粒细胞集落刺激因子改善阿尔茨海默病模型大鼠学习记忆功能及抗炎作用机制
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摘要
目的探讨粒细胞集落刺激因子(G-CSF)对阿尔茨海默病(AD)大鼠认知障碍的改善作用及其对脑内炎症反应的影响。方法 45只雄性SD大鼠,随机分为3组,包括假手术组、模型组、G-CSF组,每组15只。采用β-淀粉样蛋白1-42(Aβ1-42)复制AD大鼠模型,并给予G-CSF治疗。观察并比较大鼠的学习记忆功能及脑组织病理学变化。采用Western blotting检测核因子κB p65(NF-κB p65)、磷酸化p-38丝裂原活化蛋白激酶(p-p38MAPK)、诱导型一氧化氮合酶(i NOS)、环氧化酶-2(COX-2)、肿瘤坏死因子α(TNF-α)和白细胞介素-1β(IL-1β)蛋白的表达,采用Real-time PCR检测i NOS、COX-2、TNF-α和IL-1βmRNA水平。结果模型组大鼠神经细胞数量明显减少,细胞核固缩明显,核仁不清。G-CSF组大鼠脑组织神经细胞核固缩现象均有显著改善。假手术组、模型组、G-CSF组逃避潜伏期分别为(35.68±6.73) s、(57.92±7.35) s及(40.27±8.91) s;目标象限游泳时间分别为54.72%±4.22%、36.73%±3.21%及44.68%±4.01%;穿过平台次数分别为(8.7±2.1)次、(3.9±1.6)次及(6.5±1.7)次;与模型组相比,假手术组、G-CSF组的逃避潜伏期均显著缩短,目标象限游泳时间显著增加,穿过平台次数显著增加,差异均有统计学意义(P<0.05)。假手术组、模型组、G-CSF组NF-κB p65蛋白表达水平为0.144±0.033、0.502±0.035及0.473±0.061; pp38 MAPK蛋白水平为0.194±0.021、0.511±0.039及0.266±0.048,与模型组相比,假手术组、G-CSF组NF-κB p65、pp38MAPK蛋白水平均显著降低,差异均有统计学意义(P<0.05)。与模型组比较,假手术组、G-CSF组i NOS、COX-2、TNF-α和IL-1β的蛋白和mRNA水平显著降低(P<0.05)。结论 G-CSF具有良好的抑制AD模型大鼠脑神经细胞凋亡,改善大鼠学习记忆功能障碍的作用,其作用可能通过抑制p38MAPK和NF-κB p65的信号通路的过度激活,下调i NOS、COX-2、TNF-α和IL-1β等炎性因子的表达,抑制脑内神经炎症状态而实现。
Objective To investigate the effect of granulocyte colony-stimulating factor(G-CSF) on cognitive impairment in Alzheimer's disease(AD) in rats and its effect on brain inflammatory response.Methods Forty-five male Sprague-Dawley rats were randomly divided into 3 groups:sham operation group,model group,and G-CSF group.Rat models were induced by intracerebro ventricular injection of amyloid beta-peptides 1-42(Aβ1-42),and then G-CSF were given to G-CSF group.The learning and memory function and brain histopathological changes were observed and compared.The expression of nuclear factor κB p65(NF-κB p65),phosphorylated p38 mitogen activated protein kinase(pp38 MAPK),inducible nitric oxide synthase(i NOS),cyclooxygenase-2(COX-2),tumor necrosis factor α(TNF-α) andinterleukin-1β(IL-1β) were detected by Western blotting.The levels of i NOS,COX-2,TNF-α and IL-1β mRNA were detected by Real-time PCR.Results The number of neurons in the model group decreased significantly,the nuclear condensation was obvious and the nucleolus was unclear.Neuronal nuclear condensation was significantly improved in GCSF group.The escape latency of the sham group,model group and G-CSF group were(35.68±6.73) s,(57.92±7.35)s and(40.27±8.91) s.The swimming time of the target quadrant was 54.72% ± 4.22%,36.73% ± 3.2% and 44.68% ±4.01%.The times of crossing the platform were 8.7±2.1,3.9±1.6 and 6.5±1.7.Compared with the model group and the control group,the escape latency of G-CSF group were significantly shorter,target quadrant swimming time increased significantly,through the platform number increased significantly,the difference had statistical significance(P<0.05).In the sham group,model group and G-CSF group,the protein levels of NF-κB p65 were 0.144±0.033,0.502±0.035 and0.473±0.061,the protein levels of p-p38 MAPK were 0.194±0.021,0.511±0.039 and 0.266±0.048.Compared with the model group,the levels of NF-κB p65 and p-p38 MAPK protein in the sham group and G-CSF group decreased significantly,and the difference was statistically significant(P < 0.05).Compared with the model group,the protein and mRNA levels of i NOS,COX-2,TNF-α and IL-1β in the sham group and G-CSF group were significantly lower than those in the model group(P < 0.05).Conclusion G-CSF can improve the learning and memory dysfunction of rats with Alzheimer's disease probably through inhibiting the activation of p38 MAPK and NF-κB p65 signaling pathways and downregulating i NOS,COX-2,TNF-α and IL-1β expressions to inhibit the neuroinflammatory state of the brain.
Objective To investigate the effect of granulocyte colony-stimulating factor(G-CSF) on cognitive impairment in Alzheimer's disease(AD) in rats and its effect on brain inflammatory response.Methods Forty-five male Sprague-Dawley rats were randomly divided into 3 groups:sham operation group,model group,and G-CSF group.Rat models were induced by intracerebro ventricular injection of amyloid beta-peptides 1-42(Aβ1-42),and then G-CSF were given to G-CSF group.The learning and memory function and brain histopathological changes were observed and compared.The expression of nuclear factor κB p65(NF-κB p65),phosphorylated p38 mitogen activated protein kinase(pp38 MAPK),inducible nitric oxide synthase(i NOS),cyclooxygenase-2(COX-2),tumor necrosis factor α(TNF-α) andinterleukin-1β(IL-1β) were detected by Western blotting.The levels of i NOS,COX-2,TNF-α and IL-1β mRNA were detected by Real-time PCR.Results The number of neurons in the model group decreased significantly,the nuclear condensation was obvious and the nucleolus was unclear.Neuronal nuclear condensation was significantly improved in GCSF group.The escape latency of the sham group,model group and G-CSF group were(35.68±6.73) s,(57.92±7.35)s and(40.27±8.91) s.The swimming time of the target quadrant was 54.72% ± 4.22%,36.73% ± 3.2% and 44.68% ±4.01%.The times of crossing the platform were 8.7±2.1,3.9±1.6 and 6.5±1.7.Compared with the model group and the control group,the escape latency of G-CSF group were significantly shorter,target quadrant swimming time increased significantly,through the platform number increased significantly,the difference had statistical significance(P<0.05).In the sham group,model group and G-CSF group,the protein levels of NF-κB p65 were 0.144±0.033,0.502±0.035 and0.473±0.061,the protein levels of p-p38 MAPK were 0.194±0.021,0.511±0.039 and 0.266±0.048.Compared with the model group,the levels of NF-κB p65 and p-p38 MAPK protein in the sham group and G-CSF group decreased significantly,and the difference was statistically significant(P < 0.05).Compared with the model group,the protein and mRNA levels of i NOS,COX-2,TNF-α and IL-1β in the sham group and G-CSF group were significantly lower than those in the model group(P < 0.05).Conclusion G-CSF can improve the learning and memory dysfunction of rats with Alzheimer's disease probably through inhibiting the activation of p38 MAPK and NF-κB p65 signaling pathways and downregulating i NOS,COX-2,TNF-α and IL-1β expressions to inhibit the neuroinflammatory state of the brain.
引文
[1]Ni JZ,Chen P,Liu Q,et al.Advance reseach on strategies for the prevention of Alzheimer's disease[J].Journal of Shenzhen University(Science&Engineering),2013,30(4):331-348.(in Chinese)倪嘉缵,陈平,刘琼,等.阿尔茨海默病的防治策略研究进展[J].深圳大学学报(理工版),2013,30(4):331-348.
[2]Zhang J,Huang HCh,Jiang Zh F,et al.Progress in the mechanism ofβ-secretase inhibition in the prevention and treatment of Alzheimer disease[J].Chinese Journal of Gerontology,2013,33(21):5474-5477.(in Chinese)张姣,黄汉昌,姜招峰,等.阿尔茨海默病防治中的β-分泌酶抑制机制研究进展[J].中国老年学杂志,2013,33(21):5474-5477.
[3]Zhong X,Liu MY,Wei MJ,et al.Research progress of NLRP3 in flammatory bodies and Alzheimer disease[J].Chinese Journal of Gerontology,2016,36(13):3318-3321.(in Chinese)钟欣,刘明妍,魏敏杰,等.NLRP3炎症小体与阿尔茨海默病研究进展[J].中国老年学杂志,2016,36(13):3318-3321.
[4]Ridwan S,Bauer H,Frauenknecht K,et al.Distribution of the hematopoietic growth factor G-CSF and its receptor in the adult human brain with specific reference to Alzheimer's disease[J].JAnat,2014,224(4):377-391.
[5]Fan ZhS,Zuo J,Li Y,et al.Advances in the expression and role of G-CSF and G-CSFR in solid tumors[J].Chinese Journal of Clinical and Experimental Pathology,2015,9(10):1145-1147.(in Chinese)范志松,左静,李勇,等.实体肿瘤中G-CSF和G-CSFR的表达及作用的研究进展[J].临床与实验病理学杂志,2015,9(10):1145-1147.
[6]Bai LP,Zhao ZhH,Chen Ch,et al.Biological characteristics of endothelial progenitor cells derived from granulocyte colonystimulating factor mobilized peripheral blood[J].Journal of Clinical Rehabilitative Tissue Engineering Research,2014,8(32):5190-5196.(in Chinese)白丽萍,赵志红,陈冲,等.正常成人粒细胞集落刺激因子动员外周血内皮祖细胞的生物学特性[J].中国组织工程研究,2014,8(32):5190-5196.
[7]Qiao DD,Bian GCh,Wei XD,et al.Effects of rh G-CSF on learning and memory function of Alzheimer disease model rats induced by Aβ1-42[J].Journal of Psychiatry,2011,24(5):325-327.(in Chinese)乔冬冬,亓高超,魏贤达,等.rhG-CSF对Aβ1-42诱导阿尔茨海默病大鼠模型学习记忆功能的作用研究[J].精神医学杂志,2011,24(5):325-327.
[8]Li J,Mi YJ,Shi Zh,et al.The effect of G-CSF on brain inflammation in rats with Alzheimer's disease[J].Chinese Journal of Gerontology,2015,12(24):7027-7029.(in Chinese)李健,米永杰,石钊,等.粒细胞集落刺激因子对阿尔茨海默病大鼠脑内炎症的影响[J].中国老年学杂志,2015,12(24):7027-7029.
[9]Ding M,Zhang X,Wang W,et al.Effect of G-CSF on the expression of ChAT and cognitive function of dementia model rats induced by scopolamine[J].Practical Journal of Medicine and Pharmacy,2015,(7):635-637.(in Chinese)丁明,张迅,王威,等.G-CSF对东莨菪碱痴呆模型大鼠认知及ChAT表达的影响[J].实用医药杂志,2015,(7):635-637.
[10]Chen JB,Wang YJ,Guo Sh N,et al.Effect and mechanism of Heshowuyin on alleviating the hippocampal neurons injury induced byβ-amyloid protein[J].Acta Anatomica Sinica,2015,46(2):175-181.(in Chinese)陈靖博,王玉娟,郭胜男,等.何首乌饮减轻β-淀粉样蛋白诱导的海马神经元损伤的作用机制[J].解剖学报,2015,46(2):175-181.
[11]Gao J,Zhou R,You X,et al.Salidroside suppresses inflammation in a D-galactose-induced rat model of Alzheimer's disease via SIRT1/NF-κB pathway.[J].Metab Brain Dis,2016,31(4):771-778.
[12]Wang YW,Zheng GY,Chen XCh,et al.Ativation of gliacytes and p38 mitogen-activated protein kinase and possible mechanism of neuronal apoptosis induced by Aβ25-35injection into hippocampus in rats[J].Acta Anatomica Sinica,2014,45(5):616-621.(in Chinese)王元伟,郑关毅,陈晓春,等.海马背侧注射β-淀粉样蛋白25~35激活胶质细胞和p38丝裂原活化蛋白激酶诱导神经元凋亡[J].解剖学报,2014,45(5):616-621.
[2]Zhang J,Huang HCh,Jiang Zh F,et al.Progress in the mechanism ofβ-secretase inhibition in the prevention and treatment of Alzheimer disease[J].Chinese Journal of Gerontology,2013,33(21):5474-5477.(in Chinese)张姣,黄汉昌,姜招峰,等.阿尔茨海默病防治中的β-分泌酶抑制机制研究进展[J].中国老年学杂志,2013,33(21):5474-5477.
[3]Zhong X,Liu MY,Wei MJ,et al.Research progress of NLRP3 in flammatory bodies and Alzheimer disease[J].Chinese Journal of Gerontology,2016,36(13):3318-3321.(in Chinese)钟欣,刘明妍,魏敏杰,等.NLRP3炎症小体与阿尔茨海默病研究进展[J].中国老年学杂志,2016,36(13):3318-3321.
[4]Ridwan S,Bauer H,Frauenknecht K,et al.Distribution of the hematopoietic growth factor G-CSF and its receptor in the adult human brain with specific reference to Alzheimer's disease[J].JAnat,2014,224(4):377-391.
[5]Fan ZhS,Zuo J,Li Y,et al.Advances in the expression and role of G-CSF and G-CSFR in solid tumors[J].Chinese Journal of Clinical and Experimental Pathology,2015,9(10):1145-1147.(in Chinese)范志松,左静,李勇,等.实体肿瘤中G-CSF和G-CSFR的表达及作用的研究进展[J].临床与实验病理学杂志,2015,9(10):1145-1147.
[6]Bai LP,Zhao ZhH,Chen Ch,et al.Biological characteristics of endothelial progenitor cells derived from granulocyte colonystimulating factor mobilized peripheral blood[J].Journal of Clinical Rehabilitative Tissue Engineering Research,2014,8(32):5190-5196.(in Chinese)白丽萍,赵志红,陈冲,等.正常成人粒细胞集落刺激因子动员外周血内皮祖细胞的生物学特性[J].中国组织工程研究,2014,8(32):5190-5196.
[7]Qiao DD,Bian GCh,Wei XD,et al.Effects of rh G-CSF on learning and memory function of Alzheimer disease model rats induced by Aβ1-42[J].Journal of Psychiatry,2011,24(5):325-327.(in Chinese)乔冬冬,亓高超,魏贤达,等.rhG-CSF对Aβ1-42诱导阿尔茨海默病大鼠模型学习记忆功能的作用研究[J].精神医学杂志,2011,24(5):325-327.
[8]Li J,Mi YJ,Shi Zh,et al.The effect of G-CSF on brain inflammation in rats with Alzheimer's disease[J].Chinese Journal of Gerontology,2015,12(24):7027-7029.(in Chinese)李健,米永杰,石钊,等.粒细胞集落刺激因子对阿尔茨海默病大鼠脑内炎症的影响[J].中国老年学杂志,2015,12(24):7027-7029.
[9]Ding M,Zhang X,Wang W,et al.Effect of G-CSF on the expression of ChAT and cognitive function of dementia model rats induced by scopolamine[J].Practical Journal of Medicine and Pharmacy,2015,(7):635-637.(in Chinese)丁明,张迅,王威,等.G-CSF对东莨菪碱痴呆模型大鼠认知及ChAT表达的影响[J].实用医药杂志,2015,(7):635-637.
[10]Chen JB,Wang YJ,Guo Sh N,et al.Effect and mechanism of Heshowuyin on alleviating the hippocampal neurons injury induced byβ-amyloid protein[J].Acta Anatomica Sinica,2015,46(2):175-181.(in Chinese)陈靖博,王玉娟,郭胜男,等.何首乌饮减轻β-淀粉样蛋白诱导的海马神经元损伤的作用机制[J].解剖学报,2015,46(2):175-181.
[11]Gao J,Zhou R,You X,et al.Salidroside suppresses inflammation in a D-galactose-induced rat model of Alzheimer's disease via SIRT1/NF-κB pathway.[J].Metab Brain Dis,2016,31(4):771-778.
[12]Wang YW,Zheng GY,Chen XCh,et al.Ativation of gliacytes and p38 mitogen-activated protein kinase and possible mechanism of neuronal apoptosis induced by Aβ25-35injection into hippocampus in rats[J].Acta Anatomica Sinica,2014,45(5):616-621.(in Chinese)王元伟,郑关毅,陈晓春,等.海马背侧注射β-淀粉样蛋白25~35激活胶质细胞和p38丝裂原活化蛋白激酶诱导神经元凋亡[J].解剖学报,2014,45(5):616-621.