Investigation Of The Molecular Mechanismo Of IH Induced Hepatic Injure And The Effect Of Autophagy In Liver

Objective:Obstructive sleep apnea(OSA) is a common sleep disorder in which complete or partial airway obstruction, caused by pharyngeal collapse during sleep, determines loud snoring or choking, frequent awakenings, disrupted sleep, and excessive daytime sleepiness. OSA affects over 4% of the general population. The growing clinical relevance of OSA is due to its emerging association with diabetes mellitus, metabolic syndrome, and cardiovascular disease(CVD), independent of other traditional cardiometabolic risk factors and obesity. A lot of evidence suggests that the pathophysiological alteration in gas exchange(repetitive hypoxemic and hyper-capnic events), called chronic intermittent hypoxia(CIH), can lead to increased proinflammatory cytokine production, endothelial dysfunction, oxidative stress, metabolic dysregulation, and insulin resistance. Experimental evidence suggests that CIH may trigger liver injury, inflammation, and fibrogenesis, and, interestingly, OSAS is also believed to be one of the elements promoting the evolution of NAFLD from steatosis tononalcoholic steatohepatitis(NASH). In this study we establish intermittent hypoxia animal model, and aims to observe the structure and the pathological changes of liver tissue, preliminarily explore the inflammation response induced by intermittent hypoxia in the process of changes of hepatic cytochrome P450 enzyme and its regulating mechanism. Further, we study the effect of glococorticoid on expression of Cyp and regulation in autophagy by human LO2 cells.Method: 1. Establish intermittent hypoxia animal model, 30 rats were randomly divided in to intermittent hypoxia group and control group with the random number table(each group has 15 rats). 2. The morphology manifestations of liver tissues were evaluated. 3. The relative gene expression of inflammation factors, Cyps, NF-kappa B, nuclear receptors was evaluated through real-time quantitative polymerase chain reaction(PCR) assay. 4. The relative gene expression of autophagy associated gene was evaluated. 5. LO2 cells were maintained in RPMI 1640 medium supplemented with 10% fetal calf serum, 2m M l-glutamine, 100 units/ml penicillin and 100 μg/ml streptomycin. In the experiments as indicated, Dex was added to the culture medium at 10 n M andRU486 at 10 μM. 6. The morphology manifestations of LO2 cells were evaluated. 7. The relative gene expression of Cyps in LO2 cells were evaluated through real-time quantitative polymerase chain reaction(PCR) assay. 8. The relative gene expression of autophagy associated gene in LO2 cells was evaluated.Result: 1. Examination of liver tissues by H&E staining showed inflammatory infiltrates in rats with IH exposure as compared to control group. 2. Message RNA expression of IL-1β、IL-6、TNFα m RNA was greater in livers from IH group than those from control group. 3. Message RNA expression of NF-kappa B was higher in livers from IH group than those from control group. 4.The hepatic m RNA expression of PXR、CAR and GR was significantly reduced in IH group as compared to control group. 5. The hepatic m RNA expression level of Cyps decreased in IH group;but Cyp1A2、Cyp2D4、Cyp3A2 m RNA was significantly reduce in IH group,in contrast,the change of Cyp2C9、Cyp2C19 m RNA has non-significantly statistics. 6. The expression of m TOR, a blocker of autophagy, was decreased but not significantly in IH group as compared to control group. However, the expression of AMP-activated protein kinase(Ampk), a negative regulator of m TOR pathway, was elevated when rats were exposed to IH. And the expression of LC3 and WIPI, a positive regulator of m TOR pathway, was decreased in IH group as compared to control group. 7. The morphology of LO2 cells treated by DEX is abnormal compared with control group and DEX+antagonist group. 8. The m RNA expression level of h CYP1A2 、 h CYP2C19 、 h CYP3A4 m RNA sianificantly decreased in LO2 cells treated by DEX compared with control group, reversed by DEX antagonist; the m RNA expression level of h CYP2C9 was decreased with DEX treated and h CYP2D6 was non-significantly in the three groups. 9. The expression of autophagy associated gene almost has non-significantly except Beclin1.Conclution: 1. Intermittent hypoxia exposed can induce change of liver structure, promotethe inflammatory cytokines secretion, and regulate the expression of Cyps. 2. IH induced autophagy of hepatocytes by glucocorticoid receptor pathway,consequencely avoiding apoptosis of hepatocytes. 3. Glucocorticords influenced the morphology of human LO2 cells and regulate the expression of Cyps,but no evidence shows the inclined to induce autophagy.