Development in physiological regulation and bone metabolism of magnesium

doi:10.3969/j.issn.2095-4344.2014.03.017

Abstract

Abstract:

BACKGROUND: As a lightweight metal with mechanical properties similar to natural bone, a natural ionic presence with significant functional roles in biological systems, and in vivo degradation via corrosion in the electrolytic environment of the body, magnesium (Mg)-based implants have the potential to serve as biocompatible, osteoconductive, degradable implants for load-bearing applications. On the other hand, Mg, as a critical mediator of cellular growth and activities regulation, is the second most abundant intracellular cation where it plays an important role in enzyme function and trans-membrane ion transport.
OBJECTIVE: To retrospectively summarize the application progress, physiological role and mechanism of Mg, and to review its regulation of bone metabolism.
METHODS: Papers addressing application, physiological and bone metabolism regulation of Mg published in core periodicals were reviewed. Studies in which Mg were used as an assistance materials, or articles regarding cell physiology in which Mg were not the main research object were excluded. Literatures concerning the regulation of bone turnover by Mg were given special attention.
RESULTS AND CONCLUSION: Mg alloys are cytocompatible materials with adjustable mechanical and corrosion properties and show their potential as biodegradable implant materials. Moreover, Mg is abundantly distributed among the body and essential for maintaining physiological function of the cells. Mg deficiency has been associated with a number of clinical disorders including osteoporosis. But it remains to be determined what specific biochemical process is activated by Mg ²⁺ to regulate cellular activity and bone turnover. Further investigations of the precise mechanism are valuable to the clinical application of Mg.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

全文链接：

Key words: biocompatible materials, magnesium, corrosion, review

CLC Number:

R318

Ma Wen-hui, Zhang Ying-ze. Development in physiological regulation and bone metabolism of magnesium[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(3): 432-439.

Figures/Tables 1

2.1 纳入文献的基本情况对采纳的110篇文献归纳总结，有关镁金属的研究与应用的文献37篇，有关镁调节细胞生理活性的文献32篇，有关镁对骨代谢影响的文献41篇。 2.2 镁金属的临床应用进展最早将镁应用于临床的报道可追溯到1907年，Lambotte[9]用纯镁制成的钢板来治疗1例胫骨骨折患者，但由于纯镁降解过快，术后8 d便降解并产生了大量的皮下气体，手术未成功。在此后的几十年中，陆续有不少学者针对镁金属的应用进行了研究，虽然取得了较满意的结果，但是，镁金属在生理环境下的快速腐蚀一直是限制其临床应用的主要障碍，如不能很好地控制镁合金材料的降解速度，则其力学稳定性将不足以支撑到骨折愈合[10-15]。近十几年来，随着纳米技术和金属制造工艺的发展，在镁腐蚀速率的调整和材料性能的优化方面取得了明显进步。许多新型的具有良好耐腐蚀性和力学特性的镁合金被开发出来，例如镁-锂合金、镁-铝合金和镁-钇合金等。Gu等[16]对9种不同元素与镁形成的合金进行分析，发现合金材料的力学性质和耐腐蚀性及腐蚀产物的细胞毒性均与所加入的元素有关。Witte等[17-18]研究了4种不同镁合金植入豚鼠体内后对局部骨生理的影响，结果表明金属移植物的降解主要与其合金元素的成分有关，在金属降解表面形成活性磷酸钙的聚集，进而导致移植物周围矿物沉积率增高和成骨增加，同时，矿物的沉积也可有效减缓金属的腐蚀降解。Kannan等[19]报道镁合金经过表面钙喷涂后可明显降低其降解速率，而对金属的力学性质无明显影响。Li等[20]的研究表明，Mg-1Ca合金的力学性质和耐腐蚀性良好，是一种新型的可降解生物移植材料。Zhang等[21]报道，镁-锌合金在生理条件下的腐蚀速率明显减低，体内试验也显示此种材料具有良好的生物相容性，对重要器官无毒性作用，并且在材料周围未发现氢气的聚集。Wong等[22]在镁金属表面涂布一层可生物降解的高分子膜，借此降低金属的腐蚀速率，取得了较好的实验结果。Xin等[23]发现，在镁合金表面进行氢化非晶硅涂层可明显降低材料的腐蚀速率，增加材料的强度，是一种优化材料性质的有益选择。Xu等[24]报道，对镁-锰-锌合金进行磷酸化处理后可明显改善材料的抗腐蚀性。Zberg等[25]的研究发现，镁-锌-钙合金可明显减少氢的形成，并且具有良好的组织相容性。但有研究发现，一些稀土金属元素可能会引起细胞毒性，尤其是长期作用后会产生意想不到的全身反应，例如阿尔茨海默病可能与铝有关[26]。另外，人们在多功能降解材料的研究方面也进行一些有益的探索[27-28]。Tie等[29]报道了一种多功能的可降解镁合金材料-镁-银合金，这种材料不仅改善了金属的腐蚀和力学性能，而且通过利用银在化学状态下的抗菌活性使合金具有了良好的抗感染能力。Paton[30]和Necula等[31]的研究也表明银涂层或含有银的镁合金材料具有较好的性能。Bosetti[32]和Hardes等[33]报道，含有银的合金并没有细胞毒性，相反，加入少量的银，会改善金属的细胞相容性。尽管银在人体内的聚集和代谢机制还不甚清楚，但是Drake等[34]证实含银金属在体内释放的阴离子较低，对健康造成危害的风险很小，因此镁-银合金相较于含有其他重金属的镁合金来说更安全，可以用作生物移植材料。对金属移植物在体内腐蚀过程的认识是现代生物材料科学的重要研究内容，但是体内环境的复杂性和多变性是此类研究所面临的巨大挑战，目前大多数实验均是在体外模拟生理条件下分析材料的腐蚀性质和机制，这也被认为是了解材料生理腐蚀行为的基础[35-36]。Witte等[17]将镁合金在动物体内的腐蚀过程和体外标准环境下的腐蚀过程进行了比较，结果发现，金属在两种条件下的腐蚀速率趋势有所不同，在体内的腐蚀速率明显较体外的低。Jang等[37]分析了3种主要生理体液成分及其浓度对镁合金腐蚀行为的影响，结果表明模拟体液成分的差异会导致镁合金腐蚀行为和腐蚀产物的不同。当溶液中有HPO42- 和Cl-存在时，磷酸盐会诱导形成致密的非结晶磷酸镁腐蚀层；而当HPO42- 、Cl-和Ca2+共存时，则会在镁合金表面形成磷酸钙和羟基磷灰石沉淀，从而可限制金属局部的腐蚀，增加材料的耐腐蚀性；另外，加入HCO32-可加速材料的腐蚀速率，并随着碳酸氢盐浓度的增加而加快；当HPO42- 、HCO32-和Ca2+同时存在时，会由于不溶性羟基磷灰石的形成而降低金属的腐蚀速率。 2.3 镁的生理作用及机制镁是一种对人体无害的微量元素，其中有50%-60%存在于骨组织中，细胞外间质中的含量不足1%，其余的均位于细胞内，是细胞内含量第二的阳离子。而在细胞内的镁中，有95%与带负电荷的分子相连，如核糖体、质膜和ATP等。有超过300种酶的功能受镁的调控，包括与ATP螯合成MgATP、为酶提供催化作用、以及为酶提供特殊的结构等[38]。镁还可参与离子通道的调节，例如它能在钾和钙进出细胞的跨膜转运中发挥重要作用。因此，不难想象，镁缺乏可能会导致多种疾病，如神经肌肉系统疾病、心血管系统疾病和骨矿物代谢疾病[39]。成年男性镁的推荐日摄入量是420 mg/d，而实际的平均摄入量是323 mg/d，成年女性镁的推荐日摄入量是320 mg/d，但实际的平均摄入量只有228 mg/d，因此日常生活中镁摄入量不足的情况是普遍存在的[40]，从青春期的少年到老年人都存在镁摄入不足。据估计，10%的老年女性镁的摄入量低于 136 mg/d。老年人胃肠道和肾脏的保镁功能较年轻人明显减退可能也是引起其低镁的重要原因之一[41-42]。另外，与镁丢失有关的疾病(如糖尿病、酒精中毒、吸收不良)和导致肾脏失镁的药物都会加重人体镁的缺乏[39]。随着20世纪中期单层细胞培养技术的出现，对于细胞增殖和生理的研究取得了巨大进展。这种细胞培养技术使人们能更好地观察和计数细胞，更容易研究不同成分对细胞行为的影响。生长因子和非特异性刺激因子刺激细胞的增殖和活性是通过一系列的协同反应来实现的，而镁在协同反应许多关键步骤中起着非常重要的作用，例如磷酸化反应中磷酸果糖激酶和丙酮酸激酶的活化需要MgATP的参与。另外，镁对于启动蛋白合成及接下来的DNA合成和细胞分裂都是至关重要的[43-44]。Rubin 等[45-46]观察到焦磷酸可与镁结合，随着焦磷酸浓度的增加，游离镁的量明显减少，进而细胞蛋白、RNA和DNA的合成会减少。另外，他还发现，钙减少可抑制大鼠成纤维细胞的DNA合成，但是加入镁后会纠正这种状况，相反，由于镁缺乏引起的细胞DAN合成抑制并不能被钙所纠正。因此，作者认为镁是细胞增殖中的必需元素，并且它对细胞蛋白和DAN合成的影响存在一个最佳的浓度。Schreier等[47]的研究也表明，镁对细胞蛋白和DNA合成的影响呈钟形曲线，类似于纯化核糖体无细胞合成蛋白的生成曲线。Terasaki等[48]发现，蛋白合成速率对于镁浓度的变化非常敏感，细胞中游离镁浓度的微小改变即会导致蛋白合成速率的变化，并且镁引起的蛋白合成变化要较DNA合成改变提前数小时。当细胞内镁浓度轻度增加时，细胞会通过增加蛋白合成以及随后的DNA合成来主动地将镁浓度降低到正常水平，这样既刺激了细胞增殖，又调整了镁的平衡，突出显示了镁在细胞生长这一经济运行过程中的重要性。如果镁浓度过高，则细胞就不能通过蛋白和DNA的合成来降低胞内镁浓度，进而会引起细胞死亡。值得注意的是，镁对DNA的合成并没有直接影响，因为在不同细胞，DAN合成增加前存在一个5-12 h的延迟期。在生长因子刺激细胞的早期主要出现尿苷的磷酸化反应，而这一反应是细胞尿苷摄入速率的主要限制因素。镁可以增加细胞对尿苷的摄取，增强其调节细胞生长的作用，并且能反馈抑制尿苷激酶[49]。研究显示，镁是通过蛋白激酶的级联反应来调节细胞蛋白和DNA合成的。促细胞生长因子与细胞膜受体结合，激活膜上胞质一侧的酪氨酸激酶，活化PI3-K激酶，进而激活mTOR激酶，后者可引起参与转录起始的两种蛋白的磷酸化反应，增加核糖体蛋白和延伸因子的合成，从而刺激蛋白合成和随后的DNA合成。上述反应中的大部分蛋白激酶的活化都需要镁的参与。镁的升高可增加MgATP，激活mTOR，因此促进转录的启始，加速细胞通过G1期，开始DNA的合成和有丝分裂[50]。Lau 等[51]将镁注射入非洲爪蟾蜍的卵母细胞来研究其对蛋白合成的影响，发现注入低浓度的镁即可刺激蛋白合成，并且随着镁浓度的增加，刺激作用增强，其整个作用过程表现为钟样曲线，这一结果反映了镁在激活细胞蛋白合成中的重要作用。细胞内游离的镁仅占总镁含量的很少部分，细胞总镁的增高会导致游离镁的增加，并且后者增加的幅度与前者不成比例。生长因子的刺激可使细胞的总镁含量增加10%-20%，并且在细胞周期的G1和S期会一直保持这种增高的趋势[52]。游离镁的量可利用细胞内镁敏感受体Mag-fura-2来测定。Grubbs[53]发现，肌细胞经表皮生长因子刺激后，游离镁可在20 min内由最初的 0.32 mmol/L升高到1.4 mmol/L。Ishijima 等[54]的研究也显示，利用胰岛素刺进3T3细胞，会使镁在30-60 min内由起始的0.22 mmol/L增高的0.35 mmol/L。尽管由于Mag-fura-2从细胞内的渗出，会导致超过1 h后所测的游离镁浓度不可靠，但在整个细胞周期中游离镁会始终保持增高的水平。游离镁的增加可能主要是由连于细胞膜内表面的镁释放而来的，而总镁的增高可能是由于细胞膜上钠/钾ATP酶泵活性增加，加强了对镁的转运。镁是许多生物反应的重要辅助因子，钠和镁的反向转运对于维持体内镁的动态平衡有重要作用。另外，最近研究表明，细胞内镁离子的水平还受细胞外镁内流量的调节。促有丝分裂因素的刺激可激活细胞外的镁内流，使细胞内的镁含量增加到启动蛋白合成所需的最佳水平。因此，细胞外钙离子和镁离子的内流可促使细胞内的离子平衡，这可能有助于细胞增殖[55-59]。TRPM是近期发现的瞬时受体电位家族亚群的成员，在哺乳动物细胞，它是一类非电压依赖的钙离子通道，有8个不同的表型，分别被命名为TRPM1-8[60-63]。而TRPM6和TRPM7是参与镁调节的两种主要跨膜蛋白，它们兼具激酶和离子通道两种活性，可在细胞内镁和ATP水平的调节下激活阳离子(钙、镁和其他微量元素)的流入，其中TRPM6蛋白负责整个机体镁平衡的调节，而TRPM7则可能主要调节细胞内的镁水平。再者，TRPM7与细胞增殖和生存有关，增加细胞外镁含量可改善因TRPM7缺乏而造成的细胞生长障碍[64-67]。此外，TRMP6基因突变的患者会由于镁吸收障碍而表现出遗传性的低镁症[68]。概括来说，对于生长因子刺激细胞增殖的机制尚无统一认识，可能的机制是增加细胞摄入更多的营养物质；激活细胞内钾、钠、镁、钙这4种主要阳离子的功能；或是改变细胞内的pH值[69]。而镁在细胞增殖和各种生理反应的调控中发挥着重要作用，但到底镁通过哪些特定的生理化过程来加速蛋白合成的启动和激活酶的作用，目前还不能确定，其详细的机制还有待于进一步的研究。 2.4 镁对骨生理的调节镁占骨骼矿物质含量的0.5%-1%，对于骨骼来说，并不属于微量元素。它既可通过参与骨代谢调节的激素和因子来影响矿物质和基质的代谢，也可直接作用于骨本身。不同物种的骨骼中，镁含量差异较大，每公斤骨粉中的含镁量为150- 440 mmol。人类骨镁含量大概为每公斤骨粉中含镁 200 mmol左右，比大鼠的低30%-40%。人类髂嵴皮质骨单位质量的镁含量较松质骨低10%-20%[70]。目前还不清楚骨镁含量是否存在地区差异，但饮食中镁的摄入量直接影响骨骼中镁的含量，并且年龄与骨镁含量呈负相关。镁缺乏会对骨代谢产生不利影响。研究显示，低镁会导致成骨细胞数量减少和细胞活性降低，进而引起骨生长停止。另外，低镁还会导致成骨减少、骨质疏松，增加骨脆性。镁缺乏导致骨骺生长板变薄，同时由于低镁引起骨盐沉积结晶增加，从而使生长板中骨盐的百分比增加[71]。但是，在鸡和大鼠，并未发现镁缺乏会引起骨代谢的异化作用，相反，却会抑制骨吸收，增加皮质骨厚度，有时还会诱发骨膜下增生或骨膜纤维瘤，这种严重低镁引起的不寻常反应可能是继发于甲状旁腺素和维生素D代谢因子分泌减少和/或骨反应降低的一种间接作用结果。高镁也会对骨生理造成不利影响。Leidi等[72]的研究显示，高镁会明显抑制成骨细胞矿物基质的沉积，并降低成骨细胞分化标志物-碱性磷酸酶的活性，他们认为这可能是由于高浓度的镁通过对转运蛋白的竞争抑制改变了细胞内包括钙离子在内的多种阳离子的浓度，进而产生对细胞代谢的影响。骨盐沉积结晶障碍、成骨细胞活性降低、生长板增宽、骨骼变得短粗，导致骨软化样表现。也有报道显示，镁过量会刺激小鼠的骨吸收，而与甲状旁腺素无关[70]。Yun等[73]发现，高镁会抑制成骨细胞的增殖和贴壁，并且随着镁浓度的增高，细胞的矿化水平明显降低。关于镁缺乏和过量对人类骨生理的影响还需要更深入的研究。流行病学研究表明镁摄入量缺乏与骨质疏松有关。关于镁缺乏引起骨量减少的机制存在不同的解释。镁可促进骨细胞的分裂生长，因此它的缺乏可能引起骨形成减少。镁还能够影响晶体的形成，低镁可导致更大、更完整的晶体形成，从而影响骨的强度。另外，镁影响甲状旁腺素分泌的模式通常与钙的作用方式类似，它可与甲状旁腺细胞上钙敏感的受体相结合，引起细胞内钙的增加，进而减少甲状旁腺素的分泌[74]。Fatemi等[75]的研究发现，血清中甲状旁腺素的水平随着红细胞内游离镁水平的降低而减低。除此之外，低镁还可扰乱磷酸肌醇系统和/或降低腺苷酸环化酶活性，导致骨和肾脏的抗PTH作用，而通过镁的增加，可使机体的甲状旁腺素反应恢复正常[76-78]。以上这些资料表明，镁缺乏不仅破坏甲状旁腺素的分泌，还损害PTH对肾脏和骨的作用。文献[75，79]报道，低镁患者的血清1,25(OH)2-vit D水平也会降低，可能的原因是PTH作为1,25(OH)2-vit D合成的主要调节因子，它的分泌障碍导致其刺激 1，25(OH)2-vit D合成的作用丧失。Risco等[80]的研究还发现，肾脏1α-羟化酶是一种镁依赖性的酶，因此，低镁也可直接破坏1,25(OH)2-vit D的代谢。此外，低镁引起的血清1,25(OH)2-vit D水平降低，可减少钙的肠吸收，引起钙缺乏，增加发生骨质疏松的风险。Saggese等[81]报道，对于存在低镁的糖尿患者，低钙并不能引起血清1，25(OH)2-vit D水平的增高，补充镁后可改善这种状况。Welsh等[82]利用大鼠模型所做的实验也发现，在镁缺乏的情况下，血清1，25(OH)2-vit D水平对低钙并没用反应。因此，目前已经清楚，镁缺乏可破坏体内钙的平衡，引起血清PTH和1，25(OH)2-vit D水平的下降，补充镁可纠正这些生化方面的异常，由于甲状旁腺素和1，25(OH)2-vit D能刺激成骨细胞活性以及骨钙素与胶原的合成，因而，纠正低镁可增加骨的生成。正常情况下骨的重建处于动态平衡状态，破骨细胞分解旧骨，同时成骨细胞合成新骨进行替代。这种吸收和再建的平衡保证了总骨量的稳定，维持了骨的完整和强度。成年人每年有大约25%的松质骨被吸收和重建，而皮质骨只有3%被重建[83]。Gruber等[84]的研究显示，由于松质骨的表面积和和骨转化率均较皮质骨高，因此它对低镁的敏感性更强。成骨细胞不仅可分泌骨基质成分保证骨生成和组织矿化，并且还可产生破骨细胞分化所必需的蛋白，因此，它的增殖、迁移、分化和分泌功能的协调对于维持骨生理的动态平衡是非常重要的。任何损害成骨细胞功能和活性的因素都会破坏这种平衡，引起骨量减少和骨脆性增加，更易于发生骨折[85]。Park等[86-87]的研究显示，镁离子可促进成骨细胞附壁，增加碱性磷酸酶和骨钙素的合成，另外，还使成骨细胞转化因子以及主要的成骨细胞基因表达增加。Jeon等[88]发现，免疫抑制剂他克莫司可抑制ERK1/2通道导致细胞内镁的减少，进而影响骨的矿化，导致骨量减少。该作者还发现牛磺酸通过激活ERK1/2通道引起细胞内镁离子增高，从而促进成骨细胞的增殖和活性[89]。研究表明骨的生长修复及再建都受神经系统影响，神经系统可能是通过不同的神经肽类递质调节骨组织的代谢活动[90]。镁可抑制N-甲基-D-天冬氨酸受体(NMDA)，而NMDA受体的活化可诱导神经递质的释放，例如P物质[91]。细胞外镁的减少将降低受体激活所必需氨基酸的阈值水平，引发神经源性反应，导致T淋巴细胞释放炎性细胞因子(诸如肿瘤坏死因子α、白细胞介素1β、白细胞介素6)[92-97]。P物质是神经肽类物质之一，它可增加骨髓细胞释放白细胞介素1β和白细胞介素6[98]。这些细胞因子，无论是系统释放的，还是局部骨的微环境产生的，都会刺激破骨细胞的生成和活性，增加骨吸收[99-100]。许多研究发现在镁缺乏的大鼠或小鼠，体内P物质、白细胞介素1β和肿瘤坏死因子α的量会明显增加，这些细胞因子会增加破骨细胞的骨吸收，破坏骨吸收和骨生成的动态平衡[101-103]。破骨细胞的形成与3种成分有关，即核因子kB活化受体配体、核因子kB活化受体、骨保护素。核因子kB活化受体配体与核因子kB活化受体结合可刺激破骨细胞前体的分化，增加破骨细胞生成，而骨保护素在骨内主要由成骨细胞产生，可阻止核因子kB活化受体配体与核因子kB活化受体结合，抑制破骨细胞生成[104-105]。Rude 等[106]的研究发现，低镁可使小鼠体内核因子kB活化受体配体含量增加，骨保护素含量减少，导致破骨细胞增加，骨量丢失。Crespi等[107]研究表明，镁可引起成骨细胞分化标志物cbfa1的表达增高，刺激成骨细胞合成骨钙素和Ⅰ型胶原，同时降低核因子kB活化受体配体/骨保护素比率，减少破骨细胞的生成。 Abed等[108-110]发现，TRPM7通道可参与调节成骨细胞内镁的平衡，而血小板源性生长因子可刺激成骨细胞对镁离子通道TRPM7的表达，促进镁离子内流，增加成骨细胞增殖和迁移。并且，细胞外的镁浓度对于血小板源性生长因子的功能是至关重要的，低镁会抑制血小板源性生长因子诱导的成骨细胞增殖和迁移。另外，细胞外镁浓度还可直接调节TRPM7的表达和活性，后者参与维持细胞内镁离子的平衡并调节成骨细胞的增殖，因此，低镁会抑制人成骨细胞的增殖。"

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