| Part1: The dynamic changes of Cox-2 mRNA and protein expression in radiation-induced brain injury in rats modelsObjective: To observe and explore the rule of dynamics changes of Cox-2 mRNA and protein expression in radiation-induced brain injury in rats models.Methods:A total of 112 healthy SD rats were randomly assigned into the control group(only anesthesia not irradiated,56) and the irradiation group [Irradiated with electron beam(5MeV)20Gy to establish an animal model of radiation-induced brain injury,56]. All rats beheaded and take out the brain tissue at the time points of 3 h, 12 h, 1 d, 3 d, 7 d, 15 d, 30 d after irradiation respectively. The pathological changes were observed by the hematoxylin-eosin staining. The QRT-PCR was used to detect the levels of Cox-2 mRNA expression changes. Western-Blot and immunocytochemistry methods were used to measure the change trend of total Cox-2 protein in brain tissue.Results:1.The HE staining showed the rats’ brain cells experienced edema,fatty degeneration,nuclear condensation,cytoplasmic vacuoles,capillary disorder of pathological changes after irradiated.2.In the irradiation group,the expression of Cox-2 mRNA increased gradually at first, and thendecreased after the peak,compared with the blank,the expression at 3h,12 h,1d,3d,7d were statistically significant(P<0.05).3.The Western-Blot analysis showed the expression of the Cox-2 protein increased after irradiation and reached the peak at the third day,then decreased to the normal at the 30 th day.4.The immunohistochemical analysis showed the expression trend of Cox-2 protein in rats’ hippocampus of irradiation group wasconsistent with the results of Western-Blot.Conclusions: The expression of the Cox-2 mRNA and protein increased in the radiation-induced brain injury models, which may play the role in the development of inflammation,vascular injury of brain tissue. Part2:The study on the protective mechanism of the Cox-2 inhibitors in the radiation-induced brain injury models.Objective:To observe whether celecoxib, a selective Cox-2 inhibitor, could alleviate the RBI in the rat’s models, and to explore the protective mechanisms.Methods:A total of 240 healthy rats were randomly assigned into the control group(only anesthesia,n=80), the irradiation group [Irradiated with electron beam(5MeV)20Gy to establish animal models 0f radiation-induced brain injury,n=80] and irradiation + drug group[Irradiated with electron beam(5MeV)20Gy, an given 30mg/kg celecoxib everyday from the day before irradiation to the day after irradiation by lavage.n=80]. All rats were terminated and taken out the brain tissue at the time points of 3h,12 h,1d,3d,7d,15 d,30d respectively. The hematoxylin-eosin staining was used to observe the pathological changes.Brain water contents(BWC) were determined by wet/dry weight method. Determination of blood brain barrier integrity with detection brain EB contents.The QRT-PCR was used to detect the levels ofNF-κB and VEGF mRNA expression changes. Western-Blot and immunocytochemistry methods were used to measure the change trend of totalNF-κB and VEGF protein in brain tissue.Results:1.The HE staining showed that the nerve cells in the irradiation group and the irradiation+drug group all experienced edema, fatty degeneration, the nuclear pycnosis, cytoplasmic vacuoles, caoillaries gap widened, micro thrombosis, pathological changes after radiation when compared with the control group. But the brain injury in the irradiation+drug group were more mildthan that in the irradiation group significantly.2.The EBanalysis showedthe content of EB in the irradiation group and the irradiation+drug group were both increased after irradiation and reached the peak at the first day, then decreased to the normal at the 30 th day. And at each time points, the content of EB in the irradiation+drug group were lower than irradiation group. Compared with the control group, the results at all the time points of the irradiation group and the irradiation+drug group were statistically significant( P<0.05). Compared with the irradiation group, the content of EB in the irradiation+drug group at 3h,12 h,1d,3d,7d werelower statistically significant(P<0.05).3.TheBWC analysisshowed the content of water in brain tissue increased after irradiation and reached the peak at the seventh day, then decreased gradually. Whether in the irradiation group or the irradiation+drug group, the content of water in the brain at the all time points were larger than that in the control group(P<0.05). Conpared with the irradaiton group, the content of water in the irradiation+drug group were less statistically significant at the time points of 12 h, 1d, 3d, 7d, 15d(P<0.05).4.The QRT-PCR analysis showed the expression of the NF-κB mRNAin the irradiation group increased after irradiation and reached the peak at the third day, then decreased to the normal at the 30 th day. While in the irradiation+drug group, the expression of theNF-κB mRNA were reached the peak at seventh day, and compared with the irradiation group, the expression at the time points of 3h, 12 h, 1d, 3d, 15 d were down-regulated statistically significant(P<0.05).The expression of the VEGF mRNAin the irradiation group and the irradiation+drug group were both increased after irradiation and reached the peak at the seventh day, then decreased to the normal at the 30 th day. Compared with the irradiation group, the expression in the irradiation+drug group at the time points of 3h, 12 h, 3d, 7d were down-regulated significantly(P<0.05).5.The Immunohistochemistry and Western-Blot analysisshowed that compared with the irradiation group, the expression of NF-κB and VEGF protein were down-regulated significantly in the irradiation+drug group.Conclusions: The selective Cox-2inhibitor Celecoxib could delay and reduce theradiation-induced injury in the brain tissue, cerebral edema, blood brain barrier damage.And it could reducehe extent of radiation induced inflammation and vascular damage in the brain via inhibiting the expression of NF-κB and VEGF mRNA and protein. |
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