| Drug metabolism by cytochrome P450 enzymes is an oxygen-dependent process. At high altitudes, the body stays in a low-pressure and low-oxygen environment. Under such harsh conditions, the activities of all CYP450 enzymes in vivo are affected differently. Changes of CYP450 enzyme may directly affect drug efficacy and adverse reactions may occur. Therefore, systematic study on changes of CYP450 at different altitudes is of important significance. This experiment mainly studies the activity and protein expression changes of CYP450 isoforms after acute exposure to plateau, investigates the mechanism and observe activity changes of CYP450 enzyme under drug intervention (enzyme inducers and inhibitors). It explores activity changes of CYP450 enzyme under the dual intervention of hypoxia and drugs, detects changes in biochemical parameters of rats and observes changes in liver pathology and electron microscopy to evaluate the effect of hypoxia on rats’ liver injury after acute exposure to high altitude.Healthy adult Wistar rats aged 7-8 weeks, weighing 200±20g, were obtained from the ShangHai SLAC Laborytory Animal CO.LTD, and were randomly assigned into three groups according to their weight, respectively. Group A (plain area,55m above sea level), Group B (high altitude,4300m above sea level) and Group C (hypobaric hypoxia cabin,4300m above sea level) and with each group containing 10 animals. The rats were used for experiments and all efforts were made to minimize any animal suffering, and kept from eating for twelve hours before anatomical examination. The main blood gas levels and biochemical criterion were detected and the liver tissue pathological slices and electron microscopic observation were observed. The enzymatic activity of CYP450 isoforms as well as the total protein, total CYP450, cytochrome b5 and the protein expression of CYP450 isoforms were detected. PXR CAR AhR expressions and cytokines in the liver were assayed with western blot and elisa kits. Plain area and 4300m above sea level groups were given phenobarbitale (A1 B1), dexamethasone(A2 B2), armazide(A3 B3) and cimetidine(A4 B4) drug intervention and detected activity of CYP450 and cytokines in the liver.Blood results showed that:compared with group A, the pCO2, pO2, SO2%decreased significantly, HCO3-, SBC, BE and BB significantly decreased by group B. Rats acute exposure to high altitude had a combination of respiratory alkalosis and metabolic acidosis in high altitude environment, suggesting that the rats’ renal and cardiopulmonary function were likely to impaired and they suffered chronic liver injury with inflammation. Biochemical results showed:compared with group A, AST and ALT increased significantly, ALP and DBIL significantly decreased by group B, suggesting that the rats’ suffered liver injury, which has been demonstrated by the pathological results at the same time. Compared with Group A enzymatic activity of CYP1A2 and CYP2C11 were significantly reduced respectively and CYP3A1 increased significantly. The protein expression of CYP450 isoforms CYP1A2 and CYP2C11 were significantly reduced respectively and CYP3A1, CYP2D4 had no significant changes. The activity and expressions of CYP450 have changed in high altitude, we detected nuclear receptor content and cytokines, to discuss mechanism of them. The expression of PXR was significantly decreased compared with the plain group. Determination of cytokine levels showed that:after exposure to high altitude, the IFN-y, IL-2, IL-1β and IL-6 content were significantly increased compared with group A. The activity of CYP450 enzyme changes and cytokine levels change have good corresponding relation.After giving CYP450 enzyme inducers intervention:Compare with group A, the activity of CYP2C11 and CYP3A1 increased significantly in group A1; Compare with group B, the activity in group B, of CYP1A2,2C11 and 2D4 were significantly increased, but the CYP3A1 was significantly decreased by 40.63% after phenobarbital intervention. After dexamethasone induced, the activity of CYP3A1 and CYP2D4 were significantly increased in groupA2 compare with group A; Compare with group B, CYP2C11 and CYP2D4’s activity were significantly increased in group B2, the CYP3A1 activity was significantly decreased 65.24%. After giving CYP450 inhibitors intervention, isoniazid inhibits CYP1A2 activity in A3 group. Isoniazid inhibits this CYP1A2 activity in the plain group. Compare with group A, the activity of CYP1A2 was significantly decreased by 39.63%in group A4, compare with group B, the activity of CYP2C11 and CYP3A1 were significantly decreased in group B4 after cimetidine intervention.In 55 m altitude, compared with the blank group, Phenobarbital makes the content of IL-2 and IL-6 increased significantiy; Dexamethasone raised IFN-y, IL-2 and IL-6; Isoniazid and cimetidine increased IFN-y, IL-2 and IL-6; All groups of the expression of IL-1β is significantly decreased. In high altitude, compared to group B, phenobarbital makes IFN-y, IL-1β and IL-2 levels significantly decreased; Dexamethasone down-regulate IL-1β and IL-2; Isoniazid makes IFN-y, IL-1β and IL-2 levels significantly decreased; Cimetidine increased the content of IL-6, and IFN-y and IL-1β significantly decreased.A lot of research work has confirmed that high altitude environment has significant influence on CYP450. A lot of adverse drug reaction is basically connected with the usage of conventional medicines and drug metabolism is closely related to enzyme P450. This experiment is conducted in high altitude environment. Compared with simulation chamber, there exist more factors, such as radiation and coldness, therefore the experimental data can better reflects the real condition of high altitude and has good reference. The study of enzyme P450’s content expression and activity changes in high altitude environment provides theoretical foundation for basic research in drug metabolism and offers references for the study of clinical rational drug use in high altitude area. |
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