中樞神經(jīng)系統(tǒng)腦衰反應(yīng)調(diào)節(jié)蛋白2的研究進(jìn)展

邢進(jìn) 孫兆良 馮東福

(上海交通大學(xué)醫(yī)學(xué)院附屬第九人民醫(yī)院神經(jīng)外科，上海 201900)

·綜述·

中樞神經(jīng)系統(tǒng)腦衰反應(yīng)調(diào)節(jié)蛋白2的研究進(jìn)展

邢進(jìn) 孫兆良 馮東福*

(上海交通大學(xué)醫(yī)學(xué)院附屬第九人民醫(yī)院神經(jīng)外科，上海 201900)

腦衰反應(yīng)調(diào)節(jié)蛋白2; 神經(jīng)元軸突; 中樞神經(jīng)系統(tǒng)疾病

腦衰反應(yīng)調(diào)節(jié)蛋白2( collapsin response mediator protein2,CRMP2)又稱二氫嘧啶酶相關(guān)蛋白2，屬于CRMPs蛋白家族的5種亞型之一。在生理情況下CRMP2可作用于細(xì)胞骨架成分(如微管、微絲等)，通過對生長錐的生長、塌陷等行為的調(diào)節(jié)參與神經(jīng)元的生長發(fā)育過程。此外，CRMP2還與多種中樞神經(jīng)系統(tǒng)疾病的發(fā)生、進(jìn)展密切相關(guān)。

一、CRMP2的結(jié)構(gòu)及在神經(jīng)系統(tǒng)中的表達(dá)

CRMP2是CRMPs蛋白家族中最先發(fā)現(xiàn)的成員。1995年Goshima等在研究腦信號蛋白3A(Semaphorin 3A,Sema3A)介導(dǎo)雞背根神經(jīng)節(jié)細(xì)胞突起坍塌時(shí)克隆出CRMP2，隨后CRMPs家族的其他成員CRMP1、3、4、5陸續(xù)被發(fā)現(xiàn)。CRMP2具有同源四聚體結(jié)構(gòu)，可分為A、B兩種亞型。A亞型分子量約為75 kDa，存在于神經(jīng)元的胞體和軸突中。B亞型是CRMP2的主要亞型，分子量約為62～66 kDa，存在于軸突和樹突中[1]。CRMP2在中樞神經(jīng)系統(tǒng)的各個階段均可表達(dá)，其中在發(fā)育期的腦組織中表達(dá)量最多。在中樞神經(jīng)發(fā)育過程中隨著神經(jīng)干細(xì)胞出現(xiàn)分化，CRMP2的表達(dá)即開始上調(diào)。在成年腦組織中，CRMP2是CRMPs蛋白家族表達(dá)最多的成員，主要存在于嗅覺系統(tǒng)、小腦及海馬組織中。

二、中樞神經(jīng)系統(tǒng)CRMP2的生理功能

1.調(diào)控神經(jīng)元軸突生長：中樞神經(jīng)系統(tǒng)發(fā)育的一個重要環(huán)節(jié)是神經(jīng)元樹突和軸突通過突觸連接形成和重塑神經(jīng)纖維網(wǎng)絡(luò)。而樹突和軸突之所以能準(zhǔn)確到達(dá)其特定靶結(jié)構(gòu)并與之建立結(jié)構(gòu)和功能聯(lián)系，則取決于突起末端生長錐的運(yùn)動。

CRMP2作為Ras相似物家族的鳥苷三磷酸酶(Ras homologue family of small GTPases,Rho GTPases)下游信號和作用底物參與介導(dǎo)軸突生長、塌陷及導(dǎo)向過程。高表達(dá)的CRMP2可誘導(dǎo)神經(jīng)元形成多個軸突，并使樹突軸突化，而內(nèi)源性CRMP2缺失則可抑制軸突形成[2]。此外，神經(jīng)元突起的形成和生長延伸還需要微管運(yùn)動來實(shí)現(xiàn)。CRMP2既能以三磷酸鳥苷酶(guaanosine triphosphate,GTP)活性蛋白的形式激活GTP酶，對微管組裝和突起生成過程進(jìn)行調(diào)節(jié)，也可與微管蛋白二聚體結(jié)合促進(jìn)微管裝配從而調(diào)節(jié)突起生長[3]。

CRMP2亦是糖原合成酶激酶-3β(glycogen synthase kinase-3β,GSK-3β)的下游信號之一，而這一信號通路在神經(jīng)元突起生長過程中具有重要作用。有研究發(fā)現(xiàn)GSK-3β可以通過磷酸化CRMP2的蘇氨酸509、514和絲氨酸518位點(diǎn)降低其與微管的親和力，從而抑制軸突生長[2]。還有研究發(fā)現(xiàn)腦源性神經(jīng)營養(yǎng)因子可以通過磷酸化GSK-3β使非磷酸化的CRMP2增多，從而促進(jìn)突起生長[4]。

CRMP2不僅能通過信號通路調(diào)控突起的生長，還能與一些細(xì)胞信號分子相互作用介導(dǎo)生長錐塌陷。磷脂酶D2是細(xì)胞信號轉(zhuǎn)導(dǎo)中一種重要的酶分子，能夠影響Sema3A 介導(dǎo)的生長錐塌陷這一信號通路，在細(xì)胞骨架肌動蛋白的重排中有重要意義。CRMP2可特異性抑制磷脂酶D2活性從而直接干擾Sema3A介導(dǎo)的信號通路，引起肌動蛋白解聚，導(dǎo)致生長錐的塌陷[5]。另一種介導(dǎo)生長錐塌陷的細(xì)胞外信號分子-溶血磷脂酸，其與受體結(jié)合后會激活RhoA激酶，隨后下游的Ras相似物激酶(Ras homologue,Rho)活性增強(qiáng)，進(jìn)而導(dǎo)致CRMP2 蘇氨酸555位點(diǎn)磷酸化從而引起生長錐塌陷[3]。

2.調(diào)控N型鈣離子通道：N型電壓門控鈣通道(N-type Ca2+channels,CaV2.2)是神經(jīng)遞質(zhì)釋放的關(guān)鍵介體，活化后可引起鈣離子內(nèi)流和多種神經(jīng)遞質(zhì)(如谷氨酸、P物質(zhì)和降鈣素基因相關(guān)肽等)的釋放，對傷害感受的傳導(dǎo)起著重要作用。CRMP2是CaV2.2的一種調(diào)節(jié)子，與CaV2.2相結(jié)合后可增強(qiáng)其功能[6]。CRMP2過表達(dá)不僅可引起細(xì)胞膜表面CaV2.2數(shù)量增加和Ca2+電流增強(qiáng)[7]，還可增加降鈣素基因相關(guān)肽從背根神經(jīng)節(jié)中的釋放，進(jìn)而加重慢性炎性疼痛、提高神經(jīng)性傷害刺激的敏感性[8]。而CRMP2-CaV2.2解偶聯(lián)可降低Ca2+電流和減少神經(jīng)遞質(zhì)釋放，從而減輕慢性炎性疼痛、降低神經(jīng)性傷害刺激的敏感性[8]。轉(zhuǎn)錄反式激活因子-鈣通道結(jié)合域3能降低CRMP2與CaV2.2結(jié)合的能力，從而抑制炎性和神經(jīng)性疼痛[9]。

3.介導(dǎo)神經(jīng)元遷移：哺乳動物大腦發(fā)育過程中，多種信號通路介導(dǎo)CRMP2的高表達(dá)，誘導(dǎo)神經(jīng)元遷移，調(diào)控大腦皮層發(fā)育。位于Rho GAP信號通路中的α2嵌合蛋白可調(diào)節(jié)CRMP2的活性、影響神經(jīng)元內(nèi)CRMP2的聚集部位，調(diào)控生長錐及突起生長的方向，進(jìn)而影響神經(jīng)元遷移[10]。CRMP2除受α2嵌合蛋白調(diào)節(jié)外，還受到支架蛋白Axin的調(diào)控。Axin蛋白被細(xì)胞周期蛋白依懶性激酶5(cyclin dependent kinase 5,CDK5)磷酸化后導(dǎo)致GSK-3β活性降低，進(jìn)而引起CRMP2去磷酸化減少，最終使新生神經(jīng)元軸突增多[11]。此外，在皮層發(fā)育過程中，CRMP2的轉(zhuǎn)錄翻譯還受到骨形成蛋白的嚴(yán)格調(diào)控。骨形成蛋白通過調(diào)控轉(zhuǎn)錄因SMAD蛋白1抑制CRMP2的表達(dá)，減少新生神經(jīng)元的數(shù)量，影響腦皮質(zhì)板層結(jié)構(gòu)的形成[12]。

四、CRMP2與中樞神經(jīng)系統(tǒng)疾病

1.阿爾茲海默?。喊柶澓Ｄ∈且环N常見的神經(jīng)退行性疾病，淀粉樣蛋白斑和神經(jīng)原纖維纏結(jié)(neurofibrillary tangles,NFTs)是其最顯著的兩個病理特征。β-淀粉樣前體蛋白(β-amyloid precursor protein,β-APP)裂解成Aβ并聚集形成淀粉樣蛋白斑，NFTs則由微管結(jié)合蛋白tau過磷酸化后聚集產(chǎn)生。有研究發(fā)現(xiàn)在各種神經(jīng)退行性疾病中，高磷酸化的CRMP2只特異性地出現(xiàn)在阿爾茲海默患者腦中[13]。CRMP2的過度磷酸化可在淀粉樣蛋白斑和NFTs形成之前被檢測到，這表明它可能是阿爾茲海默病進(jìn)展的早期病理特征[14]。在阿爾茨海默病患者腦組織中可發(fā)現(xiàn)CRMP2在蘇氨酸509，色氨酸518、522 等特定位點(diǎn)高度磷酸化[15]。有研究表明，CRMP2過度磷酸化是由Cdk5和GSK3β信號通路介導(dǎo)[16]。一個有吸引力的假說是CRMP2過度磷酸化降低了功能性CRMP2的數(shù)量，導(dǎo)致軸突運(yùn)輸缺陷、微管動力學(xué)損害。而在阿爾茨海默病中，過量的β-淀粉樣前體蛋白是通過軸突運(yùn)輸?shù)姆绞竭\(yùn)輸?shù)?，上述過程將損害β-淀粉樣蛋白的運(yùn)輸和處理，使β-APP裂解形成Aβ斑，最終導(dǎo)致神經(jīng)元的死亡[3]。

2.CRMP2與缺血缺氧性腦損傷：在缺血缺氧(hypoxic-ischemic,HI)腦損傷后神經(jīng)元死亡的過程中，是由多種信號通路介導(dǎo)CRMP2去磷酸化發(fā)揮了重要作用。在新生大鼠HI腦損傷模型中發(fā)現(xiàn)Akt表達(dá)升高，同時(shí)伴隨GSK-3β磷酸化及CRMP2去磷酸化過程。而抑制Akt表達(dá)后GSK-3β磷酸化減少、CRMP2磷酸化增加[17]。CRMP2的磷酸化狀態(tài)除受Akt的調(diào)控外，還受CDK5活性影響。對腦室周圍白質(zhì)軟化癥小鼠行HI處理48h后即可檢測到腦組織中CRMP2水解片段和去磷酸化的CRMP2。進(jìn)一步通過光譜分析和western blot檢測發(fā)現(xiàn)HI處理后CRMP2的低磷酸化是由于CDK5活性降低介導(dǎo)的。此外，腦低氧預(yù)適應(yīng)是一種內(nèi)源性保護(hù)機(jī)制，能提高組織器官對低氧-缺血的耐受。蛋白激酶Cγ(protein kinase Cγ,cPKCγ)激活是預(yù)適應(yīng)形成的關(guān)鍵因素，并可與CRMP2相互作用發(fā)揮保護(hù)效應(yīng)。在腦缺血小鼠皮質(zhì)內(nèi)磷酸化CRMP2明顯增多，而注射cPKCγ抑制劑后CRMP2蛋白水解片段增多、CRMP2處于低磷酸化狀態(tài)[18]。

3.CRMP2與創(chuàng)傷性腦損傷：創(chuàng)傷性腦損傷后N-甲基-D-天冬氨酸受體過度活化導(dǎo)致Ca2+大量內(nèi)流，激活了神經(jīng)毒性級聯(lián)反應(yīng)。而CRMP2可能通過抑制這一過程從而減少神經(jīng)元死亡。此外，在創(chuàng)傷性腦損傷后神經(jīng)組織修復(fù)過程中再生的突觸異常增多，導(dǎo)致突觸間相互作用增加、神經(jīng)元興奮性增加從而引起癲癇發(fā)作。通過抑制CRMP2介導(dǎo)的微管聚合反應(yīng)影響突觸再生數(shù)量，使神經(jīng)元興奮性降低，可減少創(chuàng)傷性腦損傷所致的癲癇發(fā)作。

4.CRMP2與其他：CRMP2還與其他神經(jīng)系統(tǒng)疾病的發(fā)生、發(fā)展有密切關(guān)系。核磁共振結(jié)果顯示精神分裂癥和抑郁癥患者額葉皮質(zhì)內(nèi)，CRMP2表達(dá)較正常人顯著減少[19]。在1型神經(jīng)纖維瘤病中，CRMP2的磷酸化受神經(jīng)纖維瘤蛋白調(diào)節(jié)，其表達(dá)減少可能導(dǎo)致神經(jīng)元細(xì)胞的功能受損[20]。而在帕金森患者中，有研究發(fā)現(xiàn)發(fā)現(xiàn)CRMP2可調(diào)控Sema3A介導(dǎo)的神經(jīng)元凋亡過程[21]。CRMP2(蘇氨酸555位點(diǎn))的磷酸化過程還受到生長抑制因子-A/生長抑制因子-66受體信號通路的調(diào)控，導(dǎo)致軸突變性誘發(fā)自身免疫性腦脊髓炎和多發(fā)性硬化癥[22]。

終上所述，CRMP2在中樞神經(jīng)系統(tǒng)中發(fā)揮著重要的功能，通過多種信號通路調(diào)控神經(jīng)元軸突生長、N型電壓門控鈣通道、神經(jīng)元遷移。此外，CRMP2的磷酸化水平還與多種中樞神經(jīng)系統(tǒng)疾病的發(fā)生發(fā)展過程密切相關(guān)。隨著研究的不斷深入、相關(guān)信號通路的不斷闡明，CRMP2可以為神經(jīng)系統(tǒng)疾病的早期發(fā)現(xiàn)和尋找新的治療靶點(diǎn)提供重要思路。

1Soutar MP,Thornhjll P,Cole AR,et al. Increased CRMP2 phosphorylation is observed in Alzheimer's disease:does this tell us anything about disease development ? [J]. Curr Alzheimer Res,2009,6(3):269-278.

2Yoshimura T,Kawano Y,Arimura N,et al. GSK-3beta regulates phosphorylation of CRMP-2 and neuronal polarity [J]. Cell,2005,120( 1):137-149.

3Hensley K,Venkova K,Christov A,et al. Collapsin response mediator protein-2:an emerging pathologic feature and therapeutic target for neurodisease indications [J]. Mol Neurobiol,2011,43(3):180-191.

4Namekata K,Harada C,Guo X,et al. Dock3 stimulates axonal outgrowth via GSK-3β-mediated microtubule assembly [J]. J Neurosci,2012,32(1):264-274.

5Arimura N,Inagaki N,Chihara K,et al. Phosphorylation of CRMP-2 by Rho-kinase. Evidence for two separate signaling pathways for growth cone collapse [J]. J Biol Chem,2000,275(31):23973-23980.

6Brittain JM,Piekarz AD,Wang Y,et al. An atypical role for collapsin response mediator protein2 (CRMP-2)in neurotransmitter release via interaction with presynaptic voltage-gated Ca2+channels [J]. J Biol Chem,2009,284(45):31375-31390.

7Wang Y,BriItain JM,Wilson SM,et al. Emerging roles of collapsin response mediator proteins(CRMPs)as regulators of voltage-gated calcium channels and synaptic transmission [J]. Commun Integr Biol,2010,3(2):172-175.

8Feldman P,Khanna R. Challenging the catechism of therapeutics for chronic neuropathic pain:Targeting CaV2.2 interactions with CRMP2 peptides [J]. Neurosci Lett,2013,557 (Pt A):27-36.

9Wilson SM,Brittain JM,Piekarz AD,et al. Further insights into the antinociceptive potential of a peptide disrupting the N-type calcium channel-CRMP-2 signaling complex [J]. Channels (Austin),2011,5(5):449-456.

10Ip JP,Shi L,Chen Y,et al. Alpha2-chimaerin controls neuronal migration and functioning of the cerebral cortex through CRMP-2 [J]. Nat Neurosci,2011,15(1):39-47.

11Fang WQ,Ip JP,Li R,et al. Cdk5-mediated phosphorylation of Axin directs axon formation during cerebral cortex development [J]. J Neurosci,2011,31(38):13613-13624.

12Sun Y,Fei T,Yang T,et al. The suppression of CRMP2 expression by bone morphogenetic protein (BMP)-SMAD gradient signaling controls multiple stages of neuronal development [J]. J Biol Chem,2010,285(50):39039-39050.

13Williamson R,van Aalten L,Mann DM,et al. CRMP2 hyperphosphorylation is characteristic of Alzheimer's disease and not a feature common to other neurodegenerative diseases [J]. J Alzheimers Dis,201l,27(3):615-625.

14Cole AR,Noble W,van Aalten L,et al. Collapsin response mediator protein-2 hyperphosphorylation is an early event in Alzheimer's disease progression [J]. J Neurochem,2007,103(3):1132-1144.

15Gu Y,Hamajima N,Ihara Y. Neurofibrillary tangle-associated collapsin response mediator protein-2 (CRMP-2) is highly phosphorylated on Thr-509,Ser-518 and Ser-522 [J]. Biochemistry,2000,39(15):4267-4275.

16Cheung ZH,Ip NY. Cdk5:a multifaceted kinase in neurodegenerative diseases [J]. Trends Cell Biol,2012,22(3):169-175.

17Xiong T,Tang J,Zhao J,et al. Involvement of the Akt/GSK-3β/CRMP-2 pathway in axonal I njury after hypoxicischemic brain damage in neonatal rat [J]. Neuroscience,2012,216:123-132.

18劉燕燕,楊璇,韓松,等. cPKCγ參與低氧預(yù)適應(yīng)對小鼠腦缺血皮質(zhì)內(nèi)cRMP2水解和磷酸化的調(diào)節(jié) [J]. 基礎(chǔ)醫(yī)學(xué)與臨床,2012,32(1):25-30.

19Glausier JR,Lewis DA. Dendritic spine pathology in schizophrenia [J]. Neuroscience,2013,251:90-107.

20Patrakitkomjorn S,Kobayashi D,Morikawa T,et al. Neurofibromatosis type 1 (NF1) tumor suppressor,neurofibromin,regulates the neuronal differentiation of PC12 cells via its associating protein,CRMP-2 [J]. J Biol Chem,2008,283(14):9399-9413.

21Barzilai A,Zilkha-Falb R,Daily D,et al. The molecular mechanism of dopamine-induced apoptosis:identification and characterization of genes that mediate dopamine toxicity [J]. J Neural Transm Suppl,2000,60(60):59-76.

22Petratos S,Ozturk E,Azari MF,et al. Limiting multiple sclerosis related axonopathy by blocking Nogo receptor and CRMP-2 phosphorylation [J]. Brain,2012,135(Pt 6):1794-1818.

J Neurosurg. 2016 Dec 2:1-6. [Epub ahead of print]

Size and location of ruptured intracranial aneurysms:consecutive series of 1993 hospital-admitted patients

KorjaM1,KivisaariR1,RezaiJahromiB1,LehtoH1

1DepartmentofNeurosurgery,UniversityofHelsinkiandHelsinkiUniversityHospital,Helsinki,Finland

ObjectiveLarge consecutive series on the size and location of ruptured intracranial aneurysms (RIAs) are limited,and therefore it has been difficult to estimate population-wide effects of size-based treatment strategies of unruptured intracranial aneurysms. The authors' aim was to define the size and location of RIAs in patients diagnosed with subarachnoid hemorrhagedue to aneurysm rupture in a high-volume academic center.MethodsConsecutive patients admitted to a large nonprofit academic hospital with saccular RIAs between 1995 and 2009 were identified,and the size,location,and multiplicity of RIAs were defined and reported by patient sex.ResultsIn the study cohort of 1993 patients (61% women) with saccular RIAs,the 4 most common locations of RIAs were the middle cerebral (32%),anterior communicating (32%),posterior communicating (14%),and pericallosal arteries (5%). However,proportional distribution of RIAs varied considerably by sex;for example,RIAs of the anterior communicating artery were more frequently found in men than in women. Anterior circulation RIAs accounted for 90% of all RIAs,and 30% of the patients had multiple intracranial aneurysms. The median size (measured as maximum diameter) of all RIAs was 7 mm (range 1～43 mm),but the size varied considerably by location. For example,RIAs of the ophthalmic artery had a median size of 11 mm,whereas the median size of RIAs of the pericallosal artery was 6 mm. Of all RIAs,68% were smaller than 10 mm in maximum diameter.ConclusionsIn this large consecutive series of RIAs,83% of all RIAs were found in 4 anterior circulation locations. The majority of RIAs were small,but the size and location varied considerably by sex. The presented data may be of help in defining effective prevention strategies.

1671-2897(2016)15-564-03

國家自然科學(xué)基金資助項(xiàng)目(81372047)

邢進(jìn)，碩士研究生，E-mail:xj980906260@163.com

*通訊作者：馮東福，主任醫(yī)師，E-mail:dffeng@21cn.com

R 651.15

A

2015-10-05)