烟酰胺单核苷酸: 一种潜在分子的多种治疗应用探索

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Nicotinamide Mononucleotide: Exploration of Diverse Therapeutic Applications of a Potential Molecule

Abstract

摘要

Nicotinamide mononucleotide (NMN) is a nucleotide that is most recognized for its role as an intermediate of nicotinamide adenine dinucleotide (NAD+) biosynthesis. Although the biosynthetic pathway of NMN varies between eukaryote and prokaryote, two pathways are mainly followed in case of eukaryotic human—one is through the salvage pathway using nicotinamide while the other follows phosphorylation of nicotinamide riboside. Due to the unavailability of a suitable transporter, NMN enters inside the mammalian cell in the form of nicotinamide riboside followed by its subsequent conversion to NMN and NAD+. This particular molecule has demonstrated several beneficial pharmacological activities in preclinical studies, which suggest its potential therapeutic use. Mostly mediated by its involvement in NAD+ biosynthesis, the pharmacological activities of NMN include its role in cellular biochemical functions, cardioprotection, diabetes, Alzheimer’s disease, and complications associated with obesity. The recent groundbreaking discovery of anti-ageing activities of this chemical moiety has added a valuable essence in the research involving this molecule. This review focuses on the biosynthesis of NMN in mammalian and prokaryotic cells and mechanism of absorption along with the reported pharmacological activities in murine model.

烟酰胺单核苷酸(NMN)是一个核苷酸,最公认的作用是作为一个中间烟酰胺腺嘌呤二核苷酸(NAD +)生物合成。虽然 NMN 的生物合成途径在真核生物和原核生物之间存在差异,但在真核生物中主要有两条途径: 一条是通过烟酰胺的补救途径,另一条是烟酰胺核苷的磷酸化途径。由于没有合适的转运体,NMN 以烟酰胺核糖苷的形式进入哺乳动物细胞,随后转化为 NMN 和 NAD + 。这种特殊的分子已经证明了一些有益的药理活动在临床前研究,这表明它的潜在治疗用途。NMN 的药理活性主要通过其参与 NAD + 生物合成而介导,包括其在细胞生化功能、心脏保护、糖尿病、阿尔茨海默病以及与肥胖相关的并发症中的作用。最近对这种化学单元抗衰老活性的突破性发现为这种分子的研究增添了宝贵的精华。本文综述了 NMN 在哺乳动物和原核细胞中的生物合成、吸收机制以及在小鼠模型中报道的药理活性。Keywords: 关键词:ageing, Alzheimer’s disease, diabetes, ischemic preconditioning, nicotinamide mononucleotide, obesity 老化,阿尔茨海默病,糖尿病,缺血预适应,烟酰胺单核苷酸,肥胖Go to: 去:

1. Introduction

1. 引言

Nicotinamide mononucleotide (NMN) or Nicotinamide-1-ium-1-β-D-ribofuranoside 5′-phosphate is a type of bioactive nucleotide which is naturally formed by the reaction between a phosphate group and a nucleoside containing ribose and nicotinamide [1]. Generally, it exists in two anomeric forms namely alpha and beta. The beta anomer is the active form between these two with a molecular weight of 334.221 g/mol [2]. NMN is naturally abundant in various types of food [3]. Vegetables like broccoli, cabbage contain 0.25–1.12 and 0.0–0.90 mg NMN/100 gm, fruits like avocado, tomato contain 0.36–1.60 and 0.26–0.30 mg NMN/100 gm, whereas raw beef has 0.06–0.42 mg NMN/100 gm [3]. NMN is also used as a substrate for prokaryotic enzymes like NadM in Methanobacterium thermoautotrophicum, NadR in Haemophilus influenza, NadM/Nudix in Francisella tularensis [4].

烟酰胺单核苷酸(NMN)或烟酰胺 -1- 核糖 -1- d- 呋喃糖苷5′-磷酸酯是一类生物活性核苷酸,它是由一个磷酸基团与一个含有核糖和烟酰胺的核苷[1]反应而自然形成的。一般来说,它以两种不规则形式存在,即 α 和 β。β 异构体是介于两者之间的活性形式,分子量为334.221 g/mol [2]。NMN 在各种类型的食物中自然含量丰富[3]。西兰花、卷心菜等蔬菜含有0.25-1.12和0.0-0.90 mg NMN/100克,鳄梨、番茄等水果含有0.36-1.60和0.26-0.30 mg NMN/100克,而生牛肉含有0.06-0.42 mg NMN/100克。NMN 还被用作原核酶的底物,如土伦病法兰西斯氏菌中的 NadM、流感嗜血杆菌中的 NadR、纳达姆/nudix 等。

In human cells, NMN is available as a source of cellular energy. Not long ago, this molecule was only known for its activity as an intermediate in nicotinamide adenine dinucleotide (NAD+) biosynthesis. During this biosynthetic process of NAD+, NMN acts as an important substrate for enzymes like nicotinamide mononucleotide adenylyltransferase 1 or NMNAT 1 of nuclear origin and NMNAT 3 of mitochondrial origin that helps in the enzymatic conversion to NAD+ in human [5]. Recently, preclinical studies have demonstrated diversified pharmacological activities of NMN in cardiac and cerebral ischemia, Alzheimer’s disease, diet- and age-induced type 2 diabetes, and obesity, all of which are linked up to the deficiency of NAD+ [6,7,8]. Camacho-Pereira et al. have shown that an increased level of NAD+ consuming enzymes e.g., NAD+ dependent acetylase (Sirtuins), poly ADP-ribose polymerase (PARP), NADase (CD38) contribute to the decline of NAD+ with age [9]. In mammalian cells, CD38, a type of cell surface NADase enzyme, causes breakdown of NAD+ to form nicotinamide and (cyclic-)ADP-ribose [10]. On the other hand, expenditure of NAD+ helps PARP to produce branched ADP-ribose polymers that help in DNA repairing [11]. Another group of NAD+ consuming enzymes, sirtuins (SIRT 1-7) performs different functions by consuming NAD+. Apart from deacetylation, which is the most common NAD+ mediated function of sirtuins, other functions like desuccinylase, demalonylase, lipoamidase, and deglutarylase enzymatic activity are noteworthy that help in the cellular adaptation of energy deficit and improvement of metabolic function [12]. Administration of NMN can compensate for the deficiency of NAD+ caused by these NAD+ consuming enzymes.

在人类细胞中,NMN 可以作为细胞能量的来源。不久前,这种分子仅仅因为其作为烟酰胺腺嘌呤二核苷酸生物合成中间体的活性而为人所知。在 NAD + 的生物合成过程中,NMN 作为核起源的烟酰胺单核苷酸腺苷酰转移酶1或 NMNAT 1等酶的重要底物,并作为线粒体起源的 NMNAT 3参与人体内 NAD + 的酶转化[5]。近年来,临床前研究表明,NMN 在心脑缺血、阿尔茨海默病、饮食和年龄诱发的2型糖尿病、肥胖等疾病中具有多种药理活性,这些疾病都与 NAD + 缺乏有关[6,7,8]。Camacho-Pereira 等人的研究表明,NAD + 消耗酶如 NAD + 依赖性乙酰化酶(Sirtuins)、多 ADP-ribose 聚合酶(PARP)、 NADase (CD38)水平的增加有助于 NAD + 随年龄的降低[9]。在哺乳动物细胞中,CD38,一种细胞表面的 NADase 酶,引起 NAD + 的分解,形成烟酰胺和(环状 -) adp- 核糖[10]。另一方面,NAD + 的消耗有助于 PARP 产生支化的 adp 核糖聚合物,有助于 DNA 修复[11]。另一组 NAD + 消耗的酶,sirtuins (SIRT 1-7)通过消耗 NAD + 发挥不同的功能。除了去乙酰化是 NAD + 介导的 sirtuins 最常见的功能之外,其他功能如脱琥珀酸化酶、脱孤独酶、脂酰酶和脱戊二酸酶活性也是值得注意的,它们有助于细胞适应能量缺乏和改善代谢功能[12]。NMN 可以弥补这些 NAD + 消耗酶引起的 NAD + 缺乏。

NMN shares similar properties like other NAD+ precursors- nicotinamide riboside (NR), nicotinic acid, and nicotinamide [13]. Unlike NMN, nicotinic acid, and nicotinamide have several disadvantages in terms of their therapeutic application. Nicotinamide may cause hepatotoxicity or flushing, while a recent preclinical study suggests that it resides in the rat body for a shorter period of time compared to NMN [14,15]. Niacin or nicotinic acid is associated with adverse effects like cutaneous flushing when administered as an immediate release formulation whereas the sustained release formulations may cause hepatotoxicity [16]. Among the NAD+ precursors, NR and NMN are exceptions as fewer unfavorable side effects have been reported for these two metabolites [17]. Moreover, nicotinamide riboside is also orally bioavailable like NMN. Considering these, NMN could be proposed as a preferable therapeutic option that can be supported by several ongoing clinical trials (NCT03151239, UMIN000021309, UMIN000030609, and UMIN000025739).

NMN 与其他 NAD + 前体——烟酰胺核糖苷(NR)、烟酸和烟酰胺具有相似的性质[13]。与 NMN 不同,烟酸和烟酰胺在治疗应用方面有几个缺点。烟酰胺可能导致肝毒性或脸红,而最近的一项临床前研究表明,它在大鼠体内停留的时间比 NMN 短[14,15]。烟酸或烟酸作为即时释放配方使用时会引起皮肤潮红等不良反应,而缓释配方可能引起肝毒性[16]。在 NAD + 前体中,NR 和 NMN 是例外,因为这两种代谢产物的不良副作用较少报道[17]。此外,烟酰胺核糖苷也像 NMN 一样是口服生物可利用的。考虑到这些,NMN 可以作为一个更好的治疗选择,可以支持几个正在进行的临床试验(NCT03151239,UMIN000021309,UMIN000030609,和 umin00025739)。

Here, in this review, the biosynthetic routes and absorption of NMN are discussed followed by a comprehensive analysis of the preclinically reported pharmacological properties with their underlying mechanism of actions. This will provide an insight into the possibility of converting these successful preclinical results for the treatment of human diseases.

本文综述了 NMN 的生物合成路线和吸收,并对临床前报道的药理学性质及其作用机制进行了综合分析。这将为将这些成功的临床前结果转化为治疗人类疾病的可能性提供深入的了解。Go to: 去:

2. Biosynthesis and Mechanism of Absorption

2. 生物合成和吸收机理

As NMN is an intermediate product of NAD+ biosynthesis, first we need to focus on the biosynthesis of NAD+ for a proper understanding of NMN synthesis. This particular biosynthetic route is important to clarify the mechanism by which the deficiency of NAD+ can be compensated. NAD+ is synthesized by three different pathways in mammalian cells-1) de novo pathway-synthesis from tryptophan, 2) salvage pathway-synthesis from either nicotinamide or nicotinic acid, or 3) conversion of NR [18]. Among these, the latter two pathways will be discussed in this review (Figure 1), as NMN is an intermediate byproduct here.

由于 NMN 是 NAD + 生物合成的中间产物,我们首先需要关注 NAD + 的生物合成,才能正确理解 NMN 的合成。这种特殊的生物合成路线对于阐明 NAD + 缺陷的补偿机制具有重要意义。NAD + 在哺乳动物细胞中有三种不同的合成途径: 1)色氨酸从头合成; 2)烟酰胺或烟酸补救合成; 3) NR [18]转化。其中,后两种途径将在本文中讨论(图1) ,因为 NMN 是这里的中间副产品。

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Figure 1 图1

Biosynthetic pathway of nicotinamide mononucleotide in mammalian cells.

哺乳动物细胞内烟酰胺单核苷酸的生物合成途径。

The salvage pathway is mostly predominant in mammalian cells [19,20]. In this pathway, the intermediate degradative products of NAD+ e.g., nicotinic acid and nicotinamide are reused to produce new NAD+. Most commonly, this pathway involves the conversion of the nicotinic acid to nicotinic acid mononucleotide by nicotinate phosphoribosyltransferase 1 followed by adenylation to nicotinic acid adenine dinucleotide in presence of nicotinamide mononucleotide adenylyl transferase 1, 3. Sometimes, nicotinic acid is directly converted to nicotinic acid adenine dinucleotide by nicotinic acid phosphoribosyltransferase. It is then converted to NAD+ with the help of NAD+ synthetase 1. This NAD+ is degraded to nicotinamide by NAD+ consuming enzymes, followed by conversion to NMN by the catalytic activity of nicotinamide phosphoribosyltransferase. The enzymes nicotinate phosphoribosyltransferase 1 and nicotinamide phosphoribosyltransferase both catalyze the transfer of a phosphoribosyl residue from phosphoribosylpyrophosphate (Figure 1) [21,22].

补救途径主要是在哺乳动物细胞[19,20]。在这个途径中,NAD + 的中间降解产物,例如烟酸和烟酰胺被重复使用产生新的 NAD + 。最常见的是,这一途径涉及到在烟酰胺单核苷酸腺苷酰转移酶1,3存在下,烟酸通过烟酸磷酸核糖转移酶1将烟酸转化为烟酸单核苷酸,然后再通过腺苷酸化转化为烟酸腺苷二核苷酸。有时,烟酸被烟酸磷酸核糖转移酶直接转化为烟酸腺嘌呤二核苷酸。然后在 NAD + 合成酶1的帮助下转化为 NAD + 。这种 NAD + 通过 NAD + 消耗酶降解为烟酰胺,然后通过烟酰胺磷酸核糖转移酶的催化活性转化为 NMN。烟酸磷酸核糖基转移酶1和烟酰胺磷酸核糖基转移酶两种酶都催化磷酸核糖焦磷酸中磷酸核糖基残基的转移(图1)[21,22]。

In their study with yeast and human cells, Bieganowski et al. discovered another NAD+ precursor molecule, NR, which is converted to NMN by phosphorylation with the help of nicotinamide riboside kinase (NRK1 and NRK2) [1]. The NMN formed is then enzymatically converted to NAD+ (Figure 1).

在他们对酵母和人类细胞的研究中,Bieganowski 等人发现了另一种 NAD + 前体分子 NR,它在烟酰胺核糖苷激酶(NRK1和 NRK2)的帮助下通过磷酸化转化为 NMN。形成的 NMN 然后酶转化为 NAD + (图1)。

Use of prokaryotic bacteria, e.g., lysates from Escherichia coli (E. coli) with specific genotyping is shown to be a simple and cost-effective way to produce NMN [23]. In bacteria, NAD+ biosynthesis takes place in a slightly different way. Most bacteria depend on both de novo and salvage pathway (e.g., Bacillus anthracis) whereas some depend on either of the pathways (e.g., Helicobacter pylori) [24]. Some bacteria, for example, Francisella tularensis, a gram-negative aerobic bacterium that is the causative agent for tularemia, follows a slightly different route for NAD+ synthesis. Here, amidation of nicotinic acid mononucleotide to NMN first takes place with the help of NMN synthetase followed by adenylation to NAD+ by NMN adenyltransferase [24].

利用原核细菌,例如大肠杆菌的裂解物,并进行特定的基因分型,是产生 NMN 的一个简单和具成本效益的方法。在细菌中,NAD + 的生物合成以一种稍微不同的方式进行。大多数细菌依赖于新创和补救途径(如炭疽桿菌) ,而一些细菌则依赖于这两种途径中的一种(如幽门螺杆菌)[24]。一些细菌,例如土伦病法兰西斯氏菌,一种革兰氏阴性的需氧细菌,是兔热病的病原体,遵循一个略有不同的 NAD + 合成路线。在这里,烟酸单核苷酸酰胺化 NMN 首先发生的帮助下 NMN 合成酶,其次是腺苷酸化的 NAD + 由 NMN 腺苷酸转移酶[24]。

Except Nostoc punctiforme and Synechocystis, most of the cyanobacteria follow a biosynthetic pathway for NAD+ production which do not involve NMN [25]. In N. punctiforme, nicotinamide riboside is taken up by a PnuC-like transporter and undergoes consecutive conversion to NMN by ribosylnicotinamide kinase and then to NAD+ by nicotinamide nucleotide adenylyltransferase. On the other hand, the pathway followed by Synechocystis is similar to humans. Here, nicotinamide is converted to NMN and subsequently to NAD+ by the catalytic activity of nicotinamide phosphoribosyltransferase and nicotinamide mononucleotide adenylyltransferase, respectively [25].

除了点状念珠藻和集胞藻之外,大多数蓝藻都遵循生物合成途径生产 NAD + ,而且不涉及 NMN [25]。在 n. punctiforme 中,烟酰胺核苷酸被一个类烟酰胺转运体吸收,经过核糖基烟酰胺激酶连续转化为 NMN,然后经过烟酰胺核苷酸腺苷酰转移酶连续转化为 NAD + 。另一方面,集胞藻所遵循的路径与人类相似。在这里,烟酰胺分别通过烟酰胺磷酸核糖转移酶和烟酰胺单核苷酸腺苷酰转移酶的催化活性转化为 NMN,然后转化为 NAD + [25]。

The question that follows the biosynthesis is the mechanism of absorption of NMN after oral administration. After biosynthesis, NAD+ is readily absorbed through the gut wall. With the help of the murine model, it was found that the absorption of NMN from the gut into blood circulation starts within 2–3 min and within 15 min, it is completely absorbed into tissue. Then it is converted and stored immediately as NAD+ in tissues like liver, skeletal muscle, and cortex. This increase in hepatic NAD+ content persists for about 30 min [3]. After six months of NMN administration, this spiked concentration of NAD+ can be observed in the liver and brown adipose tissue, however, not in skeletal muscle and white adipose tissue [3]. Before entering the mammalian cells, NMN undergoes dephosphorylation to produce NR. Extracellular receptor CD73, with pyrophosphatase and 5’-ectonucleotidase activity, carries out the reaction. Mammalian cells have Equilibrative Nucleoside Transporters or ENT which facilitate the entry of the NR. The newly-formed NR then acts as an exogenous NAD+ precursor within the mammalian cells (Figure 2). Following this, the ubiquitously expressed NRK1 helps the subsequent conversion of NR to NMN [26].

生物合成之后的问题是 NMN 在口服给药后的吸收机制。生物合成后,NAD + 很容易通过肠壁被吸收。在小鼠模型的帮助下,发现 NMN 从肠道吸收进入血液循环在2-3分钟内开始,在15分钟内完全被组织吸收。然后在肝脏、骨骼肌和大脑皮层等组织中转化和储存为 NAD + 。肝脏 NAD + 含量的增加持续约30分钟[3]。服用 NMN 六个月后,可以在肝脏和褐色脂肪组织中观察到 NAD + 的浓度达到峰值,但是在骨骼肌和白色脂肪组织中却没有。在进入哺乳动物细胞之前,NMN 经过脱磷酸化生成 NR。胞外受体 CD73具有焦磷酸酶和5’-核苷酸酶活性,可以进行反应。哺乳动物细胞具有平衡型核苷转运蛋白或 ENT,它们促进 NR 的进入。新形成的 NR 然后充当哺乳动物细胞内外源 NAD + 的前体(图2)。在此之后,NRK1的泛在表达有助于随后 NR 转换为 NMN [26]。

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Figure 2 图2

Schematic of absorption of NMN in mammalian cells. NMN: nicotinamide mononucleotide; NR: nicotinamide riboside; NRK 1: nicotinamide riboside kinase 1; ENT: Equilibrative nucleoside transporters.

哺乳动物细胞吸收 NMN 示意图。NMN: 烟酰胺单核苷酸; NR: 烟酰胺核苷; NRK 1: 烟酰胺核苷激酶1; ENT: 平衡核苷转运蛋白。

Considering all the differences in NMN biosynthetic pathway between prokaryote and eukaryote and rapid absorption pattern of it, NMN can be conferred as one of the metabolites that have paramount significance in the turnover of NAD+.

考虑到原核生物与真核生物 NMN 生物合成途径的差异以及 NMN 的快速吸收模式,NMN 可以被认为是 NAD + 转化过程中最重要的代谢产物之一。Go to: 去:

3. Pharmacological Activities

3. 药理活性

NMN could open up a new horizon in modern therapeutics. This biomolecule has demonstrated numerous beneficial pharmacological activities in several preclinical disease models including myocardial and cerebral ischemia, neurodegenerative disorders like Alzheimer’s disease, and diabetes [6,7,27]. The most recent discovery of its anti-ageing, life-span prolonging property in the murine model has made NMN more attractive as a potential therapeutic candidate [28]. Most of its pharmacological actions take place by facilitating NAD+ synthesis, as direct NAD+ administration in higher dose exhibits side effects like insomnia, fatigue, and anxiety sometimes and it has poor penetration capability through plasma membrane compared to NMN [29].

NMN 为现代治疗学开辟了新的视野。这种生物分子已经证明了许多有益的药理活动在几个临床前疾病模型,包括心肌和脑缺血,神经退行性疾病如阿尔茨海默病,和糖尿病[6,7,27]。最近在小鼠模型中发现的抗衰老、延长寿命的特性使得 NMN 作为一个潜在的治疗候选者更具吸引力[28]。其药理作用主要是通过促进 NAD + 的合成而发生的,与 NMN 相比,更高剂量的 NAD + 直接给药具有失眠、疲劳、焦虑等副作用,其穿透血浆的能力较差[29]。

3.1. Ischemia-Reperfusion Injury

3.1. 缺血-再灌注损伤

Due to ischemic events, the amount of oxygen, as well as adenosine triphosphate (ATP) level in the cardiac muscle cells or cardiomyocytes, decrease. Upon further aggravation, these cardiac muscle cells undergo necrosis [30]. Reperfusion, also known as the reoxygenation process, is the event of resupplying blood to the tissue, which has previously undergone ischemia. Reperfusion causes blood to re-enter the tissue cells leading to calcium (Ca2+) overload by microvascular injury and production of ROS. These consecutive events cause severe tissue damage [31]. Ischemia followed by reperfusion is a deadly condition which is counteracted by a mechanism of the human body known as ischemic preconditioning or IPC [32]. IPC, an endogenous mechanism of the body, helps to revert this condition by stimulating multiple signaling mediators [33]. IPC induces activation of sirtuin1 (SIRT1) [34]. SIRT1 is a NAD-dependent class-III histone deacetylase protein which causes deacetylation of lysine residues of FoxO transcription factor that is responsible for generating oxygen free radicals. Therefore, providing a counteracting mechanism to protect the body from oxidative stress and injury due to ischemia and reperfusion. SIRT1 depends on intracellular NAD+ for its deacetylase activity [35]. Yamamoto et al. investigated the connection between nicotinamide phosphoribosyltransferase, a rate-limiting enzyme of the NAD+ salvage pathway, and IPC via activation of SIRT1 [7]. By using nicotinamide phosphoribosyltransferase +/− mice, it was found that nicotinamide phosphoribosyltransferase has a positive role of attenuation of myocardial injury following ischemia and reperfusion. Since nicotinamide phosphoribosyltransferase is the rate-limiting enzyme of NAD+ production, administration of the intraperitoneal NMN at 500 mg/kg, either 30 min before the onset of ischemia or every 6 hours during the reperfusion period for 24 hours, causes significant amelioration of ischemia-reperfusion injury by reducing the infarct size by 44% and 29%, respectively, compared to the phosphate buffer saline control. But, when this intervention was repeated against cardiac-specific KO mice, NMN intervention proved unsuccessful. Thus, it can be asserted that NMN, being the intermediate product of NAD+ biosynthesis, has the capability to activate SIRT1 (Figure 3) and thus mimics the action of IPC to ameliorate ischemia-reperfusion injury [7].

由于缺血事件,心肌细胞或心肌细胞的含氧量,以及三磷酸腺苷(ATP)水平下降。在进一步恶化时,这些心肌细胞发生坏死[30]。再灌注,也称为复氧过程,是组织再供血的过程,此前已经发生了缺血。再灌注导致血液重新进入组织细胞导致钙超载的微血管损伤和产生活性氧。这些连续发生的事件会造成严重的组织损伤。缺血再灌注是一种致命的情况,被称为缺血预处理或 IPC 的人体机制抵消。IPC 是机体的一种内源性机制,通过刺激多种信号传导介质来帮助恢复这种状态。IPC 诱导 sirtuin1(SIRT1)的活化。SIRT1是一种依赖于 nad- 的 iii 类组蛋白脱乙酰酶蛋白,它能使 FoxO 转录因子的赖氨酸残基脱乙酰化,从而产生氧自由基。因此,提供一个抵消机制,以保护机构免受氧化应激和损伤,由于缺血和再灌注。SIRT1的脱乙酰基酶活性依赖于细胞内 NAD + [35]。山本等人通过激活 SIRT1[7] ,研究了 NAD + 补救途径中的限速酶——烟酰胺磷酸核糖基转移酶与 IPC 之间的联系。用烟酰胺磷酸核糖基转移酶 +/-小鼠,发现烟酰胺磷酸核糖基转移酶对心肌缺血再灌注损伤有明显的减轻作用。由于烟酰胺磷酸核糖基转移酶是产生 NAD + 的限速酶,与磷酸盐缓冲盐对照组相比,腹腔注射500mg/kg 的 NMN,在缺血开始前30min 或再灌注期间每6小时注射一次,可显著改善缺血再灌注损伤,分别缩小梗死面积44% 和29% 。但是,当对心脏特异性 KO 小鼠重复进行这种干预时,NMN 干预被证明是不成功的。因此,可以断言 NMN 作为 NAD + 生物合成的中间产物,具有激活 SIRT1的能力(图3) ,从而模拟 IPC 改善缺血再灌注损伤的作用[7]。

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Figure 3 图3

Mechanism of SIRT1 mediated pharmacological activities of NMN. NMN is converted to NAD+ intracellularly which performs physiological functions via SIRT1. (a) SIRT1 causes deacetylation of lysine residues of FOXO transcription factor that stimulates catalase enzyme to inhibit ROS and the chain reactions leading to ischemia-reperfusion injury. (b) In normal condition, p65 subunit of NFκB transcription factor complex, in its acetylated form, expresses ROS which is also responsible for insulin resistance. SIRT1, due to its inherent deacetylation activity, deacetylates p65-NFκB and thus inhibits production of ROS, which is responsible for the occurrence of type-2 diabetes mellitus. (c) SIRT1 also deacetylates protein PGC-1α and stimulates the expression of proteins responsible for mitochondrial biogenesis, which can be used for the treatment of Alzheimer’s disease.

SIRT1介导 NMN 药理作用机制的研究。NMN 通过 SIRT1转化为细胞内 NAD + ,并发挥生理功能。(a) SIRT1引起 FOXO 转录因子的赖氨酸残基去乙酰化,刺激过氧化氢酶抑制活性氧和导致缺血再灌注损伤的链式反应。在正常情况下,nf b 转录因子复合物的 p65亚基以乙酰化形式表达活性氧,这也是胰岛素抵抗的原因。由于 SIRT1固有的去乙酰化活性,去乙酰化 p65-nf b,从而抑制 ROS 的产生,而 ROS 是2型糖尿病的产生原因。(c) SIRT1还能使 pgc-1蛋白去乙酰化,刺激线粒体生物发生蛋白的表达,可用于治疗阿尔茨海默病。

Apart from this SIRT1 mediated mechanism, another pathway shown to be responsible for this cardioprotective activity is the stimulation of glycolysis or acidosis depending on the timing of NMN delivery in respective to ischemic incidence [36]. If NMN is provided before the occurrence of an ischemic event, then glycolysis is increased that facilitates ATP production during ischemic events, thus promotes cardioprotection. In contrast, when NMN is given during reperfusion, it protects the heart by inducing acidosis, contributed by the cardiac lactate and pyruvate mainly. This causes a shutdown of mitochondrial permeability transition pore and, therefore, ensures cardioprotection [37].

除了这个 SIRT1介导的机制,另一个被证明负责这种心脏保护活性的途径是糖酵解或酸中毒的刺激,这取决于 NMN 分娩的时间在各自的缺血发生率[36]。如果 NMN 是在缺血事件发生之前提供的,那么糖酵解增加,促进缺血事件中 ATP 的产生,从而促进心脏保护。相比之下,在再灌注期间给予 NMN,可以通过诱导酸中毒来保护心脏,而酸中毒主要由心脏的乳酸和丙酮酸贡献。这导致线粒体通透性转换孔关闭,因此,确保心脏保护[37]。

NMN has also shown its therapeutic potential for the treatment of cerebral ischemia in preclinical studies. In a recent investigation by Park et al., NMN was introduced at a dose of 62.5 mg/kg in transient forebrain ischemic mice to find out the expression of different biomarkers of post-ischemic event like depletion of hippocampal NAD+ levels, accumulation of poly-ADP-ribosylation (PAR). In comparison to the effect of the control, neurologic outcome and hippocampal CA1 neuronal death after reperfusion were significantly improved by NMN treatment. Simultaneously, PAR formation and NAD+ catabolism were reduced and body temperature remained unaffected which proved that NMN treatment was solely responsible for this protective effect against ischemic brain injury [27].

在临床前研究中,NMN 也显示了其治疗脑缺血的潜力。在 Park 等人最近的一项研究中,以62.5 mg/kg 的剂量给短暂性前脑缺血小鼠注射 NMN,观察缺血后海马 NAD + 水平缺失、多聚 adp- 核糖基化(PAR)积累等不同生物标志物的表达情况。与对照组相比,NMN 治疗可明显改善再灌注后神经功能和海马 CA1区神经元死亡。同时,PAR 的形成和 NAD + 分解代谢降低,体温不受影响,这证明 NMN 治疗是唯一负责这种保护作用对缺血性脑损伤[27]。

Looking at the current scenario, there are only a few available intervention strategies for protecting the heart and brain from ischemia-reperfusion injury due to insufficient clinical trial data and lack of pharmacokinetic and pharmacodynamic study [38]. Again, the complex sequential events leading to this condition also limit the benefits of different surgical interventions like coronary artery bypass graft surgery and percutaneous coronary intervention [39]. NMN, being a successful entity in preclinical studies, could act as an alternative therapeutic strategy in this disorder.

从目前的情况来看,由于临床试验数据不足,缺乏药代动力学和药效学研究,保护心脏和大脑免受缺血再灌注损伤的干预策略很少[38]。同样,导致这种情况的复杂的连续事件也限制了不同外科手术的好处,如冠状动脉搭桥手术和冠状动脉再成形术。NMN 是临床前研究中的一个成功实体,可以作为该疾病的一种替代治疗策略。

3.2. Neurological Disorders: Alzheimer’s Disease and Intracerebral Hemorrhage

3.2. 神经系统疾病: 老年痴呆症和脑内出血

NMN has also shown promising activity in the treatment of Alzheimer’s disease as demonstrated by Long et al. It has the capability to treat underlying causes of Alzheimer’s disease, e.g., morphological abnormalities of mitochondria, decrease in oxygen consumption rates (OCR) and NAD+ content [8]. As the currently available pharmacological interventions like memantine or the cholinesterase inhibitor galantamine only aim at providing symptomatic treatment. Moreover, they cause side effects like anorexia and bradycardia, hence, NMN can play an important role since it directly targets the etiology of this disease [40]. NAD+ is recognized for its catalytic activity in aerobic respiration where oxygen is consumed by mitochondria [41]. Due to both aging and pathophysiology of Alzheimer’s disease, the availability of NAD+ decreases which leads to the decline in OCR by brain and muscle cell mitochondria [8]. Again, mitochondrial dynamics entirely depend upon fission and fusion of mitochondria. Consequently, mitochondria tend to undergo more fragmentation, i.e., increased fission and decreased fusion, leading to abnormal morphology and function of mitochondria [42,43]. In a published study by Long et al. NMN was introduced in AD chimeric APP(swe)/PS1(ΔE9) (AD-Tg) mice for mitochondrial OCR assay and on mice having fluorescent proteins tagged to neuronal mitochondria (CaMK2a-mito/eYFP) to assess mitochondrial morphology [8]. NMN was found to be effective in increasing the mitochondrial maximal OCR in transgenic AD-Tg mice compared to AD-Tg-vehicle and nontransgenic vehicle-treated group by efficiently crossing the blood-brain barrier and thus compensating the deficiency of NAD+ in mitochondria. In addition to increasing maximal OCR, NMN was successful to increase OCR significantly following the addition of ADP (for the initiation of state 3 respiration) in AD-Tg NMN treated mice compared to non-transgenic one, although the basal OCRs were similar. NMN also activated SIRT1 via NAD+ biosynthesis by salvage pathway, which, in turn, stimulated the deacetylation of target protein PGC-1α (Figure 3), responsible for mitochondrial biogenesis [44]. Again in CaMK2a-mito/eYFP mice, the NMN treated group showed reduced fragmentation and increased length of mitochondria, evidenced by an increase in rod shape and tubular length, which ensured the integrity of mitochondria [8].

正如 Long 等人所证明的那样,NMN 在治疗阿尔茨海默病方面也显示出有希望的活性。它有能力治疗阿尔茨海默病的根本原因,例如,线粒体的形态异常,降低耗氧率(OCR)和 NAD + 含量[8]。由于目前可用的药物干预,如美金刚胺或胆碱酯酶抑制剂加兰他敏,只是为了提供对症治疗。此外,他们引起副作用,如厌食和心动过缓,因此,NMN 可以发挥重要作用,因为它直接针对这种疾病的病因[40]。NAD + 因其在有氧呼吸中的催化活性而被公认,在有氧呼吸中线粒体消耗氧气[41]。由于阿尔茨海默病的老化和病理生理,NAD + 的可用性降低,导致大脑和肌细胞线粒体的 OCR 下降[8]。同样,线粒体动力学完全依赖于线粒体的分裂和融合。因此,线粒体趋向于进行更多的碎片化,即增加裂变和减少融合,导致线粒体的形态和功能异常[42,43]。在 Long 等人发表的一项研究中。将 NMN 引入 AD 嵌合 APP (swe)/PS1(e9)(AD-tg)小鼠,用于线粒体 OCR 测定,用荧光蛋白标记的神经元线粒体(CaMK2a-mito/eYFP)测定线粒体形态[8]。结果表明,NMN 通过血脑屏障有效地跨越血脑屏障,从而弥补了线粒体 NAD + 的缺乏,使转基因 AD-Tg 小鼠的线粒体最大 OCR 比非转基因组和转基因组小鼠有显著提高。与非转基因小鼠相比,经 AD-Tg NMN 处理的小鼠在添加 ADP (启动状态3呼吸)后,除了增加最大 OCR 外,NMN 还显著增加 OCR,尽管基础 OCR 相似。NMN 还通过补救途径通过 NAD + 生物合成激活 SIRT1,从而刺激靶蛋白 pgc-1的去乙酰化(图3) ,负责线粒体的生物发生[44]。在 CaMK2a-mito/eYFP 小鼠中,NMN 治疗组小鼠线粒体断裂减少,线粒体长度增加,杆状和管状线粒体长度增加,保证了线粒体的完整性[8]。

The amyloid beta oligomer, also known as Aβ or A-beta, is a type of neurotoxic protein which forms amyloid plaques in the brains of Alzheimer patients [45]. It is also responsible for inducing depression of hippocampal long-term potentiation or LTP [46]. NMN successfully inhibited Aβ oligomers induced LTP by 140% compared to that of baseline. On organotypic hippocampal slice cultures (OHCs), NMN decreased Aβ oligomers induced cell death by 65% in Aβ oligomer infusion AD model rats (dose 500 mg/kg, intraperitoneally) indicating the improvement in cognitive function [45]. This finding confirms the previous studies where NMN has been proved for its potential therapeutic application in Alzheimer’s disease. With further studies regarding the dosage regimen and subsequent clinical trials, it could be a promising therapeutic intervention for Alzheimer’s disease in human.

淀粉样 β 低聚体,也称为 a 或 a-β,是一种神经毒性蛋白,在阿尔茨海默病患者的大脑中形成淀粉样斑块[45]。它还负责诱导海马长时程增强作用或 LTP [46]。与基线相比,NMN 成功抑制了寡聚体诱导的 LTP 140% 。在器官型海马脑片培养(ohc)方面,NMN 使 a 寡聚体诱导的 AD 模型大鼠(剂量500mg/kg,腹腔注射)的细胞死亡率降低了65% ,表明认知功能得到改善[45]。这一发现证实了先前的研究,其中 NMN 已被证明其潜在的治疗应用阿尔茨海默病。随着对给药方案和随后临床试验的进一步研究,它可能成为治疗人类阿尔茨海默病的一种有前途的治疗方法。

Intracerebral hemorrhage (ICH) is another neurological disorder, responsible for about 10–15% of all strokes, where NMN can improve the disease condition [47]. The injury caused by ICH occurs in two stages: at first, mechanical damage to neighboring tissues takes place by hematoma formation that is followed by secondary damage via pathological alterations (e.g., cytotoxicity, excitotoxicity, activation of inflammatory pathways causing neuroinflammation) caused by hematoma [48]. A study on collagenase-induced ICH mouse model, upon exposure to 300 mg/kg intraperitoneal dose of NMN after 30 min of ICH episode, showed that NMN treatment increased intracerebral NAD+ concentration at two and six-hour post-ICH and provided protection against amyotrophic lateral sclerosis and ischemic stroke. Furthermore, NMN treatment resulted into significant improvement, measured by a decrease in edema, neuronal death, ROS content, neurological inflammation, expression of intercellular adhesion molecule-1, neutrophil infiltration and microglia activation in the affected area of the brain. Although this intervention failed to reduce hematoma volume and hemoglobin content [49], it successfully improved the conditions mediated by ICH which suggests that NMN can be used for the treatment of ICH.

脑内出血是另一种神经系统疾病,约占所有中风病例的10-15% ,NMN 可以改善中风病情[47]。脑出血引起的损伤分为两个阶段: 首先,血肿形成引起邻近组织的机械性损伤,然后是血肿引起的病理改变引起的继发性损伤(如细胞毒性、兴奋毒性、炎症通路的激活引起神经炎症)。对胶原酶诱导的 ICH 小鼠模型的研究表明,NMN 治疗增加了 ICH 后2小时和6小时脑内 NAD + 的浓度,对肌萎缩性嵴髓侧索硬化症和缺血性脑卒中有保护作用。此外,NMN 治疗导致显着改善,衡量减少水肿,神经元死亡,活性氧含量,神经炎症,表达细胞间粘附分子 -1,中性粒细胞浸润和小胶质细胞活化的影响区域。虽然这种干预未能减少血肿体积和血红蛋白含量[49] ,但它成功地改善了脑出血介导的条件,提示 NMN 可以用于治疗脑出血。

3.3. Diabetes

3.3. 糖尿病

NMN has also shown potential to be used as a therapeutic for the treatment of diabetes. Insulin resistance is the characteristic feature of type 2 diabetes which occurs due to oxidative stress, augmented inflammatory response, impaired lipid metabolism- all of which can be ameliorated by NAD+ [6]. High-fat diet and ageing contribute to the predisposition of this particular type of diabetes, one of the common mechanisms being the reduction of NAD+ [6]. High-fat diet, consisting mainly of saturated fats, can decrease nicotinamide phosphoribosyltransferase level and ultimately reduce NAD+ in the liver and white adipose tissue [6]. On the other hand, ageing influences the decrease of NAD+ level in the pancreas, white adipose tissue, skeletal muscle, and liver to a greater extent compared to the younger population [6]. This NAD+ acts as a safeguard against various physiological disorders. Glutathione S-transferase, a protective agent against oxidative stress from lipid peroxidation products, is restored by NAD+ [50]. Furthermore, expression of interleukin 1β (IL-1β) and S100 calcium-binding protein A8 and A9, the two targets of immune and inflammatory mediator nuclear factor kappa B (NF-kB), are downregulated by NAD+ [51]. This NF-kB is also responsible for causing insulin resistance. Activation of SIRT1 by NAD+ promotes deacetylation of p65 component of NF-kB (Figure 3) and thus prevents insulin resistance [52]. To revert the high-fat diet and ageing induced type 2 diabetes, Yoshino and his associates administered NMN, at 500 mg/kg/day intraperitoneally for 7 and 10 consecutive days in high-fat diet fed female and male diabetic mice, respectively. They found a significant improvement in insulin intolerance in both female and male mice, while female mice showed improvement to a greater extent [6]. In the case of age-induced diabetes, the same single dose and 11 consecutive doses of intraperitoneal NMN ameliorated glucose intolerance in diabetic male and female mice respectively. NMN did not hamper glucose homeostasis in non-diabetic normal mice whereas, energy utilization solely from glucose and improvement of hyperlipidemia in aged diabetic mice were ensured by this intervention. It also improved inflammation mediated islet cell dysfunction by increasing extracellular nicotinamide phosphoribosyltransferase concentration through suppression of IL-1β from islets cells and thereby restored insulin secretion from beta cells [53].

NMN 还显示出潜力,可用作糖尿病的治疗。胰岛素抵抗是2型糖尿病的典型特征,发生于氧化应激、炎症反应增强、脂质代谢受损-所有这些都可以通过 NAD + [6]改善。高脂肪饮食和衰老导致了这种特殊类型糖尿病的易感性,其中一个常见的机制是降低 NAD + [6]。高脂肪饮食,主要是饱和脂肪,可以降低烟酰胺磷酸核糖转移酶水平,最终降低肝脏和白色脂肪组织的 NAD + 。另一方面,与年轻人相比,衰老对胰腺、白色脂肪组织、骨骼肌和肝脏 NAD + 水平的影响更大[6]。这种 NAD + 作为防止各种生理疾病的保障。谷胱甘肽S-转移酶,一种从脂质过氧化产品中提取的防止氧化应激的保护剂,通过 NAD + [50]进行修复。此外,NAD + [51]可降低免疫和炎症介质核转录因子 -κb (NF-kB)的作用靶点白细胞介素1(il-1)和 S100钙结合蛋白 A8和 A9的表达。这个 NF-kB 也是导致胰岛素抵抗的原因。NAD + 激活 SIRT1促进 NF-kB p65成分脱乙酰化,从而防止胰岛素抵抗[52]。为了逆转高脂肪饮食和老化诱导的2型糖尿病,吉野和他的同事分别在高脂肪饮食喂养的雌性和雄性糖尿病小鼠腹腔内注射500毫克/千克/天的 NMN,连续7天和10天。他们发现雌性和雄性小鼠的胰岛素耐受性都有显著改善,而雌性小鼠的改善程度更大。在年龄诱发糖尿病的情况下,相同的单次剂量和连续11次剂量腹腔注射 NMN 分别改善了糖尿病雄性和雌性小鼠的葡萄糖耐受不良。NMN 对非糖尿病正常小鼠的血糖稳态无明显影响,而对老年糖尿病小鼠的能量利用和高脂血症的改善有明显的保护作用。它还通过抑制胰岛细胞的 il-1而增加细胞外烟酰胺磷酸核糖转移酶的浓度,从而改善炎症介导的胰岛细胞功能障碍,从而恢复 β 细胞的胰岛素分泌[53]。

3.4. Obesity and Its Related Complications

3.4. 肥胖及其相关并发症

NMN can reduce age-associated weight gain in a dose-dependent manner, supported by a research on mice [3]. In this study, 100 and 300 mg/kg dose of NMN, applied throughout a duration of 12 months, was capable of reducing body weight by 4% and 9%, respectively, compared to the control group without any compromise to growth and appetite. There is an interconnection between the pathology of obesity and diabetes. Obesity exerts negative health effects through alteration in biochemical pathways that cause mitochondrial dysfunction. The decreased ATP production through changes in NAD+ and NADH level by dysfunctional muscle and hepatic mitochondria leads to insulin resistance and type-2 diabetes [54,55,56]. NAD+ helps to replenish the cellular energy level by stimulating mitochondria to generate ATP [57]. As mentioned before, SIRT1 utilizes NAD+ as a cofactor to improve mitochondrial biogenesis, (Figure 3) which is hampered due to obesity [58]. In a recent study, NMN was administered in high-fat diet (HFD) induced obese mice to assess and compare the capability of improving the NAD+ content with that of treadmill exercise, at an average of 15 m/min taken for 45 min and continued six days per week for six weeks. NMN treatment at a dose of 500 mg/kg body weight daily for 17 days was successful in increasing the NAD+ content in both muscle and liver, which was previously reduced by HFD-induced obesity, whereas exercise increased NAD+ in muscle only [59]. NMN treatment also improved metabolic disorders like glucose intolerance and reduced hepatic citrate synthase activity, similar to that of exercise [59]. In light of the above discussion, it can be inferred that NMN can improve certain metabolic complications associated with obesity, although this effect on many other obesity-related metabolomic complications like the non-alcoholic fatty liver disease is yet to be explored [59].

NMN 可以以剂量依赖的方式减少年龄相关的体重增加,这一结论得到了小鼠研究的支持[3]。在这项研究中,100和300毫克/千克剂量的 NMN,在整个12个月期间,分别能够减少体重4% 和9% ,与对照组相比,没有任何对生长和食欲的损害。肥胖和糖尿病的病理学之间有着相互联系。肥胖通过改变引起线粒体功能障碍的生化途径而对健康产生负面影响。功能失调的肌肉和肝线粒体通过改变 NAD + 和 NADH 水平降低 ATP 的产生,导致胰岛素抵抗和2型糖尿病[54,55,56]。NAD + 通过刺激线粒体产生 ATP 来帮助补充细胞能量水平。如前所述,SIRT1利用 NAD + 作为一个辅助因子来改善线粒体的生物合成(图3) ,这是由于肥胖而阻碍[58]。在最近的一项研究中,以高脂饮食(HFD)诱导的肥胖小鼠为研究对象,评估并比较了平均15m/min 运动和跑台运动对提高 NAD + 含量的能力,平均15m/min 运动45min,连续6周每周6天。NMN 治疗剂量500毫克/千克体重每天17天,成功地增加了 NAD + 含量在肌肉和肝脏,这是以前减少了高强度手足口病诱导的肥胖,而运动只增加了 NAD + 在肌肉[59]。NMN 治疗也改善了代谢紊乱,如葡萄糖耐受不良和减少肝脏柠檬酸合酶活性,类似于运动[59]。根据上述讨论,可以推断 NMN 可以改善与肥胖相关的某些代谢并发症,虽然这对许多其他与肥胖相关的代谢并发症,如非酒精性脂肪性肝病的影响还有待探讨[59]。

3.5. Ageing

3.5. 衰老

Geroscience is the field that deals with the relationship between ageing and the diseases accompanying these [60]. The process of ageing comes along with some age-related complications. Ageing is a natural human phenomenon characterized by downregulated energy production by mitochondria, as previously mentioned, owing to the depletion of NAD+ in multiple organs like pancreas, skeletal muscle, liver, skin, adipose tissue and brain [3,61,62]. Aside from the decrease in the mitochondrial function, ageing is also associated with other biological alterations like-DNA damage, cognitive impairment, sirtuin gene inactivation which can be retreated by NAD+ [63]. The age-related depletion of the NAD+ count, specifically that of nuclear origin, is also accounted for disruption of mitochondrial regulation of PGC-1α/β-independent pathway of oxidative-phosphorylation, leading to pseudohypoxia. This can be reversed by increasing the amount of NAD+ [64]. NAD+ has been reported for its ability to regenerate cells like muscle stem cells in older mice. Another precursor of NAD+, NR, has also been attributed for its ability to induce neurogenesis, halt the decrease in melanocyte stem cells and increase the lifespan in mice slightly [65]. Being the precursor of NAD+, NMN can provide these beneficial effects as well.

老年科学是研究衰老和伴随这些疾病的关系的领域[60]。衰老的过程伴随着一些与年龄有关的并发症。正如前面提到的,衰老是一种人类自然现象,由于胰腺、骨骼肌、肝脏、皮肤、脂肪组织和大脑等多个器官中 NAD + 的损耗,线粒体的能量产生拥有属性下降。除了线粒体功能下降,衰老还与其他生物学改变有关,如 dna 损伤、认知障碍、 sirtuin 基因失活,NAD + [63]可以消除这些改变。年龄相关的 NAD + 计数的减少,特别是核起源的 NAD + 计数的减少,也可以解释线粒体调节 pgc-1/-非依赖的氧化-磷酸化途径的中断,导致假缺氧。这可以通过增加 NAD + [64]的数量来逆转。据报道,NAD + 具有在老年老鼠体内再生肌肉干细胞等细胞的能力。NAD + 的另一个前体 NR 也被认为具有诱导神经发生、阻止黑素细胞干细胞减少和稍微延长老鼠寿命的能力[65]。作为 NAD + 的前体,NMN 也可以提供这些有益的作用。

Decreasing NAD+ level is also associated with age-related DNA damage. NAD+ binds to the nudix homology domain of various proteins. Although the specific function of nudix homology domain is yet to be found, studies have shown that the Deleted in Breast Cancer protein1 (DBC1) possesses this domain, which inhibits the DNA repairing protein PARP1 [66]. NAD+ binds to nudix domain of DBC1, thus the inhibition on PARP1 is reversed. With ageing, NAD+ level declines, so PARP1 loses its capability to repair DNA. This phenomenon was assessed on old (30-month-old) and young mice (6-month-old) with gamma radiation-induced DNA damage. The mice were treated with an intraperitoneal NMN at 500 mg/kg/day for one week. After the experimental period, it was found that NMN increased hepatic NAD+ concentration as well as PARP1 activity in repairing DNA damage [67].

降低 NAD + 水平也与年龄相关的 DNA 损伤有关。NAD + 与许多蛋白质的 nudix 同源结构域相结合。虽然 nudix 同源结构域的具体功能尚未发现,但研究表明,乳腺癌蛋白1(DBC1)中的缺失结构域具有这一结构域,它可以抑制 DNA 修复蛋白 PARP1[66]。NAD + 与 DBC1的 nudix 结构域结合,从而逆转了对 PARP1的抑制作用。随着年龄的增长,NAD + 水平下降,因此 PARP1失去了修复 DNA 的能力。这种现象是在老年(30个月大)和幼年(6个月大)小鼠中用伽马辐射引起的 DNA 损伤进行评估的。小鼠腹腔注射 NMN 500mg/kg/d,持续一周。实验结束后,发现 NMN 能增加肝组织 NAD + 浓度和 PARP1活性以修复 DNA 损伤[67]。

With the course of ageing, the pathophysiological changes in different cells and tissues like- eye and bone have become apparent. The lacrimal gland becomes less capable of producing tears, whereas, in some cases, light-colored spots in the fundus of the eye due to an age-dependent pileup of subretinal microglia and macrophages takes place [67,68].NMN, at a dose of 100 and 300 mg/kg/day, was found to be effective in reducing these spots in the fundus of the transgenic mouse strain C57BL/6N which contains the rd8 mutation, responsible for these abnormalities [3]. A 12-month NMN administration also increased tear production in a dose-dependent manner, in which the 300mg/kg/day dose was capable to increase the level of tear production comparable to that of maximal tear production of mice lifetime [3]. Thus, it substantiates its capability of reverting the optical abnormalities due to ageing. In addition, bone density depletion, a characteristic attribute of age-associated physiological change, was reverted significantly by NMN in a dose-dependent manner [3].

随着年龄的增长,眼、骨等不同组织和细胞的病理生理改变逐渐明显。泪腺分泌眼泪的能力下降,而在某些情况下,由于视网膜下小胶质细胞和巨噬细胞的年龄依赖性堆积,眼底出现浅色斑点[67,68]。NMN,100和300mg/kg/天,被发现有效地减少这些斑点在眼底的转基因小鼠株 C57BL/6N,其中包含 rd8突变,负责这些异常[3]。12个月的 NMN 治疗也以剂量依赖的方式增加了泪液生成,300毫克/千克/天的剂量能够提高泪液生成水平,与小鼠一生中最大泪液生成水平相当[3]。因此,它证实了其还原老化引起的光学异常的能力。此外,骨密度损耗,一个年龄相关的生理变化的特征性属性,显着扭转 NMN 的剂量依赖的方式[3]。

Senescence of the vascular system is another common occurrence during the process of aging. It is accompanied by oxidative stress imposed by the free radicals. NMN was found to be effective in reversing these conditions when tested on mice model. De Picciotto and his co-researchers conducted a study on this particular field to analyze the efficacy of NMN [69]. Vascular functionality was assessed in terms of carotid artery endothelium-dependent dilation (EDD); while nitric oxide-mediated EDD was used to analyze vascular oxidative stress. The older control mice (26–28 months) showed diminished functions on both of these parameters compared to the young control (4–8 months). Following an 8-week oral NMN intervention at 300 mg/kg/day on male C57B1/6 mice, both carotid artery EDD and nitric oxide EDD were restored to normal level, like that of young control mice. Again, the conditions of older mice like the abundance of oxidative stress marker nitrotyrosine, reduced elastin and vascular SIRT1 activity, all were reversed by NMN administration. Moreover, when arteries were incubated with NMN for 48 hours, oxidative stress was reduced due to a 50% increase in manganese superoxide dismutase and a threefold increase in NAD+ levels [69].

血管系统的衰老是衰老过程中的另一个常见现象。它伴随着自由基施加的氧化应激。在小鼠模型实验中发现 NMN 能有效地逆转这些情况。De Picciotto 和他的合作研究人员在这一特定领域进行了一项研究,以分析 NMN 的疗效[69]。用颈动脉内皮依赖性舒张功能评价血管功能,用一氧化氮介导的内皮依赖性舒张功能评价血管氧化应激。与年轻的对照组(4-8个月)相比,年长的对照组(26-28个月)在这两个参数上的功能都有所下降。雄性 C57B1/6小鼠口服 NMN 300mg/kg/d 8周后,颈动脉 EDD 和一氧化氮 EDD 均恢复到正常水平,与年轻对照组小鼠相似。同样,老年小鼠的情况,如氧化应激标记硝基酪氨酸的丰富,弹性蛋白和血管 SIRT1活性的减少,都被 NMN 治疗所逆转。此外,当动脉与中性粒细胞共同孵育48小时后,氧化应激降低是由于超氧化物歧化酶增加了50% ,而 NAD + 水平增加了3倍[69]。

The expressions of certain genes in metabolic organs like skeletal muscle, white adipose tissue, liver and immunological functions also start to decline with ageing. Mills et al., while conducting microarray assessment, found that 76.3% of 300 compromised genes of skeletal muscle, 73.1% of 360 compromised genes of white adipose tissue, and 41.7% of 513 compromised ones from liver were upregulated by NMN treatment on mice [3]. Not only this, the increased expression of immune cells from the immunometabolic system, specifically that of white adipose tissue, improvement in hematological conditions like a decrease in neutrophils and increase in lymphocyte, cytokine activity, leukocyte activation, are the outcomes of NMN treatment upon aged mice [3,70]. As stated earlier, increase in body weight and obesity-related complications like a decrease in energy metabolism and locomotor activity, age-dependent insulin insensitivity and higher triglyceride levels are also associated with ageing. These conditions were reversed by 12-month NMN intervention [3].

随着年龄的增长,骨骼肌、白色脂肪组织、肝脏和免疫功能等代谢器官中某些基因的表达也开始下降。米尔斯等人,在进行微阵列评估时,发现300个受损的骨骼肌基因中的76.3% ,360个受损的白色脂肪组织基因中的73.1% ,以及513个受损的肝脏基因中的41.7% 在小鼠上调 NMN 治疗[3]。不仅如此,增加免疫代谢系统的免疫细胞表达,特别是白色脂肪组织的免疫细胞表达,改善血液条件,如中性粒细胞减少和淋巴细胞增加,细胞因子活性,白细胞活化,都是 NMN 治疗老年小鼠的结果。如前所述,体重增加和与肥胖有关的并发症,如能量代谢和运动活动减少、年龄依赖性胰岛素不敏感性和甘油三酯水平升高,也与衰老有关。通过12个月的 NMN 干预,这些情况被逆转[3]。

The success of NMN in preclinical study instigated the researchers of Keio University School of Medicine in Tokyo and Washington University School of Medicine of St. Louis to initiate a collaborative research program for the phase I NMN clinical trial [28]. The goal of this study was to assess the safety and to find out the bioavailability of NMN in the clinical model. A positive outcome from this study could curve a new direction for the long-awaited treatment strategy of ageing.

NMN 在临床前研究中的成功促使东京庆应大学医学院和圣路易斯华盛顿大学医学院的研究人员为 i 期 NMN 临床试验启动了一个合作研究项目[28]。本研究的目的是评估 NMN 在临床模型中的安全性和生物利用度。这项研究的一个积极结果可以为期待已久的老龄化治疗策略指明一个新的方向。Go to: 去:

4. Nicotinamide Mononucleotide or Nicotinamide Riboside: Which One is Better

4. 烟酰胺单核苷酸还是烟酰胺核苷酸: 哪个更好

Among the diverse type of NAD+ precursors, until now only NMN and NR presented better pharmacokinetic and pharmacological property. Accordingly, these two intermediates are now widely utilized for clinical trials. Still, the question remains: Which one is better? Researchers in favor of both the intermediates have their supporting arguments. NR is highly available in normal human diet and its cellular permeation is simple as NR does not require any conversion to other intermediates. There are safety studies for NR administration while NMN is yet to prove the safety of human consumption. In a 12-week study of 2000 mg/day NR supplementation to assess the efficacy of improvement of glucose metabolism and insulin sensitivity in obese patients, although NR did not improve glucose metabolism and insulin sensitivity, NR treatment was safe [71]. On the other hand, NMN has quite a few strong advantages of its own. While assessing the efficiency to treat Friedreich’s Ataxia (FRDA), a rare inherited childhood heart disease, NMN was found successful where NR treatment failed [72,73]. In this disease, increased acetylation of a mitochondrial protein namely frataxin and impaired SIRT3 activity results in cardiac hypertrophy. An intervention of 500 mg/kg NMN 2 times per week for six weeks on FXN-KO mice caused improvement in diastolic function and normalized systolic function compared to that of normal saline-treated control mice. This positive effect is mediated by increasing the deacetylase activity of SIRT3 on frataxin [72], while in another study, NR at a dose of 10 mg/kg for five weeks on FXN KO mice neither improved SIRT3 activity nor cardiac function [73].

在不同类型的 NAD + 前体中,迄今为止只有 NMN 和 NR 具有较好的药代动力学和药理学性质。因此,这两种中间体现在被广泛用于临床试验。然而,问题依然存在: 哪一个更好?支持这两种中间产物的研究人员有他们的论据。NR 是高度可用于正常人的饮食,其细胞渗透是简单的,因为 NR 不需要任何转换到其他中间体。目前已有关于天然橡胶安全性的研究,而 NMN 尚未证实人类消费的安全性。在一项为期12周的2000mg/日补充 NR 的研究中,评估肥胖患者改善糖代谢和胰岛素敏感性的效果,尽管补充 NR 没有改善糖代谢和胰岛素敏感性,但补充 NR 是安全的[71]。另一方面,NMN 自身也有一些强大的优势。在评估治疗一种罕见的儿童遗传性心脏病——弗里德里希共济失调症(FRDA)的有效性时,NMN 被发现在 NR 治疗失败的情况下是成功的[72,73]。在这种疾病,增加乙酰化的线粒体蛋白质,即 frataxin 和损害 SIRT3活性导致心肌肥大。与生理盐水处理的对照组小鼠相比,500mg/kg NMN 每周2次,连续6周对 FXN-KO 小鼠的舒张功能和收缩功能均有改善。这种积极作用是通过增加 SIRT3对 frataxin [72]的去乙酰化酶活性而介导的,而在另一项研究中,10mg/kg 剂量的 NR 对 FXN KO 小鼠5周没有改善 SIRT3活性和心功能[73]。

In case of treatment of cognitive impairment in Alzheimer’s disease, for which β amyloid plaque is thought of as the main culprit, NMN was potent enough to reduce the burden of amyloid β plaque by reducing its production. During a 6-month long NR intervention at a dose of 12 mM in drinking water on 3xTgAD/Polβ+/− mice, it was observed that DNA damage, neuroinflammation, and apoptosis of hippocampal neurons were significantly decreased whereas SIRT3 activity in brain was increased leading to improvement of cognitive function, although, there was no impact on amyloid β accumulation [74]. In contrast to this, subcutaneous administration of 100 mg/kg NMN for 28 days reduced the loss of synapses, inflammation and improved neurobehavioral activities by reducing amyloid β build up in the brain. The mechanism behind this was the inhibition of amyloidogenic amyloid precursor protein (APP) and stimulation of nonamyloidogenic APP by NMN [75].

在阿尔茨海默病认知障碍的治疗中,NMN 被认为是淀粉样蛋白斑块的主要罪魁祸首,NMN 通过减少淀粉样蛋白斑块的产生,足以减轻其负担。在3xtgad/pol +/-小鼠饮水12mm 剂量的6个月 NR 干预期间,观察到海马神经元的 DNA 损伤、神经炎症和凋亡明显减少,而 SIRT3活性增加导致认知功能的改善,但对淀粉样蛋白的积累没有影响[74]。与此相反,皮下注射100毫克/千克 NMN 28天可减少突触损失、炎症,并通过减少大脑中淀粉样蛋白的积累改善神经行为活动。其机制是 NMN 抑制淀粉样蛋白前体蛋白(APP)和刺激非淀粉样蛋白前体蛋白(APP)。

It is very hard to delineate the borderline between the efficiency of NMN and NR. It is reasonable to conclude that both of them share some overlapping activities, as well as their own positive and negative impacts.

NMN 和 NR 的有效性之间的界限很难划分。可以合理地得出这样的结论,即它们都有一些相互重叠的活动,以及它们各自的积极和消极影响。Go to: 去:

5. Future Prospects

5. 未来前景

As a precursor molecule of NAD+ in our biological system, NMN can play an important role in the treatment of various disease states. NAD+ was found to stimulate cell survival in response to genotoxic stress like exposure to benz(a)anthracene, and benz(a)pyrene [76]. The reason behind cell death due to genotoxicity is the hyperactivation of NAD+ dependent mitochondrial DNA repair enzyme- PARP-1 which utilizes and thus decreases NAD+. It also causes translocation of apoptosis-inducing factor (AIF) from mitochondrial membrane to the nucleus [77,78,79]. By providing NAD+, cell survival can be significantly improved. Whether NMN can serve as a substitute for NAD+ to improve cell survival could be an important field of research.

NMN 作为 NAD + 在我们生物系统中的前体分子,在各种疾病的治疗中发挥着重要作用。人们发现 NAD + 可以刺激细胞对基因毒性应激的反应,比如接触苯(a)蒽和苯(a)芘[76]。遗传毒性导致细胞死亡的原因是 NAD + 依赖的线粒体脱氧核糖核酸修复酶-PARP-1的过度激活,这种酶能够利用并因此降低 NAD + 。它还引起凋亡诱导因子从线粒体膜向细胞核的转位。通过提供 NAD + ,细胞存活率可以显著提高。NMN 是否可以作为 NAD + 的替代物以提高细胞存活率可能是一个重要的研究领域。

This particular mechanism is also responsible for beta cell destruction leading to type 1 diabetes [80]. When beta cells of pancreatic islets are exposed to beta cell toxin, namely streptozocin or oxidative stress, PARP-1 utilizes the cellular NAD+ storage to revert DNA strand break. Consequently, ATP production and protein synthesis are reduced which triggers beta cell death [81,82]. NMN could be used to compensate the ATP shortage, thus enabling the beta cells to survive.

这种特殊的机制也是导致1型糖尿病的 β 细胞破坏的原因[80]。当胰岛 β 细胞暴露于 β 细胞毒素—- 链球菌素或氧化应激—- 时,PARP-1利用细胞 NAD + 存储来恢复 DNA 链断裂。因此,ATP 的产生和蛋白质的合成减少,从而引发 β 细胞死亡[81,82]。NMN 可以用来弥补 ATP 的不足,从而使 β 细胞存活。

Aside from the two aforementioned fields of research, the association of NAD+ with enzymes like mono-ADP-ribosyltransferases and sirtuin enzymes, which control apoptosis, DNA repair, stress resistance, metabolism, and endocrine signaling, could also be an area of interest [13]. As NAD+ metabolism can be a potential target for abnormalities related to these biological processes, NMN can be of therapeutic benefit.

除了上述两个研究领域外,NAD + 与单 adp- 核糖基转移酶和去乙酰化酶等酶的关联也可能是一个有趣的研究领域,这些酶控制细胞凋亡、 DNA 修复、抗逆性、代谢和内分泌信号传导。由于 NAD + 代谢可能是与这些生物学过程相关的异常的潜在目标,NMN 可能具有治疗作用。Go to: 去:

6. Conclusions

6. 结论

Although NMN has shown significant beneficial pharmacological activities in preclinical studies which the scientists have been searching for a long time, it still lacks sufficient clinical and toxicological data. The high manufacturing cost of NMN causes an increase in the ultimate price that creates a burden from the patients’ perspective. Despite having these drawbacks, NMN could still be a potential chemical entity, to be used as a therapeutic agent in Alzheimer’s, diabetes, cardiovascular diseases. Some capsule formulations of NMN is already available in the market. With advanced clinical studies along with the exploration of newer pharmacological applications, NMN could be an ‘all-in-one’ intervention strategy, transpiring a new era of therapeutic approach in medical science.

虽然 NMN 在临床前研究中表现出了显著的有益的药理活性,这是科学家们长期以来一直在寻找的,但它仍然缺乏足够的临床和毒理学数据。NMN 的高生产成本导致最终价格的上涨,从患者的角度来看,这造成了一种负担。尽管有这些缺点,NMN 仍然可能是一种潜在的化学实体,用于治疗阿尔茨海默氏症,糖尿病,心血管疾病。一些 NMN 胶囊制剂已经在市场上出售。随着先进的临床研究和新的药理应用的探索,NMN 可以成为一个“一体化”的干预策略,开启了医学治疗方法的新时代。Go to: 去:

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