEAS-2B cells with cytomix caused a significant increase in eosinophil adhesion. This was associated with an increase in expression of ICAM-1 and induced expression of VCAM-1. Treatment of eosinophils with Mn2+ and IL-5 but not eotaxin, RANTES or PAF also significantly enhanced eosinophil adhesion to medium-treated BEAS-2B cells. Using blocking mAbs we were able to demonstrate that the increased adhesion resulting from stimulation of eosinophils or BEAS-2B cells was in both cases mediated by a combination of CD18 and alpha4 integrins. CONCLUSIONS: This study demonstrates a selective role for IL-5 in mediating integrin-dependent eosinophil adhesion to airway epithelium and once again emphasizes the importance of this cytokine in controlling eosinophil activation in diseases such as asthma.     

Combination therapy: Synergistic suppression of virus-induced chemokines in airway epithelial cells

Viruses are associated with the majority of exacerbations of asthma and chronic obstructive pulmonary disease. Virus induction of neutrophil and lymphocyte chemokines in bronchial epithelium is important in exacerbation pathogenesis. Combined corticosteroid/beta2 agonists synergistically suppress airway smooth muscle chemokine production. Because bronchial epithelium expresses glucocorticoid and beta2 receptors, we investigated whether combination therapy can synergistically suppress rhinovirus-induced bronchial epithelial cell neutrophil (CXCL5, CXCL8) and lymphocyte (CCL5, CXCL10) chemokine production. We investigated modulation of rhinovirus- and IL-1beta-induced bronchial epithelial cell chemokine production by salmeterol and fluticasone propionate, used at therapeutic concentrations, alone and in combination. After 1 h pretreatment, combined treatment significantly inhibited rhinovirus 16, 1B, and IL-1beta-induced CCL5 and CXCL8 protein and mRNA production in BEAS-2B cells compared with fluticasone alone. When used 4 h after treatment, the combination significantly reduced virus-induced CCL5 but not CXCL8. Salmeterol alone had no effect; therefore, this inhibition was synergistic. Kinetic analysis demonstrated that combination therapy reduced by 5-fold the concentration of corticosteroid required to inhibit CXCL8 mRNA expression. In primary cells, salmeterol alone reduced rhinovirus-induced CCL5 and CXCL10 and increased CXCL5 production in a dose-dependent manner but had no effect on CXCL8. Fluticasone alone reduced CCL5, CXCL8, and CXCL10 but had no effect on CXCL5. Combination therapy augmented inhibition of CXCL8, CCL5, and CXCL10 but had no effect on CXCL5. Corticosteroids and beta2 agonists suppress rhinovirus-induced chemokines in bronchial epithelial cells through synergistic and additive mechanisms. This effect was greater for lymphocyte- than for neutrophil-related chemokines.     

Retinoic acid inhibits elastase-induced injury in human lung epithelial cell lines

The protective effects of retinoic acid on elastase-induced lung epithelial cell injury were studied using elastase extracted from purulent human sputum, the BEAS-2B human bronchial epithelial cell line, A549 human type II lung cell line, and primary cultures of human tracheal epithelial cells. Elastase decreased viability of BEAS-2B cells, A549 cells, and human tracheal epithelial cells in concentration- and time-dependent fashions. Elastase also induced apoptosis of BEAS-2B cells, A549 cells, and the tracheal epithelial cells detected with cell death detection enzyme-linked immunosorbent assay and terminal deoxyribonucleotidyl transferase-mediated dUTP-biotin nick-end labeling (TUNEL) methods. Retinoic acid alone did not affect the viability of BEAS-2B cells, A549 cells, or the tracheal epithelial cells, and did not induce apoptosis of the cells. However, retinoic acid prevented the decreases in the viability and reduced apoptosis of BEAS-2B cells, A549 cells, and the tracheal epithelial cells induced by elastase. Likewise, retinoic acid inhibited caspase 3 activity in BEAS-2B cells and A549 cells induced by elastase, as well as proteolytic activity of elastase. Furthermore, caspase 3 inhibitor inhibited the elastase-induced apoptosis of the cells. These findings suggest that retinoic acid may inhibit elastase-induced lung epithelial cell injury partly through the inhibition of proteolytic activity of elastase and through the inhibition of caspase 3 activity by elastase. Retinoic acid may, therefore, have protective effects against the elastase-induced lung injury and subsequent development of pulmonary emphysema.     

Airway epithelial epidermal growth factor receptor mediates hogbarn dust-induced cytokine release but not Ca2+ response

A subset of workers in swine confinement facilities develops chronic respiratory disease. An aqueous extract of dust from these facilities (hogbarn dust extract [HDE]) induces IL-6 and IL-8 release and several other responses in isolated airway epithelial cells. The cell membrane receptors by which HDE initiates these responses have not been identified. Because several other inhaled agents induce airway epithelial cell responses through epidermal growth factor receptor (EGFR) activation, we hypothesized that HDE would activate EGFRs and that EGFRs would be required for some of the responses to HDE. Exposure of Beas-2B cells to HDE caused EGFR phosphorylation and downstream ERK activation, and both responses were blocked by the EGFR-selective kinase inhibitor AG1478. AG1478 and EGFR-neutralizing antibody reduced HDE-stimulated IL-6 and IL-8 release by about half. Similar EGFR phosphorylation and requirement of EGFRs for maximal IL-6 and IL-8 release were found with primary isolates of human bronchial epithelial cells. Because HDE-stimulated IL-6 and IL-8 release involve the Ca(2+)-dependent protein kinase Cα, we hypothesized that HDE would induce intracellular Ca(2+) mobilization. HDE exposure induced intracellular Ca(2+) mobilization in Beas-2B cells and in primary cell isolates, but this response was neither mimicked by EGF nor inhibited by AG1478. Thus, HDE activates EGFRs and their downstream signaling, and EGFR activation is required for some but not all airway epithelial cell responses to HDE.     

NOD-like receptors mediated activation of eosinophils interacting with bronchial epithelial cells: a link between innate immunity and allergic asthma

Key intracytosolic pattern recognition receptors of innate immunity against bacterial infections are nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs). We elucidated the NOD1 and NOD2-mediated activation of human eosinophils, the principal effector cells for allergic inflammation, upon interacting with human bronchial epithelial BEAS-2B cells in allergic asthma. Eosinophils constitutively expressed NOD1,2 but exhibited nonsignificant responses to release chemokines upon the stimulation by NOD1 ligand γ-D-glutamyl-meso-diaminopimelic acid (iE-DAP) and NOD2 ligand muramyl dipeptide (MDP). However, iE-DAP and MDP could significantly upregulate cell surface expression of CD18 and intercellular adhesion molecule (ICAM)-1 on eosinophils and ICAM-1 on BEAS-2B cells, as well as induce chemokines CCL2 and CXCL8 release in the coculture system (all P<0.05). Both eosinophils and BEAS-2B cells were the main source for CXCL8 and CCL2 release in the coculture system upon iE-DAP or MDP stimulation. Direct interaction between eosinophils and BEAS-2B cells is responsible for CCL2 release, and soluble mediators are implicated in CXCL8 release. ERK and NF-κB play regulatory roles for the expression of adhesion molecules and chemokines in coculture. Treatment with NOD1,2 ligand could induce the subepithelial fibrosis and significantly enhance the serum concentration of total IgE, chemokine CCL5 for eosinophils and T helper type 2 (Th2) cells and asthma Th2 cytokine IL-13 in bronchoalveolar lavage fluid of ovalbumin-sensitized allergic asthmatic mice (all P<0.05). This study provides further evidence of bacterial infection-mediated activation of NOD1,2 in triggering allergic asthma via the activation of eosinophils interacting with bronchial epithelial cells at inflammatory airway.     

TNF-alpha and IFN-alpha enhance influenza-A-virus-induced chemokine gene expression in human A549 lung epithelial cells

Lung epithelial cells are the primary cellular targets for respiratory virus pathogens such as influenza and parainfluenza viruses. Here, we have analyzed influenza A, influenza B and Sendai virus-induced chemokine response in human A549 lung epithelial cells. Influenza virus infection resulted in low CCL2/MCP-1, CCL5/RANTES, CXCL8/IL-8 and CXCL10/IP-10 production at late times of infection. However, when cells were pretreated with TNF-α or IFN-α, influenza-A-virus-induced chemokine production was greatly enhanced. Cytokine pretreatment resulted in enhanced expression of RIG-I, IKKε, interferon regulatory factor (IRF)1, IRF7 and p50 proteins. Most importantly, influenza-A-virus-induced DNA binding of IRF1, IRF3, IRF7 and NF-κB onto CXCL10 ISRE and NF-κB elements, respectively, was markedly enhanced in cytokine-pretreated cells. Our results suggest that IFN-α and TNF-α have a significant role in priming epithelial cells for higher cytokine and chemokine production in influenza A virus infection.     

Chitinase activates protease-activated receptor-2 in human airway epithelial cells

Mammalian chitinase released by airway epithelia is thought to be an important mediator of disease manifestation in an experimental model of asthma. However, the intracellular signaling mechanisms engaged by exogenous chitinase in human airway epithelial cells are unknown. Here, we investigated the direct effects of exogenous chitinase from Streptomyces griseus on Ca(2+) signaling in human airway epithelial cells. Spectrofluorometry was used to measure intracellular Ca(2+) concentration ([Ca(2+)](i)) in fura-2-AM-loaded cells. S. griseus chitinase induced dose-dependent [Ca(2+)](i) increases in normal human bronchial epithelial cells and promoted [Ca(2+)](i) oscillations in H292 cells. Chitinase-induced [Ca(2+)](i) oscillations were independent of extracellular Ca(2+), suggesting that the observed [Ca(2+)](i) increases were due to Ca(2+) release from intracellular stores. Accordingly, after depleting endoplasmic reticulum (ER) Ca(2+) with the ER Ca(2+) ATPase inhibitor, thapsigargin, chitinase-mediated [Ca(2+)](i) increases were abolished. Treatment with the phospholipase C (PLC) inhibitor U73122 or the 1, 4, 5-trisinositolphosphate (IP(3)) receptor inhibitor 2-APB attenuated chitinase-induced [Ca(2+)](i) increases. Desensitization of protease-activated receptor-2 (PAR-2) by repetitive agonist stimulation or siRNA-mediated PAR-2 knock-down revealed that chitinase-mediated [Ca(2+)](i) increases were exclusively mediated by PAR-2 activation. Finally, chitinase was found to cleave a model peptide representing the cleavage site of PAR-2 and enhanced IL-8 production. These results indicate that exogenous chitinase is a potent proteolytic activator of PAR-2 that can directly induce PLC/IP(3)-dependent Ca(2+) signaling in human airway epithelial cells.     

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone, a component of tobacco smoke, modulates mediator release from human bronchial and alveolar epithelial cells

Respiratory epithelial cells are known to contribute to immune responses through the release of mediators. The aim of this study was to characterize the immunomodulatory effects of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a tobacco carcinogen, on respiratory epithelial cells and to compare two metabolic pathways, alpha-methylhydroxylation and alpha-methylenehydroxylation, involved in these effects using selective precursors, 4-(acetoxy-methylnitrosamino)-1-(3-pyridil)-1-butanone (NNKOAc) and N-nitroso (acetoxymethyl) methylamine (NDMAOAc), respectively. Human bronchial and alveolar epithelial cell lines, BEAS-2B and A549, respectively, were treated with NNK, NNKOAc and NDMAOAc for 24 h with and without tumour necrosis factor (TNF) and mediators released in cell-free supernatants were measured by enzyme-linked immunosorbent assay (ELISA). NNK significantly inhibited interleukin (IL)-8, IL-6 and monocyte chemoattractant protein-1 (MCP-1) production in both cell types. Similar results were observed with primary bronchial and alveolar epithelial cells. Although NNK increased prostaglandin E(2) (PGE(2)) production by A549 cells, its immunomodulatory effects were not mediated by PGE(2) according to the results with cyclo-oxygenase inhibitors. NNKOAc mimicked NNK effects, whereas NDMAOAc significantly inhibited IL-8 production in BEAS-2B cells and MCP-1 in both cell types. These results demonstrate that NNK and its reactive metabolites have immunosuppressive effects on respiratory epithelial cells, which could contribute to the increased respiratory infections observed in smokers and the development and/or the progression of lung cancer.     

Isorhamnetin attenuates TNF-α-induced inflammation,proliferation, and migration in human bronchial epithelial cells via MAPK and NF-κB pathways

Isorhamnetin has distinct anti-inflammatory activity and inhibits cell proliferation and migration. These effects are also involved in the pathogenesis of asthma. However, the effect of isorhamnetin on bronchial epithelial cells in patients with asthma has not been examined. Cells of human bronchial epithelial cell line BEAS-2B were cultured with isorhamnetin and tumor necrosis factor (TNF)-α. The effects of isorhamnetin on BEAS-2B cell viability were assessed using CCK8 assay. The EdU (5-ethynyl-2′-deoxyuridine) cell proliferation assay was performed to assess cell proliferation. BEAS-2B cell migration was measured using Transwell and wound healing assays. Real-time PCR and enzyme-linked immunosorbent assay were conducted to measure the expression of pro-inflammatory cytokines. Protein expression levels were determined by western blotting. Immunofluorescence was used to detect nuclear translocation of nuclear factor kappa B (NF-κB). We found that isorhamnetin at 20 and 40 μM reduced the proliferation of BEAS-2B cells induced by TNF-α. Isorhamnetin significantly decreased the expression of interleukin (IL)-1β, IL-6, IL-8, and C-X-C motif chemokine ligand 10 in BEAS-2B cells induced by TNF-α. Additionally, 10 μM isorhamnetin effectively reduced cell migration induced by TNF-α. Treatment with isorhamnetin inhibited the phosphorylation of mitogen-activated protein kinase (MAPK) and NF-κB pathways induced by TNF-α. In summary, isorhamnetin inhibited the inflammation, proliferation, and migration of BEAS-2B cells by regulating the MAPK and NF-κB signaling pathways and is a drug candidate for asthma.     

Acute application of interleukin-1beta induces Ca(2+) responses in human microglia

The effects of the pro-inflammatory cytokine interleukin-1-beta (IL-1beta) on levels of intracellular calcium [Ca(2+)](i) in cultured human microglia have been studied using the fluorescent Ca(2+) indicator fura-2. IL-1beta (2 ng/ml) caused a slow, progressive increase in [Ca(2+)](i) in standard Ca(2+)-containing physiological solution (PSS). A similar effect was observed in separate studies using Ca(2+)-free PSS, however, the mean rate of increase was significantly lower than that measured with PSS. Similar results were obtained in a separate protocol, where cells were exposed to both IL-1beta in Ca(2+)-free PSS and PSS. The slope of the IL-1beta induced increase of [Ca(2+)](i) in Ca(2+)-free PSS was not altered when adenosine triphosphate was added prior to application of the cytokine. These results suggest that IL-1beta-induced responses in human microglia involve both a Ca(2+) entry pathway and a mechanism of intracellular increase other than from IP(3)-sensitive stores.     

Adenoviral interleukin-12 gene transduction into human bronchial epithelial cells: up-regulation of pro-inflammatory cytokines and its prevention by corticosteroids

BACKGROUND: One of the potential effects of IL-12 is to restore Th1/Th2 balance. Therefore, we investigated the possibility of developing a system for local delivery of IL-12 into the airways by examining protein expression in a human bronchial epithelial cell line (BEAS-2B) after adenoviral IL-12 gene transduction. The effects of dexamethasone on the gene-modified cells were also examined. MEHODS: Adenoviral vectors AxCAegfp and Ax1CIhp40ip35 were used to transduce enhanced green fluorescence protein and IL-12 genes, respectively, into BEAS-2B cells. Wild-type and IL-12 gene-transduced BEAS-2B cells were then incubated with or without dexamethasone, and concentrations of IL-12, IFN-gamma, IL-6, IL-8, granulocyte macrophage-colony stimulating factor and chemokines (TARC and RANTES) in the supernatant were measured by ELISA. IL-12 receptor expression was analysed by flow cytometry and RT-PCR. RESULTS: The efficiency of transgene expression in BEAS-2B cells at a multiplicity of infection of 30 was approximately 80%. Gene-modified BEAS-2B cells produced biologically active IL-12, regardless of dexamethasone treatment. While IL-12 gene transduction led to increased production of IL-6 and IL-8 by BEAS-2B cells, expressions of these proteins were suppressed by dexamethasone. Addition of exogenous IL-12 failed to augment BEAS-2B cell IL-6 and IL-8 production, and IL-12 receptor expression by BEAS-2B cells was not detected. CONCLUSIONS: Our findings suggest that adenoviral IL-12 gene transduction may be effective in inducing IL-12 expression in the airways, and could be a potential approach in the management of bronchial asthma.     

Oxidant-injured airway epithelial cells upregulate thioredoxin but do not produce interleukin-8

We tested the hypothesis that oxidant-injured cells upregulate thioredoxin, whereas oxidant-stressed, but not injured, cells upregulate interleukin (IL)-8 after injury. We exposed primary human tracheobronchial epithelial cells and transformed human bronchial epithelial cells (BEAS-2B S.6) to 0, 200, 400, or 600 microM H(2)O(2) for 1 h followed by an additional 7 h of incubation. Subsequently, the cells were double-labeled with markers of injury (either Ethidium Homodimer-1 for cellular injury or MitoTracker dye for functional mitochondria) or oxidant stress (5-[and 6]-chloromethyl-2',7'-dicholorodihydrofluorescein diacetate) and antibodies specific for the chemoattractants IL-8 or thioredoxin. We found significant inverse relationships between numbers and stained chemoattractant volumes of IL-8 and thioredoxin-positive cells with increasing H(2)O(2) dose. Cells with mitochondrial injury produced thioredoxin but not IL-8, and oxidant-stressed cells were more likely to produce thioredoxin than IL-8. Isolated human neutrophils were more likely to colocalize with thioredoxin-positive BEAS-2B S.6 cells than thioredoxin-negative cells. The H(2)O(2) injury did not induce significant apoptosis in the BEAS-2B S.6 cells as measured by caspase 3 activation. We conclude that oxidant-injured and stressed airway epithelial cells upregulate thioredoxin, but produce little IL-8, which may be important in airway epithelial cell-mediated multistep navigation of neutrophils to sites of oxidant injury.     

A549肺癌细胞条件培养液促进人骨髓间充质干细胞的增殖和迁移

目的:探讨A549肺癌细胞对人骨髓间充质干细胞(hBMSCs)增殖和运动的影响.方法:以提取A549肺癌细胞和BEAS-2B正常人肺上皮细胞上清液制备条件培养基,培养的hBMSCs分别为A549组和BEAS-2B组;2组均添50 nmol/L趋化因子-8(CXCL-8),分别为A549 C组和BEAS-2B C组,若同...     

Interleukin-31 induces cytokine and chemokine production from human bronchial epithelial cells through activation of mitogen-activated protein kinase signalling pathways: implications for the allergic response

Interleukin-31 (IL-31) is a novel T-helper-lymphocyte-derived cytokine that plays an important role in allergic skin inflammation and atopic dermatitis. It has recently been implicated in bronchial inflammation. We investigated the functions and mechanisms of IL-31-induced activation of human bronchial epithelial cells. The gene and protein expressions of candidate cytokines/chemokines from IL-31-stimulated human bronchial epithelial BEAS-2B cells were first quantified by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay, respectively. The activity of different mitogen-activated protein kinases (MAPKs) in IL-31-stimulated BEAS-2B cells was assessed by Western blot. The IL-31 could significantly elevate the gene and protein expressions of epidermal growth factor (EGF), vascular endothelial growth factor (VEGF) and monocyte chemoattractant protein-1 (MCP-1/CCL2) of BEAS-2B cells in both time-dependently and dose-dependently. Combination of IL-31 with either IL-4 or IL-13 further enhanced VEGF and CCL2 production while IL-31 could synergistically augment the release of EGF, VEGF, CCL2, IL-6 and IL-8 in cocultures of BEAS-2B cells and eosinophils. In addition, IL-31 could activate p38 MAPK, extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) of BEAS-2B cells. Selective inhibitors of p38 MAPK (SB203580), ERK (PD98059), and JNK (SP600125) could differentially inhibit the production of EGF, VEGF and CCL2, thereby suggesting a role for MAPKs in IL-31 functions. In conclusion, the activation of MAPKs can be crucial for IL-31-mediated activation of bronchial epithelial cells, thereby providing an immunological role for IL-31 in bronchial inflammation, at least partly, via epithelial EGF, VEGF and CCL2 production.     

肺炎克雷伯杆菌荚膜多糖经EGFR途径诱导支气管上皮细胞分泌细胞因子

目的:探讨表皮生长因子受体(EGFR)在肺炎克雷伯杆菌(KP)荚膜多糖(CPS)诱导人正常支气管上皮BEAS-2B细胞分泌炎性细胞因子中的作用。方法:体外培养KP并提取CPS,用不同浓度的CPS刺激BEAS-2B细胞,通过ELISA检测细胞上清中肿瘤坏死因子α(TNF-α)和白细胞介素8(IL-8)的水平,并于刺激后不同时点通过Western blot检测EGFR的磷酸化水平;EGFR抑制剂AG1478预处理BEAS-2B细胞后,Western blot检测ERK磷酸化水平,间接免疫荧光染色检测p65的核转位,并观察细胞上清中TNF-α和IL-8的变化情况;最后经ERK抑制剂PD98059和NF-κB抑制剂PDTC分别预处理后用CPS刺激细胞,ELISA检测细胞上清中TNF-α和IL-8的水平。结果:10 mg/L CPS刺激BEAS-2B细胞12 h能够显著诱导其分泌TNF-α和IL-8;Western blot和间接免疫荧光染色检测结果显示,CPS刺激可显著诱导BEAS-2B细胞中EGFR和ERK的磷酸化及p65的核转位。经EGFR抑制剂AG1478预处理细胞后,ERK的磷酸化水平显著降低,NF-κB的核转位减少;而在EGFR、ERK及NF-κB抑制剂预处理的细胞的上清中,TNF-α和IL-8分泌水平均明显降低(P<0.05)。结论:肺炎克雷伯杆菌荚膜多糖能够通过EGFR激活ERK和NF-κB信号通路,进而诱导人正常支气管上皮细胞分泌炎性细胞因子TNF-α和IL-8,提示EGFR可能是KP感染引起炎症反应的关键因子。