| Objective (s):To establish the animal model to simulate obstructive sleep apnea hypopnea syndrom (obstructive sleep apnea hypopnea syndrome, OSAHS) of Intermittent Hypoxia (Intermittent Hypoxia, IH) model.Research in different concentrations of intermittent hypoxia effect on inflammatory cytokines, inflammatory cells,damage and protect factors and the influence of the interactionin in lung tissue.Methods:1. To establish animal model of different concentrations of intermittent hypoxia in rats,Thirty adult male Wistar rats of 200-220g were divided into 5 groups,6 rats in each group, blank control group (under normoxia natural exposure, NC),5% oxygen concentration group,7.5% oxygen concentration group,10% oxygen concentration group,12.5% oxygen concentration group (IH-1, IH-2, IH-3, IH-4), every day hypoxia exposure time for 9Am-5Pm and exposed for 12 weeks. After rats were exposed to 13 weeks,acquire bronchoalveolar lavage fluid and lung tissue.2. Application of ELISA method to detect lung inflammatory cytokine TNF-a, IL 6, IL-1 beta concentration.3. Detection the count of leukocytes, multinuclear cells and mononuclear cell in BALF.4. Application of lung tissue make pathological slices, observed the pathological changes of lung tissue under different concentrations of IH.5. Using Western Blot method to detect the protein levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), heme oxide synthase (HO-1) and nuclear factor E2 related factor 2 (Nrf2) in lung tissue of each group.Results:1. Markers of inflammation within intermittent hypoxia rat lung tissue:(1) Compared with NC group, With oxygen concentration decreased in lung tissue of the IH group TNF-a levels were rising trend.Comparison among groups had significant difference (F=72.611, P= 0.000), in addition to IH-4 with NC group, the differences among other groups with statistical significance (P< 0.05).(2) Compared with NC group, With oxygen concentration decreased in lung tissue of the IH group IL-6 levels were rising trend.Comparison among groups had significant difference (F=18.019, P= 0.000).IH-1 and IH-2 compared with NC group difference were statistically significant (P< 0.05), IH-1 group compared with IH-4 group difference was statistically significant (P< 0.01), there was no significant difference between the other groups.(3) Compared with NC group, With oxygen concentration decreased in lung tissue of the IH group IL-1β levels were rising trend.Comparison among groups had significant difference (F=16.30, P= 0.000).IH-1, IH-2 and IH-3 compared with NC group difference were statistically significant (P< 0.05), IH-1 group compared with IH-4 group difference was statistically significant (P< 0.01), there was no significant difference between the other groups.2. Effects of IH on the cell count of bronchoalveolar lavage fluid in rats:(1)Compared with NC group, With oxygen concentration decreased in bronchoalveolar lavage fluid of the IH group leukocytes count were rising trend.Comparison among groups had significant difference (F=74.82, P= 0.000), in addition to IH-2 with IH-3,IH-1 with IH-2 group, the differences among other groups with statistical significance (P<0.05).(2)Compared with NC group, The IH group of multinuclear cells count in bronchoalveolar lavage fluid were increased. Comparison among groups had significant difference (F=8.282, P= 0.000). IH-1, IH-2 and IH-3 compared with NC group difference was statistically significant (P< 0.05),there was no significant difference between the other groups.(3)Compared with NC group, With oxygen concentration decreased in bronchoalveolar lavage fluid of the IH group mononuclear cell count were rising trend.Comparison among groups had significant difference (F=61.517, P= 0.000), in addition to IH-1 with IH-2 group, the differences among other groups with statistical significance (P<0.05).3. The lung pathological section express:lung inflammation damage increased with the decreasing concentration of oxygen. IH-1, IH-2 group of lung tissue in addition to inflammatory cell infiltration is also accompanied by the formation of pulmonary fibrosis.4. Markers of damage and protect within intermittent hypoxia rat lung tissue:(1) Compared with NC group, With oxygen concentration decreased in lung tissue of the IH group iNOS protein expression levels were rising trend.Comparison among groups had significant difference (F= 149.01, P= 0.000), in addition to IH-4 with NC group, the differences among other groups with statistical significance (P<0.05)(2)Compared with NC group, The IH group of COX-2 protein expression levels in lung tissue were increased.When the oxygen concentration was reduced to 7.5%,the COX-2 protein expression level reached a peak. Comparison among groups had significant difference (F=79.30, P= 0.000), in addition to IH-4 with NC,IH-1 with IH-3, IH-1 with IH-2group, the differences among other groups with statistical significance (P<0.01)(3) Compared with NC group, The IH-1, IH-2 and IH-3 group of HO-1 protein expression levels in lung tissue were increased.When the oxygen concentration was reduced to 10%,the HO-1 protein expression level reached a peak. Comparison among groups had significant difference (F=9.62, P= 0.000). IH-1, IH-2 and IH-3 compared with NC group difference was statistically significant (P< 0.01), IH-1, IH-2 and IH-3 compared with IH-4 group difference was statistically significant (P< 0.01), there was no significant difference between the other groups.(4) Compared with NC group, There was no significant change in the expression level of Nrf2 protein in the cytoplasm of all IH groups, There was no significant difference Comparison among groups (F=0.892, P=0.503).(5) Compared with NC group, The expression level of Nrf2 protein in the nucleus of IH group were increased in lung tissue. When the oxygen concentration was reduced to 7.5%,the Nrf2 protein expression level reached a peak. Comparison among groups had significant difference (F=28.93, P= 0.000), IH-1 and IH-2 compared with NC group difference was statistically significant (P< 0.05), IH-1 and IH-2 compared with IH-4 group difference was statistically significant (P< 0.05), there was no significant difference between the other groups.Conclusion(s):1. IH may cause inflammation by increase inflammation factors of lung and inflammatory cells count of bronchoalveolar lavage fluid.finally, which can lead to inflammatory damage in the lung tissue.2. IH induced inflammation of lung was a certain concentration dependence of hypoxia.3. The body under the intermittent hypoxia can produce a series of inflammatory material result in lung injury. At the same time, the body to hypoxia can produce a certain degree of tolerance, also may experience initiate protective mechanism to prevent damage to endlessly expand. Through this experiment revealed iNOS and COX-2 and HO-1 and nuclear Nrf2 involved in the pathophysiological mechanism of IH. |
PDF Full Text Request