汪鴻浩 胡學(xué)強(qiáng) (中山大學(xué)附屬第三醫(yī)院神經(jīng)科,廣州 510630)

多發(fā)性硬化(Multiple sclerosis,MS)是一種中樞神經(jīng)系統(tǒng)(Central nervous system,CNS)脫髓鞘性疾病,其真正病因至今尚未明確。國內(nèi)外有大量文獻(xiàn)證實(shí)其與細(xì)胞免疫、體液免疫均有關(guān)聯(lián)。自從2000年白細(xì)胞介素21(IL-21)和IL-21受體(IL-21R)被發(fā)現(xiàn)以來[1],一直是研究的熱點(diǎn),近年來又有很多新的作用被發(fā)現(xiàn)。IL-21對CD8+T細(xì)胞、CD4+Th細(xì)胞、調(diào)節(jié)性T細(xì)胞(regulatory T lymphocyte,Treg)均有作用,因此深入研究IL-21在MS發(fā)病中的意義具有重要的價(jià)值。

1 IL-21及IL-21R的生物學(xué)特點(diǎn)

人IL-21基因定位于4q26-q27[2],共642個(gè)核苷酸,其中自42位至535位核苷酸為編碼區(qū)。鼠定位于3號染色體上,兩者有57%的同源性[1]。IL-21是一個(gè)四螺旋簇Ⅰ型細(xì)胞因子,其結(jié)構(gòu)與γ c家族其他成員如 IL-2、IL-4、IL-15高度同源[3]。在體內(nèi)主要是活化的CD4+T細(xì)胞分泌IL-21[1]。IL-21R是一個(gè)由獨(dú)特的IL-21R亞單位和γ c組成的異二聚體,其中IL-21R亞單位為配體識(shí)別結(jié)合部位,γ c為信號傳導(dǎo)單位[4],與IL-2/IL-15Rβ鏈有高度同源性,由胞外和胞內(nèi)兩個(gè)部分組成,胞外區(qū)有細(xì)胞因子信號識(shí)別模塊,胞內(nèi)區(qū)有典型的Box1、Box2信號傳導(dǎo)區(qū)。IL-21分子信號傳遞途徑尚未完全清楚,但 JAK/STAT、MAPK和P13這三個(gè)途徑研究相對較多。IL-21可以通過激活STAT1和STAT3途徑,激活T細(xì)胞、B細(xì)胞和NK細(xì)胞,對固有免疫階段發(fā)揮相應(yīng)生物調(diào)節(jié)效應(yīng)[5];當(dāng)MAPK和PI3通路被特異抑制因子阻斷后IL-21介導(dǎo)的免疫細(xì)胞增殖即被阻斷[6]。另外IL-21R有STAT5錨定位點(diǎn),可激活STAT5途徑,作用于B細(xì)胞[4]。進(jìn)一步研究發(fā)現(xiàn),IL-21R胞質(zhì)區(qū)酪氨酸510(Y510)可以有效的介導(dǎo)STAT1和STAT3磷酸化,從而激活CD4+T細(xì)胞分化,而不依賴STAT5途徑。

2 IL-21在多發(fā)性硬化發(fā)病中的作用

2.1 多發(fā)性硬化發(fā)病中的免疫學(xué)機(jī)制 多發(fā)性硬化的發(fā)病機(jī)制中涉及體液免疫和細(xì)胞免疫。以前認(rèn)為,MS的發(fā)病和Th1/Th2分化的偏移有關(guān)。一般認(rèn)為MS患者CD4+Th1細(xì)胞分泌的TNF-α和IFN-γ是導(dǎo)致MS發(fā)病的重要細(xì)胞因子,另外CD4+Th2可以下調(diào)MS的自身免疫反應(yīng)[7]。近年來通過對MS患者腦組織免疫組化研究發(fā)現(xiàn),急性病灶內(nèi)浸潤的T細(xì)胞表達(dá)IL-17及IL-17mRNA均顯著增多,高于靜止病灶、慢性病灶及正常組織[8],表明MS的發(fā)病和IL-17相關(guān),而IL-17主要由活化的 Th17產(chǎn)生,故Th17與MS的病理機(jī)制之間的關(guān)系引起了關(guān)注。動(dòng)物實(shí)驗(yàn)也證明人體來源的Th17細(xì)胞能破壞小鼠血腦屏障,在體外對人類神經(jīng)元細(xì)胞有強(qiáng)烈的細(xì)胞毒性作用[9]。另外CD4+CD25+Foxp3+Treg功能低下是MS發(fā)病的另一重要原因。MS患者體內(nèi)CD4+CD25+CD45RA+初始 Treg比例減少,CD4+CD25+CD45RO+記憶性Treg比例相對增高,表達(dá)轉(zhuǎn)錄因子Foxp3減少,抑制T細(xì)胞活化能力減弱[10-13]。早在上世紀(jì)就發(fā)現(xiàn)了MS患者IgG1、IgG3、IgG4的鞘內(nèi)合成[14],通過腦脊液(Cerebrospinal fluid,CSF)蛋白電泳可見IgG寡克隆帶(Oligoclonal bands,OCB)。IgG抗原識(shí)別呈多樣性,有的識(shí)別髓鞘蛋白成份[15];有的識(shí)別病毒如EB病毒[16],據(jù)此認(rèn)為B細(xì)胞免疫在MS發(fā)病中有重要作用。

2.2 IL-21和Th17 IL-21是可由活化的Th17分泌,而且相對其他的CD4+T細(xì)胞亞群,Th17更加有選擇的通過STAT3途徑產(chǎn)生IL-21,轉(zhuǎn)錄因子STAT3直接和Th17上IL-21基因的啟動(dòng)子結(jié)合,調(diào)控IL-21基因的表達(dá)[17]。盡管由抗原呈遞細(xì)胞分泌的IL-6、TGF-β可以通過STAT3促進(jìn)Th17的分化[18],但是抗體封閉實(shí)驗(yàn)證明Th17細(xì)胞分泌的IL-21也是Th17分化的重要途徑,甚至在IL-6缺乏的動(dòng)物模型上IL-21仍可誘導(dǎo)Th17的分化[19]。IL-21聯(lián)合IL-6通過STAT3和孤核受體(orphan nuclear receptor,RORγ t)作用促進(jìn)Th17的分化,并能增加IL-23R的表達(dá),IL-23可以促進(jìn)Th17的分化,并能維持Th17的存活[20,21]。Th17細(xì)胞分泌的IL-21的功能有重要的生物學(xué)意義,因?yàn)橛?IL-6聯(lián)合TGF-β促進(jìn)分化的 Th17其IL-21/IL21R表達(dá)下降[19,22]。在用少突膠質(zhì)細(xì)胞髓鞘磷脂糖蛋白(Myelin oligodendrocyte glycoprotein,MOG)35-55進(jìn)行實(shí)驗(yàn)變態(tài)反應(yīng)性腦脊髓炎(Experimental autoimmune encephalomyelitis,EAE)模型造模之前給予IL-21注射會(huì)加重神經(jīng)系統(tǒng)損傷,而IL-21/IL-21R基因敲除的小鼠在同樣進(jìn)行造型后分泌IL-17的細(xì)胞減少,神經(jīng)系統(tǒng)損傷較輕[19]。

2.3 IL-21和Treg IL-21并不直接作用于Treg,但是可以干擾CD4+CD25+Treg對CD4+CD25-T細(xì)胞的免疫抑制作用[23],也有研究認(rèn)為IL-21與TGF-β聯(lián)合、IL-21和IL-6聯(lián)合可抑制Foxp3的表達(dá),減少CD4+初始T細(xì)胞分化形成Treg[19]。IL-21抑制Treg的免疫調(diào)節(jié)作用具體是通過什么信號傳導(dǎo)途徑來實(shí)現(xiàn)目前還不清楚,但是無論通過什么機(jī)制減少IL-21的產(chǎn)生,增強(qiáng)Treg免疫調(diào)節(jié)功能作用,可能是一種潛在的治療自身免疫性疾病的方法[24]。雖然理論上IL-21的水平升高可以抑制CD4+CD25+Foxp3+Treg的免疫調(diào)節(jié)功能,促進(jìn)自身免疫反應(yīng),但是Piao等[25]在EAE模型鼠試驗(yàn)中卻得出了相反的結(jié)果:封閉IL-21R卻導(dǎo)致CD4+CD25+Treg的減少,他認(rèn)為試驗(yàn)結(jié)果不同可能和EAE模型鼠的遺傳背景的差異有關(guān)。

2.4 IL-21對B細(xì)胞分化的影響 IL-21對B細(xì)胞的增殖分化起重要的作用,IL-21有促進(jìn)記憶性B細(xì)胞分化為CD138+漿細(xì)胞并促進(jìn)漿細(xì)胞分泌抗體[26],該作用主要通過影響 Bcl-6、Blimp-1(B-lymphocyte-induced maturation protein-1),Pax-5的表達(dá)實(shí)現(xiàn)[27]。IL-21可以上調(diào)Blimp-1、下調(diào)Pax-5的表達(dá)使記憶性B細(xì)胞分化為漿細(xì)胞,而上調(diào)Bcl-6的作用可以使生發(fā)中心的B細(xì)胞分化為同型記憶性B細(xì)胞[28-30]。在缺乏IL-21R的小鼠中,免疫刺激后IgG1和IgG2b、IgG3的含量較免疫刺激后的正常小鼠血清含量減少,但是IgG2a、IgM的水平幾乎未受影響。也有實(shí)驗(yàn)證明IL-21聯(lián)合CD40mb對刺激B細(xì)胞分泌抗體的作用是IL-10的100倍[31,32]。造模前給予IL-21注射相對于PBS注射的小鼠行EAE造模后外周血IgG含量明顯升高,而小鼠體內(nèi)受Th2產(chǎn)生的細(xì)胞因子如:IL-4調(diào)控的IgG2a卻未見明顯變化,并且B細(xì)胞缺乏的小鼠EAE造模成功后給予IL-21則對肢體殘疾程度未見明顯影響[33]。

2.5 IL-21和IFN-γ NK細(xì)胞在MS和EAE模型發(fā)病中的意義目前不肯定,但是研究證實(shí)NK細(xì)胞可以通過殺傷靶細(xì)胞、產(chǎn)生一系列的細(xì)胞因子在自身免疫性疾病的病理機(jī)制中發(fā)揮重要的作用[34]。將小鼠NK細(xì)胞轉(zhuǎn)移至IL-18基因敲除的NK細(xì)胞相對缺乏的小鼠體內(nèi),這些小鼠保持了對自身免疫性腦脊髓炎的易感性,而將IFN-γ缺乏的NK細(xì)胞轉(zhuǎn)移至同種小鼠體內(nèi)卻沒有保持這種特性[35]。NK細(xì)胞在MS發(fā)病中的作用和該細(xì)胞分泌的IFN-γ有關(guān),IFN-γ使初始T細(xì)胞向Th1分化,促進(jìn)EAE的發(fā)病[36,37]。在EAE模型上證實(shí)了在造模過程中注射IL-21可以提高NK細(xì)胞分泌IFN-γ的水平,促進(jìn)Th1的分化[35]。

3 展望

IL-21是重要的免疫平衡調(diào)節(jié)因子,對Th17、Treg、B細(xì)胞、NK細(xì)胞的分化和功能均有影響,在自身免疫性疾病的發(fā)病中有重要意義。以往MS的免疫學(xué)研究很大部分將細(xì)胞免疫和體液免疫分開研究,未找到兩者之間的聯(lián)系。隨著IL-21研究的深入,越來越多的證據(jù)表明,Th17細(xì)胞在MS病灶處的增多并分泌IL-21等細(xì)胞因子,一方面促進(jìn)CD4+T細(xì)胞的分化,另一方面促進(jìn)B細(xì)胞的分化可能是MS病理機(jī)制的一個(gè)重要因素,因此IL-21在MS發(fā)病中的作用值得去深入研究。但是IL-21在MS和EAE模型的發(fā)病機(jī)制中仍有很多問題懸而未決,如:IL-21可以誘導(dǎo)Th1免疫反應(yīng)相關(guān)基因IFN-γ、T-bet(T box expressed in T cells)、IL-2Rα、IL-12Rβ2 、IL-18R 、髓樣分化因子(myeloid differentiation factor88,MyD88)的表達(dá)[38],但是Wurster等[39]卻得出了相反的結(jié)論認(rèn)為IL-21主要是促進(jìn)Th2反應(yīng)的細(xì)胞因子,并抑制IFN-γ的分泌;另外,近年來一種新的CD4+細(xì)胞亞群被發(fā)現(xiàn),其表型為CD4+CXCR5+CD40L+ICOS+T細(xì)胞,稱之為濾泡輔助性T細(xì)胞(T follicular helper cells,Tfh),和Th17一樣其分化依賴IL-6、IL-21,而且分泌IL-21[40],該細(xì)胞亞群在MS發(fā)病中的作用仍有待進(jìn)一步探尋。

1 Parrish Novak J,Dillon S R,Nelson A et al.Interleukin 21 and its receptor are involved in NK cell expansion and regulation of lymphocyte function[J].Nature,2000;408:57-63.

2 Mehta D S,Wurster A L,Grusby M J.Biology of IL-21 and the IL-21 receptor[J].Immunol Rev,2004;202:84-95.

3 Habib T,Senadheera S,Weinberg K et al.The common gamma chain(γ c)is a required signaling component of the IL-21 receptor and supports IL-21-induced cell proliferation via JAK3[J].Biochemistry,2002;41(27):8725-8731.

4 Asano K,Ikegami H,Fujisawa T et al.The gene forhuman IL-21 and ge-netic susceptibility to type 1 diabetes in the Japanese[J].Ann N Y Acad Sci,2006;1079:47-50.

5 Mary Collins,Matthew J,Whitters Deborah A.IL-21 and IL-21 Receptor.Immunologic Research,2003;(28):131-140.

6 Zeng R,Spolski R,Casas E et al.Themolecularbasis of IL-21mediated proliferation[J].Blood,2007;109(10):4135-4142.

7 Hans Lassmannl,Richard M Ransohoff.The CD4+Th1 model for multiple sclerosis:a crucial re-appraisal[J].TRENDS in Immunology,2004;3(25):132-137.

8 Tzartos J S,Friese M A,Craner M J et al.Interleukin-17 production in central nervous system-infiltrating T cells and glial cells is associated with active disease in multiple sclerosis[J].Am J Pathol,2008;172(1):146-155.

9 KebirH,Kreymborg K,Ifergan I et al.HumanTH17 lymphocytes promote blood-brain barrier disruption and central nervous system inflammation[J].Nat Med,2007;13(10):1173-1175.

10 Haas J,Fritzsching B,Trübswetter P et al.Prevalence of newly generated naive regulatory T cells(Treg)is critical for Treg suppressive function and determines Treg dysfunction in multiple sclerosis[J].J Immunol,2007;179(2):1322-1330.

11 Venken K,Hellings N,Broekmans T et al.Natural naive CD4+CD25+CD127low regulatory T cell(Treg)development and function are disturbed in multiple sclerosis patients:recovery of memory Treg homeostasis during disease progression[J].J Immunol,2008;180(9):6411-6420.

12 KumarM,Putzki N,Limmroth V et al.CD4+CD25+FoxP3+T lymphocytes fail to suppressmyelin basic protein-induced proliferation in patients with multiple sclerosis[J].J Neuroimmunol,2006;180(1-2):178-184.

13 Haas J,Hug A,Vieh?ver A,Fritzsching B et al.Reduced suppressive effect of CD4+CD25high regulatory T cells on the T cell immune response against myelin oligodendrocyte glycoprotein in patients with multiple sclerosis[J].Eur J Immunol,2005;35(11):3343-3352.

14 Grimaldi L M,Maimone D,Reggio A et al.IgG1,3 and 4 oligoclonal bands in multiple sclerosis and other neurological diseases[J].Ital J Neurol Sci,1986;7(5):507-513.

15 Cross A H,Trotter J L,Lyons J.B cells and antibodies in CNS demyelinating disease[J].Neuroimmunol,2001;112(1-2):1-14.

16 Lindsey J W,Patel S,Zou J.Epstein-Barr virus genotypes in multiple sclerosis[J].Acta Neurol Scand,2008;117(2):141-144.

17 Wei L,Laurence A,Elias K M et al.IL-21 is produced by Th17 cells and drives IL-17 production in a STAT3-dependent manner[J].J Biol Chem,2007;282(48):34605-34610.

18 Bettelli E,Carrier Y,Gao W et al.Reciprocal developmental pathways for the generation of pathogenic effectorTH17 and regulatory T cells[J].Nature,2006;441(7090):235-238.

19 Korn T,Bettelli E,Gao W et al.IL-21 initiates an alternative pathway to induce proinflammatory T(H)17 cells[J].Nature,2007;448(7152):484-487.

20 Zhou L,Ivanov II,Spolski R et al.IL-6 programs T(H)-17 cell differentiation by promoting sequential engagement of the IL-21 and IL-23 pathways[J].Nat Immunol,2007;8(9):967-974.

21 Stockinger B,Veldhoen M.Differentiation and function of Th17 T cells[J].Curr Opin Immunol,2007;19(3):281-286.

22 Nurieva R,Yang X O,Martinez G et al.Essential autocrine regulation by IL-21 in the generation of inflammatory T cells[J].Nature,2007;448(7152):480-483.

23 Peluso I,Fantini M C,Fina D et al.IL-21 counteracts the regulatory T cell-mediated suppression of human CD4+T lymphocytes[J].J Immunol,2007;178(2):732-739.

24 Monteleone G,Pallone F,MacDonald T T et al.Interleukin-21:a critical regulator of the balance between effector and regulatory T-cell responses[J].Trends Immunol,2008;29(6):290-294.

25 Piao W H,Jee Y H,Liu R L et al.IL-21modulates CD4+CD25+regulatory T-Cell homeostasis in experimental autoimmune encephalomyelitis[J].Scand J Immunol,2008;67(1):37-46.

26 Danijela Konforte.IL-21:an executor of B cell fate[J].J Immunol,2009;182(4):1781-1787.

27 Ozaki K,Spolski R,Ettinger R et al.Regulation of B cell differentiation and plasma cell generation by IL-21,a novel inducer of Blimp-1 and Bcl-6[J].J Immunol,2004;173:5361-5371.

28 Bryant,V L,Macs,Averiy D T et al.Cytokine-mediated regulation of human B cell differentiation into Ig-secreting cells:predominant role of IL-21 produced by CXCR5_T follicular helper cells[J].J Immunol,2007;179:8180-8190.

29 Johnson K M,Shapiro-Shelef,Tunyaplin C et al.Regulatory events in early and late B-cell differentiation[J].Mol Immunol,2005;42:749-761.

30 Scheeren F A,Marianne N,Sean D et al.STAT5 regulates the self-renewal capacity and differentiation of human memory B cells and controls Bcl-6 expression[J].Nat Immunol,2005;6:303-313.

31 Kasaian M T,Whitters M J,Carter L L et al.IL-21 limits NK cell responses and promotes antigen-specific T cell activation:a mediator of the transition from innate to adaptive immunity[J].Immunity,2002;16:559-569.

32 Ozaki K,Spolski Rosanne,Feng C G et al.A critical role for IL-21 in regulating immunoglobulin production[J].Science,2002;298:1630-1634.

33 Shi F D,LjunggrenH G,Sarvetnick N.Innate immunity and autoimmunity:from self-protection to self-destruction[J].Trends Immunol,2001;22(2):97-101.

34 VollmerT L,Liu R,Price M et al.Differential effects of IL-21 during initiation and progression of autoimmunity against neuroantigen[J].J Immunol,2005;174(5):2696-2701.

35 Shi F D,Takeda K,Akira S et al.IL-18 directs autoreactive T cells and promotes autodestruction in the central nervous system via induction of IFN-gamma by NK cells[J].J Immunol,2000;165(6):3099-3104.

36 Arase H,Arase N,Saito T.Interferon gamma production by natural killer(NK)cells and NK1.1+T cells upon NKR-P1 cross-linking[J].J Exp Med,1996;183(5):2391-2396.

37 Romagnani S.Induction of TH1 and TH2 responses:a key role for the natural′immune response?[J].Immunol Today,1992;13(10):379-381.

38 Strengell M,Sareneva T,Foster D et al.IL-21 up-regulates the expression of genes associated with innate immunity and Th1 response[J].Immunol,2002;169:3600-3605.

39 Wurster A L,Rodgers V L,Satoskar A R et al.Interleukin 21 is a T helper(Th)cell 2 cytokine that specifically inhibits the differentiation of naive Th cells into interferon gamma-producing Th1 cells[J].J Exp Med,2002;196(7):969-977.

40 Ma C S,Tangye S.Helping the Helpers![J].Immunity,2009;31(1):12-14.