ω-3多不饱和脂肪酸对脂多糖所致新生大鼠脑损伤Toll-NF-κB信号通路调节作用
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摘要
目的探讨ω-3PUFAs在脂多糖(LPS)诱导的新生大鼠脑损伤中对TLR4、NF-κB信号通路的调节作用。方法将96只SD新生大鼠按随机数字法分为对照组,ω-3组,ω-6组和LPS组。4组大鼠分别采用生理盐水或LPS腹腔注射后,立即注射生理盐水或不同的脂肪乳剂,使用时间分别为1天或5天,建立感染所致新生大鼠脑损伤模型;1天或5天后处死新生大鼠,检测TLR4mRNA、NF-κB mRNA和对应蛋白水平的变化。结果 LPS注射后的第1天、第5天,新生大鼠TLR4mRNA、NF-κB mRNA表达及其对应的蛋白水平均明显高于对照组(P均<0.05)。1d时ω-3组TLR4mRNA、NF-κB mRNA表达及其对应的蛋白水平均低于ω-6组和LPS组(P均<0.05)。5d时ω-3组TLR4mRNA、NF-κB mRNA表达及其对应的蛋白水平均低于ω-6组和LPS组(P均<0.05)。不论第1d和第5d,ω-3PUFAs均可明显降低TLR4mRNA、NF-κB mRNA表达及其对应的蛋白水平(P均<0.05);而ω-6PUFAs能提高TLR4mRNA、NF-κB mRNA表达及对应的蛋白水平(P均<0.05)。结论ω-3PUFAs能下调TLR4、NF-κB活性,在LPS所致的脑损伤中具有抗炎作用。
Objective To investigate the effects of ω-3 PUFAs on regulation of Toll-like receptor 4(TLR4)and nuclear factor-κB(NF-κB)signaling pathway in neonatal rats with brain injury induced by LPS. Methods Totally 96 neonatal Sprague-Dawley rats were divided into control group, ω-3 group, ω-6 group,and LPS group.Intraperitoneal injection with saline or LPS was administered and immediately saline or fat emulsions was injected to establish a model of rat with brain injury induced by infection and the duration time was 1 day or 5 days respectively.Those groups were sacrificed at 1 dor 5 dafter intraperitoneal injection with saline or fat emulsions.Expression levels of TLR4 mRNA and NF-κB mRNA were examined by real-time PCR,meanwhile the levels of TLR4,and NF-κB in the hippocampus were measured by Western Blot. Results The expressions of TLR mRNA and NF-κB mRNA and the protein level of TLR4 and NF-κB in the hippocampus were significantly different in four groups(all P<0.05).TLR mRNA and NF-κB mRNA and the protein expression levels in ω-3 group were lower than those in ω-6 group and LPS group on 1 dafter injection(all P<0.05).And the protein expression levels of TLR mRNA and NF-κB mRNA in ω-3 group were lower than those in ω-6 group and LPS group on five days after injection(all P<0.05).No matter the first day or the fifth day,the expression of TLR mRNA and NF-κB mRNA and the level of TLR4 and NF-κB in ω-3 group were significantly lower than those in ω-6 group(all P<0.05). Conclusion ω-3 PUFAs can down regulate the activity of signaling pathway of TLR4 and NF-κB,so it may has an anti-inflammatory effect on brain injuryinduced by LPS.
Objective To investigate the effects of ω-3 PUFAs on regulation of Toll-like receptor 4(TLR4)and nuclear factor-κB(NF-κB)signaling pathway in neonatal rats with brain injury induced by LPS. Methods Totally 96 neonatal Sprague-Dawley rats were divided into control group, ω-3 group, ω-6 group,and LPS group.Intraperitoneal injection with saline or LPS was administered and immediately saline or fat emulsions was injected to establish a model of rat with brain injury induced by infection and the duration time was 1 day or 5 days respectively.Those groups were sacrificed at 1 dor 5 dafter intraperitoneal injection with saline or fat emulsions.Expression levels of TLR4 mRNA and NF-κB mRNA were examined by real-time PCR,meanwhile the levels of TLR4,and NF-κB in the hippocampus were measured by Western Blot. Results The expressions of TLR mRNA and NF-κB mRNA and the protein level of TLR4 and NF-κB in the hippocampus were significantly different in four groups(all P<0.05).TLR mRNA and NF-κB mRNA and the protein expression levels in ω-3 group were lower than those in ω-6 group and LPS group on 1 dafter injection(all P<0.05).And the protein expression levels of TLR mRNA and NF-κB mRNA in ω-3 group were lower than those in ω-6 group and LPS group on five days after injection(all P<0.05).No matter the first day or the fifth day,the expression of TLR mRNA and NF-κB mRNA and the level of TLR4 and NF-κB in ω-3 group were significantly lower than those in ω-6 group(all P<0.05). Conclusion ω-3 PUFAs can down regulate the activity of signaling pathway of TLR4 and NF-κB,so it may has an anti-inflammatory effect on brain injuryinduced by LPS.
引文
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[9]石计朋,黄丽密,钱燕,等.脂肪乳剂对脂多糖诱导的急性肺损伤中细胞因子IL-1β和IL-6的影响[J].临床儿科杂志,2014(3):250-253.
[10]黄丽密,石计朋,余震,等.脂肪乳剂对急性肺损伤大鼠肺组织病理形态的影响[J].肠外与肠内营养,2012,19(2):107-110.
[11]黄丽密,钱燕,余震,等.3种脂肪乳剂对急性肺损伤大鼠炎症和细胞凋亡的干预作用比较[J].浙江医学,2010,32(2):195-197.
[12]宋亚辉,尤庆旺,王倩楠,等.Omega-3鱼油脂肪乳对缺氧缺血性脑损伤新生大鼠海马组织Toll样受体4和核因子κB的影响[J].中华实用儿科临床杂志,2016,31(12):923-926.
[13]李艳艳.ω-3鱼油脂肪乳和亚低温对新生大鼠缺氧缺血脑损伤保护作用探讨[D].新乡:新乡医学院,2015.
[14]Graham EM,Holcroft CJ,Rai KK,et al.Neonatal cerebral white matter injury in preterm infants is associated with culture positive infections and only rarely with metabolic acidosis[J].Am J Obstet Gynecol,2004,191(4):1305-1310.
[15]Fan LW,Tien LT,Zheng B,et al.Dopaminergic neuronal injury in the adult rat brain following neonatal exposure to lipopolysaccharide and the silent neurotoxicity[J].Brain Behav Immun,2011,25(2):286-297.
[16]Mallard C,Wang X.Infection-induced vulnerability of perinatal brain injury[J].Neurol Res Int,2012,2012:102153.
[17]Jursza E,Kowalewski MP,Boos A,et al.The role of tolllike receptors 2and 4in the pathogenesis of feline pyometra[J].Theriogenology,2015,83(4):596-603.
[18]Zhang J,Liu S,Rajendran K V,et al.Pathogen recognition receptors in channel catfish:III phylogeny and expression analysis of Toll-like receptors[J].Dev Comp Immunol,2013,40(2):185-194.
[19]Sun HN,Shen GN,Jin YZ,et al.2-cyclohexylamino-5,8-dimethoxy-1,4-naphthoquinone inhibits LPS-induced BV2microglial activation through MAPK/NF-kB signaling pathways[J].Heliyon,2016,2(7):e132.
[20]Yao L,Kan EM,Lu J,et al.Toll-like receptor 4 mediates microglial activation and production of inflammatory mediators in neonatal rat brain following hypoxia:role of TLR4in hypoxic microglia[J].J Neuroinflammation,2013,10:23.
[21]Yang S,Li R,Qu X,et al.Fosinoprilat alleviates lipopolysaccharide(LPS)-induced inflammation by inhibiting TLR4/NF-kappaB signaling in monocytes[J].Cell Immunol,2013,284(1-2):182-186.
[22]Gorina R,Font-Nieves M,Marquez-Kisinousky L,et al.Astrocyte TLR4activation induces a proinflammatory environment through the interplay between MyD88-dependent NFkappaB signaling,MAPK,and Jak1/Stat1pathways[J].Glia,2011,59(2):242-255.
[23]Zhao LL,Hu GC,Zhu SS,et al.Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway[J].Braz J Med Biol Res,2014,47(12):1062-1067.
[24]Bawadekar M,De Andrea M,Lo CI,et al.The Extracellular IFI16Protein Propagates Inflammation in Endothelial Cells Via p38 MAPK and NF-kappaB p65Activation[J].J Interferon Cytokine Res,2015,35(6):441-453.
[25]Gokcinar D,Ergin V,Cumaoglu A,et al.Effects of ketamine,propofol,and ketofol on proinflammatory cytokines and markers of oxidative stress in a rat model of endotoxemia-induced acute lung injury[J].Acta Biochim Pol,2013,60(3):451-456.
[26]Hagberg H,Peebles D,Mallard C.Models of white matter injury:comparison of infectious,hypoxic-ischemic,and excitotoxic insults[J].Ment Retard Dev Disabil Res Rev,2002,8(1):30-38.
[27]徐发林,庄方莉,白琼丹,等.细菌脂多糖对不同成熟度新生小鼠脑发育的影响[J].实用儿科临床杂志,2012,27(8):611-613,616.
[28]Berman DR,Mozurkewich E,Liu Y,et al.Docosahexaenoic acid augments hypothermic neuroprotection in a neonatal rat asphyxia model[J].Neonatology,2013,104(1):71-78.
[2]刘畅,富建华,薛辛东.早产儿脑白质损伤的早期影像学改变及其对预后的影响[J].中华儿科杂志,2012,50(10):762-766.
[3]Groenendaal F,Termote JU,van der Heide-Jalving M,et al.Complications affecting preterm neonates from 1991 to2006:what have we gained?[J].Acta Paediatr,2010,99(3):354-358.
[4]Dean JM,Shi Z,Fleiss B,et al.A critical review of models of perinatal infection[J].Dev Neurosci,2015,37(4-5):289-304.
[5]石计朋,闫建国,韩金芬,等.ω-3PUFAs、ω-6PUFAs和ω-9MUFAs对LPS诱导急性肺损伤大鼠血清IL-6及细胞凋亡的影响[J].中国儿童保健杂志,2014,22(3):271-274.
[6]石计朋,闫建国,韩金芬,等.ω-3PUFAs对LPS诱导急性肺损伤大鼠前炎症因子TNF-α、IL-1β、IL-6的影响[J].中国儿童保健杂志,2014,22(1):35-37,40.
[7]石计朋,钱燕,余震,等.持续输注脂肪乳对大鼠内毒素诱导急性肺损伤的影响及机制[J].临床儿科杂志,2013(6):557-560.
[8]石计朋,黄丽密,尚云.脂肪乳对内毒素诱导急性肺损伤大鼠肺组织白介素6分泌及肺泡上皮细胞凋亡的影响[J].中国全科医学,2014,17(6):652-655,662.
[9]石计朋,黄丽密,钱燕,等.脂肪乳剂对脂多糖诱导的急性肺损伤中细胞因子IL-1β和IL-6的影响[J].临床儿科杂志,2014(3):250-253.
[10]黄丽密,石计朋,余震,等.脂肪乳剂对急性肺损伤大鼠肺组织病理形态的影响[J].肠外与肠内营养,2012,19(2):107-110.
[11]黄丽密,钱燕,余震,等.3种脂肪乳剂对急性肺损伤大鼠炎症和细胞凋亡的干预作用比较[J].浙江医学,2010,32(2):195-197.
[12]宋亚辉,尤庆旺,王倩楠,等.Omega-3鱼油脂肪乳对缺氧缺血性脑损伤新生大鼠海马组织Toll样受体4和核因子κB的影响[J].中华实用儿科临床杂志,2016,31(12):923-926.
[13]李艳艳.ω-3鱼油脂肪乳和亚低温对新生大鼠缺氧缺血脑损伤保护作用探讨[D].新乡:新乡医学院,2015.
[14]Graham EM,Holcroft CJ,Rai KK,et al.Neonatal cerebral white matter injury in preterm infants is associated with culture positive infections and only rarely with metabolic acidosis[J].Am J Obstet Gynecol,2004,191(4):1305-1310.
[15]Fan LW,Tien LT,Zheng B,et al.Dopaminergic neuronal injury in the adult rat brain following neonatal exposure to lipopolysaccharide and the silent neurotoxicity[J].Brain Behav Immun,2011,25(2):286-297.
[16]Mallard C,Wang X.Infection-induced vulnerability of perinatal brain injury[J].Neurol Res Int,2012,2012:102153.
[17]Jursza E,Kowalewski MP,Boos A,et al.The role of tolllike receptors 2and 4in the pathogenesis of feline pyometra[J].Theriogenology,2015,83(4):596-603.
[18]Zhang J,Liu S,Rajendran K V,et al.Pathogen recognition receptors in channel catfish:III phylogeny and expression analysis of Toll-like receptors[J].Dev Comp Immunol,2013,40(2):185-194.
[19]Sun HN,Shen GN,Jin YZ,et al.2-cyclohexylamino-5,8-dimethoxy-1,4-naphthoquinone inhibits LPS-induced BV2microglial activation through MAPK/NF-kB signaling pathways[J].Heliyon,2016,2(7):e132.
[20]Yao L,Kan EM,Lu J,et al.Toll-like receptor 4 mediates microglial activation and production of inflammatory mediators in neonatal rat brain following hypoxia:role of TLR4in hypoxic microglia[J].J Neuroinflammation,2013,10:23.
[21]Yang S,Li R,Qu X,et al.Fosinoprilat alleviates lipopolysaccharide(LPS)-induced inflammation by inhibiting TLR4/NF-kappaB signaling in monocytes[J].Cell Immunol,2013,284(1-2):182-186.
[22]Gorina R,Font-Nieves M,Marquez-Kisinousky L,et al.Astrocyte TLR4activation induces a proinflammatory environment through the interplay between MyD88-dependent NFkappaB signaling,MAPK,and Jak1/Stat1pathways[J].Glia,2011,59(2):242-255.
[23]Zhao LL,Hu GC,Zhu SS,et al.Propofol pretreatment attenuates lipopolysaccharide-induced acute lung injury in rats by activating the phosphoinositide-3-kinase/Akt pathway[J].Braz J Med Biol Res,2014,47(12):1062-1067.
[24]Bawadekar M,De Andrea M,Lo CI,et al.The Extracellular IFI16Protein Propagates Inflammation in Endothelial Cells Via p38 MAPK and NF-kappaB p65Activation[J].J Interferon Cytokine Res,2015,35(6):441-453.
[25]Gokcinar D,Ergin V,Cumaoglu A,et al.Effects of ketamine,propofol,and ketofol on proinflammatory cytokines and markers of oxidative stress in a rat model of endotoxemia-induced acute lung injury[J].Acta Biochim Pol,2013,60(3):451-456.
[26]Hagberg H,Peebles D,Mallard C.Models of white matter injury:comparison of infectious,hypoxic-ischemic,and excitotoxic insults[J].Ment Retard Dev Disabil Res Rev,2002,8(1):30-38.
[27]徐发林,庄方莉,白琼丹,等.细菌脂多糖对不同成熟度新生小鼠脑发育的影响[J].实用儿科临床杂志,2012,27(8):611-613,616.
[28]Berman DR,Mozurkewich E,Liu Y,et al.Docosahexaenoic acid augments hypothermic neuroprotection in a neonatal rat asphyxia model[J].Neonatology,2013,104(1):71-78.