熊果酸改善精神分裂症小鼠脱髓鞘和脑组织间液引流紊乱

doi:10.19723/j.issn.1671-167X.2024.03.016

摘要/Abstract

摘要：

目的: 探讨精神分裂症(schizophrenia, SZ)的髓鞘和脑组织间液(interstitial fluid, ISF)相关发病机制，探究熊果酸(ursolic acid, UA)对SZ中髓鞘损伤及其继发的ISF异常引流的治疗效果。方法: 使用30只6~8周雌性C57BL/6J小鼠，体质量(20±2) g，随机分为UA治疗组、SZ模型组、对照组3组，每组10只：(1)对照组：腹腔注射(intraperitoneal injection, ip)生理盐水，灌胃(intragastric, ig)给予1%(质量分数)羧甲基纤维素钠(carboxymethylcellulose sodium, CMC-Na)；(2)SZ模型组：ip给与2 mg/kg的地卓西平(dizocilpine maleate，MK-801)，ig给与1% (质量分数)CMC-Na；(3)UA治疗组：ig给与25 mg/kg的UA，ip给与2 mg/kg的MK-801。治疗组先ig给药UA预治疗一周，然后对3组小鼠进行为期两周的药物干预。在造模完成后依次通过旷场测试和前脉冲抑制实验对小鼠进行行为学评价。行为测试后，通过向脑区注射荧光示踪剂来探究各组ISF分区引流的改变；通过免疫荧光探究各组脑内水通道蛋白4(aquaporin 4, AQP4)极性分布的改变以及蛋白表达的变化；通过激光共聚焦显微镜(laser scanning confocal microscope, LSCM)髓鞘反射光成像研究鼠脑内髓鞘的变化。采用单因素方差分析(one-way ANOVA)对计量数据进行组间比较，使用TukeyHSD进行组间两两比较。结果: 旷场测试发现，模型组在旷场中运动的总路程[(7 949.39±1 140.55) cm vs. (2 831.01±1 212.72) cm, P < 0.001]和中央区域停留时间[(88.43±22.06) s vs. (56.85±18.58) s, P=0.011]显著高于对照组，而治疗组在旷场中运动总路程[(2 415.80±646.95) cm vs. (7 949.39±1 140.55) cm, P < 0.001]和中央区域停留时间[(54.78±11.66) s vs. (88.43±22.06) s, P=0.007]较模型组显著降低。前脉冲抑制实验发现，模型组给与预脉冲时对震惊反射的抑制率均显著低于对照组(P均 < 0.001)；治疗组上述指标较模型组显著增加(P均 < 0.001)。髓鞘反射光检测发现，模型组小鼠脑内存在显著的脱髓鞘，治疗组脑内脱髓鞘情况得到逆转。荧光示踪发现，模型组示踪剂向头侧皮层区的扩散面积和向尾侧丘脑区的返流面积均显著大于对照组[(13.93±3.35) mm² vs. (2.79±0.94) mm², P < 0.001; (2.48±0.38) mm² vs. (0.05±0.12) mm², P < 0.001]，且脑区间引流分区明显破坏；治疗组示踪剂向头侧皮层区的扩散面积和向尾侧丘脑区的返流面积均较模型组显著减小[(7.93±2.48) mm² vs. (13.93±3.35) mm², P < 0.001; (0.50±0.30) mm² vs. (2.48±0.38) mm², P < 0.001]。免疫荧光染色发现，模型组小鼠脑内AQP4极性分布遭到破坏，且模型组AQP4蛋白表达量较对照组明显下降[(3 663.88±733.77) μm² vs. (13 354.92±4 054.05) μm², P < 0.001]；治疗组较模型组AQP4极性分布得到改善，AQP4蛋白表达量较模型组显著升高[(11 104.68±3 200.04) μm² vs. (3 663.88±733.77) μm², P < 0.001]。结论: 一定剂量的UA干预可以改善SZ小鼠的行为学表现，这种改善表现为运动亢进和焦虑症状得到缓解，感觉运动门控功能得到恢复；其机制可能是通过改善SZ小鼠的脱髓鞘病理改变及ISF分区引流紊乱。

关键词: 精神分裂症, 脑组织间液, 脱髓鞘, 熊果酸

Abstract:

Objective: To unveil the pathological changes associated with demyelination in schizophrenia (SZ) and its consequential impact on interstitial fluid (ISF) drainage, and to investigate the therapeutic efficacy of ursolic acid (UA) in treating demyelination and the ensuing abnormalities in ISF drainage in SZ. Methods: Female C57BL/6J mice, aged 6-8 weeks and weighing (20±2) g, were randomly divided into three groups: control, SZ model, and UA treatment. The control group received intraperitoneal injection (ip) of physiological saline and intragastric administration (ig) of 1% carboxymethylcellulose sodium (CMC-Na). The SZ model group was subjected to ip injection of 2 mg/kg dizocilpine maleate (MK-801) and ig administration of 1% CMC-Na. The UA treatment group underwent ig administration of 25 mg/kg UA and ip injection of 2 mg/kg MK-801. The treatment group received UA pretreatment via ig administration for one week, followed by a two-week drug intervention for all the three groups. Behavioral assessments, including the open field test and prepulse inhibition experiment, were conducted post-modeling. Subsequently, changes in the ISF partition drainage were investigated through fluorescent tracer injection into specific brain regions. Immunofluorescence analysis was employed to examine alterations in aquaporin 4 (AQP4) polarity distribution in the brain and changes in protein expression. Myelin reflex imaging using Laser Scanning Confocal Microscopy (LSCM) was utilized to study modifications in myelin within the mouse brain. Quantitative data underwent one-way ANOVA, followed by TukeyHSD for post hoc pairwise comparisons between the groups. Results: The open field test revealed a significantly longer total distance [(7 949.39±1 140.55) cm vs. (2 831.01±1 212.72) cm, P < 0.001] and increased central area duration [(88.43±22.06) s vs. (56.85±18.58) s, P=0.011] for the SZ model group compared with the controls. The UA treatment group exhibited signifi-cantly reduced total distance [(2 415.80±646.95) cm vs. (7 949.39±1 140.55) cm, P < 0.001] and increased central area duration [(54.78±11.66) s vs. (88.43±22.06) s, P=0.007] compared with the model group. Prepulse inhibition test results demonstrated a markedly lower inhibition rate of the startle reflex in the model group relative to the controls (P < 0.001 for both), with the treatment group displaying significant improvement (P < 0.001 for both). Myelin sheath analysis indicated significant demyelination in the model group, while UA treatment reversed this effect. Fluorescence tracing exhibited a significantly larger tracer diffusion area towards the rostral cortex and reflux area towards the caudal thalamus in the model group relative to the controls [(13.93±3.35) mm² vs. (2.79±0.94) mm², P < 0.001 for diffusion area; (2.48±0.38) mm² vs. (0.05±0.12) mm², P < 0.001 for reflux area], with significant impairment of drainage in brain regions. The treatment group demonstrated significantly reduced tracer diffusion and reflux areas [(7.93±2.48) mm² vs. (13.93±3.35) mm², P < 0.001 for diffusion area; (0.50±0.30) mm² vs. (2.48±0.38) mm², P < 0.001 for reflux area]. Immunofluorescence staining revealed disrupted AQP4 polarity distribution and reduced AQP4 protein expression in the model group compared with the controls [(3 663.88±733.77) μm² vs. (13 354.92±4 054.05) μm², P < 0.001]. The treatment group exhibited restored AQP4 polarity distribution and elevated AQP4 protein expression [(11 104.68±3 200.04) μm² vs. (3 663.88±733.77) μm², P < 0.001]. Conclusion: UA intervention ameliorates behavioral performance in SZ mice, Thus alleviating hyperactivity and anxiety symptoms and restoring sensorimotor gating function. The underlying mechanism may involve the improvement of demyelination and ISF drainage dysregulation in SZ mice.

Key words: Schizophrenia, Brain interstitial fluid, Demyelination, Ursolic acid

中图分类号:

R285.5

龙仁,毛鑫,高天姿,解倩,谈瀚博,李子寅,韩鸿宾,袁兰. 熊果酸改善精神分裂症小鼠脱髓鞘和脑组织间液引流紊乱[J]. 北京大学学报（医学版）, 2024, 56(3): 487-494.

Ren LONG,Xin MAO,Tianzi GAO,Qian XIE,Hanbo TAN,Ziyin LI,Hongbin HAN,Lan YUAN. Ursolic acid improved demyelination and interstitial fluid drainage disorders in schizophrenia mice[J]. Journal of Peking University (Health Sciences), 2024, 56(3): 487-494.

图/表 7

图1

图2

图3

图4

图5

图6

图7

参考文献 33

1	Stewart LA .Chemotherapy in adult high-grade glioma: A systematic review and meta-analysis of individual patient data from 12 randomised trials[J].Lancet,2002,359(9311):1011-1018. doi: 10.1016/S0140-6736(02)08091-1
2	Falkai P , Raabe F , Bogerts B , et al.Association between altered hippocampal oligodendrocyte number and neuronal circuit structures in schizophrenia: A postmortem analysis[J].Eur Arch Psychiatry Clin Neurosci,2020,270(4):413-424. doi: 10.1007/s00406-019-01067-0
3	Stępnicki P, Kondej M, Kaczor AA. Current concepts and treatments of schizophrenia[J/OL]. Molecules, 2018, 23(8) (2018-08-20)[2023-03-01]. https://www.mdpi.com/1420-3049/23/8/2087.
4	Koshiyama D , Fukunaga M , Okada N , et al.White matter microstructural alterations across four major psychiatric disorders: Nega-analysis study in 2937 individuals[J].Mol Psychiatry,2020,25(4):883-895. doi: 10.1038/s41380-019-0553-7
5	Karlsgodt KH .White matter microstructure across the psychosis spectrum[J].Trends Neurosci,2020,43(6):406-416. doi: 10.1016/j.tins.2020.03.014
6	Wang A , Wang R , Cui D , et al.the drainage of interstitial fluid in the deep brain is controlled by the integrity of myelination[J].Aging Dis,2019,10(5):937-948. doi: 10.14336/AD.2018.1206
7	赵越, 李昀倩, 李怀业, 等.荧光及磁示踪法观测脑组织液的引流分区特征[J].北京大学学报(医学版),2017,49(2):303-309. doi: 10.3969/j.issn.1671-167X.2017.02.021
8	Wang R , Han H , Shi K , et al.The alteration of brain interstitial fluid drainage with myelination development[J].Aging Dis,2021,12(7):1729-1740. doi: 10.14336/AD.2021.0305
9	Mccutcheon RA , Abi-Dargham A , Howes OD .Schizophrenia, dopamine and the Striatum: From biology to symptoms[J].Trends Neurosci,2019,42(3):205-220. doi: 10.1016/j.tins.2018.12.004
10	Kashyap D , Tuli HS , Sharma AK .Ursolic acid (UA): A metabolite with promising therapeutic potential[J].Life Sci,2016,146,201-213. doi: 10.1016/j.lfs.2016.01.017
11	Yamamoto S , Sakemoto C , Iwasa K , et al.Ursolic acid treatment suppresses cuprizone-induced demyelination and motor dysfunction via upregulation of IGF-1[J].J Pharmacol Sci,2020,144(3):119-122. doi: 10.1016/j.jphs.2020.08.002
12	Ramos-Hryb AB , Pazini FL , Kaster MP , et al.Therapeutic potential of ursolic acid to manage neurodegenerative and psychiatric diseases[J].CNS Drugs,2017,31(12):1029-1041. doi: 10.1007/s40263-017-0474-4
13	He J , Zu Q , Wen C , et al.Quetiapine attenuates schizophrenia-like behaviors and demyelination in a MK-801-induced mouse model of schizophrenia[J].Front Psychiatry,2020,11,843.
14	Nakamoto C , Kawamura M , Nakatsukasa E , et al.GluD1 knockout mice with a pure C57BL/6N background show impaired fear memory, social interaction, and enhanced depressive-like behavior[J].PLoS One,2020,15(2):e0229288. doi: 10.1371/journal.pone.0229288
15	Kraeuter AK , Mashavave T , Suvarna A , et al.Effects of beta-hydroxybutyrate administration on MK-801-induced schizophrenia-like behaviour in mice[J].Psychopharmacology (Berl),2020,237(5):1397-1405. doi: 10.1007/s00213-020-05467-2
16	Schilke ED , Tremolizzo L , Appollonio I , et al.Tics: Neurological disorders determined by a deficit in sensorimotor gating processes[J].Neurol Sci,2022,43(10):5839-5850. doi: 10.1007/s10072-022-06235-0
17	Ruggiero RN , Rossignoli MT , De ross JB , et al.Cannabinoids and vanilloids in schizophrenia: Neurophysiological evidence and directions for basic research[J].Front Pharmacol,2017,8,399. doi: 10.3389/fphar.2017.00399
18	Iliff JJ , Wang M , Liao Y , et al.A paravascular pathway facilitates CSF flow through the brain parenchyma and the clearance of interstitial solutes, including amyloid β[J].Sci Transl Med,2012,4(147):147r.
19	Chen S , Tang Y , Fan X , et al.The role of white matter abnor-mality in the left anterior corona radiata: In relation to formal thought disorder in patients with schizophrenia[J].Psychiatry Res,2022,307,114302. doi: 10.1016/j.psychres.2021.114302
20	Di biase MA , Zhang F , Lyall A , et al.Neuroimaging auditory verbal hallucinations in schizophrenia patient and healthy populations[J].Psychol Med,2020,50(3):403-412. doi: 10.1017/S0033291719000205
21	Walch E , Fiacco TA .Honey, I shrunk the extracellular space: Measurements and mechanisms of astrocyte swelling[J].Glia,2022,70(11):2013-2031. doi: 10.1002/glia.24224
22	Walch E , Bilas A , Bebawy V , et al.Contributions of astrocyte and neuronal volume to CA1 neuron excitability changes in elevated extracellular potassium[J].Front Cell Neurosci,2022,16,930384. doi: 10.3389/fncel.2022.930384
23	DoiT, Fan Y, Gold J I, et al. The caudate nucleus contributes causally to decisions that balance reward and uncertain visual information[J]. Elife, 2020, 9(2020-06-22)[2023-03-01]. https://elifesciences.org/articles/56694.
24	Kirino E , Tanaka S , Fukuta M , et al.Functional connectivity of the caudate in schizophrenia evaluated with simultaneous resting-state functional MRI and electroencephalography recordings[J].Neuropsychobiology,2019,77(4):165-175. doi: 10.1159/000490429
25	Teng Z , Wang A , Wang P , et al.The effect of aquaporin-4 knockout on interstitial fluid flow and the structure of the extracellular space in the deep brain[J].Aging Dis,2018,9(5):808-816. doi: 10.14336/AD.2017.1115
26	Tekieh T , Robinson P A , Postnova S .Cortical waste clearance in normal and restricted sleep with potential runaway tau buildup in Alzheimer's disease[J].Sci Rep,2022,12(1):13740. doi: 10.1038/s41598-022-15109-6
27	Carotenuto A , Cacciaguerra L , Pagani E , et al.Glymphatic system impairment in multiple sclerosis: relation with brain damage and disability[J].Brain,2021,145(8):2785-2795.
28	Aggleton JP , O'mara SM .The anterior thalamic nuclei: Core components of a tripartite episodic memory system[J].Nat Rev Neurosci,2022,23(8):505-516.
29	Halassa MM , Sherman SM .Thalamocortical circuit motifs: A general framework[J].Neuron,2019,103(5):762-770. doi: 10.1016/j.neuron.2019.06.005
30	Byne W , Hazlett EA , Buchsbaum MS , et al.The thalamus and schizophrenia: Current status of research[J].Acta Neuropathol,2009,117(4):347-368. doi: 10.1007/s00401-008-0404-0
31	Zhang Y , Li X , Ciric B , et al.A dual effect of ursolic acid to the treatment of multiple sclerosis through both immunomodulation and direct remyelination[J].Proc Natl Acad Sci USA,2020,117(16):9082-9093. doi: 10.1073/pnas.2000208117
32	Faden J , Citrome L .Schizophrenia: One name, many different manifestations[J].Med Clin North Am,2023,107(1):61-72. doi: 10.1016/j.mcna.2022.05.005
33	Flint J , Munafò M .Schizophrenia: Genesis of a complex disease[J].Nature,2014,511(7510):412-413. doi: 10.1038/nature13645

Metrics

Viewed

Full text

Abstract

Cited

Shared

Discussed

[1]	刘小璇,张朔,马妍,孙阿萍,张英爽,樊东升. 腓骨肌萎缩症1A型患者和慢性炎性脱髓鞘性多发性神经病患者F波改变的比较[J]. 北京大学学报（医学版）, 2023, 55(1): 160-166.
[2]	王雪萍,张于亚楠,卢天兰,卢喆,康哲维,孙瑶瑶,岳伟华. 首发精神分裂症肠道微生物多态性与临床症状及血清代谢组学的关联[J]. 北京大学学报（医学版）, 2022, 54(5): 863-873.
[3]	白宁，杨凌飞，安丽华，王雯，李昀倩，盛荟，王彤，李华侃，袁兰. 精神分裂症模型鼠腹腔中的中性粒细胞趋化运动的动态可视化分析[J]. 北京大学学报(医学版), 2018, 50(2): 226-230.
[4]	郑丽娜, 郭茜, 李惠, 李春波, 王继军. 不同重复经颅磁刺激模式对精神分裂症认知功能和精神症状的影响[J]. 北京大学学报（医学版）, 2012, 44(5): 732-736.
[5]	韩雪, 杨磊, 程章, 张涛, 原岩波, 于欣. 首次发作精神分裂症患者及独立样本未患病一级亲属的神经认知功能：横断面研究[J]. 北京大学学报（医学版）, 2010, 42(6): 681-686.
[6]	漆学良, 陈静, 郑日亮, 李颖, 黄一宁, 袁云. 可缓解和复发的瘤样炎性脱髓鞘病1例[J]. 北京大学学报（医学版）, 2009, 41(5): 588-589.
[7]	汤宜朗, 王玉凤, 蔡焯基, 周儒伦, 汪冰, 周朝凤. 儿茶酚-○-甲基转移酶基因多态性与精神分裂症发病特点的关系[J]. 北京大学学报（医学版）, 2002, 34(4): 342-344.