| Objective:To investigate the expression and relationship of nuclear factor-kappaB (NF-κB) and inflammatory mediators Tumor Necrosis Factor-alpha(TNF-a), nitric oxide synthase(NOS) in hippocampus in a rat model of chronic constrictive injury (CCI).so as to research the effect of NF-κB and inflammatory mediators TNF-a, NOS in chronic pain rat.Methods:Models of neuropathic pain were made by ligating the sciatic nerve of rats. Seventy-six male Wister rats were randomly divided into 2 groups (n=38), the CCI group in which the rats suffered the chronic constriction injury and the sham group in which rats suffered the sham operation. The models of neuropathic pain were established according to the Bennett and Xie s method. The mechanical and thermal nociceptive thresholds were assessed with paw withdrawal latency (PWL) to von Frey filaments and radiant heat at 1d before CCI, 1d,4d,7d,14d,21d, and 28d (n=5,respectively) after CCI. Afterwards rats were sacrificed and the hippocampus was taken. The mRNA expression of each factor was tested by Real time polymerase chain reaction (RT-PCR).Then according to the result of RT-PCR, another six male Wistar rats (n=3 for each group) were sacrificed at 7d after operation and hippocampus were taken for cryosections for immunofluorescence histochemical staining for the expression of NF-κB and TNF-a in hippocampus.Results:1) The changes of thresholds of machine and therm:In CCI group, the time of limb-strike was shorter in the operated side, and the right foot twisted and limped. Occasionally protective phenomenon occurred in hind limbs, such as licking and hanging. However none of them happened in sham group and untreated limbs.There was no significant difference of the pain threshold at 1d before CCI(P>0.05). When compared with before CCI, thresholds of machine and therm in CCI group began to decrease 1 day after operation (P>0.05). Mechanical thresholds declined to the lowest point at 7d after operation (P<0.01), while thermal thresholds got to the lowest at 4d post-operatively (P<0.01). Subsequently both of them began to recover, but were still in a low level at 28d. Compared with sham group, thresholds of machine and therm of the operated legs in CCI group were always lower from 1d to 28d after CCI (P＜0.05). Thresholds of machine and therm in unoperated legs appeared no difference between the two groups (P>0.05).2) The expression of qualitation of protein:At 7d after CCI, the expression of NF-κB and TNF-αreached the top. The expression of NF-κB in the dentate gyrus of the hippocampus was obviously more other regions. It was mainly manifested as the increasing number of positive neurons and the deeper staining, both of which located in the nuclear of neurons. The expression of TNF-αin cytoplasm of dentate gyrus of the hippocampus also increased but was not so significant as NF-κB.3) Variation of mRNA:The quantity of mRNA in every indicator before CCI was set as 1. The mRNA of NF-κB in normal rats'hippocampus showed low expression. In CCI group, the mRNA of NF-κB of hippocampus began to increase 1 day after injury, but was not significant (P>0.05). Then it increased markedly at 4d (P<0.05), and reached peak at 7d which was significantly different when compared with the sham group (P＜0.05). Despite a decrease at 28d, it still maintained at a higher level than it in sham group.(P<0.05). The expression of TNF-αstarted to increase at 1d after injury, reached peak at 7d, and declined rapidly since then. At 4d and 7d, the level of mRNA of TNF-αin CCI group was higher than that in sham group (P<0.01). Compared with sham group, the expression of nNOS mRNA began to increase at 4d after injury, but expression of iNOS mRNA began to increase at 7d after injury. Both of them reached peak at 21d, and still maintained at a high level at 28d (P<0.05).Conclusions:CCI induced NF-κB activation and regulation up in rat hippocampus, so did the expression of TNF-α, iNOS and nNOS. All the four factors were involved in the progress of hippocampus responses to neuropathic pain.
|
PDF Full Text Request