In order to investigate the effect of vitamin A (VA) on the secretion of IFN-gamma and IL-4 in Mycoplasma Pneumoniae (MP)-induced A549 cells, A549 cells were co-cultured with MP for different time lengths and then the levels of IFN-gamma and IL-4 in the cell culture supernatants were detected before and after treatment with different concentrations of VA by using the enzyme-linked immunosorbent assay (ELISA). The results showed that the level of IFN-gamma and IL-4 in the supernatants of MP-induced A549 cells was much higher than that in non-induced cells (P<0.01). After application of VA, IL-4 level was not increased until the concentration of VA was up to 0.5x10(-5) mol/L (P<0.01). However, with concentration of VA increased up to 1x10(-4) mol/L, IL-4 was significantly suppressed (P<0.01). It was concluded that MP could induce the secretion of IFN-gamma and IL-4 in A549 cells. VA could inhibit the secretion of IFN-gamma and increase the IL-4 level in MP-induced A549 cells. However, high concentration of VA had an inhibitory effect on the secretion of IL-4 as well as on the IFN-gamma. These data provided a theoretical basis for the application of VA in MP pneumonia in the clinical practice.
Cell Line, Tumor ; Coculture Techniques ; Culture Techniques ; Interferon-gamma/*secretion ; Interleukin-4/*secretion ; Lung Neoplasms/pathology ; Mycoplasma pneumoniae/growth & development ; Mycoplasma pneumoniae/*physiology ; Vitamin A/*pharmacology

Purpose: Although Mycoplasma pneumoniae (M. pneumoniae) infection can cause wheezing in non-asthmatic children, the mechanisms of this symptom remain unclear. Vascular endothelial growth factor (VEGF) is a major mediator of angiogenesis and vascular permeability, and is also known to be elevated in cases of chronic pulmonary disease such as asthma. We hypothesized that VEGF may increase in children with acute M. pneumoniae pneumonia and wheezing. Methods: Nine patients with clinical and laboratory evidence of acute M. pneumoniae pneumonia were enlisted from children admitted to Korea University Hospital. They had had more than one episode of wheezing during the illness, which was confirmed by a physician; they comprised the wheezer group. The individuals with M. pneumoniae pneumonia without wheezing were 63 in number, and they comprised the non-wheezer group. Patients with a history of asthma or who had received asthma medications were excluded. Serum concentrations of VEGF, total IgE, eosinophil cationic protein (ECP), and peripheral blood eosinophil counts were measured. Results: The serum VEGF concentrations were higher in the wheezer group (mean+/-SD; 650.2+/-417.9 pg/mL) than in the non-wheezer group (376.5+/-356.2 pg/mL, P=0.049). M. pneumoniae antibody (1:1,380 vs. 1:596, P=0.048) and serum total IgE (591.8 IU/mL vs. 162.2 IU/mL, P=0.032) were higher in the wheezer group than in the non-wheezer group. There were no differences between the two groups in terms of serum ECP concentration or blood eosinophil count. Conclusion: In the presence of wheezing, serum VEGF concentrations were higher in the children with M. pneumoniae pneumonia. This finding suggests that VEGF may associate with wheeze-related symptoms in children with acute M. pneumoniae pneumonia.
Asthma ; Capillary Permeability ; Child ; Eosinophil Cationic Protein ; Eosinophils ; Humans ; Immunoglobulin E ; Korea ; Lung Diseases ; Mycoplasma ; Mycoplasma pneumoniae ; Pneumonia ; Pneumonia, Mycoplasma ; Respiratory Sounds ; Vascular Endothelial Growth Factor A

PURPOSE: Mycoplasma pneumoniae is a common causative agent of community acquired pneumonia in children and is well known to cause various respiratory and extrapulmonary diseases. We determined whether growth factors, including transforming growth factor (TGF)-beta1 and platelet-derived growth factor (PDGF-BB) that regulate airway fibrosis and remodeling, are increased in young children with M. pneumoniae pneumonia and also investigated if there was any difference in relation to the clinical status of the patients. METHODS: Fifty-two patients (3 to 6 years of age) who were admitted with M. pneumoniae pneumonia were enrolled and divided into 2 groups: the patients with frequent wheezing episodes (group A, n=28) and the patients with no previous history of wheeze (group B, n=24). The former group included the patients who had recurrent wheeze more than 3 times before admission. Fifteen children admitted with minor surgical problems were also studied as controls. TGF-beta1 and PDGF-BB were measured in the plasma samples collected on admission using ELISA in both patient groups and controls. RESULTS: Plasma TGF-beta1 and PDGF-BB levels were increased significantly in the patients with M. pneumoniae pneumonia as compared to the controls (P<0.01, respectively). TGF-beta1 and PDGF-BB were higher in group A than in group B, but the difference was not statistically significant (P=0.08 vs. P=0.05). In group A, TGF-beta1 was significantly higher in atopic patients than in non-atopic patients (P<0.05). CONCLUSION: Our study showed significantly increased TGF-beta1 and PDGF-BB in patients with M. pneumoniae pneumonia. It is suggested that these growth factors may play an important role in the pathogenesis of lower airway infection by M. pneumoniae.
Child ; Enzyme-Linked Immunosorbent Assay ; Fibrosis ; Humans ; Intercellular Signaling Peptides and Proteins ; Mycoplasma ; Mycoplasma pneumoniae ; Plasma ; Platelet-Derived Growth Factor ; Pneumonia ; Pneumonia, Mycoplasma ; Proto-Oncogene Proteins c-sis ; Respiratory Sounds ; Transforming Growth Factor beta1 ; Transforming Growth Factors

PURPOSE: Previous studies have outlined mechanisms by which Mycoplasma pneumonia (M. pneumonia) infection may promote allergic lung inflammation and airway remodeling, and increasing evidence from human studies suggests that atypical bacterial infections contribute to asthma exacerbation, chronic asthma, and disease severity with changes in cytokine expression. The present study evaluated changes in serum levels of vascular endothelial growth factor (VEGF) and interleukin (IL)-5 in atopic children with Mycoplasma pneumoniae pneumonia. METHODS: We recruited a total of 72 children with pneumonia. The patients were divided into 4 groups: atopic children with M. pneumonia pneumonia (group I, n=24), non-atopic children with M. pneumonia pneumonia (group II, n=23), atopic children with viral pneumonia (group III, n=13), and non-atopic children with viral pneumonia (group IV, n=12). Serum levels of IL-5, IL-13, VEGF, and tumor necrosis factor-alpha were measured at admission and at recovery using enzyme-linked immunosorbent assays. RESULTS: Serum levels of VEGF and IL-5 were elevated in group I compared with the other groups at both admission phase and clinical recovery phase. In group I, serum levels of VEGF and IL-5 were higher at recovery phase than at admission phase (VEGF: 1,102.2+/-569.4 vs. 874.9+/-589.9 pg/mL, respectively; IL-5: 150.5+/-63.9 vs. 120.2+/-46.7 pg/mL, respectively). CONCLUSIONS: The serum levels of VEGF and IL-5 were more increased in atopic children with M. pneumonia pneumonia than in the other groups. In this group, the serum levels of VEGF and IL-5 were more increased at recovery phase than at admission phase. The results of this study suggest that increases in VEGF and IL-5 may contribute to the development of hypersensitivity during M. pneumonia infection. These cytokines may act through their respective pro-inflammatory pathways to aggravate the allergic status and induce airway hypersensitivity during M. pneumonia pneumonia in atopic children.
Airway Remodeling ; Asthma ; Bacterial Infections ; Child ; Cytokines ; Humans ; Hypersensitivity ; Interleukin-13 ; Interleukin-5 ; Interleukins ; Mycoplasma ; Mycoplasma pneumoniae ; Pneumonia ; Pneumonia, Mycoplasma ; Pneumonia, Viral ; Tumor Necrosis Factor-alpha ; Vascular Endothelial Growth Factor A

PURPOSE: There has been an increasing amount of literature concerning the association between Mycoplasma pneumoniae and asthma pathogenesis. Interleukin(IL)-6 stimulates the differentiation of monocytes, and can promote Th2 differentiation and simultaneously inhibit Th1 polarization. IL-8 is a potent chemoattractant and, it has been suggested, has a role in asthma pathogenesis. Nitric oxide (NO) synthesized by airway epithelium may be important in the regulation of airway inflammation and reactivity. Vascular endothelial growth factor(VEGF) has been reported to be a mediator of airway remodeling in asthma. We investigated the effects of M. pneumoniae on IL-6, IL-8, NO and VEGF production in human respiratory epithelial cells. METHODS: A549 cells were cultured and inoculated with M. pneumoniae at a dose of 20 cfu/cell. After infection, the presence of M. pneumoniae in epithelial cell cultures was monitored by immunofluorescence and confirmed by polymerase chain reaction(PCR) detection. IL-6, IL-8 and VEGF were determined by an enzyme-linked immunosorbent assay and reverse transcriptase-polymerase chain reaction. NO was measured using the standard Griess reaction. RESULTS: In A549 cells, M. pneumoniaeinduced IL-6, IL-8, NO and VEGF release in time-dependent manners. It also induced mRNA expression of IL-6, IL-8 and VEGF in similar manners. CONCLUSION: These observations suggest that M. pneumoniae might have a role in the pathogenesis of the allergic inflammation of bronchial asthma.
Airway Remodeling ; Asthma ; Enzyme-Linked Immunosorbent Assay ; Epithelial Cells ; Epithelium* ; Fluorescent Antibody Technique ; Humans ; Inflammation ; Interleukin-6 ; Interleukin-8 ; Monocytes ; Mycoplasma pneumoniae ; Mycoplasma* ; Nitric Oxide ; Pneumonia ; Pneumonia, Mycoplasma ; RNA, Messenger ; Vascular Endothelial Growth Factor A

OBJECTIVETo investigate the effect of annexin A2 (AnxA2) on epithelial growth factor receptor (EGFR)/nuclear factor-κB (NF-κB) signal transduction and mucin expression in human airway epithelial H292 cells treated with Mycoplasma pneumoniae (MP).

METHODSH292 cells were divided into control group, MP group, NC-siRNA+MP group, and AnxA2 siRNA+MP group. The cells in the MP group were incubated with 5 μg/mL MP antigen for 2 hours. The cells in the NC-siRNA+MP and AnxA2 siRNA+MP groups were transfected with NC-siRNA and AnxA2 siRNA for 24 hours, followed by MP antigen stimulation for 2 hours. The MTT method was used to measure cell viability; quantitative real-time PCR was used to measure the mRNA expression of AnxA2; Western blot was used to measure the protein expression of AnxA2, phosphorylated EGFR (p-EGFR), and phosphorylated p65 NF-κB (p-p65 NF-κB); ELISA was used to measure the secretion of mucin 5AC (MUC5AC) and mucin 5B (MUC5B).

RESULTSThe MP and NC-siRNA+MP groups had lower cell viability than the control group (P<0.05). The AnxA2 siRNA+MP group had higher cell viability than the MP and NC-siRNA+MP groups and lower cell viability than the control group (P<0.05). The MP and NC-siRNA+MP groups had significantly higher mRNA and protein expression of AnxA2 than the AnxA2 siRNA+MP group (P<0.05). Compared with the control group, the MP and NC-siRNA+MP groups had significant increases in the protein expression of p-EGFR, p-p65 NF-κB, MUC5AC, and MUC5B (P<0.05); the AnxA2 siRNA+MP group had lower protein expression than the MP and NC-siRNA+MP groups, but higher protein expression than the control group (P<0.05).

CONCLUSIONSAnxA2 is involved in the airway lesion induced by MP antigen via mediating EGFR/NF-κB signaling activation and mucin expression in human airway epithelial cells.

Annexin A2 ; physiology ; Bronchi ; physiology ; Cells, Cultured ; Epithelial Cells ; microbiology ; Humans ; Mucins ; analysis ; Mycoplasma pneumoniae ; pathogenicity ; NF-kappa B ; physiology ; Receptor, Epidermal Growth Factor ; physiology ; Signal Transduction ; physiology

PURPOSE: Mycoplasma pneumoniae is an extracellular pathogen that attaches to and destroys the ciliated epithelial cells of the respiratory tract. The vascular endothelial growth factor (VEGF) is a critical angiogenic factor that manages the formation and function of vascular networks. Thus, we examined whether M. pneumoniae lysate (MPL) induces VEGF and MPL-induced VEGF expression is regulated by the activation of mitogen-activated protein kinase (MAPK) pathways in airway epithelial cells. METHODS: Cells were treated with MPL in dose and time dependent manners or pretreated with chemical inhibitors of MAPK signaling molecules before the addition of MPL. The supernatants were measured by a specific human VEGF enzyme-linked immunosorbent assay (ELISA). The RNAs were extracted and synthesized into cDNAs for VEGF gene expression by polymerase chain reaction. RESULTS: MPL considerably increased VEGF mRNA 2 hours after treatment, which was gradually reduced thereafter. On the other hand, VEGF protein was continuously amplified for 12 hours after both 5 and 10 microg/mL MPL treatment. Pretreatment with U0126 (a specific extracellular signal-regulated kinase inhibitor) and SB202190 (a specific p38 inhibitor) abolished MPL-stimulated VEGF protein close to basal level (-85%), whereas JNK inhibitor II (a specific c-Jun N-terminal kinase inhibitor) partially decreased VEGF protein (57%). CONCLUSION: We concluded that MPL induces VEGF expression through the activation of MAPK signaling molecules (ERK, p38 and JNK) in airway epithelial cells.
Angiogenesis Inducing Agents ; Butadienes ; DNA, Complementary ; Enzyme-Linked Immunosorbent Assay ; Epithelial Cells ; Gene Expression ; Hand ; Humans ; Imidazoles ; JNK Mitogen-Activated Protein Kinases ; Mycoplasma ; Mycoplasma pneumoniae ; Nitriles ; Phosphotransferases ; Pneumonia ; Pneumonia, Mycoplasma ; Polymerase Chain Reaction ; Protein Kinases ; Pyridines ; Respiratory System ; RNA ; RNA, Messenger ; Vascular Endothelial Growth Factor A

This study investigated the serum vascular endothelial growth factor (VEGF) levels in children with community-acquired pneumonia. Serum VEGF levels were measured in patients with pneumonia (n=29) and in control subjects (n=27) by a sandwich enzyme-linked immunosorbent assay. The pneumonia group was classified into bronchopneumonia with pleural effusion (n=1), bronchopneumonia without pleural effusion (n=15), lobar pneumonia with pleural effusion (n=4), and lobar pneumonia without pleural effusion (n=9) groups based on the findings of chest radiographs. We also measured serum IL-6 levels and the other acute inflammatory parameters. Serum levels of VEGF in children with pneumonia were significantly higher than those in control subjects (p<0.01). Children with lobar pneumonia with or without effusion showed significantly higher levels of serum VEGF than children with bronchopneumonia. For lobar pneumonia, children with pleural effusion showed higher levels of VEGF than those without pleural effusion. Children with a positive urinary S. pneumonia antigen test also showed higher levels of VEGF than those with a negative result. Serum IL-6 levels did not show significant differences between children with pneumonia and control subjects. Serum levels of VEGF showed a positive correlation with the erythrocyte sedimentation rate in the children with pneumonia. In conclusion, VEGF may be one of the key mediators that lead to lobar pneumonia and parapneumonic effusion.
Vascular Endothelial Growth Factor A/*blood ; Streptococcus pneumoniae/growth & development/immunology ; Pneumonia, Bacterial/*blood/complications/microbiology ; Pleural Effusion/*blood/complications/microbiology ; Mycoplasma pneumoniae/growth & development/immunology ; Male ; Interleukin-6/blood ; Infant ; Humans ; Female ; Enzyme-Linked Immunosorbent Assay ; Community-Acquired Infections/blood/microbiology ; Child, Preschool ; Child ; Antigens, Bacterial/immunology ; Antibodies, Bacterial/immunology ; Adolescent