| Backgrouds:Currently,a large number of studies show that aseptic inflammatory response plays an important role in the secondary brain injury after traumatic brain injury(TBI).It's well known that nuclear factor kappa B(NF-?B)functions as a key regulator in the inflammatory response.Although there is lots of research focusing on NF-?B,it is still obscure about the specific role in inflammatory response.It is reported that there are double peaks of cerebral NF-?B activation after hypoxia-ischemia and subarachnoid hemorrhage in animal experiments,showing brain injury at the early stage and brain repair at the late stage respectively.Our previous studies found that NF-?B activity was up-regulated in early stage and remained elevated at day 7 after TBI.However,data are lacking regarding the panoramic view of NF-?B activity and expression of NF-?B subunits after TBI.Accordingly,in the current study,we designed to detect the time course of NF-?B activation and expression of NF-?B p65 and c-Rel subunits around the contused cortex during 14 day following TBI,and to observe the effects of pyrrolidine dithiocarbamate(PDTC)admirustration in secondary brain injury at different time points after TBI.Methods:An experimental TBI model was induced in male Sprague-Dawley(SD)rats,and they were randomly divided into sham and TBI groups at different time points on day 1,day 3,day 5,day 7,day 10 and day 14,respectively.Then the NF-?B DNA-binding activity around the contused brain was detected by electrophoretic mobility shift assay(EMSA),and the expression of subunits p65 and c-Rel was detected by western blot and immunohistochemical studies.The concentrations of tumor necrosis factor-?(TNF-?)and interleukin-1?(IL-1?)were measured by enzyme linked immunosorbent assay(ELISA).PDTC was given an intraperitoneal injection at a dose of 100mg/d/kg at different stages after TBI according to the different peaks of NF-?B activation.EMSA was taken to detect the NF-?B activity,and Western blot was also performed to measure the protein levels of p65 and c-Rel.ELISA was used to measure the contents of TNF-? and IL-1?.Moreover,Nissl staining was used to detect the number of surviving cells,and modified neurological severity scores(mNNS)were used to evaluate the neurological function.Results:There are double peaks of cerebral cortical NF-?B activity at 3 d and 10 d post-injury,respectively.Additionally,protein levels of p65 were found to be elevated and peaked at 3 d and 10 d after TBI.Furthermore,p65 protein expression at 3 d was much higher than that at 10 d,and it decreased to the baseline at 14 d after TBI;while levels of c-Rel were significantly elevated during the late phase of injury,and still showed a trend of increase at 14 d following TBI.TNF-? and IL-1? concentrations also showed a biphasic increase,and they were enhanced immediately at 1 d,and then went down to baseline at 7 d,followed by another increase till 14 d after TBI.NF-?B DNA binding activity and protein levels of p65 and c-Rel were down-regulated after PDTC administration as compared to the TBI group,and the p65 protein expression decreased more sharply than c-Rel.The contents of TNF-?and IL-1? were also lowered after PDTC administration.Furthermore,the number of surviving cells markedly increased,and the neurological severity recovery was much better after PDTC administration as compared with that in TBI group.Conclusions:Our findings suggested a biphasic activation of cerebral cortical NF-?B after experimental TBI in rats.Meanwhile,we found that the different subunits p65 and c-Rel could be activated at different phases.PDTC administration could significantly inhibit the DNA binding activity of NF-?B,and reduce the protein levels of p65 and c-Rel.Moreover,PDTC can also significantly improve the cell survival during both early and late stages after TBI.In conclusion,the results indicated that PDTC might serve as a novel therapeutic target with a wide time window for TBI. |
PDF Full Text Request