| Objective1.Research the expression and correlation of phosphatidylinositol3kinase serine/threonine protein kinase (PI3K/AKT) and gamma glutamyl cysteine synthase (r-GCS) in Chronic airway inflammation lung tissue of rats, then prove that PI3K/AKT pathway may be involved in the oxidation of chronic obstructive pulmonary disease antioxidant imbalance and invest more oxidation and anti-oxidation mechanism about COPD, and provide new ideas for diagnosis and treatment of COPD.2. Use carbocisteine to intervention the airway inflammation rats, explore its affect on PI3K/AKT and r-GCS pathway, then describe its antioxidant and anti-inflammatory effects. Finally, provide experimental basis for clinical application of carbocisteine.Methods30clean-level male SD rats in the Experimental Animal Center of Zhengzhou University were divided into three groups according to the random number table: normal control group (A), model group (B) and carbocisteine treatment group (C)(n=10). Rat models of chronic airway inflammation were made using the method of twice injection lipopolysaccharide in the trachea plus smoking twice a day in model group and carbocisteine treatment group. Carbocisteine treatment group were given carbocisteine (500mg/kg) orally half an hour before smoking in17-30days morning; saline instead of lipopolysaccharide was injected into the tracheal and normal saline1.5ml was poured into the stomach in normal group. Anesthetized and sacrificed rats in31th days, lavage and cell were counted; Lung tissue were stained with HE; The expression of y-glutamyl cysteine synthase (y-GCS) and the phosphorylation of serine/threonine protein kinase (P-AKT) were determined by immunohistochemical methods; The expression of phosphatidyl inositol3kinase (PI3K) mRNA and y-GCSmRNA were determined by reverse transcription polymerase chain reaction (PCR) semi-quantitative (RT-PCR). All the data were used SPSS17.0software for statistical analysis.Results1. Symptoms of rats:the modeled group appear the symptoms of cough and asthma; carbocisteine treatment group compared with the model group were improved; normal control group showed no abnormal.2. Lung pathology in rats:The airway epithelial cells in normal control group rat was complete, lung tissue structure was normal; There were epithelial cell shedding, severe necrosis and inflammatory cell infiltration in model group; In treatment group, epithelial cell shedding, necrosis and inflammatory cells infiltration were reduced compared with model group.3. Count of the lavage fluid in rats:The total cells in BALF of normal control group, model group, carbocisteine treatment group were (1.65±0.38)×105,(7.32±0.77)×105,(4.20±0.61)×105respectively; The proportion of monocytes macrophages cell in BALF were (81.13±2.58)%,(65.15±4.85)%,(78.15±5.41)%respectively; The proportion of neutrophils in BALF were (13.21±1.76)%,(30.43±4.54)%,(16.14±3.94)%respectively; The proportion of lymphocytes in BALF were (5.66±0.91)%,(4.42±1.19)%,(5.71±2.37)%respectively.4. The expression Of γ-GCS, P-AKT in the lung tissue of rats:the expression of y-GCS in the lung tissue of normal control group, model group and carbocisteine treatment group were113.87±3.49,145.09±9.58and126.51±4.76respectively; the expression of P-AKT in the lung tissue were106.64±5.91,151.33±11.09and130.48±4.86respectively.5. The expression of PI3KmRNA, γ-GCSmRNA in the lung tissues of rats:The expression of γ-GCSmRNA in lung tissue of normal control group, model group and carbocisteine treatment group were0.15±0.02,0.45±0.05and0.21±0.04respectively; The expression of PI3KmRNA in the lung tissue were0.45±0.07,1.17±0.11and0.87±0.09respectively.6. The expression of GSH, ROS, T-AOC in the lung tissue of rats:the activity of GSH in model group was twice times than the control group and was also significantly increased compared with the treatment group, while the ROS increased more significantly, but the T-AOC was significantly lower than that of the treatment group and control group.7. Statistical analysis:total cells and neutrophils in BALF were model group> treatment group> normal control group, but lymphocyte and monocytes macrophages were normal control group> treatment group> model group (the difference were statistically, P<0.05); the expression of γ-GCS, P-AKT, PI3KmRNA, γ-GCSmRNA in lung tissue were the model group> treatment group> normal control group (the difference were statistically, P<0.05);the expression of P-AKT, and γ-GCS and γ-GCSmRNA in the chronic airway inflammation lung tissue of rat were highly significant positive correlation (r=0.812,0.741, P<0.001).Conclusionl.The expression of P-AKT, and γ-GCS and γ-GCSmRNA were relevant, PI3K/AKT pathway may be involved in and promote the expression of γ-GCS protein.2. Carbocisteine is possible improve the oxidant antioxidant imbalance, and exert an antioxidant effect through the effect of PI3K/AKT signal pathway and regulation of γ-GCS expression.3.Carbocisteine can reduce the infiltration of inflammatory cells, clear ROS to play the role of anti-infective, antioxidant effect. Which provide the experimental basis for clinical application of carbocisteine. |
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