促红细胞生成素对大鼠急性脊髓损伤后核因子-κB及炎症因子表达的影响
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摘要
第一部分大鼠急性脊髓损伤后核因子-κB及炎症因子表达的时相变化
目的:
观察大鼠急性脊髓损伤后脊髓组织中核因子-κB(NF-κB)活性及炎症细胞因子如肿瘤坏死因子-α(TNF-α)、白介素-1β(IL-1β)、白介素-6(IL-6)表达水平的时相变化,探讨其与大鼠急性脊髓损伤后继发性损伤的关系,也为下一步观察促红细胞生成素(EPO)治疗大鼠脊髓损伤提供合理的时间窗。
方法:
30只成年SD大鼠体重(250+10)g,随机分成5组(对照组、6h组、24h组、72h组、7d组)。对照组大鼠仅椎板切除,其余大鼠分别用30g力量动脉瘤夹从两侧夹闭脊髓30s造成脊髓损伤模型,分别于术后6h、24h、72h、7d处死取材。用EMSA法检测各组脊髓组织中NF-κB活性,ELISA法检测TNF-α、IL-1β、IL-6的表达,比较分析各组差别。
结果:
损伤后6h组脊髓组织中NF-κB活性明显增加,TNF-α、IL-1β、IL-6表达明显上调,与对照组比较有明显差异(p<0.01)。NF-κB活性最高值出现在损伤后72h组,TNF-α、IL-1β表达在损伤后24h组达到高峰,IL-6表达高峰出现在损伤后72h组。7d后各组因子表达均已下降,但仍高于对照组水平(p<0.01)。
结论:
TNF-α、IL-1β、IL-6等炎症因子的高表达可能是导致炎症反应、加重脊髓损伤的重要因素,调控NF-κB活性表达可以为临床治疗脊椎损伤提供新的策略。NF-κB活性与各组炎症因子表达高峰期均在24h至72h间,这也为我们观察EPO治疗脊髓损伤提供了时间窗。
第二部分促红细胞生成素对大鼠急性脊髓损伤后核因子-κB及炎症因子表达的影响
目的:
观察促红细胞生成素(EPO)对大鼠急性脊髓损伤后脊髓组织中核因子-κB(NF-κB)表达及炎症细胞因子如肿瘤坏死因子(TNF-α)、白介素1β(IL-1β)、白介素6(IL-6)表达水平的影响,探讨其减轻大鼠急性脊髓损伤后继发伤的机制。
方法:
18只体重为(250+10)g的成年SD大鼠随机分成3组:对照组,损伤组和治疗组,每组6只。对照组大鼠仅椎板切除,其余大鼠用30g力量动脉瘤夹从两侧夹闭脊髓30s造成脊髓损伤模型,治疗组予术后1h、1d、2d、3d分别予EPO 5000u/kg腹腔注射,对照组与损伤组相同时间予腹腔注射同等剂量生理盐水。所有动物于术后3d处死取材,用EMSA法检测脊髓组织中NF-κB活性,ELISA法检测TNF-α、IL-1β、IL-6的表达,脊髓含水量测定判断脊髓水肿程度,BBB评分评价大鼠后肢功能恢复情况,比较3组间差异。
结果:
3d后EPO治疗组脊髓组织中NF-κB、TNF-α、IL-1β、IL-6的表达水平均比损伤组显著降低(P<0.05),而治疗组大鼠脊髓水肿程度及后肢功能变化较损伤组均无明显统计学意义。
结论:
EPO能下调急性脊髓损伤后脊髓组织中炎性信号因子的表达从而抑制炎症反应,对继发性脊髓损伤起到一定的保护作用,而对于减轻脊髓水肿促进脊髓功能恢复方面作用不明显。
Part I
Expression of nuclear factor-κB and inflammatory cytokines after acute spinal cord injury in rats
Objective
To observe the expression level of nuclear factor-κB(NF-κB) and inflammatory factor of tumor necrosis factor (TNF-α), interleukin(IL)-1βand IL-6 after acute spinal cord injury in rats, and to explore its possible relationship with acute spinal cord injury,also observed for the next step of erythropoietin (EPO) treatment of spinal cord injury in a reasonable time window.
Methods
30 SD rats weighting(250+10)g were randomly divided into 5 groups(control group, 6h group, 24h group, 72h group, 7d group), SD rat model of acute spinal cord injury were established by aneurysm clip with 30g power cord clamping from both sides of spinal cord for 30s. Rats after operation were sacrificed respectively after 6h, 24h, 72h and 7d. The expression of NF-κB was detected by EMSA assay and the expression of TNF-α, IL-1βand IL-6 were examined with ELISA in spinal cord tissue.
Result
The activity of NF-κB and the expression of TNF-α, IL-1βand IL-6 were significantly higher than control group 6 hours after spinal cord injury(P<0.01). The highest NF-κB activity in 72h after operation, TNF-α, IL-1βexpression peaked at 24h and IL-6 expression peaked at 72h after operation. Factor expression in each group after 7 days had fallen, but still higher than the level of control group(P<0.01).
Conclusion
The high expression of TNF-α, IL-1βand IL-6 may lead to inflammation and increase the important factors in spinal cord injury, regulation of the expression of NF-κB activity can provide a new clinical treatment strategy for spinal cord injury. NF-κB activity and expression of inflammatory cytokines in each group wre in the peak period between 24h to 72h, this provided a time window to observe the EPO treatment of spinal cord injury.
Part II
The effect of erythropoietin on the expression of NF-κB and inflammatory cytokines after acute spinal cord injury in rats
Objective
To observe the influence of erythropoietin (EPO) administration on expression of nuclear factor-kappa B(NF-κB) and inflammatory cytokines of tumor necrosis factor-α(TNF-α), interleukin(IL)-1β, IL-6 following acute spinal cord injury and to explore the possible mechanisms of reducing secondary spinal cord injury.
Methods
18 SD rats weighting(250+10)g were randomly divided into 3 groups: Control group, injury group and treatment group. Treatment group were given intraperitoneal injection of EPO(5000u/kg) at 1h, on the 1st, 2nd and 3rd day after establishment of spinal cord injury model. All rats were sacrificed after 3days. The expression of NF-κB was detected by EMSA assay and the expression of TNF-α, IL-1βand IL-6 were examined with ELISA, the spinal cord edema was evaluated by spinal cord water content, and the hindlimb function in rats was evaluated by the BBB Rating.
Result
The levels of NF-κB, TNF-α, IL-1β, IL-6 in spinal cord tissue of treatment group were significantly lower than those in injury model group(P<0.05), and the levels of spinal cord edema and indlimb function were no statistically significant.
Conclusion
EPO administration can down-regulate the expression of the inflammatory cytokines, inhibit the inflammatory response and play a role in protecting the secondary spinal cord injury, but the Functional recovery of spinal cord was no significant change.
目的:
观察大鼠急性脊髓损伤后脊髓组织中核因子-κB(NF-κB)活性及炎症细胞因子如肿瘤坏死因子-α(TNF-α)、白介素-1β(IL-1β)、白介素-6(IL-6)表达水平的时相变化,探讨其与大鼠急性脊髓损伤后继发性损伤的关系,也为下一步观察促红细胞生成素(EPO)治疗大鼠脊髓损伤提供合理的时间窗。
方法:
30只成年SD大鼠体重(250+10)g,随机分成5组(对照组、6h组、24h组、72h组、7d组)。对照组大鼠仅椎板切除,其余大鼠分别用30g力量动脉瘤夹从两侧夹闭脊髓30s造成脊髓损伤模型,分别于术后6h、24h、72h、7d处死取材。用EMSA法检测各组脊髓组织中NF-κB活性,ELISA法检测TNF-α、IL-1β、IL-6的表达,比较分析各组差别。
结果:
损伤后6h组脊髓组织中NF-κB活性明显增加,TNF-α、IL-1β、IL-6表达明显上调,与对照组比较有明显差异(p<0.01)。NF-κB活性最高值出现在损伤后72h组,TNF-α、IL-1β表达在损伤后24h组达到高峰,IL-6表达高峰出现在损伤后72h组。7d后各组因子表达均已下降,但仍高于对照组水平(p<0.01)。
结论:
TNF-α、IL-1β、IL-6等炎症因子的高表达可能是导致炎症反应、加重脊髓损伤的重要因素,调控NF-κB活性表达可以为临床治疗脊椎损伤提供新的策略。NF-κB活性与各组炎症因子表达高峰期均在24h至72h间,这也为我们观察EPO治疗脊髓损伤提供了时间窗。
第二部分促红细胞生成素对大鼠急性脊髓损伤后核因子-κB及炎症因子表达的影响
目的:
观察促红细胞生成素(EPO)对大鼠急性脊髓损伤后脊髓组织中核因子-κB(NF-κB)表达及炎症细胞因子如肿瘤坏死因子(TNF-α)、白介素1β(IL-1β)、白介素6(IL-6)表达水平的影响,探讨其减轻大鼠急性脊髓损伤后继发伤的机制。
方法:
18只体重为(250+10)g的成年SD大鼠随机分成3组:对照组,损伤组和治疗组,每组6只。对照组大鼠仅椎板切除,其余大鼠用30g力量动脉瘤夹从两侧夹闭脊髓30s造成脊髓损伤模型,治疗组予术后1h、1d、2d、3d分别予EPO 5000u/kg腹腔注射,对照组与损伤组相同时间予腹腔注射同等剂量生理盐水。所有动物于术后3d处死取材,用EMSA法检测脊髓组织中NF-κB活性,ELISA法检测TNF-α、IL-1β、IL-6的表达,脊髓含水量测定判断脊髓水肿程度,BBB评分评价大鼠后肢功能恢复情况,比较3组间差异。
结果:
3d后EPO治疗组脊髓组织中NF-κB、TNF-α、IL-1β、IL-6的表达水平均比损伤组显著降低(P<0.05),而治疗组大鼠脊髓水肿程度及后肢功能变化较损伤组均无明显统计学意义。
结论:
EPO能下调急性脊髓损伤后脊髓组织中炎性信号因子的表达从而抑制炎症反应,对继发性脊髓损伤起到一定的保护作用,而对于减轻脊髓水肿促进脊髓功能恢复方面作用不明显。
Part I
Expression of nuclear factor-κB and inflammatory cytokines after acute spinal cord injury in rats
Objective
To observe the expression level of nuclear factor-κB(NF-κB) and inflammatory factor of tumor necrosis factor (TNF-α), interleukin(IL)-1βand IL-6 after acute spinal cord injury in rats, and to explore its possible relationship with acute spinal cord injury,also observed for the next step of erythropoietin (EPO) treatment of spinal cord injury in a reasonable time window.
Methods
30 SD rats weighting(250+10)g were randomly divided into 5 groups(control group, 6h group, 24h group, 72h group, 7d group), SD rat model of acute spinal cord injury were established by aneurysm clip with 30g power cord clamping from both sides of spinal cord for 30s. Rats after operation were sacrificed respectively after 6h, 24h, 72h and 7d. The expression of NF-κB was detected by EMSA assay and the expression of TNF-α, IL-1βand IL-6 were examined with ELISA in spinal cord tissue.
Result
The activity of NF-κB and the expression of TNF-α, IL-1βand IL-6 were significantly higher than control group 6 hours after spinal cord injury(P<0.01). The highest NF-κB activity in 72h after operation, TNF-α, IL-1βexpression peaked at 24h and IL-6 expression peaked at 72h after operation. Factor expression in each group after 7 days had fallen, but still higher than the level of control group(P<0.01).
Conclusion
The high expression of TNF-α, IL-1βand IL-6 may lead to inflammation and increase the important factors in spinal cord injury, regulation of the expression of NF-κB activity can provide a new clinical treatment strategy for spinal cord injury. NF-κB activity and expression of inflammatory cytokines in each group wre in the peak period between 24h to 72h, this provided a time window to observe the EPO treatment of spinal cord injury.
Part II
The effect of erythropoietin on the expression of NF-κB and inflammatory cytokines after acute spinal cord injury in rats
Objective
To observe the influence of erythropoietin (EPO) administration on expression of nuclear factor-kappa B(NF-κB) and inflammatory cytokines of tumor necrosis factor-α(TNF-α), interleukin(IL)-1β, IL-6 following acute spinal cord injury and to explore the possible mechanisms of reducing secondary spinal cord injury.
Methods
18 SD rats weighting(250+10)g were randomly divided into 3 groups: Control group, injury group and treatment group. Treatment group were given intraperitoneal injection of EPO(5000u/kg) at 1h, on the 1st, 2nd and 3rd day after establishment of spinal cord injury model. All rats were sacrificed after 3days. The expression of NF-κB was detected by EMSA assay and the expression of TNF-α, IL-1βand IL-6 were examined with ELISA, the spinal cord edema was evaluated by spinal cord water content, and the hindlimb function in rats was evaluated by the BBB Rating.
Result
The levels of NF-κB, TNF-α, IL-1β, IL-6 in spinal cord tissue of treatment group were significantly lower than those in injury model group(P<0.05), and the levels of spinal cord edema and indlimb function were no statistically significant.
Conclusion
EPO administration can down-regulate the expression of the inflammatory cytokines, inhibit the inflammatory response and play a role in protecting the secondary spinal cord injury, but the Functional recovery of spinal cord was no significant change.
引文
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