Expression of IFN-γ, IL-4 and IL-17A in asthmatic mice vaccinated with BCG and HepB in neonatal period

AIM: To investigate the expression of IFN-γ, IL-4 and IL-17A in asthmatic mice vaccinated with bacillus Calmette-Guérin (BCG) and hepatitis B (HepB) in the neonatal period. METHODS: BALB/c mice were randomly divided into BGG+HepB+ovalbumin (OVA) group (B/H/O group), B/O group, H/O group, B/H group, OVA group, BCG group, HepB group and normal saline (NS) group (n=6). The mice in B/H/O group and B/H group at 0, 7 and 14 d received subcutaneous injection of 1×10⁵ CFU BCG for 3 times, while at 0 and 28 d received intramuscular injection of 1.5 μg HepB on the hindlimb twice. The mice in other groups were individually vaccinated with BCG or HepB. OVA sensitization and aerosol inhalation were performed to establish the asthma model. The lung tissues were collected for HE staining. Bronchoalveolar lavage fluid (BALF) and peripheral blood (PB) were collected, and the number of eosinophils (EOS) in BALF was counted. The serum levels of IFN-γ and IL-4, and the level of IL-17A in lung tissue homogenate were measured by ELISA. RESULTS: The pathological changes of the lung in OVA group, B/O group, B/H/O group and H/O group were observed. There were extensive inflammatory cell infiltration around the bronchus, and epithe-lial cell hypertrophy. Those in B/H/O group and H/O group were worse than those in OVA group, while those in B/O group was better than those in OVA group. Total BALF cell counts in B/H/O group, B/O group and H/O group were decreased (P<0.05) as compared with OVA group. The BALF EOS count in B/H/O group was higher than that in B/H group, that in B/O group was higher than that in BCG group, and that in H/O group was higher than that in HepB groups (P<0.05). Compared with H/O group, OVA group and NS group, the serum IFN-γ/IL-4 ratio in HepB group was increased (P<0.05), and compared with B/H/O group, B/O group, OVA group and NS group, that in B/H group was also increased (P<0.05). Compared with OVA group, the level of IL-17A in the lung tissues of B/H/O group and B/O group was decreased (P<0.05), and compared with B/O group, that in B/H/O group was further decreased (P<0.05). CONCLUSION: Combined vaccination of BCG and HepB reduces the inflammotory responses in the lung tissues of asthmatic mice. The mechanism may be related with the decrease in the release of IL-4, the increase in IFN-γ/IL-4, and the inhibition of IL-17A expression.     

Levels of IL-17, IL-8 and VEGF in bronchoalveolar lavage fluid in asthma children

AIM: To investigate the mechanism of airway inflammation in children with asthma by determining the levels of IL-17, IL-8 and vascular endothelial growth factor(VEGF) in bronchoalveolar lavage fluid (BALF). METHODS: Eighty-eight children were enrolled in the study and divided into asthma group (n=52), pneumonia group (n=25) and control group (n=11). BALF were collected from all 88 cases. The levels of IL-17, IL-8, VEGF, IL-4 and IFN-γ in BALF were measured by ELISA. The cell types in BALF were determined. RESULTS: Compared with the control, the levels of IL-17 and IL-8 were significantly elevated in asthma group and pneumonia group (all P<0.05). The level of IL-8 (P<0.05) in the patients with asthma was lower than that in the pneumonia patients. No statistical difference of the IL-17 level between asthma group and pneumoniae group was observed (P>0.05). Compared with pneumonia group and control group, the level of VEGF was significantly increased in asthma group (all P<0.01), and the VEGF level among control group and pneumonia groups was almost similar (P>0.05). The levels of IL-4 and IFN-γ, and ratio of IL-4/IFN-γ among groups were not statistically different. The percentage of neutrophils in BALF was significantly higher in asthma group and pneumonia group than that in control group (all P<0.01). CONCLUSION: IL-17, IL-8 and VEGF play important roles in airway inflammation in children with asthma. Th17 cells may participate in the pathogenesis of asthma in children.     

Isorhamnetin inhibits ovalbumin-induced pulmonary inflammation in asthmatic mice

AIM To investigate the effect of isorhamnetin on pulmonary inflammation in asthmatic mice and to analyze its primary mechanism. METHODS BALB/c mice were randomly divided into normal control group, asthma model group, isorhamnetin low-dose （50 mg/kg） treatment group and isorhamnetin high-dose （150 mg/kg） treatment group. Ovalbumin （OVA） was used to establish a mouse asthma model. HE staining and PAS staining were used to observe the pathological changes of lung tissue. The contents of cysteinyl leukotriene1 （CysLT1）, cystelinyl leukotriene receptor 1 （CysLTR1）, interleukin-4 （IL-4）, IL-5, IL-13 and tumor necrosis factor-α （TNF-α） in bronchoalveolar lavage fluid （BALF） were measured by ELISA. The expression of nuclear factor of activated T cells 4（NFATc4） in lung tissue was detected by immunohistochemical staining. The protein expression of NFATc4, intercellular cell adhesion molecule-1 （ICAM-1） and vascular cell adhesion molecule-1 （VCAM-1） was determined by Western blot. RESULTS Isorhamnetin improved histopathological changes in OVA-induced asthma model mice, reduced the contents of CysLT1, CysLTR1, IL-4, IL-5, IL-13 and TNF-α in BALF, and reduced NFATc4, ICAM-1 and VCAM-1 expression in lung tissue （P<0.05）. CONCLUSION Isorhamnetin inhibits the inflammatory response of lung tissue in asthmatic model mice, and the mechanism may be related to the inhibition of the calcineurin/NFATc4 signaling pathway.     

Effects of ginsenoside Rh1 on expression of inflammatory factors in mouse asthma model

AIM: To observe the effects of ginsenoside Rh1 on the levels of inflammatory factors in serum and bronchoalveolar lavage fluid (BALF), and the pathological changes of the lung tissues in an experimentally induced mouse asthma model. METHODS: Male BALB/c mice (n=40) were divided into 4 groups:normal control group, asthma mo-del group, and low-dose (40 mg·kg^-1·d^-1) and high-dose (80 mg·kg^-1·d^-1) ginsenoside Rh1 groups. The bronchial asthma mouse model was established by the method of ovalbumin induction and excitation, and during the excitation period, the mice were daily treated with ginsenoside Rh1 for 2 weeks. At 24 h after the final dose of ginsenoside Rh1, the mice were sacrificed. The number of eosinophils (EOS) and the concentrations of interleukin (IL)-4, IL-5 and interferon (IFN)-γ in BALF were determined. The levels of IgG and IgE in serum were measured, and the expression of transforming growth factor (TGF)-β1 and the pathological changes in lung tissues were evaluated. RESULTS: Ginsenoside Rh1 inhibited the increases in the number of EOS and the concentrations of IL-4, IL-5, IFN-γ and IgE, reversed the increased expression of TGF-β1, and improved the pathological changes of the lung tissues in asthmatic mice. CONCLUSION: Ginsenoside Rh1 improves the immuno-inflammatory profile and pathological changes in the experimentally induced mouse asthma model, implying its potential therapeutic effect on asthma.     

Inhibitory effect of T-bet gene transfer on airway inflammation in a established murine allergic asthmatic model

AIM: To investigate the effect of T-bet plasmid gene transfer to airway on allergen induced airway inflammation in a murine asthmatic model. METHODS: A mouse asthma model was established by sensitization with ovalbumin (OVA). Forty C57BL/6 mice were divided into 4 groups (10 mice in each group): the normal control group (group A), the asthmatic model group (group B), the pcDNA3 plasmid group (group C), and the pcDNA3-T-bet group (group D). The animals in group B, C and D were sensitized and challenged with OVA. The animals in group A were applied with normal saline. pcDNA3 plasmid at dose of 50 μg was intranasally administered at 24 h before intranasal challenges to the mice in group C, and the 50 μg pcDNA3-T-bet plasmid for the mice in group D. Bronchial alveolar lavage fluid (BALF) was collected and lung tissues were resected at 48 h after OVA challenge for later assay. RESULTS: After administration with pcDNA3-T-bet plasmid, high level of T-bet expression at 48 h was detected in the lung tissue by Western blotting. In pcDNA3-T-bet treated asthmatic models, histological evaluation revealed the significant suppression of eosinophil peribronchial and perivascular infiltration, and reduction of epithelial damage. The numbers of eosinophils, neutrophils and lymphocytes in BALF from pcDNA3-T-bet treated mice were significantly reduced compared to those in asthmatic control group (P<0.05). The level of IL-4 in BALF was significantly decreased in pcDNA3-T-bet group compared to that in asthmatic control group (P<0.05), while the level of IFN-γ in BALF was significantly increased in pcDNA3-T-bet group. No significant change of inflammation cells and cytokines in pcDNA3 plasmid group and asthmatic control group was observed (P>0.05). CONCLUSION: Intranasal pcDNA3-T-bet plasmid transfer inhibits asthmatic airway inflammation in the murine asthmatic model, suggesting a new therapeutic strategy for allergic asthma.     

IgE-FcεRI crosslink accelerates atherosclerosis development in allergic asthmatic mice

AIM:To investigate whether allergic asthma accelerates the development of atherosclerosis in mice related to Th2 cells and interleukin-4 (IL-4), and the roles of activation of macrophages by immunoglobulin E (IgE)-Fc ε receptor I (FcεRI) crosslink during the process. METHODS:Six-week-old ApoE^-/- mice were sensitized and challenged by ovalbumin to establish the allergic asthma model, and then assigned to 3 groups:control group, asthmatic placebo group and asthmatic IL-4 monoclonal antibody (mAb) intervention group (intervention for 8 weeks). The lesion area was measured by oil red O staining. The percentages of Th2 cells in the splenocytes of the mice were analyzed by flow cytometry. The mRNA expression of IL-4 and the macrophage-related inflammatory factors, monocyte chemotactic protein 1 (MCP-1), macrophage inflammatory protein-1α (MIP-1α) and IL-6, in the spleen was detected by real-time PCR. Local IgE and FcεRIα expression in the plaque was evaluated by immunofluorescence/immunohistochemical staining, and the circulating IL-4 and IgE were measured by ELISA. RESULTS:Accompanied by aggravated atherogenesis in asthmatic ApoE^-/- mice, the proportion of Th2 cells and IL-4 mRNA in the spleen, IgE and FcεRIα expression in the aortic root, and the mRNA expression of MCP-1, MIP-1α and IL-6 were markedly increased. After 8-week treatment with IL-4 mAb, the lesion area in the aortic root of asthmatic ApoE^-/- mice was markedly decreased, the elevated IgE and FcεRIα expression was significantly decreased, and the mRNA expression of macrophage-related inflammatory factors was also decreased. CONCLUSION:Allergic asthma accelerates the atherosclerosis in ApoE^-/- mice, which is associated with the increased Th2 cells and IL-4, and the activation of macrophages by IgE-FcεRI crosslink.     

Effects of glucocorticoid on miRNA-155 expression in CD4+T cells of asthmatic mice

AIM:To investigate the effects of glucocorticoid on the regulation of microRNA-155 (miRNA-155) expression in the CD4⁺ T cells of asthmatic mice. METHODS:The ovalbumin (OVA)-induced asthma mouse model was established and the mice were treated with glucocorticoid. The effects of glucocorticoid on the pulmpnary histopathological changes, the expression of miRNA-155 in the lung tissues and CD4⁺T cells, and the levels of cytokines in the bronchoal-veolar lavage fluid (BALF) were evaluated. RESULTS:The results of RT-qPCR showed that the expressions of miRNA-155 in the lung tissues and CD4⁺T cells from the spleen of asthmatic mice were significantly increased, and the level of miRNA-155 in the CD4⁺T cells was significantly increased with the increase in the allergen exposure time (P<0.01). HE and PAS staining showed that OVA significantly increased inflammatory cell infiltration as compared with control group, and the peribronchial and perivascular inflammation and mucus secretion of proliferative goblet cells were significantly reduced after glucocorticoid treatment. Glucocorticoid treatment inhibited the increase in the proportion of CD4⁺ CD8^- cells in the spleen and decreased the accumulation of CD4⁺ T cells in the lung tissues of asthmatic mice (P<0.01). After glucocorticoid treatment, the levels of interleukin-4 (IL-4), IL-5 and IL-13 in BALF were decreased, while the level of interferon-γ was increased significantly (P<0.01). CONCLUSION:Glucocorticoid reduces the accumulation of CD4⁺ T cells and inhibits the expression of miRNA-155 in the lung tissues and spleen CD4⁺ T cells of asthmatic mice.     

Influence of Jiaomu oil A2 on eosinophil manifold and CD34+ with marrow granule system mobilization in bronchia asthma mice

AIM: To observe the influence of Jiaomu oil A2 on eosinophil manifold and CD34+ with marrow granule system mobilization in bronchial asthma mice. METHODS: The asthmatic mouse model was established by sensitization and challenge of the animals with 20% Al(OH)3+10% ovalbumen (OVA). After the mice were excitated for 10 d and giving medical therapy at the same time, the mice were executed, the bronchial-alveolar lavage inflammatory cells and the hemocytopoiesis cells were examined using Wrish-Giemsa staining. The expressions of interleukin-5 (IL-5) and eotaxin (EON) in lung tissue and marrow were examined by in situ hybridization. The expression of IL-5 and EON albumen in lung tissue and marrow were detected by immunohistochemistry. The inflammatory cell infiltration and CD34+ cells in lung tissue were also observed by HE and immunofluorescence staining. RESULTS: Both Jiaomu oil A2 and prednisone significantly attenuated the pathological process degree of bronchial inflammatory reaction such as tissue swelling, reconstruction, hyperplasia, and epithelial cell shedding, and inhibited eosinophil count and infiltration caused by stimulation of allergic effect. Moreover, the two drugs markedly lessen the eosinophil density in tracheal surrounding tissue and marrow in asthma mice and depressed the differentiation of marrow myelocyte to eosinophil. Finally, the apparent decrement of CD34+IL-5, CD34+IL-5R, CD34+CCR-3 cells in bronchial tissue and marrow showed some relationship with the downregulation of IL-5, IL-4, GM-CSF in lung tissue. CONCLUSION: The effect of Jiaomu oil on airway inflammation in asthma mice is associated with inhibiting the mobilization of eosinophil and marrow granule system.     

Effect of Kechuanning on airway remodeling and protein level of p-ERK1/2 in lung tissues of asthmatic rats induced by virus

AIM:To investigate the effect of Kechuanning on airway remodeling and the protein level of p-ERK1/2 in lung tissues of asthmatic rats induced by virus. METHODS:The asthmatic rat model induced by respiratory syncytial virus was established. The experimental rats were divided into normal group, asthma model group, low dose (0.33 mL/kg), middle dose (3.0 mL/kg) and high dose (10 mL/kg) of Kechuanning groups, and PD98059 (3 mg/kg) group. The airway responsiveness of the rats was measured by animal ventilator. The pathological changes of the lung tissues were observed by HE staining. PAS staining and Masson staining were used to observe goblet epithelial cells metaplasia and airway collagen deposition. The expression of matrix metalloproteinases-9 (MMP-9) and tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) in the lung tissues of the rats was detected by immunohistochemical staining. The protein levels of ERK1/2 and p-ERK1/2 were determined by Western blot. RESULTS:Compared with model group, the airway responsiveness of the rats in middle dose and high dose of Kechuanning groups was significantly decreased (P<0.01), the injury of lung tissues was significantly decreased, the goblet epithelial cells metaplasia and airway collagen deposition were significantly reduced (P<0.01), and the expression of MMP-9 and TIMP-1 in the lung tissues was also significantly decreased (P<0.01). In addition, the protein level of p-ERK1/2 in high dose of Kechuanning group was significantly decreased compared with model group (P<0.01). CONCLUSION:Kechuanning may treat asthma by regulating the expression of p-ERK1/2 in the lung tissues and improving the airway remodeling symptoms of asthmatic rats induced by virus.     

Effects of interferon-γ, tumor necrosis factor-α and interleukin-1β on apoptosis of human airway smooth muscle cellsin vitro

AIM:To clarify if interferon-γ(IFN-γ), tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)can induce apoptosis of human airway smooth muscle cells (ASMCs) in vitro.METHODS:Human ASMCs were isolated and cultured in DMEM containing 10% fetal bovine serum. Passage 4-6 cell was used in the experiment. IFN-γ,TNF-α and IL-1β, were used separately or together in the treatment of human ASMCs. The effects of IFN-γ,TNF-α and IL-1β on the growth of the cells was detected by MTT method at the hour 0,24,48 and 72. Light microscopy and electron microscopy were used to examine the morphological change. DNA fragmentation was analyzed by agarose gel electrophoresis. SP immunohistological staing method was performed to detect the change of expressions of p 53, bcl- 2 and bax gene. The apoptosis cell percentage were detected by in situ end labeling technique (TUNEL)of fragmental DNA. RESULTS:(1)IFN-γ or IFN-γ together with TNF-α and IL-1β decreased the number of viable cells in a time dependent manner. (2) Light and electron microscopic examination showed cell shrinkage, membrane blebbing, nuclear contraction, chromatin condensation and nuclear fragmentation in human ASMCs. (3) Agarose gel electrophoresis showed a characteristic"ladder"of DNA bands representing integer multiples of the internucleosomal fragments (about 180-200 bp) in cytokine cotreated human ASMCs. (4)The expression of p 53 and bax gene in cytokine cotreated group was significantly higher than in control group, but the expression of bcl-2 gene was lower than in control group. (5)Stimultaneous treatment with IFN-γ(4×10⁵ U/L),TNF-α(4×10⁵ U/L)and /or IL-1β (10×10⁴ U/L) induced apoptosis of human ASMCs. Apoptotic index of human ASMCs in cytokine co-treated group was significantly higher than in control group (P＜0.01).CONCLUSION:Stimultaneous treatment with IFN-γ,TNF-α and /or IL-1β induced apoptosis of human ASMCs. These immune cytokines may play an important role in airway remodeling of asthma and of chronic obstructive pulmonary disease.     

Effect of microRNA-7 knockdown on ConA-induced acute liver injury in mice

AIM: To study the effect of microRNA-7 (miR-7) knockdown (KD) on concanavalin A (ConA)-induced acute liver injury (ALI) in mice.METHODS: Wild type (WT) mice and miR-7KD mice were received ConA (30 mg/kg) to induced acute liver injury model by intraperitoneal injection, and the morphological changes, liver weight and weight index were measured 48 h later. The pathological changes of the liver tissues were observed by HE staining. The levels of serum alanine aminotransferase (ALT), IL-4 and IFN-γ were detected by ELISA. The proportional changes of CD4⁺ T cells and the relative levels of IL-4 and IFN-γ were analyzed by flow cytometry.RESULTS: The color of the liver tissue became lighter, and the weight and weight index were changed significantly in miR-7KD mice compared with control group (P<0.05). HE staining showed that the inflammatory cell infiltration was increased in the liver of miR-7KD mice. Moreover, the level of serum ALT was significantly increased (P<0.05). The serum level of IFN-γ elevated significantly (P<0.01), while the IL-4 levels decreased significantly (P<0.01) in the serum of miR-7KD mice. Furthermore, the proportion of CD4⁺ T cells and relative IFN-γ cells increased obviously (P<0.01).CONCLUSION: miR-7 knockdown promotes the pathogenesis of the ConA-induced acute liver injury in mice.     

Effects of specific immunotherapy on allergic rhinitis children accompanied with asthma

AIM: To explore the effects of Dermatophagoides pteronyssinus allergen-specific immunotherapy (SIT) on the serum interleukin (IL)-13,IL-4,interferon (IFN)-γ, nasal symptoms and pulmonary functions in allergic rhinitis children accompanied with asthma. METHODS: Fifty-eight cases of allergic rhinitis children accompanied with asthma participated in this study. Their allergens were Dermatophagoides pteronyssinus. Thirty-five children received SIT were SIT group, and the other 23 children received local glucocorticoid treatment were medical group. The serum levels of IL-13, IL-4 and IFN-γ were examined, and the nasal symptoms and pulmonary functions were checked before treatment and one year after treatment. RESULTS: There was a significant difference in nasal symptoms between the two groups one year after treatment (P<0.05). The patients in SIT group had fewer symptoms. The serum levels of IL-4 and IL-13 were clearly reduced. IFN-γ and the ratio of IFN-γ/IL-4 were significantly increased (P<0.05). The pulmonary functions were significantly improved in SIT group (P<0.05). Meanwhile in medical group, the serum levels of IL-4 and IL-13 had less change (P>0.05), and the pulmonary functions were poorly improved (P>0.05). CONCLUSION: SIT may regulate the imbalance of Th1/Th2 cells in allergic rhinitis accompanied with asthma by reducing the serum levels of IL-4 and IL-13 and increasing IFN-γ and the ratio of IFN-γ/IL-4, resulting in reducing the nasal symptoms and improving the pulmonary functions.     

Effects of zanthoxylum seed oilA2 on NF-кB signaling pathway and inflammatory factor in lung tissue of asthmatic mice

AIM: To investigate the dynamic influence of zanthoxylum seed oilA2 (ZSOA2) on NF-кB signaling pathway and inflammatory factor in pulmonary tissue of asthmatic mice. METHODS: The suspensoid (0.2 mL containing 20% albumin hydroxide and 10% ovalbumin) was administered by intraperitoneal injection to sensitize the BALB/c mice on day 1, then 0.4% ovalbumin solution (50 μL in phosphate buffer fluid) was dripped into the respiratory tract through nasal cavity to establish the asthmatic mouse model. After dripped ovalbumin for 24 h, 48 h, 3 d, 7 d and 14 d, the mice were killed at specified time points. The contents of interleukin-4 (IL-4), interleukin-5 (IL-5) and interferon-γ (IFN-γ) in bronchoalveolar lavage fluid (BALF) were determined by ELISA. The pathological changes of the lung tissues were observed with HE staining. The inflammatory cell counts were conducted by Eosin staining. The protein levels of adhesion molecule and the molecules of NF-κB signaling pathway in lung tissues were determined by Western blotting. RESULTS: In ZSOA2 treated mice, the pathological injury of the lung was significantly attenuated as compared to the model mice, the counts of eosinophils and lymphocytes were reduced obviously in lung bronchial area of asthmatic mice at all observed time points (P<0.05). The levels of IL-5 and IL-4 decreased and IFN-γ increased in BALF. The results of Western blotting showed that ZSOA2 down-regulated the protein levels of intercellular adhesion molecule-1, vascular cell adhesion molecule-1, I kappa B kinase alpha-α and phosphorylation inhibitory-κB. ZSOA2 also up-regulated the protein levels of tumor necrosis factor receptor 1 and phosphorylation nuclear factor-kappaB in lung tissue at all observed time points. CONCLUSION: ZSOA2 has therapeutic effect on asthma by down-regulating the protein expression of IκB-β and p-IκB, inhibiting the releases of cytokines and chemotactic factors, and attenuating the infiltration of inflammatory cells in the lungs of ovalbumin challenged asthma mice.     

Effects of eosinophils deletion on sPLA2-X in bronchial asthma model mice

AIM To investigate the association between soluble phospholipase A2-X（sPLA2-X） and eosinophils in bronchial asthma, and to provide new insight and strategies for the treatment of bronchial asthma. METHODS Female Babl/c mice （n=48） of SPF grade and 6~8 weeks old were divided into 4 groups （with 12 in each group： healthy control group,asthma control group, eosinophil deletion group, and asthma /eosinophil deletion isotype control group）. The mouse model of bronchial asthma was constructed. The mice in healthy control group were intraperitoneally injected with saline on days 0, 7, and 14. The mice in other groups were intraperitoneally injected with 50 μg OVA and 2 mg aluminum hydroxide gel（soluble in 200 μL saline.On the 21st d and 26 th d, eosinophil deletion antibody （anti-CCR3） and isotype control were intraperitoneally injected and intranasally respectively, and then the lungs function test was conducted within 48 h after the end of nebulization.Half of the mice in each group were subjected to whole lung lavage, the remaining half were used for lung tissue section with HE staining, the whole blood was used to measure serum IgE, the supernatant of broncho alveolar lavage fluid （BALF） was used to measure cytokines, and total number of cells in the bronchoalveolar lavage fluid was analyzed for cell classification and flow cytometry. RESULTS （1）Compared with asthma control group,the airway and alveolar inflammatory responses in asthma/eosinophil deletion group was significantly alleviated.（2） Compared with asthma control group, anti-CCR3 successfully deleted eosinophils, and the percentage of eosinophils in bronchoalveolar lavage fluid in asthma/eosinophil deletion group was significantly reduced （P<0.05）.（3） The airway hyperresponsiveness in asthma/eosinophil deletion group was significantly decreased as compared with asthma control group（P<0.05）.（4） The levels of sPLA2-X in the serum and BALF was significantly reduced in asthma/eosinophil deletion group as compared with asthma control group（P<0.05）.（5）Compared with asthma control group,the levels of IL-4, IL-5, and IL-13 in the BALF of the mice in asthma/eosinophil deletion group were significantly reduced, and the serum level of IgE was also decreased （P<0.05）. CONCLUSION Eosinophils in bronchial asthma are importantly associated with sPLA2-X.     

Inhalation of inactivated Mycobacterium phlei down-regulates expression of nuclear factor-kappa B and intercellular adhesion molecule-1 in asthmatic mice

AIM:To investigate the effect of inhalation of inactivated Mycobacte-rium phlei on the expression of nuclear factor-kappa B (NF-κB), intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 in the lung tissues of asthmatic mice. METHODS:Male BALB/c mice (n=24) were randomly divided into normal control group (A), asthmatic model group (B), and inactivated Mycobacterium phlei inhalation group (C). Asthmatic model was made by inhalation of chicken ovalbumin. The mice in group C were treated with inactivated Mycobacterium phlei for 5 d. The lung tissues and bronchoalveolar lavage fluid (BALF) were harvested. HE and AB-PAS staining were used to measure the lung inflammation and mucus production. The inflammatory cells in the BALF were counted. The mRNA expression of NF-κB, ICAM-1 and VCAM-1 was detected by real-time fluorescence quantitative PCR. RESULTS:Mycobacterium phlei treatment alleviated lung inflammation, attenuated mucus production, and reduced the percentage of eosinophils in the BALF. The mRNA levels of NF-κB and ICAM-1 were significantly decreased after treated with Mycobacterium phlei. However, no significant difference of VCAM-1 mRNA expression was found before and after treatment. The correlation between NF-κB mRNA and ICAM-1 mRNA, and between NF-κB mRNA and VCAM-1 mRNA was not found. CONCLUSION:Inhalation of inactivated Mycobacterium phlei attenuates asthmatic airway inflammation. NF-κB participates in the pathogenesis of asthma. NF-κB signal pathway may be associated with the therapeutic mechanism. Another important mechanism is the reduction of adhesion molecule expression.     

Effects of Foxp3 transfection on splenocyte functions in asthma mice

AIM: To study the expression and the effects of Foxp3 on the immunologic functions by transfecting the Foxp3 eukaryotic expression plasmid into the splenocytes of the asthma mice. METHODS: The mice were sensitized and challenged by ovalbumin to make asthma model. The splenocytes were harvested and cultured. The Foxp3 expression vector pcDNA3.1(-)-Foxp3 was transfected into the splenocytes with electroporation. The splenocytes transfected with empty vector and control splenocytes (non-transfected) were also set up. The expression of Foxp3 at mRNA and protein levels was detected by RT-PCR and Western blotting, respectively. The proportion of CD4⁺CD25⁺ Treg cells/CD4⁺ cells was measured by flow cytometry. Proliferation of the splenocytes was analyzed with MTT assay. ELISA was used to determine the levels of interleukin 4 (IL-4) and interferon γ (IFN-γ) in the supernatant of the splenocytes. RESULTS: The expression of Foxp3 at mRNA and protein levels in transfection group was significantly higher than that in empty vector group and control group. The proportion of CD4⁺CD25⁺Treg cells/CD4⁺ cells in transfection group was higher than that in empty vector group and control group. The proliferation of transfected cells was markedly inhibited compared with empty vector group and control group. The levels of IL-4 and IFN-γ were significantly lower in transfection group than those in empty vector group and control group. CONCLUSION: The transfected Foxp3 gene overexpresses in the splenocytes of asthma mice. Foxp3 increases the number of CD4⁺CD25⁺ T cells and inhibits the proliferation and production of Th1/Th2 cytokines in splenocytes.     

Cucurbitin E relieves airway inflammation by inhibiting MAPKs/NF-κB signaling pathways in asthmatic mice

AIMTo investigate the effects of cucurbitacin E on airway inflammation and the signaling pathways of MAPKs and NF-κB in asthmatic mice. METHODSHealthy mice (n=40) were randomly divided into control group, model group, low-dose cucurbitacin E group, high-dose cucurbitacin E group and dexamethasone group. Ovalbumin sensitization was used to induce asthma in the mice. The protein levels of p-JNK, p-ERK1/2, p-p38 MAPK and p-p65 in the lung tissues were determined by Western blot. RESULTSCompared with control group, the numbers of inflammatory cells, such as eosinophils, lymphocytes and neutrophils, were significantly increased in model group, and the activity of MAPKs and NF-κB signaling pathway-related proteins was significantly enhanced. Cucurbitin E at high dose attenuated airway inflammation in asthmatic mice, and significantly inhibited the activity of MAPKs and NF-κB signaling pathway-related proteins. Histopathological results showed proliferation of goblet cells and bronchial mucosal epithelial cells, infiltration of inflammatory cells in the alveoli, and narrow alveolar cavity in model group, while the pathological changes were significantly alleviated in cucurbitin E treatment groups. CONCLUSION Cucurbitin E improves airway inflammation in asthmatic mice, and its mechanism may be related to the inhibition of MAPKs and NF-κB signaling pathways.     

Expression of nuclear factor-κB in asthmatic guinea pigs and the effect of erigeron breviscapus on it

AIM:To explore the expression of nuclear factor-κB(NF-κB)in asthmatic guinea pigs,and the effect of erigeron breviscapus,a protein kinase C(PKC)inhibitor,on the expression of nuclear factor-κB(NF-κB).METHODS:48 guinea pigs were randomly divided into 6 groups(n=8).Airway resistance and eosinophilic inflammation of airway wall were examined,the expression of NF-κB in the lung tissue was detected by immunohistochemical staining.RESULTS:The expression of NF-κB was mainly found in airway epithelium,all the asthmatic animals showed significantly higher optical densities than that of the normal control group(P＜0.01),and the rats subjected therapeutic treatment for two weeks showed significantly lower NF-κB expression than those of the asthmatic groups(P＜0.01).Positive correlation exist between the airway resistance and the percentage of cells expressing NF-κB in epithelium,and between the amount of eosinophil in airway wall and the percentage of cells expressing NF-κB in epithelium(P＜0101).CONCLUSION:The increased expression of NF-κB in airway epithelium of the asthmatic guinea pigs suggested that NF-κB may be involved in asthma.And result that the increased expression of NF-κB was inhibited significantly by the treatment of the erigeron breviscapus suggested that PKC may play a significant role in the pathogenesis of asthma through NF-κB.