参枝苓口服液对APP/PS1双转基因小鼠海马α7-nAChRs和mGluR5表达的影响
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摘要
目的探讨参枝苓口服液对APP/PS1双转基因小鼠海马神经元突触信号传递相关蛋白α7-nAChRs和mGluR5表达的影响。方法将雄性3月龄APP/PS1双转基因小鼠随机分成5组,模型组、多奈哌齐组[0.92 mg/(kg·d)],参枝苓大剂量组[50 g/(kg·d)]、参枝苓中剂量组[25 g/(kg·d)]、参枝苓小剂量组[12.5 g/(kg·d)]。正常组为同月龄同背景野生型C57BL/6J小鼠。连续灌胃3个月,进行行为学跳台实验。实验结束后取海马组织,运用蛋白印迹法(Western blot)和免疫组织化学方法检测小鼠海马CA1区突触相关蛋白α7-nAChRs和mGluR5的表达。结果 Western blot检测显示,模型组小鼠海马α7-nAChRs的蛋白条带比正常组小鼠明显变窄,颜色变浅(P<0.01)、而mGluR5蛋白条带明显增宽,颜色变深(P<0.01);与模型组小鼠相比,各干预组α7-nAChRs蛋白条带增宽、颜色变深(P<0.05或P<0.01),mGluR5蛋白条带变窄、颜色变浅(P<0.05或P<0.01)。免疫组织化学染色检测小鼠海马α7-nAChRs和mGluR5的阳性细胞表达结果与Western blot检测结果一致。结论参枝苓口服液可能是通过调节小鼠海马CA1区突触中的关键蛋白,改善突触功能发挥减缓阿尔茨海默病(AD)大脑损伤,维持记忆获得能力,从而发挥抗痴呆作用。
Objective To discuss the influence of Shenzhilin(Ginseng, Cassia Twig and Indian Bread) Oral Solution(SZL) on expressions of hippocampal α7-nAChRs and mGluR5(neuronal synapse signaling related proteins) in APP/PS1 double transgenic mice. Methods Male APP/PS1 double transgenic mice(3 months old) were randomly divided into 5 groups: model group, donepezil group [0.92 mg/(kg·d)], high-dose SZL group [50 g/(kg·d)], mid-dose SZL group [25 g/(kg·d)] and low-dose SZL group [12.5 g/(kg·d)]. Wild type C57 BL/6 J mice with the same age and background were chosen into normal group. After gavage for 3 months continuously, behavioral platform test was carried out in all groups, and then hippocampal tissue samples were collected for detecting the expressions of α7-nAChRs and mGluR5 in hippocampal CA1 zone by using Western blotting assay and immunohistochemistry technique. Results The results of Western blotting assay showed that size of protein bands of α7-nAChRs were significantly narrower and color was lighter(P<0.01), and size of protein bands of mGluR5 were wider and color was darker(P<0.01) in model group than those in normal group. Compared with model group, size of protein bands of α7-nAChRs were wider and color was darker(P<0.05 or P<0.01), and size of protein bands of mGluR5 were narrower and color was lighter(P<0.05 or P<0.01) in all medicated groups. The positive cell expressions of α7-nAChRs and mGluR5 detected by using immunohistochemistry technique were consistent with those detected by using Western blotting assay. Conclusion SZL may play an anti-dementia role through regulating the key synaptic proteins in hippocampal CA1 zone, improving synapse function for slowing down brain damage due to Alzheimer disease, and maintaining memory acquisition ability.
Objective To discuss the influence of Shenzhilin(Ginseng, Cassia Twig and Indian Bread) Oral Solution(SZL) on expressions of hippocampal α7-nAChRs and mGluR5(neuronal synapse signaling related proteins) in APP/PS1 double transgenic mice. Methods Male APP/PS1 double transgenic mice(3 months old) were randomly divided into 5 groups: model group, donepezil group [0.92 mg/(kg·d)], high-dose SZL group [50 g/(kg·d)], mid-dose SZL group [25 g/(kg·d)] and low-dose SZL group [12.5 g/(kg·d)]. Wild type C57 BL/6 J mice with the same age and background were chosen into normal group. After gavage for 3 months continuously, behavioral platform test was carried out in all groups, and then hippocampal tissue samples were collected for detecting the expressions of α7-nAChRs and mGluR5 in hippocampal CA1 zone by using Western blotting assay and immunohistochemistry technique. Results The results of Western blotting assay showed that size of protein bands of α7-nAChRs were significantly narrower and color was lighter(P<0.01), and size of protein bands of mGluR5 were wider and color was darker(P<0.01) in model group than those in normal group. Compared with model group, size of protein bands of α7-nAChRs were wider and color was darker(P<0.05 or P<0.01), and size of protein bands of mGluR5 were narrower and color was lighter(P<0.05 or P<0.01) in all medicated groups. The positive cell expressions of α7-nAChRs and mGluR5 detected by using immunohistochemistry technique were consistent with those detected by using Western blotting assay. Conclusion SZL may play an anti-dementia role through regulating the key synaptic proteins in hippocampal CA1 zone, improving synapse function for slowing down brain damage due to Alzheimer disease, and maintaining memory acquisition ability.
引文
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[7] 吴艺琼,陈芳,盛宁,等.参枝苓口服液对APPswe/PSldE9小鼠学习记忆和突触超微结构的影响[J].北京中医药大学学报,2018,41(10):821-827.Wu YQ, Chen F, Sheng N, et al. Effects of Shenzhiling Oral Liquid on learning and memory and synaptic structure in APPswe/PSldE9 mice[J]. Journal of Beijing University of Traditional Chinese Medicine, 2018, 41(10): 821-827.
[8] Shinohara M, Koga S, Konno T, et al. Distinct spatiot -emporal accumulation of N-truncated and full-length amyloid-β42 in Alzheimer's disease[J]. Brain A Journal of Neurology, 2017, 140(12): 3301-3316.
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[10] Takata K, Amamiya T, Mizoguchi H, et al. Alpha7 nicotinic acetylcholine receptor-specific agonist DMXBA (GTS-21) attenuates Aβ accumulation through suppression of neuronal γ-secretase activity and promotion of microglial amyloid-β phagocytosis and ameliorates cognitive impairment in a mouse model of Alzheimer’s disease[J]. Neurobiol Aging, 2018, 62: 197-209.
[11] Chen L, Yamada K, Nabeshima T, et al. α7 Nicotinic acetylcholine receptor as a target to rescue deficit in hippocampal LTP induction in β-amyloid infused rats[J]. Neuropharmacology, 2006, 50(2): 254-268.
[12] Huang M, Felix AR, Flood DG, et al. The novel α7 nicotinic acetylcholine receptor agonist EVP-6124 enhances dopamine, acetylcholine, and glutamate efflux in rat cortex and nucleus accumbens[J]. Psychopharmacology, 2014, 231(23): 4541-4551.
[13] Raka F, Sebastiano AR, Kulhawy SC, et al. Ca(2+)/Calmodulin-dependent protein Kinase II interacts with group I Metabotropic Glutamate and facilitates Receptor Endocytosis and ERK1/2 signaling: role of β-Amyloid[J]. Molecular Brain, 2015, 8(1): 1-15.
[14] Um JW, Kaufman AC, Kostylev M, et al. Metabotropic glutamate receptor 5 is a coreceptor for Alzheimer Aβ oligomer bound to cellular prion protein[J]. Neuron, 2013, 79(5): 887-902.
[15] Shrivastava AN, Kowalewski JM, Renner M, et al. β-amyloid and ATP-induced diffusional trapping of astrocyte and neuronal metabotropic glutamate type-5 receptors[J]. Glia, 2013, 61(10): 1673-1686.
[16] Overk CR, Cartier A, Shaked G, et al. Hippocampal neuronal cells that accumulate α-synuclein fragments are more vulnerable to Aβ oligomer toxicity via mGluR5-implications for dementia with Lewy bodies[J]. Mol Neurodegener., 2014, 9(1): 1-18.
[2] Gouras GK, Willén K, Faideau M. The inside-out amyloid hypothesis and synapse pathology in Alzheimer’s disease[J]. Neurodegener Dis, 2014, 13(2-3): 142-146.
[3] Elnagar MR, Walls AB, Helal GK, et al. Probing the putative α7 nAChR/NMDAR complex in human and murine cortex and hippocampus: Different degrees of complex formation in healthy and Alzheimer brain tissue[J]. Plos One, 2017, 12(12): e0189513.
[4] 杜威,郭英,袁维秀. 毒蕈碱乙酰胆碱M2/M4受体亚型在调节脊髓背角神经元谷氨酸能递质释放中的作用[J]. 南方医科大学学报,2013,33(6):838-841.Du W, Guo Y, Yuan WX. Role of muscarinic cholinergic receptor subtypes in regulating glutamatergic synaptic transmission in rat spinal dorsal horn[J]. J South Med Univ, 2013, 33(6): 838-841.
[5] Krajcs N, Hernádi L, Elekes K, et al. Excitatory neurotransm-itters in the tentacle flexor muscles responsible for space positioning of the snail olfactory organ[J]. Invert Neurosci, 2014, 14(1): 59-69.
[6] 陈久林,孙申维,郁志华,等.参枝苓口服液治疗轻度认知功能损害的临床疗效观察[J]. 中西医结合心脑血管病杂志,2017,15(3):365-368.Chen JL, Sun SW, Yu ZH, et al. Clinical observation on Shenzhiling Oral Liquid in the treatment of mild cognitive impairment[J]. Chinese Journal of Integrative Medicine Cardio-/Cerebrovascular Disease, 2017, 15(3): 365-368.
[7] 吴艺琼,陈芳,盛宁,等.参枝苓口服液对APPswe/PSldE9小鼠学习记忆和突触超微结构的影响[J].北京中医药大学学报,2018,41(10):821-827.Wu YQ, Chen F, Sheng N, et al. Effects of Shenzhiling Oral Liquid on learning and memory and synaptic structure in APPswe/PSldE9 mice[J]. Journal of Beijing University of Traditional Chinese Medicine, 2018, 41(10): 821-827.
[8] Shinohara M, Koga S, Konno T, et al. Distinct spatiot -emporal accumulation of N-truncated and full-length amyloid-β42 in Alzheimer's disease[J]. Brain A Journal of Neurology, 2017, 140(12): 3301-3316.
[9] Lewis AS, Schalkwyk GI, Bloch MH. Alpha-7 nicotinic agonists for cognitive deficits in neuropsychiatric disorders: A translational meta-analysis of rodent and human studies[J]. Prog Neuropsychopharmacol Biol Psychiatry, 2017, 75(Complete): 45-53.
[10] Takata K, Amamiya T, Mizoguchi H, et al. Alpha7 nicotinic acetylcholine receptor-specific agonist DMXBA (GTS-21) attenuates Aβ accumulation through suppression of neuronal γ-secretase activity and promotion of microglial amyloid-β phagocytosis and ameliorates cognitive impairment in a mouse model of Alzheimer’s disease[J]. Neurobiol Aging, 2018, 62: 197-209.
[11] Chen L, Yamada K, Nabeshima T, et al. α7 Nicotinic acetylcholine receptor as a target to rescue deficit in hippocampal LTP induction in β-amyloid infused rats[J]. Neuropharmacology, 2006, 50(2): 254-268.
[12] Huang M, Felix AR, Flood DG, et al. The novel α7 nicotinic acetylcholine receptor agonist EVP-6124 enhances dopamine, acetylcholine, and glutamate efflux in rat cortex and nucleus accumbens[J]. Psychopharmacology, 2014, 231(23): 4541-4551.
[13] Raka F, Sebastiano AR, Kulhawy SC, et al. Ca(2+)/Calmodulin-dependent protein Kinase II interacts with group I Metabotropic Glutamate and facilitates Receptor Endocytosis and ERK1/2 signaling: role of β-Amyloid[J]. Molecular Brain, 2015, 8(1): 1-15.
[14] Um JW, Kaufman AC, Kostylev M, et al. Metabotropic glutamate receptor 5 is a coreceptor for Alzheimer Aβ oligomer bound to cellular prion protein[J]. Neuron, 2013, 79(5): 887-902.
[15] Shrivastava AN, Kowalewski JM, Renner M, et al. β-amyloid and ATP-induced diffusional trapping of astrocyte and neuronal metabotropic glutamate type-5 receptors[J]. Glia, 2013, 61(10): 1673-1686.
[16] Overk CR, Cartier A, Shaked G, et al. Hippocampal neuronal cells that accumulate α-synuclein fragments are more vulnerable to Aβ oligomer toxicity via mGluR5-implications for dementia with Lewy bodies[J]. Mol Neurodegener., 2014, 9(1): 1-18.
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