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锌在糖皮质激素诱导性青光眼中的作用机制与治疗途径

The role of Zinc in glucocorticoid-Induced glaucoma: mechanisms and therapeutic approaches

来源期刊: 眼科学报 | 2024年6月 第39卷 第6期 275-284 发布时间:2024-06-28 收稿时间:2024/8/28 16:35:40 阅读量:974
作者:
关键词:
糖皮质激素青光眼小梁网视神经保护
glucocorticoid glaucoma zinc trabecular meshwork optic nerve protection
DOI:
10.12419/24063002
收稿时间:
2024-05-03 
修订日期:
2024-05-22 
接收日期:
2024-06-03 
糖皮质激素(glucocorticoid, GC)由于其抗炎特性被广泛用于治疗眼部炎症,而G C诱导性青光眼(glucocorticoid-induced glaucoma, GIG) 作为一种常见并发症,其发病机制长期受到关注。文章综述了锌在GIG中的关键作用及其调控机制,揭示了锌在青光眼发病机制中的重要角色。锌作为人体中含量第二丰富的过渡金属,对蛋白质结构、酶催化和细胞信号调节至关重要。GC对锌分布的调控作用在不同组织和细胞类型中表现出复杂性,影响锌的摄取和释放,进而参与青光眼的病理过程。锌通过影响小梁网细胞外基质(extracellular matrix, ECM)的降解和重塑,以及视网膜神经节细胞的存活和轴突再生,在GIG的发病机制中发挥着复杂的作用。文章同时介绍了体内锌调控的现有途径,包括补充锌和减少锌的策略,提供了潜在的治疗途径。未来的研究应深入探索锌在青光眼中的作用机制以及与GC的相互作用,评估锌补充或螯合在青光眼治疗中的安全性和有效性,以及开发新型锌递送和螯合系统,有助于全面揭示锌在青光眼中的作用及治疗潜力,以实现更加精准的防治方案,改善患者预后。
Glucocorticoid (GC) is widely used in the treatment of ocular inflammation for its anti-inflammatory propery. However, glucocorticoid-induced glaucoma (GIG) is a common complication, and its pathogenesis has been extensively studied. This review summarizes the crucial role of zinc in GIG and its regulatory mechanisms, highlighting zinc's significant involvement in the pathogenesis of glaucoma. Zinc, the second most abundant transition metal in the human body, is essential for protein structure, enzyme catalysis, and cell signaling regulation. The effects of GC on zinc distribution vary across different tissues and cell types, affecting zinc uptake and release, which may contribute to the pathological processes of glaucoma. Zinc influences the degradation and remodeling of the trabecular meshwork extracellular matrix and the survival and axonal regeneration of retinal ganglion cells, playing complex roles in the pathogenesis of GIG. We discuss available strategies for regulating zinc in vivo, including zinc supplementation and reduction strategies, providing potential therapeutic approaches. Future research should explore the mechanisms of zinc's role in glaucoma and its interaction with glucocorticoids, evaluate the safety and efficacy of zinc supplementation or chelation in glaucoma treatment, and develop novel zinc delivery and chelation systems. These efforts will help fully elucidate the role of zinc in glaucoma and its therapeutic potential, enabling more precise prevention and treatment strategies to improve patient outcomes.

文章亮点

1. 关键发现

      研究发现,锌不仅影响小梁网细胞外基质(ECM) 的降解和重塑,还参与视网膜神经节细胞(RGCs) 的存活和轴突再生。这些发现强调了锌在青光眼病理过程中的关键角色。

2. 意义与改变

      通过深入理解锌在青光眼中的作用机制,研究者可以开发新的治疗方法,如锌补充或螯合剂的使用,以调节眼内锌水平,减轻青光眼的病理进程。此外,这些发现还可能促进个体化治疗的发展,通过识别对糖皮质激素更敏感的个体,实现更加精准的治疗。

1 概述

       在眼科治疗中,糖皮质激素(glucocorticoid, GC)因其强大的抗炎作用而被广泛应用,但随之而来的眼压升高却成为不容忽视的并发症。锌作为生命活动中不可或缺的微量元素,在青光眼的病理过程中扮演着重要角色。本文旨在探讨锌在GC诱导性青光眼(glucocorticoid-induced glaucoma, GIG)中的作用机制,并探索通过调节锌水平来治疗青光眼的可能性,以期为青光眼的防治提供新的视角和策略。

1.1 GC在眼科治疗中的应用与风险

      GC是由肾上腺产生的两种天然类固醇皮质激素之一(另一种为盐皮质激素),在应激时被释放,可调节炎症反应。因此,眼科医生通常将其用于局部治疗眼部炎症或水肿,如角结膜炎、葡萄膜炎、年龄相关性黄斑变性、糖尿病视网膜病变、视网膜静脉阻塞[1-2]。GC可穿过胞膜与细胞内受体结合,启动信号级联反应,影响数百个基因表达[3]。因此GC的使用可引起高度个体化的反应,也包括易感人群的不良反应,其中最为显著的为长期眼局部使用GC而引起的眼压(intraocular pressure, IOP)升高,这被称为GC诱导的高眼压(glucocorticoid-induced ocular hypertension, GIOH)[3-4]GIOH的发展取决于GC效力、剂量、给药方法、治疗时间和个体对GC的易感性等因素,地塞米松(dexamethasone, DEX)在提高IOP方面效果最为显[3-5]。个体对GC的易患性各不相同:约90%的青光眼患者是激素反应者,而在普通人群中这一比例为30%~40%[4-5]。在离体灌注培养的人眼球中,DEX治疗会在约30%的眼球中诱发高眼压[6]。如果不进行干预,长期的IOP升高可能会导致青光眼性视神经病变和永久性视力丧失[7],即GIG[3-4]

1.2 锌是人体重要金属并受到严格调控

      锌是人体中含量第二丰富的过渡金属,是蛋白质结构、酶催化和细胞信号调节的重要组成部分[8]。人体中约有10%的蛋白质被归类为锌结合蛋白[9]。锌金属酶中占比最多的为蛋白酶,细胞内游离锌离子(Zn2+)水平的波动调节这类蛋白质的生物活性[10]。细胞Zn2+水平通过各种机制波动,包括锌波、锌火花以及锌金属伴侣、锌转运蛋白(Zn transporters, ZnTs)和Zrt,Irt相关蛋白(Zrt/Irt-related proteins, ZIPs)的合成和亚细胞定[8, 11-12]。Zn2+通过ZnTs从细胞质分别转运到细胞器、小泡或细胞外,或通过ZIP转运到细胞质[8, 13]。胞浆中几乎所有的Zn2+都与金属硫蛋白(metallothioneins, MTs)结合,所以胞浆中游离Zn2+水平非常低[14]。然而,与其他锌结合蛋白相比,MT不具有最高的 Zn2+亲和力,使得Zn2+容易与其他具有更高亲和力的结合位点交[14-15]。最小锌配额是指让细胞处于最佳生长状态所需的最小锌量,每个哺乳动物细胞大约需要108 个锌原子,这可能取决于蛋白质上锌结合位点的数量。当锌缺乏时,细胞通过增加摄取、减少外排和从细胞内储存位点调动锌等机制,或降低特异性锌结合蛋白的水平,以减少细胞对锌的需求,将细胞锌水平维持于最低锌配额[16]

2 GC诱导性青光眼中的锌调控异常

2.1 青光眼致病机制中的锌失调

     青光眼发病机制复杂,其中锌水平的变化已有文献报道。研究显示5月龄青光眼前期DBA/2J小鼠视网膜中的锌水平高于10月龄青光眼期的DBA/2J小[17]。前期研究揭示,Zn2+水平在视神经损伤后1 h内在无长突细胞(amacrine cells, ACs)中迅速升高,这可能是继发于视网膜中一氧化氮的迅速而持续的升高,随后Zn2+水平经锌转运蛋白ZnT3进入ACs突触囊泡并释放至突触间隙,跨突触转运至神经节细胞(retinal ganglion cells, RGCs)引起其过量锌积累,介导RGCs凋亡并抑制轴突再生[18-20]。而在眼前段中,前期研究发现原发性青光眼患者及GIOH小鼠的房水中锌水平较对照组显著升高,且随眼压升高,房水中锌水平呈上升趋势[21]。而在房水流出通道中,小梁网细胞(trabecular meshwork cells, TMCs)在GC影响下锌转运蛋白表达发生改变,降低了胞内Zn2+水平[22]。这些结果均提示在GIG中存在锌调控相关发病机制。

2.2 GC对锌分布的调控作用

      GC对锌分布的影响通过调控MTs、ZnTs与ZIPs实现,并已在多个系统中得到证实。GC可使TMCs、视网膜色素上皮细胞、成骨细胞、皮肤或肝脏等组织的MT蛋白表达增加[22-26]。应激状态下或经GC处理后,海马神经元表现出Zn2+传输增加,即在突触前释放和摄取以及突触后摄取均增加[27-28],这其中涉及多种锌转运蛋白如ZnT1、ZnT3、ZnT10、ZIP4与ZIP6,可能导致海马功能障碍,影响学习和记忆[29-30]。甲泼尼龙是一种合成皮质类固醇,可增加小胶质细胞中ZIP8表达并升高胞内Zn2+,继而引起小胶质细胞活化降低及自噬性死亡[31]。GC可引起山羊结肠、大鼠肝脏及小鼠乳腺上皮ZIP14表达升高,促进其Zn2+摄取[32-34]。DEX可诱导骨髓间充质干细胞中ZnT7的水平显著降低并促进其向成骨细胞分化[35]。然而,在胰腺腺泡细胞中,DEX可诱导其顶膜上的ZnT1和酶原颗粒上的ZnT2的上调继而引起其胞质Zn2+减少[36]。在TMCs中,GC下调了ZIP8与ZIP10表达水平从而引起胞内Zn2+降低。这些发现表明Zn2+在调节GC诱导的生理和病理变化中起着复杂的作用。

3 锌调控对青光眼病理进程的影响

3.1 锌可通过影响基质金属蛋白酶活性调节ECM

      小梁网(trabecular meshwork, TM)中ECM 成分增加可导致房水流出阻力增加,引起眼压升高[37]。而GC处理会增加TM中ECM沉积,使TM细胞层间呈现出指纹状基底膜样、细纤维样物质沉积,且Schlemm's 管内壁内皮下基底膜更为连续及致密[38-39]
    
       基质金属蛋白酶(matrix metalloproteinases, MMPs)是一类能够靶向ECM内多种蛋白质并降解ECM和基底膜的酶[40-41]。近管区TMCs可通过机械形变感知眼压升高,通过激活和释放多种MMPs以局灶性降解ECM,从而降低房水流出阻力,允许更多的房水流出[42-44]。已有诸多研究表明,GC治疗可以抑制TM中MMPs的表达[3, 45]。已开发多种药物及蛋白质或GC诱导病毒载体用于上调MMPs的表达以抵抗TM纤维化,促进房水外排及降低眼压[46-51]。但GC对TM中MMPs的具体调控机制仍无定论。

      MMPs的活性受多种因素调节,包括转录、翻译、分泌、细胞区域定位、酶原活化、底物可及性、降解及抑制物的影响[52]。在MMPs降解底物过程中,其催化活性中心中的Zn2+与底物氧原子及MMPs自身谷氨酸氧原子结合,随后谷氨酸氧原子发起亲核攻击,最终导致底物分解并释放水[40, 53]。Zn2+在这一过程中起着不可或缺的作用。在Zn2+缺乏的状态下,MMPs可能发生错误金属化或保持未金属化,导致活性丧失或合成减少[11, 54]
图1 基质金属蛋白酶MMP-2三维重建示Zn2+结合位点与催化结构域
Figure 1 Three dimensional reconstruction of MMP-2 reveals Zn2+ binding sites and catalytic domains

      在炎症介质的诱导下,软骨、脊柱髓核等组织细胞内Zn2+水平的增加,可能通过影响MMPs的表达来介导ECM的降解[55-57]。锌转运蛋白的异常表达也可导致ECM紊乱,正常发育的小鼠胚胎心脏内皮细胞表达ZIP8,该基因缺失小鼠心脏表现出显著的ECM沉积及数种金属蛋白酶的表达减少[58]。在特发性肺纤维化患者及衰老小鼠的2型肺上皮细胞中同样发现ZIP8的缺乏,ZIP8的缺乏导致细胞自我更新受损及肺纤维化增强[59]。这些发现表明,细胞内Zn2+在ECM降解和重塑中发挥作用,维持细胞存活和功能[60-62]。在TM中,GC影响下的TMCs胞内Zn2+减少,导致MMP-2表达降低,可能促进ECM沉积,补充锌后小鼠TM内蓝染的胶原蛋白含量降低,表明锌稳态异常可能是GIOH发生和发展机制之一[22]

3.2 锌调控在视神经保护中的作用

      视神经损伤后RGCs发生氧化应激,影响其存活。已开发活性材料用于增强清除视神经损伤后的活性氧(reactive oxygen species, ROS),以挽救RGCs[63-64]。而锌和氧化还原信号在多个方面上错综复杂地相互作用,Zn2+超载诱导ROS产生和清除失调,引起氧化应激[65]。研究表明,在视神经损伤后螯合Zn2+,可激活RGCs中的核转录因子红系2相关因子2(nuclear factor erythroid 2-related factor 2, Nrf2)及其下游抗氧化反应蛋白表达,抑制ROS产生,降低自噬水平,从而保护RGCs并促进轴突再生[66]。又由于在视网膜中,RGCs的树突和轴突分别与视网膜和大脑中的神经元连接,神经突触需要大量的能量来合成神经递质、组装突触囊泡、形成离子梯度和缓冲钙流。线粒体在满足RGCs突触结构高耗能性方面起至关重要的作用[67- 68]。而线粒体参与广泛的细胞功能,包括ROS的产生与清除[69]。高水平ROS可诱导线粒体依赖性细胞凋亡[70]。锌是否通过ROS调节线粒体继而影响RGCs存活尚需要进一步研究。

      在线粒体内部,研究表明线粒体中的Zn2+超载可导致神经元线粒体去极化、过度分裂和通透性增加等异常[71-73],并激活线粒体介导的促凋亡因子,最终导致神经元死亡[74]。m-AAA蛋白酶1同源物(overlapping with the m-AAA protease 1 homolog, OMA1)是一种线粒体内膜蛋白酶,具有Zn2+结合位点,且该位点与酶活性域空间关系紧密[75]。在视神经损伤后,RGCs内增多的Zn2+可过度激活OMA1的表达与活性,继而触发线粒体应激,螯合Zn2+可有效抑制OMA1活性,同时保护RGCs线粒体结构,促进RGCs存活[19]

图2 与m-AAA蛋白酶1同源物OMA1三维重建示Zn2+结合位点与催化结构域
Figure 2 Three dimensional reconstruction of OMA1 reveals Zn2+ binding sites and catalytic domains

4 体内锌调控的策略

4.1 补充锌策略

      目前体内锌的补充主要依靠于饮食主动吸收的方式[76-77]。补锌对血浆及多种器官内锌水平的调节已有诸多报道[78-79],而关于改变具有多重屏障的脑部及眼部锌水平的相关报道较少。研究表明,在小鼠饮用水中添加锌可升高其大脑海马区域Zn2+水平[80]。对头部损伤患者补锌有助于其神经功能的恢复并降低病死率[81]。表达ZIP8蛋白的基因突变可导致血脑屏障完整性降低,加剧免疫及炎症反应[82-83]。眼内锌主要集中于视网膜、视网膜色素上皮及脉络膜中,补锌可延缓退行性视网膜疾病的进展[84-85]。最新研究表明,肠道微生物可通过破坏的肠道屏障及血视网膜屏障移行至视网膜,这也为饮食对眼内的影响提供了间接证据[86]。膳食补锌可升高房水内锌水平,表明膳食补锌可影响眼内锌稳态[22]

      除膳食补锌外,在动物实验中可采用腹腔注射进行补充。GC可导致小鼠抑郁表型,如活动性降低,但腹腔注射葡萄糖酸锌可以逆转这些改变[87-88]。腹腔注射硫酸锌可通过增加RGCs和外侧膝状体中热休克蛋白72表达,保护慢性高眼压大鼠模型中的RGCs和外侧膝状体免受损伤[89]
    
      体内锌水平还可通过药物或调节锌转运蛋白水平来升高。在卒中大鼠模型中,使用吲哚美辛(一种非选择性环氧酶抑制剂)腹腔注射进行慢性预处理,可诱导神经元锌及突触间隙锌水平的适度升高,提高机体对缺血性损伤的耐受性[90]。上调低锌饮食小鼠小肠中ZIP4的表达,可促进其肠道锌摄取[91]。使用小干扰RNA敲低骨髓间充质干细胞中ZnT7表达水平可提高细胞内Zn2+水平,促进其向成骨细胞分化及细胞外基质矿化[35]

      锌还可与其他化合物结合,以提高整体吸收率与疗效。肌肽是一种内源性二肽,大量存在于骨骼肌和大脑中,具有抗氧化、金属螯合、抗交联和抗糖基化等多种有益作用,肌肽和锌的复合物,广泛用于锌补充和溃疡的治疗,对神经退行性疾病具有保护作用[92]。姜黄素是一种从姜黄根茎中分离的多酚类化合物,具有抗氧化、抗炎、抗微生物、增强免疫等特性。然而,由于姜黄素不溶于水、不稳定、生物利用度差,其疗效受到限制。姜黄素作为配体与锌形成的稳定配合物具有更强的抗氧化特性,还显示出肝保护、胃保护、神经保护、心脏保护和成骨等功效[93]

4.2 减少锌策略

      诸多研究表明,锌在神经元中积累可导致缺血性神经元损伤,已开发诸多减少锌积累的策略。在局灶性缺血性脑梗死中,静脉注射EDTA-Ca(ethylene diamine tetraacetic acid dicalcium)已被用于螯合体内大脑的Zn2+以发挥保护作用[94]。其他常用Zn2+螯合剂还包括:ZX1,EGTA(ethylene glycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic acid),BAPTA[1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid],EDPA(ethylenediamine-N,N'-diacetic-N,N'-di-β-propionic),DPESA(4-{[2-(bis-pyridin-2-ylmethylamino)ethylamino]methyl}phenyl)methanesulfonic acid,TPESA(4-({[2-(bis-pyridin-2-ylmethylamino)ethyl] pyridin-2-ylmethylamino}-methyl)phenyl]methanesulfonic acid)等,但这些均不具备细胞膜通透性[95]。Zhao等[96]通过腹腔内注射具有高亲和力及细胞膜渗透性的Zn2+螯合剂TPEN[N, N, N0, N0-tetra(2-pyridylmethyl) ethylenediamine]以减少脑神经元缺血性凋亡。TPEN眼内注射可减少视神经损伤后RGCs死亡,并促进其轴突再生[18]。Huang等[97]开发了一种多功能仿生纳米酶材料Ce/Zeo NMs,通过尾静脉注射后吸附过量Zn2+并催化清除ROS以减轻大鼠缺血再灌注后脑梗死面积,并保护血脑屏障,减少神经血管的缺血性损伤。还有研究者使用常压高氧环境以减少缺血性脑损伤组织的锌积累而发挥保护作用[98]

表1 各锌离子螯合剂的金属解离常数 (Kd) 及优缺点[95]
Table 1 Metal dissociation constants (Kd) and advantages and disadvantages of various zinc ion chelating agents


项目
TPEN
EDTA
EDPA
CaEDTA
ZX1
DPESA
TPESA
Kd Zn
0.7 fMa
0.4 fMa
0.8 pMa
2.0 nMb
1.0 nMb
1.6 pMc
0.5 pMc
Kd Ca
68 μMa
19 nMa
0.9 μMa
未检出
63 μMc
3.3 mMc
优点
Zn2+的选择性
高亲和力
快速动力学;高金属
亲和力
高金属亲和力
Zn2+
选择
Zn2+的选择性;  快速动力学
Zn2+的选择性;
高金
属亲和力
Zn2+的选择性;
高金属亲和力
缺点
去除蛋白金属
非选择性;
影响
钙水平
非选择性;
慢动
力学
慢动力学
尚需进一步探究
尚需进一步探究;  慢动力学
尚需进一步探究

    通过改变锌转运蛋白的水平,如敲低或敲除ZIP8,可以调节Zn2+内流及MMPs活性与表达水平以减轻软骨、脊柱髓核等组织由于炎症刺激和过度机械应力诱导的ECM降解,保护骨关节组织[56, 99]。ZnT3负责神经突触间的Zn2+转运[100]。靶向ACs敲除ZnT3可减少其在视神经损伤后向RGCs转运Zn2+,以保护RGCs[101]

5 展望

      锌在GIG中的作用不容忽视,其调控异常可能是青光眼发病的关键因素之一,这一认识有望协助我们开发新的治疗及干预措施。首先,进一步的分子生物学和细胞生物学研究将有助于揭示锌和锌转运蛋白在青光眼中的具体作用机制,以及它们如何响应GC的刺激。这些研究将为开发新的药物和治疗方法提供理论基础。
      其次,临床研究应关注于评估锌补充剂或锌螯合剂在青光眼治疗中的安全性和有效性。通过精心设计的临床试验,我们可以确定最佳的锌补充及锌螯合剂量和治疗方案,以及它们对青光眼患者眼压控制和视神经保护的长期影响。此外,结合遗传学和表观遗传学研究,我们可能能够识别出对GC更敏感的个体,从而实现个体化治疗。
      最后,跨学科合作将是推动治疗进步的重要途径。结合材料科学、纳米技术和生物工程的进展,开发新型锌递送及锌螯合系统,精确靶向特定细胞,提高治疗效果并减少不良反应。总之,随着对锌在青光眼中作用机制的深入研究和治疗策略的不断创新,可在未来为青光眼患者提供更加有效和个性化的治疗方案。

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1、国家自然科学基金(81870657);广东省自然科学基金(2022A1515012168);广东省基础 与应用基础研究基金(2024A1515013296);广州市科技计划项目(202201020492);眼科国家重点实验室开放研究基 金(2023KF01)。
This work was supported by the National Natural Science Foundation of China (81870657), the Natural Science Foundation of Guangdong Province of China (2022A1515012168), Guangdong Basic and Applied Basic Research Foundation (2024A1515013296), Science and Technology Program of Guangzhou of China (202201020492), and the Open Research Funds of the State Key Laboratory of Ophthalmology (2023KF01).()
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