Résumé
Background@#In consideration of characteristics and functions, extra-cellular signal-regulated protein kinase 5 (ERK5) signaling pathway could be a new target for spinal cord injury (SCI) treatment. Our study aimed to evaluate the roles of ERK5 signaling pathway in secondary damage of SCI.@*Methods@#We randomly divided 70 healthy Wistar rats into five groups: ten in the blank group, 15 in the sham surgery + BIX02188 (sham + B) group, 15 in the sham surgery + dimethyl sulfoxide (DMSO; sham + D) group, 15 in the SCI + BIX02188 (SCI + B) group, and 15 in the SCI + DMSO (SCI + D) group. BIX02188 is a specific inhibitor of the ERK5 signaling pathway. SCI was induced by the application of vascular clips (with the force of 30 g) to the dura on T10 level, while rats in the sham surgery group underwent only T9-T11 laminectomy. BIX02188 or DMSO was intra-thecally injected at 1, 6, and 12 h after surgery or SCI. Spinal cord samples were taken for testing at 24 h after surgery or SCI.@*Results@#Expression of phosphorylated-ERK5 (p-ERK5) significantly increased after SCI. Application of BIX02188 indeed inhibited ERK5 signaling pathway and reduced the degree of spinal cord tissue injury, neutrophil infiltration and proinflammatory cytokine expression, nuclear factor-κB (NF-κB) activation and apoptosis (measured by TdT-mediated 2′-deoxyuridine 5′-triphosphate nickend labeling, expression of Fas-ligand, BCL2-associated X [Bax], and B-cell lymphoma-2 [Bcl-2]). Double immunofluorescence revealed activation of ERK5 in neurons and microglia after SCI.@*Conclusion@#ERK5 signaling pathway was activated in spinal neurons and microglia, contributing to secondary injury of SCI. Moreover, inhibition of ERK5 signaling pathway could alleviate the degree of SCI, which might be related to its regulation of infiltration of inflammatory cells and release of inflammatory cytokines, expression of NF-κB and cell apoptosis.
Résumé
Objective Through observation of the expression and activity of extracellular regulated protein kinase 5 (ERK5) and its relationship with the learning and memory ability in rats with chronic fluorosis,to further study the pathogenesis of chronic fluorosis in nervous system.Methods Thirty SD rats were divided into 3 groups according to body weight by means of a random number table (10 rats in each group,half male and half female).The rats in control group were fed with free drinking tap water containing less than 0.5 mg/L fluoride (NaF);the rats in low fluoride group with 10.0 mg/L fluoride;the rat in high dose fluoride group with 50.0 mg/L fluoride.After 6months of experiment,rat brain tissue was took,mRNA expression level of ERK5 was detected by real-time fluorescence quantitative PCR (real-time PCR),protein expression level and activity of ERK5 were detected by Western blotting;the learning and memory ability of rats with chronic fluorosis were detected by Morris water maze test.Results The rat in groups exposed to fluoride exhibited different degrees of dental fluorosis and the fluoride content in urine of rats increased gradually with increase of fluoride doses (F =164.10,P ＜ 0.05).The protein levels of phosphor-ERK5 in the control group,low fluoride group and high fluoride group were 0.13 ± 0.03,0.29 ± 0.10and 0.43 ±0.17,respectively,the difference was statistically significant (F=11.96,P＜ 0.05),and low fluoride group and high fluoride group were higher than control group (all P ＜ 0.05).The total protein levels of ERK5 in control group,low fluoride group and high fluoride group were 0.32 ± 0.11,0.37 ± 0.13 and 0.49 ± 0.16,respectively,the difference was statistically significant (F =3.45,P ＜ 0.05),and high fluoride group was higher than control group (P ＜ 0.05).The expression of ERK5 mRNA in rat brains between groups was not significantly different (F =0.81,P ＞ 0.05).The second,third,and forth days of directional navigation experiment,the time of escape latency and the number of crossing the platform between groups were statistically significant (H =28.20,29.90,26.47,27.23,35.34,27.62,all P ＜ 0.01);the fifth day of space exploration experiment,the difference of the time of the first crossing platform and the number of crossing the platform between groups were statistically significant (H =31.41,30.80,all P ＜ 0.01);the protein level of phosphor-ERK5 in brain tissue of rats was negatively correlated with the number of the first crossing platform (r =-0.470,P ＜ 0.01),while positively related to escape latencies at the fifth day of the test (r =0.591,P ＜ 0.01).Conclusion The changes of ERK5 signaling pathway in rat brain tissue caused by chronic fluorosis are found,which are related to the decrease of leaming and memory ability of animals with chronic fluorosis.