Objective To observe the eotaxin expression of rat airway smooth muscle cells ( ASMCs) induced by serum from asthmatic rats, and explore the possible mechanism. Methods ASMCs isolated fromrat tracheas were cultured in vivo. Then they were treated with serum from asthmatic rats, or treated with serum and dexamethasone simultaneously. The level of eotaxin protein in supernatant and eotaxin mRNA in ASMCs were measured by ELISA and reverse transcription-polymerase chain reaction. The expression of cAMP in ASMCs was examined by radioimmunoassay. Results After the treatment with sensitized serum, the eotaxin level in supernatant and mRNA expression in ASMCs were significantly higher [ ( 107. 09 ±7. 12) ng/L vs. ( 0. 63 ±0. 56) ng/L, P lt; 0. 05; 1. 39 ±0. 04 vs. 0. 05 ±0. 01, P lt;0. 05] , and the level of cAMP in ASMCs was significantly lower compared with the control group [ ( 17. 58 ±3. 62) ng/L vs. ( 32. 39 ±3. 36) ng/L, P lt; 0. 05] . After intervened by the sensitized serum and dexamethasone simultaneously, the protein and mRNA expressions of eotaxin were lower compared with those intervened by sensitized serumalone [ ( 64. 18 ±4. 04) ng/L and 0. 77 ±0. 19] . The level of eotaxin in supernatant was negatively correlated with cAMP level in ASMCs ( r = - 0. 788, P lt; 0. 01) . Conclusions There is anautocrine function in ASMCs as inflammatory cells after stimulation with sensitized serum. Eotaxin may play an important roll in the pathogenesis of asthma via a cAMP-dependent pathway.
ObjectiveTo investigate the expressions of IL-10,tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ) in serum and lung tissue of COPD rats in order to elucidate the potential mechanism of airway inflammation. MethodsForty-five healthy adult male SD rats were randomly divided into a COPD model group (n=30) and a normal control group (n=15). The COPD rat model was established by intratracheal instillation of lipopolysaccharide (LPS) and exposure to cigarette smoke for 28 days. The concentrations of IL-10,TNF-α and IFN-γ in serum and lung tissue were measured by ELISA. ResultsTNF-α level of serum and lung tissue in the COPD model group increased significantly compared with the control group(P<0.05),while the levels of IFN-γ and IL-10 decreased significantly[serum:(44.68±8.67) ng/L vs. (75.96±10.59) ng/L;lung tissue:(64.55±9.03) ng/L vs. (94.06±8.71) ng/L,P<0.01]. The level of IL-10 in serum and lung tissue was negatively correlated with TNF-α (serum:r=-0.67,lung tissue:r=-0.80,P<0.01). The level of IL-10 in serum and lung tissue was positively correlated with IFN-γ (serum:r=0.64,lung tissue:r=0.72,P<0.01). The level of IL-10 in serum and lung tissue was negatively correlated with the percentage of neutrophils(serum:r=-0.70,lung tissue:r=-0.67,P<0.01). ConclusionIn COPD rats,down regulation of IL-10 plays an important role in regulation of airway inflammation.
Objective To investigate the mRNA and protein expression of human β-defensin-2 (hBD-2) induced by lipopolysaccharide (LPS)，IL-1β and TNF-α in human airway primary epitheliums.Methods The bronchial primary epitheliums from human were stimulated with LPS，IL-1β and TNF-α respectively and then were harvested for hBD-2 expression detection.The mRNA expression of hBD-2 was detected by RT-PCR，and the protein expression by immunocytochemistry and western blot.Results There was a small expression of hBD-2 mRNA in human airway primary epitheliums before stimulation.The hBD-2 mRNA expression was significantly increased after 3 hours of LPS，IL-1β and TNF-α stimulation respectively and the expression increasement was in a dose dependent manner.The hBD-2 protein could be detected in cytoplasm after 4 hours of LPS (0.1 μg/mL)，IL-1β (1 ng/mL) and TNF-α (10 ng/mL) stimulation.Conclusions LPS and proinflammatory cytokines can induce the mRNA and protein expression of hBD-2 in a short time.The expression of hBD-2 may play an initial defense role against bacterial invasion.
Objective To explore the effects of Aspergillus fumigatus（A. fumigatus） spores on airway inflammation and responsiveness in asthmatic rats.Methods Seventy male Wistar rats were randomly divided into Ⅰ and Ⅱ groups（n=35 in each group），then Group Ⅰ and Group Ⅱ were subdivided into a normal control group（n=5），an asthma group（n=10），a spores-treated control group（n=10），and a spores-treated asthma group（n=10）.The rats were sensitized to ovalbumin（OVA） and challenged with aerosol OVA to establish the asthma model.The effects of A. fumigatus spores on asthmatic rats before and after OVA aerosol challenging were investigated in Group Ⅰ and Group Ⅱ，respectively.The parameters associated with bronchial epithelial damage were observed by total protein concentration in BALF measured by BCA method.Total and differential cell counts in BALF were also counted.The airway resistance and airway responsiveness were calculated by transpulmonary pressure and gas flow rate.Results In Group Ⅰ，the total protein in BALF in the asthma group treated with A. fumigatus spores before OVA challenging（Group CA） was increased remarkably compared to the asthma group（Group A1）[（1.125±0.254）μg/mL vs（0.825±0.173）μg/mL，Plt;0.01].The nonspecific airway resistances induced by different concentration of acetylcholine in Group CA ［（0.997±0.196）cm H2O•mL-1•s-1，（1.123±0.142）cm H2O•mL-1•s-1，（1.130±0.197）cm H2O•mL-1•s-1］were increased significantly compared to Group A1 [（0.655±0.089）cm H2O•mL-1•s-1，（0.687±0.048）cm H2O•mL-1•s-1，（0.821±0.043）cm H2O•mL-1•s-1]（all Plt;0.05）.In Group Ⅱ，however，the above parameters in the asthma group treated with A. fumigatus spores after OVA challenging（Group AC） were not dramatically increased compared with the asthma group（Group A2）（all Pgt;0.05）.The differences in the total and differential cell counts in BALF in Group CA were not remarkable compared to other subgroups in Group Ⅰ（all Pgt;0.05）.But the BALF neutrophil count in Group AC was increased obviously compared to Group A2 ［（2.488±0.420）×106 vs （0.936±0.459）×106，Plt;0.05］.Conclusion These data indicate that exposure to A. fumigatus spores before challenging causes aggravated epithelial damage and increased airway resistance in an asthma rat model.
Objective To explore the role of nuclear factor kappa B(NF-KB)in the pathogenesis of chronic obstructive pulmonary disease(COPD)and the therapeutic efects of glucocorticoid．Methods Twenty-four Wistar rats were randomly divided into three groups，ie．normal control，COPD model and prednisone preventive treatment group．Rat COPD model Was established by exposing the rats to cigarette smoke daily．Prednisone Was given through stomachal injection on altemate days．After COPD model Was set up，bronchoalveolar lavage(BAL)Was performed．Total cell counts and neutrophil counts in BALF were examined．Pathological changes of lung tissue Was observe0 by hematoxylin-eosin staining．The morphological indices of pulmonary emphysema(MLI，MAN and PAA)Was measured by a computerizedimage analyzer and compared in three groups．NF-KB expression in lung tissues were detected by immunohistochemistry assay．Rults Emphysema Was confirmed by three morphological indices in COPD model group compared to those of normal control group[MLI：(97.97±11.10)×10-6m vs (47．23±2．80)×10-6 m，MAN：(95.98±l4.89)×106 ／m vs (164.21±9．30)×106 ／m ，PAA：(64 ±5.7)％vs (44±2.7)％，Plt;0.01]．Total cell counts and neutrophil counts in BALF of COPD model group were significantly higher than those of control group[（5.76±0.29）×108/L vs （1.64±0.12）×108/L，（1.26±0.25）×108/L vs （0.099±0.065）×108/L，Plt;0.01]．After the preventive treatment with prednisone，MLI，MAN and PAA were significantly changed[（57.66±4.62）×10－6mvs （97.97±11.10）×10－6m，（111.40±16.92）×106个/m2 vs （95.98±14.89）×106个/m2，Plt;0.01；（58±6.1）% vs （64±5.7）%，Plt;0.05]，which indicated that airway inflammation and emphysematous injury in preventive treatm ent group were milder than those of COPD mode1．Total ceil counts and neutrophil countsin BALF were found in preventive treatment group as compared to those of COPD model[[（3.18±0.29）×108/L vs （5.76±0.29）×108/L，（0.57±0.12）×108/L vs （1.26±0.25）×108/L，Plt;0.01].The percentage of positive cells of NF-KB nuclear staining in bronchiolar epithelial ceils was significantly increased in the COPD group than that in the control group[（29.02±1.25）% vs （12.17±1.13）%，Plt;0.01]，but was significantly decreased in the preventive treatment group[(19.23±1.18)％vs (29.02±1.25)％，Plt;0．01]．Conclusions NF-KB may be responsible for the persistence and amplification of inflammation in COPD through neutrophil recruitment and activation．Prednisone may suppress airwayinflammation in COPD by inhibiting NF-KB．
Objective To investigate the relationship of small airway function with airway sensitivity and reactivity and assess the factors influencingairway hyperresponsiveness (AHR).Methods Data of consecutive subjects with suspected asthma who had a≥20% reduction in FEV1 after ≤12.8 mmol/L cumulative doses of methacholine were analyzed from January 2005 to April 2006.Airway sensitivity was assessed by the cumulative dose of methacholine required to cause 20% reduction in FEV1 (PD20).Airway reactivity was analyzed using the slope of the dose-response curve (DRS).The DRS was defined as the reduction in FEV1 from baseline after the final dose of methacholine inhaled divided by the cumulative dose inhaled.Because of their highly skewed distribution，DRS was logarithmically transformed (log10) for all analysis.Results A total of 184 consecutive subjects aged 16 to 80 years was enrolled.There were 70 male (38.0%) and 114 female (62.0%) subjects.Subjects with higher airway sensitivity，indicated by lower PD20，also had a lower Vmax50% and Vmax25%，and vise versa.PD20 was negatively correlated wit log10DRS (r=-0.874，Plt;0.01).In a simple linear regression model，log10DNS was significantly correlated with FEV1%，Vmax50% or Vmax25% respectively (the determinant r2 were 0.062，0.097 and 0.085，respectively，all Plt;0.01).In a multiple linear regression model that included age，height，and percentage of predicted FEV1，Vmax50% and Vmax25% accounted for 3.9% and 2.6%，respectively，of variability in airway reactivity.The association between Vmax50% and log10DNS was significant in both male and female subjects.The r2 was higher in male subjects.The subjects were divided into three age groups and the association between Vmax50% or Vmax25% and log10DNS was higher in female than in male for age≤25 years，higher in male than in female for 25 -45 years.No association was found for agegt;45 years in both males and females.Conclusions Impaired small airway function is associated with higher airway sensitivity and reactivity to methacholine in subjects with suspected asthma.
Objective To examine the effects of Pseudomonas aeruginosa(PA)quorum-sensing systems on airway mucus hypersecretion．Methods Sixty Sprague-Dawley rats were intubated with a silicone tube pre-coated with PAO1(wild-type PA strain)，PAO1-JP2(quorum-sensing-mutant strain)or saline in the bronchus．After 28 days，the mRNA and protein expression of MUC5AC in the rats’bronchial epithelia were detected by RT-PCR，alcian blue／periodic acid—Schif(AB／PAS)staining and enzyme linked immunosorbent assay(ELISA)．Results In the PAO1 group，bronchiolar epithelium goblet cells by AB／PAS staining was significantly more than those in the PAO1-JP2 and control groups(both Plt;0．05)．The expression level of MUC5AC mRNA in the PAO1 group was significantly higher than those in the PAO1-JP2 and control groups(both P lt;0．05)．The ELISA showed that the concentration of MUC5AC protein in bronchoalveolar lavage fluid(BALF)in the PAO1 group was much higher than that in the PAO1-JP2 group(P lt;0．05)．Conclusion PA quorum-sensing system plays an important role in airway mucus hypersecretion
Objective To study the intervention effect of ginkgo biloba extract（GBE） on airway and vascular remodeling in rat model of chronic obstructive pulmonary disease(COPD).Methods Forty wistar rats were randomly divided into group A，B，C and D.The rat model of COPD were established by intratracheally injection of lipopolysaccharide and exposure to cigarette smoke in groups B，C and D.Groups C and D were given intraperitoneally injection with 40 mg/kg GBE respectively from day1 to day14 and day29 to day42.Forty-three days later，the rats were sacrificed for lung pathological examination.Results Group B，C and D all showed pathological changes characteristic of COPD to different extent.The average area and standard number of alveoli showed significant difference between each groups（all Plt;0.01）.The structure of bronchiole walls in group C and D show mild changes.The ratio of bronchial smooth muscle thickness to bronchial wall thickness and bronchial wall area to bronchial area of group C and D showed significant difference when compared with group A and B（all Plt;0.01）.The vascular smooth muscle cell of group C and D had mild hyperplasia and the vascular wall had slightly thickened.The ratio of vascular smooth muscle thickness to vascular wall thickness and vascular wall area to vascular area in group C and D showed significant difference when compared with group A and B（all Plt;0.01）.Conclusion GBE has inhibitory effects on airway and vascular remodeling in rats model of COPD.
Objective To explore the effects of prolonged Aspergillus fumigatus spores inhalation on airway inflammation and remodeling in rats with chronic obstructive pulmonary disease（COPD）.Methods Fifty Wistar rats were randomly divided into group A，B，C，D and E，(n=10 in each group) and group E was served as normal control.In group A，B，C and D，COPD models were established by intratracheal administration of lipopolysaccharide (LPS) combined with cigarette smoke exposure.The rats in group A，B and C were given intranasal inhalation of 1×106cfu spores，1×103cfu spores and 100 mL saline twice a week for consecutive 5 weeks，respectively，while the rats in group D were given no treatment.Bronchoalveolar lavage fluid(BALF) were collected for total and differential cell count，and interleukin-8（IL-8） and transforming growth factor-b(TGF-b) concentration measurement.The pathologic changes of lung tissue were observed by HE，PAS and Masson stainings.Results Pathological changes characteristic of COPD were found in group D.The total cell count，the percentage of neutrophile and lymphocyte in BALF in group A and B were higher than those in group C and D(all Plt;0.01).IL-8 and TGF-b in BALF in group A and B were higher than those in group C and D(all Plt;0.01).The pathologic score of airway inflammation in group A was higher than those in group B，C and D(all Plt;0.01)：The thickness of airway wall(WAt/Pbm) and airway smooth muscles(WAm/Pbm)，the collagen deposition in the total airway wall(WCt/Pbm) and in the outer airway wall(WCo/Pbm) and the percentage of goblet cells to epithelial cells in group A and B were higher than those in group C and D(all Plt;0.01).In group A and B，IL-8 was positively correlated with the percentage of neutrophile(r=0.856，Plt;0.01)，the pathologic score of airway inflammation(r＝0.884，Plt;0.01)，and the percentage of goblet cells to epithelial cells (r=0.702，Plt;0.05)，respectively.TGF-b was positively correlated with WAt/Pbm，WCt/Pbm，WCo/Pbm and the ratio of goblet cells to epithelial cells (r=0.706，Plt;0.05：r=0.802，Plt;0.01：r=0.876，Plt;0.01：r=0.713，Plt;0.05).Conclusion Prolonged inhalation of Aspergillus fumigatus spores can aggravate the airway inflammation and remodeling in rats with COPD.
Objective To investigate the role of endogenous Hydrogen Sulfide ( H2S) in airway inflammation and responsiveness in a rat model of chronic passive-smoking. Methods Male SD rats were randomly divided into a control group ( breathing fresh air) and a passive smoking group [ cigarette smoking( CS) passively] , with 18 rats in each group. Six rats in each group were randomly intraperitoneally injected with normal saline, sodium hydrosulfide ( NaHS) or propargylglycine ( PPG, an irreversible inhibitor of cystathionine- γ-lyase) . The animals were divided into six subgroups, ie. Con group, NaHS group, and PPG group, CS group, CS+ NaHS group, and CS + PPG group. After 4 months, lung histological change and airway tension were measured. The H2S levels of plasma and lung tissue were analyzed by the sensitive sulphur electrode assay. The expression of cystathionine-γ-lyase ( CSE) was measured by western blot. Results Compared with the Con group, CSE protein expression in lung tissues was increased in CS group( P lt;0. 05) ; the H2 S levels of plasma were significantly higher in CS group, NaHS group and CS + NaHS group, and much lower in PPG group ( P lt; 0. 05, respectively) . Compared with CS group, the H2S levels of plasma were significantly higher in CS + NaHS group, and much lower in CS + PPG group( P lt; 0. 05, respectively) . The H2S level of lung tissue in each group had no significant difference ( P gt; 0. 05) . Compared with Con group,score of lung pathology was significant elevated, and the responsiveness of airway smooth muscles to ACh and KCl was significant augmented in CS group. Compared with CS group, the score of lung pathology was decreased, and the responsiveness of airway smooth muscles was decreased in CS +NaHS group( P lt;0. 05) , and vise versa in CS + PPG group( P lt; 0. 01) . Conclusion H2S can alleviate airway inflammation and hyperresponsiveness induced by CS, and administration of H2S might be of clinical benefit in airwayinflammation and airway responsiveness.