NMN Plus – Dosage Recommendations

DO NOT TAKE MORE THAN ONE CAPSULE ON YOUR FIRST EXPERIENCE WITH NMN PLUS!

We include a very small dose of Niacin in the formula for NMN PLUS.

NIACIN  is well known to cause an uncomfortable “Niacin Flush in some people.  

We used a dosage that is about 1/2 of what  research shows affects less than 1% of users.  Read more about this, and what you can do to minimize niacin flush.

In addition to dosage, avoid taking NMN PLUS on an empty stomach to further minimize the chance you might experience a flush.

After the first trial or 2, you can increase to 2 capsules at a time if you desire.

Tolerance to Niacin Flush builds up rapidly. The few people that do experience it find it diminishes and no longer occurs after 4-7 days of continued use with a particular dosage, and can be increased further.

For those that want ONLY NMN, we offer  NMN PURE, which contains 125 mg of NMN per capsule and nothing else, but that product will not be available to ship until Dec 8, 2017.

Time of Day

Humans have a natural Circadian rhythm with a peak NAD+ levels around Noon, and a second, smaller peak in the middle of the night.

It is not yet known if supplementing with 2 doses to emulate the 2 natural peaks is beneficial, or, just one peak in the middle of the day. Experts such as Dr Brenner, Dr Sinclair, and others are on both sides of this issue.

If you chose to take NMN Plus twice a day, we recommend:

  • 1 Capsule between 8-10am
  • 1 Capsule before bedtime

If you chose to take NMN Plus once per day, we recommend

  • 1 Capsule between 8-10 am, until you are sure you don’t experience Niacin Flush
  • after acclimated for a few days

  • 2 Capsules between 8-10 am

INGREDIENTS  in NMN PLUS

In addition to NMN, which is the  IMMEDIATE PRECURSOR used by our bodies to produce NAD+, we include the other 3  major precursors:

  • NAM (Nicotinamide)
  • Tryptophan
  • NA (Niacin)

Obviously, it would be easier to produce only the single ingredient NMN PURE, but we are so convinced the proven NAD+ boosting ability of the secondary precursors make this product more effective, we have gone many months of research, development, and testing, in addition to the expense, to offer this Complete NAD+ Booster.

to read more about NMN  and other NAD+ precursors  Click here

to read more about NAD+  Click here

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Best Anti-aging supplements other than NR and NMN

22 Best Anti-Aging Supplements

Geroprotectors are substances that support healthy aging, slow aging, or extend healthy life. Sometimes people refer to them as “aging suppressants,” “anti-aging drugs,” “gerosuppressants,” “longevity therapeutics,” “senolytics,” or “senotherapeutics.” They include various foods, nutraceuticals (supplements), and pharmaceuticals (drugs). Unfortunately none comes close to realizing the age-old aspiration of ending aging altogether (yet), but some may make a practical difference for many people.

I’ve used several geroprotectors for years. And I’m exploring ways to incorporate others into my diet, if they’re applicable to my personal situation and meet a few general criteria:

First, I look for geroprotectors supported by multiple studies on humans – not just anecdotal evidence, one study, or studies on non-human animals. Although I’ve nothing against the health benefits of placebo, I prefer knowing that something more than only placebo is at work.

Second, I look for geroprotectors with the highest ratios of efficacy to expense. Given innumerable options and a limited budget, I want to do more than just empty my wallet.

Third, I look for geroprotectors that are legal and generally safe. If it’ll put me in a hospital or a prison, it’s not worth it.

Based on those criteria, I’ve compiled a list of top tier natural geroprotectors. These are, to the best of my knowledge, the most well-researched and effective geroprotectors available in the United States without a prescription. I’ve excluded from this list any geroprotectors that are primarily nootropic geroprotectors (such as ginkgo and melatonin), which you can find in my list of top tier nootropics. This information is for educational purposes only. It is not medical advice. Please consult a physician before and during use of these and other geroprotectors.

1) Berberine

Barberry

Berberine is a compound of extracts from herbs such as barberry. Supplementation may provide a strong decrease to blood glucose, and a notable decrease to total cholesterol, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Berberine may also provide a subtle increase to HDL-C; and a subtle decrease to insulin, LDL-C, and triglycerides. Evidence for these effects may not be as reliable. See the Berberine article  for more studies and details.

2) Blueberry

Blueberry

Blueberry is the fruit of a perennial flowering plant native to North America. Supplementation may provide a notable decrease to DNA damage, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

See the Blueberry article at Examine.com for more studies and details.

3) Boswellia Serrata (Frankincense)

Frankincense

Boswellia Serrata is a plant native to India and Pakistan. Supplementation may provide notable support for long-term joint function, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

See the Boswellia Serrata article at Examine.com for more studies and details.

4) Cocoa

Cocoa

Cocoa comes from the seeds of evergreen trees native to tropical regions of Central and South America. Supplementation may provide a notable increase to blood flow, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Cocoa may also provide a subtle increase to insulin sensitivity, and photoprotection; and a subtle decrease to general oxidation, platelet aggregation, and LDL-C. Evidence for these effects may not be as reliable.

5) Coenzyme Q10

Coenzyme Q10

Coenzyme Q10 is a molecule found in the mitochondria of humans and other organisms. Supplementation may provide a notable decrease to lipid peroxidation, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Coenzyme Q10 may also provide a subtle increase to blood flow, endothelial function, and exercise capacity; and a subtle decrease to blood pressure, exercise-induced oxidation, and general oxidation. Evidence for these effects may not be as reliable. See the Coenzyme Q10 article at Examine.com for more studies and details.

6) Creatine

Creatine

Creatine is a nitrogenous organic acid that occurs naturally in vertebrates. Supplementation may provide a strong increase to power output and a notable increase to hydration, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Creatine may also provide a subtle increase to anaerobic running capacity, lean mass, bone mineral density, muscular endurance, testosterone, VO2 max, and glycogen resynthesis; and a subtle decrease to blood glucose, lipid peroxidation, and muscle damage. Evidence for these effects may not be as reliable. See the Creatine article at Examine.com for more studies and details.

7) Curcumin

Turmeric

Curcumin is the bioactive in Turmeric, which is a perennial plant native to Southern Asia. Supplementation may provide a notable increase to antioxidant enzyme profile and a notable decrease to inflammation and pain, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Curcumin may also provide a subtle increase to HDL-C, and functionality in the elderly or injured; a subtle decrease to blood pressure, general oxidation, lipid peroxidation, and triglycerides; and subtle support for long-term joint function. Evidence for these effects may not be as reliable. See the Curcumin article for more studies and details.

8) DHEA (Dehydroepiandrosterone)

DHEA

DHEA is a natural hormone in humans and other animals. Supplementation may provide a notable increase to estrogen or testosterone (depending on the need of the body), according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

See the Dehydroepiandrosterone article at Examine.com for more studies and details.

9) Fish Oil

Fish

Fish Oil, as the name suggests, is an oil that accumulates in the tissues of some fish species. Supplementation may provide a strong decrease to triglycerides, thereby supporting a healthy cardiovascular system, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Fish Oil may also provide a subtle increase HDL-C, endothelial function, and photoprotection; and a subtle decrease to blood pressure, inflammation, natural killer cell activity, platelet aggregation, and LDL-C. Evidence for these effects may not be as reliable. See the Fish Oil article at Examine.com for more studies and details.

10) Garlic

Garlic

Garlic is a bulbous plant native to Central Asia. Supplementation may provide a notable increase to HDL-C and a notable decrease to LDL-C, total cholesterol, and blood pressure, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Garlic may also provide a subtle decrease to triglycerides and a strong decrease to rate of sickness. Evidence for these effects may not be as reliable. See the Garlic article at Examine.com for more studies and details.

11) Horse Chestnut (Aesculus Hippocastanum)

Horse Chestnut

Horse Chestnut is a deciduous flowering tree native to South East Europe. Supplementation may provide notable support to long-term circulatory function, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Horse Chestnut may also provide a subtle decrease to pain. Evidence for this effect may not be as reliable. See the Horse Chestnut article at Examine.com for more studies and details.

12) Magnesium

Magnesium

Magnesium is an essential dietary mineral found in food like nuts, cereals, and vegetables. Supplementation may provide a notable decrease to blood pressure (only in cases of high blood pressure), according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Magnesium may also provide a subtle increase to insulin sensitivity, aerobic exercise, and muscle oxygenation; and a subtle decrease to blood glucose, and insulin. Evidence for these effects may not be as reliable. See the Magnesium article at Examine.com for more studies and details. also check out my article on Magnesium Glycinate supplementation. Magnesium is an ingredient in Thrivous Serenity.

13) Nitrate

Beetroot

Nitrate is a molecule produced in the body in small amounts and available in vegetables like beetroot. Supplementation may provide a notable decrease to blood pressure, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Nitrate may also provide a notable increase to anaerobic running capacity; and a notable decrease to oxygenation cost of exercise. Evidence for these effects may not be as reliable.

14) Olive Leaf

Olive Leaf

Olive Leaf comes from an evergreen tree native to the Mediterranean, Africa, and Asia. Supplementation may provide a notable decrease to blood pressure and oxidation of LDL, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Olive Leaf may also provide a subtle increase to HDL-C; and a subtle decrease to LDL-C, total cholesterol, cell adhesion factors, and DNA damage. Evidence for these effects may not be as reliable. See the Olive Leaf Extract article at Examine.com for more studies and details.

15) Pycnogenol (Pine Bark)

Maritime Pine

Pycnogenol is an extract from bark of the maritime pine, native to the Mediterranean. Supplementation may provide a notable increase to blood flow, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Pycnogenol may also provide a subtle decrease to leg swelling; and subtle support for long-term joint function. Evidence for these effects may not be as reliable. See the Pycnogenol article at Examine.com for more studies and details.

16) Salacia Reticulata

Salacia Reticulata

[“Kothala Himbutu” by Satheesan.vn under CC BY-SA 3.0 / cropped]

Salacia Reticulata is a plant native to the forests of Sri Lanka. Supplementation may provide a notable decrease to blood glucose and insulin, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

See the Salacia Reticulata article at Examine.com for more studies and details.

17) SAMe (S-Adenosyl Methionine)

SAMe

SAMe is a naturally-occurring compound found in most tissues and fluids of the human body. Supplementation may provide notable support for long-term joint function, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with SAMe may also provide a subtle increase to functionality in elderly or injured; and a notable decrease to pain. Evidence for these effects may not be as reliable. See the S-Adenosyl Methionine article at Examine.com for more studies and details.

18) Spirulina

Spirulina

[“Spirulina” by Lara Torvi under CC BY 2.0 / cropped]

Spirulina is a blue-green algae. Supplementation may provide a notable decrease to lipid peroxidation and triglycerides, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Spirulina may also provide a strong decrease to allergies, nasal congestion, and liver fat; a notable increase to power output; a notable decrease to blood pressure and general oxidation; a subtle increase to HDL-C and muscular endurance; and a subtle decrease to LDL-C and total cholesterol. Evidence for these effects may not be as reliable. See the Spirulina article at Examine.com for more studies and details.

19) TUDCA (Tauroursodeoxycholic Acid)

TUDCA

TUDCA is a bile acid found naturally in trace amounts in humans and in large amounts in other animals like bears. Supplementation may provide a notable decrease to liver enzymes, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with TUDCA may also provide a notable increase to insulin sensitivity. Evidence for this effect may not be as reliable. See the Tauroursodeoxycholic Acid article at Examine.com for more studies and details.

20) Vitamin B3 (Niacin)

Niacin

Vitamin B3, also known as Niacin, is an essential dietary vitamin found in foods like liver, chicken, beef, fish, peanuts, cereals, and legumes. Supplementation may provide a strong increase to HDL-C and a notable decrease to LDL-C and triglycerides, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Vitamin B3 may also provide a subtle increase to blood glucose and insulin; and a subtle decrease to insulin sensitivity and vLDL-C. Evidence for some of these effects may not be as reliable. See the Vitamin B3 article at Examine.com for more studies and details.

21) Vitamin D

Vitamin D3

Vitamin D is an essential dietary vitamin naturally synthesized in the skin from sun exposure. Supplementation may provide a notable decrease to risk of falls, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Vitamin D may also provide a notable increase to functionality in elderly or injured; and a subtle decrease to blood pressure, bone fracture risk, and fat mass. Evidence for some of these effects may not be as reliable. See the Vitamin D article at Examine.com for more studies and details.

22) Vitamin K

Vitamin K1

Vitamin K is an essential dietary vitamin found in foods like leafy green vegetables and some fruits. Supplementation may provide a notable increase to bone mineral density, according to multiple peer-reviewed, double-blind, placebo-controlled studies in humans:

Supplementation with Vitamin K may also provide a notable decrease to bone fracture risk. Evidence for this effect may not be as reliable. See the Vitamin K article at Examine.com for more studies and details.

Nicotinamide mononucleotide inhibits JNK activation to reverse Alzheimer disease.

Study published here

Highlights

  •   NMN improved behavioral measures of cognitive impairments in AD-Tg mice.
  •   NMN decreased β-amyloid production, amyloid plaque burden, synaptic loss, and inflammatory responses in AD-Tg mice.
  •   NMN reduced JNK activation in AD-Tg mice.
  •   NMN regulated the expression of APP cleavage secretase in AD-Tg mice.

Abstract

Amyloid-β (Aβ) oligomers have been accepted as major neurotoxic agents in the therapy of Alzheimer’s disease (AD). It has been shown that the activity of nicotinamide adenine dinucleotide (NAD+) is related with the decline of Aβ toxicity in AD. Nicotinamide mononucleotide (NMN), the important precursor of NAD+, is produced during the reaction of nicotinamide phosphoribosyl transferase (Nampt). This study aimed to figure out the potential therapeutic effects of NMN and its underlying mechanisms in APPswe/PS1dE9 (AD-Tg) mice. We found that NMN gave rise to a substantial improvement in behavioral measures of cognitive impairments compared to control AD-Tg mice. In addition, NMN treatment significantly decreased β-amyloid production, amyloid plaque burden, synaptic loss, and inflammatory responses in transgenic animals. Mechanistically, NMN effectively controlled JNK activation. Furthermore, NMN potently progressed nonamyloidogenic amyloid precursor protein (APP) and suppressed amyloidogenic APP by mediating the expression of APP cleavage secretase in AD- Tg mice. Based on our findings, it was suggested that NMN substantially decreases multiple AD- associated pathological characteristically at least partially by the inhibition of JNK activation.

Introduction

As a chronic neurodegenerative disorder, Alzheimer’s disease (AD) is clinically featured by progressive pattern of cognitive deficits and memory impairment. Disturbed energy metabolism in the brain and oxidative stress are two potential factors leading to neural degeneration and cognitive impairments [1]. Aβ oligomers are found to be associated with the pathology of AD [2]. Recent studies indicates that Aβ oligomers inhibit synaptic transmission prior to neuronal cell death [3] and LTP (long-term potentiation), an experimental model for synaptic plasticity and memory [4]. In addition, Aβ oligomers are also found to be relevant to the producing of the free oxygen radical. So far, there is no curative treatment for AD [5]. Considering the varied and well-defined pathologies of AD, new therapies with the functions of reducing pathologies are needed to prevent or slow disease progression.

Nicotinamide adenine dinucleotide (NAD), oxidized (NAD+) or reduced (NADH), plays a key role in many metabolic reactions, for both forms of NAD regulate transfer of hydrogens metabolic reactions, oxidative or reductive [6], as well as mitochondrial morphological dynamics in brain [7]. Among these two forms, oxidized NAD is particularly important to mitochondrial enzyme reactions and cellular energy metabolism [8, 9]. In normal conditions, as people ages, the level of NAD+ drops [6], inhibiting cellular respiration and further causing decreased mitochondrial ATP and possibly cellular death. NAD+ serves a substrate for enzymes that depend on NAD+, such as ADP-ribosyl cyclase (CD38), poly(ADP-ribose) polymerase 1 (PARP1), and Sirtuin 1 (SIRT1) [10].

To treat neurodegenerative diseases, NAD+ depletion and cellular energy deficits need to be prevented for protecting nerves [10]. There are four pathways synthesizing NAD+ in mammals. The salvage pathway (primary route) way is to use nicotinamide, nicotinic acid, nicotinamide riboside, or the de novo pathway with tryptophan [11]. As an essential precursor of NAD+, Nicotinamide mononucleotide (NMN) is produced during the reaction of nicotinamide phosphoribosyltransferase (Nampt). Nampt is essential to regulating NAD+ synthesis [12], for it stimulates phosphoribosyl components to separate from phosphoribosyl pyrophosphates and to combine with nicotinamides. In this way, NMN is generated and with NMN adenylyltransferase, NMN is converted to NAD+. However, the potential therapeutic effects of NMN on AD remain unclear.

c-Jun N-terminal kinases (JNKs) are a family of protein kinases that play a central role in stress signaling pathways implicated in gene expression, neuronal plasticity, regeneration, cell death, and regulation of cellular senescence [13]. Activation of JNK has been identified as a key element responsible for the regulation of apoptosis signals and therefore, it is critical for pathological cell death associated with neurodegenerative diseases and, among them, with Alzheimer’s disease (AD) [14].

As suggested, NAD+ may be essential to brain metabolism and might influence memory and learning. According to recent studies, the stimulation of NAD level is relevant to the reduced amyloid toxicity in AD animal models [15]. Therefore, in this study, the potential therapeutic effects of NMN and the mechanisms of its action regulated in JNK in APPswe/PS1dE9 mice with AD were investigated.

Materials and Methods

Animals

The Institutional Animal Experiment Committee of Tongji University, China, approved all procedures conforming to the Animals’ Use and Care Policies. APPswe/PS1dE9 transgenic mice (6 months old) were purchased from Beijing Bio-technology, China. All animals were maintained in an environment that was pathogen-free. During the experimental period, water and food were accessible to all mice, and the body weight of mice and the intake of food and water were identified at the beginning of the study and then on a weekly basis. In addition, all mice that receive the treatment were observed for their general health. APPswe/PS1dE9 transgenic mice (AD-Tg) and their nontransgenic wild-type mice (NTG) were randomly assigned into four groups with six mice in each group, and each type was treated by NMN and vehicle, respectively Subcutaneous adiministration of NMN (100 mg/kg, Sigma N3501) in sterile (Phosphate Buffered Saline) PBS (200 μl) was applied to each mouse of NMN-treated groups every other day for 28 days. Each mouse with vehicle treatment subcutaneously received sterile PBS (200 μl) every other day for 28 days.

Behavioral Tests

Behavioral tests were carried out by 2 experimenters who were blinded to the treatments twelve weeks after the treatments.

Memory and spatial learning test

To evaluate the memory and spatial learning of all animals, a Morris water navigation task was performed as described previously [16]. Generally, a tracking system (Water 2020; HVS Image, Hampton, UK) was utilized to monitor the trajectory of all mice. During the training trials, a platform with the diameter of 5cm was hidden 1.5 cm below the surface of water and maintained at the same quadrant. In every trial, all mice had at most 1 minute to find the hidden platform and climb onto it. If one mouse cannot find the platform within 1 minute, the experimenters would manually guide the mouse to the platform and kept it there for 10 seconds. The trial was carried out 4 times daily for 6 days. The escape latency referring to the time that a mouse spent in finding the platform is considered as spatial learning score. Following the last training trial, the probe trial was carried out for spatial memories by allowing animals to take a free swim in the pool with the platform removed for 1 minute (swim speeds are equal). The time that each animal took to reach the previously platform-contained quadrant was measured for spatial memories.

Measurement of Passive Avoidance

To assess contextual memories, passive avoidance test was carried out, which was described in the previous studies [17]. Briefly speaking, a two-compartment apparatus with one brightly lit and one dimly lit was used. During the training trial, the animal was put into the light lit compartment. After 60 seconds, the door between the two compartments was opened. The acquisition latency refers to the first latency time of mice to ran into the dimly lit compartment. After coming into the lit compartment, mice were exposed to a mild foot shock (0.3mA) for 3 seconds with the door closed. After 5 seconds, the animals were taken out of the compartment. One day later after the acquisition trial, the mice underwent a retention test. Like in the acquisition test, the latency to go into the dark compartment without foot shock was regarded as retention latency to test retention memory. Longer latency indicates better retention.

Tissue Preparation

Following the two behavioral tests, 24 mice were first anesthetized and then infused with icy normal saline in a transcardial way. The brains were taken out and cut into 2 hemibrains along the midsagittal plane. One of the hemispheres was kept in PBS with 4% paraformaldehyde. Following the xylene treatment, the other fixed hemisphere was maintained in the paraffin for immunohistochemical tests. Then the cerebral cortex and the hippocampus were separated quickly from the hemisphere on the ice. For biochemical tests, they were maintained at −80°C following the separation. The hippocampus, brain cortex, and as well as the whole brain were weighed, respectively.

Immunohistochemistry

Immunohistochemical staining was carried out as described [16]. Briefly speaking, 10 μm brain slices were deparaffinized and rehydrated. To retrieve antigens, proteinase K (200μg/ml) was treated for the staining of Aβ, and sodium citrate (0.01M, pH 6.0) was for the staining of microglia and astrocyte. Sections were blocked through incubation with fetal bovine serum (2%) and Triton X-100 (0.1%) for nonspecific binding. For immunohistochemical analysis, the section was incubated at 4°C for a night with anti-Aβ1-16 monoclonal antibody (1:600; Cell Signaling Technology, Massachusetts, USA) and monoclonal antibody anti-Iba1 (1:1,000; Osaka Wako Pure Chemical Industries, Japan) for rabbits and also monoclonal anti-Aβ antibody (1:200; Billerica, MA) for mouse.

Olympus (Tokyo, Japan) microscope with a connection to a digital microscope camera was applied to capture the images for quantitative analyses. The plaques in μm2s and the proportion of area kept by plaques positive to Aβ1–16 respectively, microglia positive to Iba1 were obtained with imaging software (Bethesda Media Cybernetics, MD). The mean value of every parameter was obtained from 6 sections with an equidistant interval of 150μm through the hippocampal region of each mouse in all groups. All measurements were blindedly conducted.

Enzyme-Linked Immunosorbent Assay (ELISA)

As described before, soluble Aβ fractions and insoluble ones were obtained from both the cortex and hippocampi of brain homogenates of mouse using RIPA (Radioimmunoprecipitation assay buffer) buffer and formic acid, respectively [18]. The levels of both the insoluble and soluble Aβ were identified using the ELISA kits (Camarillo Invitrogen, CA). Besides, concentrations of oligomeric Aβ of brain homogenates treated with RIPA were obtained employing an ELISA kit for amyloid β oligomer (Gunma Immuno-Biochemical Laboratories, Japan).

Proinflammatory Cytokines Measurement

As described, mouse brain proinflammatory cytokine was evaluated [19]. The expressions of TNFα, IL-6, and IL-1β were identified with immunoassay kits (Minneapolis R&D Systems, Minnesota, USA) which is for measuring these factors in mouse.

Western blotting (WB) analysis

The cortex and hippocampus tissue was homogenized with icy PBS and the lysate was for Western blot. At first SDS-PAGE (Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis) was applied to divide the proteins on NuPage Bis-Tris gel (12%, Invitrogen). The separated protein was subsequently transferred to nitrocellulose membrane which was blocked with 5% nonfat milk and probed overnight at 4°C with anti-p-JNK, anti-JNK, anti-APP, anti-sAPPα, anti- sAPPβ, anti-phosphorylated APP (p-APP, Thr668), anti-ADAM10, anti-BACE1 (CA Santa

Cruz Biotechnology, USA), anti-CDK5, anti–p-CDK5, anti–p-GSK3β, anti-GSK3β, anti-SYP, anti–postsynaptic density-95, anti–β-actin Abcam (Cambridge, MA, USA). The membrane was cleaned with TBS/0.05% Tween-20 and incubated at room temperature with secondary antibodies conjugated with horseradish peroxidase for 60 minutes, following incubation with primary antibodies. Enhanced chemiluminescence reagents (Pierce, Rockford, IL, USA) were used for detecting signals.

Statistical Analysis

The data were expressed as mean ± SD. The comparisons in the speed of swimming and escape latency between the groups during the test of memory and spatial learning were made employing two-way ANOVA with repeated measures. Next, post hoc least significant difference (LSD) test was used for multiple comparison. Before post hoc LSD test or Student t test, 1-way ANOVA was employed for the rest data. Statistical analyses were carried out with Prism version 5. A P < 0.05 was considered statistically significant.

Results

NMN Treatment Rescues Cognitive impairments in AD-Tg Mice

The test of memory and spatial learning has shown that 1-year-old AD-Tg mice have experienced impairment in memory and spatial learning [16]. In our study, comparied to the vehicle-/NMN-treated wild-type (WT) mice, the vehicle-treated AD-Tg mice had a longer escape latency, which showed severe impairment of spatial learning in the test (Fig. 1A). However, with shorter escape latency, NMN treatment greatly improved the impairment of spatial learning in vehicle-treated AD-Tg animals (Fig. 1A). Besides, it was also identified that compared with two wild-type groups, vehicle-treated AD-Tg animals spent less time in the target quadrant during the probe trial (p < 0.01), suggesting severe spatial memory impairment. AD-Tg mice treated with NMN spent longer time in the target quadrant, which indicates marked alleviation of the spatial learning impairments present in AD-Tg mice treated with vehicle (p<0.01) (Fig. 1B).

To further identify the alleviation of memory deficits by NMN treatment in AD-Tg mouse, contextual memories were evaluated employing the measurement of passive avoidance [17]. As illustrated in Fig. 1C, retention latency was decreased compared with two wild-type mice groups (p < 0.01), suggesting impaired contextual memories in the AD-Tg animals treated by vehicle. In contrast, NMN-treated mice exhibited longer retention latency compared with those treated with vehicles (p < 0.01), demonstrating outstanding reversal of NMN in contextual memories. All these data indicate that NMN treatment markedly improves cognitive impairments in AD-Tg animals.

NMN Suppresses JNK Phosphorylation in AD-Tg Mice

JNK, also called a protein kinase activated by stress, is said to play a role in a couple of pathophysiological processes in AD [13]. Therefore, in this study, we tested the inhibitory effects of NMN on the activation of JNK through Western blotting. It was revealed by quantitative analysis that p-JNK level was significantly grown in hippocampus and cerebral cortex in the vehicle-treated AD-Tg mice when contrasting to two wild-type groups (Fig. 2A; p < 0.01), whereas NMN gave rise to a sharp decline in p-JNK in hippocampus and cerebral cortex with a comparison to the vehicle-treated AD-Tg mice (Fig. 2B; P< 0.01). Both reductions symbolized a reverse to the wild-type level. But the whole expression of JNK kept unchanged in all the 4 groups. Conclusively, all data indicate that NMN treatment has an inhibitory effects on JNK activation in AD-Tg mice.

NMN Treatment Decreases the Level of Aβ and Deposition in AD-Tg Mice

The role of reduced activation of JNK in the changes of the Aβ level and deposition was studied in AD-Tg mice through employing histological and biochemical analyses. As presented in Figs. 3A-D, it was found that NMN-treated AD-Tg mice had a sharp reduction in the levels of Aβ when comparing to the vehicle treatment group (p < 0.01). Comparing to the vehicle treatment group, NMN treatment gave rise to a marked decrease in Aβ oligomers (p < 0.01) (Fig. 3E). Immunohistochemical staining identified this observation, indicating the lessened diffuse plaques and also the shrinked area taken by diffuse plaques in AD-Tg mice treated by NMN compared to the vehicle treatment group (Figs. 4A-D). Thus, on the basis of the findings, it was demonstrated that the generation of Aβ in the brain of AD-Tg mice is effectively decreased by the inhibited activation of JNK with NMN treatment.

NMN Treatment Changes the Processing of APP in AD-Tg Mice

To study the mechanism of inhibition on the production of Aβ and deposition, the effects of NMN on the processing of APP were examined by Western blotting. As presented in Figs. 5A-C, the level of full-length APP expression was greatly increased in the brain of AD-Tg mouse treated with vehicle compared with wild-type ones (p < 0.01). However, they kept unaltered between the group treated with vehicle and that with NMN. Importantly, it was found that NMN treatment remarkably lowered the increased levels of p-APP in the AD-Tg mice treated by vehicle (p < 0.01). Besides, α-secretase cleaved sAPPα and β-secretase cleaved sAPPβ in the brain tissues of Tg mice were tested via Western blotting. It was shown by quantitative analyses that NMN treatment led to a remarkable elevation of sAPPα (p < 0.01) and a marked decline in sAPPβ (p < 0.01) compared with the transgenic mice treated by vehicle (Figs. 5D-F). Based on these data, it was indicated that NMN treatment is strongly effective in suppressing the phosphorylation of APP, improving cleaving of APP by α-secretase, and decreasing the cleaving of APP by β-secretase in AD-Tg mice brains.

NMN Treatment Improves Inflammatory Responses in AD-Tg Mice

Since JNK activation is indicated to play a role in the inflammatory response induced by Aβ in previous studies [20], whether reduced activation of JNK influences neural inflammation in AD- Tg animals was investigated. The role of NMN on the neural inflammation was identified by measuring proinflammatory cytokines that were in the lysates of cortical tissues. It was found that the level of IL-6, IL-1β, and TNFα were sharply declined in the AD-Tg mice treated by NMN relative to those by vehicle (Figs. 6A-C). According to these findings, NMN treatment is indicated to be potently effective in the amelioration of neural inflammation in AD-Tg mice brains.

NMN Treatment Ameliorates Synaptic Loss in AD-Tg Mice

The loss of synapse is an important pathological characteristic of AD and said to be relevant to the cognitive impairments of AD [21]. The changes in SYP (presynaptic marker) level and PSD- 95 level (postsynaptic marker) were investigated via Western blotting. It was showed by quantitative analysis that SYP levels and the levels of PSD-95 expression substantially reduced in hippocampus and brain cortex of AD-Tg mice treated with vehicle relative to WT ones

(p < 0.01), whereas NMN treatment significantly elevated SYP levels and the levels of PSD-95 expression in hippocampus and brain cortex relative to AD-Tg mice treated with vehicle (p < 0.01) (Fig. 6D-F). This finding suggest that NMN treatment sharply ameliorates the loss of synapse in AD-Tg mice brains.

Discussion

In the present study, it is mainly found that NMN treatment substantially improves primary pathological characteristics of the AD-modeled AD-Tg mice, including cognitive impairments, neuroinflammation, Aβ pathology, and synaptic loss, which consistent with a recent study [22]. It was also found that NMN treatment inhibited JNK activation and amyloidogenic processing of APP by mediating the expression of APP-cleavage secretase, and also facilitated APP processing in AD-Tg mice. The data prove that NMN treatment greatly reduces multiple AD-associated pathological characteristics, at least partially by the inhibition of JNK activation.

Numerous studies have reported the increase of abnormal activation of JNK in both the transgenic AD mice models and the AD patients [23-25]. Conforming to the above previous studies, we also found that the level of phosphorylated JNK in AD-Tg mice treated by vehicle was higher than that in the wild-type group, but NMN treatment in AD-Tg mice potently suppressed the phosphorylation of JNK to the basic level of WT groups. The controlled activation of JNK through NMN gave rise to a substantial decrease of Aβ pathology in AD-Tg animals. According to the studies before, active JNK is proved to engage in BACE1 expressions and PS1 expressions [26, 27]. In addition, the increased BACE1 and PS1 in AD-Tg mice treated by vehicle were found to be greatly suppressed by NMN to the basic level of WT groups (data not shown). More interestingly, it was also observed that NMN treatment led to substantially elevated sAPPα and reduced sAPPβ. It was notable that according to the previous studies, APP phosphorylation at the site of Thr668 is proved to promote the β-secretase cleavage of APP to grow Aβ generation in vitro [28]. In present study, we also found that the administration of NMN in AD-Tg mice significantly declined the elevated phosphorylation of APP to the primary level of WT controls, indicating an in vivo inhibition mechanism of Aβ pathology through NMN treatment. Collectively, all these findings indicate that the potent effects of NMN on the marked decrease in Aβ pathology in the brains of AD-Tg mice may be responsible for its enhancement of nonamyloidogenic APP processing. What we found is consistent to a recent study demonstrating that genetic depletion of JNK3 in 5XFAD mice is attributed to a significant decrease in the levels of Aβ and the total plaque loads [29]. Recently numerous studies suggested energy failure and accumulative intracellular waste also play a causal role in the pathogenesis of several neurodegenerative disorders and Alzheimer’s disease (AD) in particular regulated by potential role of several metabolic pathways Wnt signaling, 5′ adenosine monophosphate-activated protein kinase (AMPK), mammalian target of rapamycin (mTOR), Sirtuin 1 (Sirt1, silent mating-type information regulator 2 homolog 1), and peroxisome proliferator-activated receptor gamma co- activator 1-α (PGC-1α) [30, 31]. It will be warrant to study if NMN also participate in regulation of these signaling pathways.

Some recent studies indicate that enhanced neuroinflammation is essential to the development of AD [32-34]. In our study, a marked decline in the proinflammatory cytokines levels (IL-6, IL-1β, and TNFα) proved that NMN treatment effectively controlled the neuroinflammatory responses of the brain of AD-Tg mouse. Considering the key role of oligomeric and fibrillar Aβ for activation of microglia cells and astrocytes with the subsequent generation of proinflammatory cytokines [34], the reduction in neuroinflammatory responses may be less important to the substantial reduction in Aβ pathologies presented in the AD-Tg mice treated by NMN. Several previous studies had proved that JNK represents an important mediator for activation of glial cell and proinflammatory cytokines [35, 36]. Thus, the favorable effects of NMN on lowered inflammatory responses in AD-Tg groups can be largely responsible for its direct control of inflammation by inhibiting JNK activation. Based on the previous reports, it was proved that some proinflammatory cytokines (ie, IL-1β, interferon gamma, and TNFα) may elevate the expression of β-secretase and γ-secretase to ameliorate amyloidogenic APP processing and Amyloid-β production by an in vitro JNK-mediated pathway [33]. Hence, we have reasons to believe that the reduced proinflammatory cytokines through NMN treatment may be effective in reducing the production of Aβ in vivo. Moreover, in our study, it was demonstrated that NMN treatment ameliorates cognitive impairments in AD-Tg mouse models. An increasing evidence has proved that grown Aβ levels, neuroinflammation, synaptic dysfunction and loss are closely related to the cognitive dysfunction in AD [37]. In addition, our data confirms the finding of a recent research, which revealed that genetic down-regulation of JNK3 gives rise to a remarkable amelioration of cognitive impairments in 5XFAD mice [29]. Collectively, our findings, along with all the previous research, demonstrate that the inhibited JNK activaty by NMN is potently effective in ameliorating AD-associated cognitive deficits.

Synaptic loss is a major pathological change of AD and is tightly associated with AD-related cognitive impairments [37]. It was presented that PSD-95, a biomarker of postsynaptic density, is essential to synapse maturation and synaptic plasticity [38], and that SYP, a presynaptic protein, also acts as an integral membrane protein in the synapse and it plays a key role in plasticity of synapses [39]. Therefore, it can be soundly supposed that the greatly lowered expression of PSD- 95 and SYN presented in the study may suggest the impairment of synaptic integrity and  plasticity in AD-Tg mice treated by vehicle. Intriguingly, the treatment of NMN in AD-Tg animals substantially elevated the lowered PSD-95 and SYN expression level back to the primary level of WT controls. Since it was demonstrated by several studies that Amyloid-β- induced synaptic loss and dysfunction are regulated through the JNK activation [40, 41], the possible mechanisms behind NMN treatment leading to the elevated expression of PSD-95 and SYN in AD-Tg animals may be responsible for its inhibitory effects on JNK activation. Thus, it is possible that the treatment of NMN may ameliorate the impaired synaptic plasticity which is caused by toxic Aβ species in AD-Tg mice.

In summary, this study provides essential preclinical evidences that NMN takes effects in reversing cognitive deficits and substantially lowering the burden of amyloid plaque, neuroinflammation, cerebral amyloid-β concentrations, and loss of synapse in middle-aged AD- Tg mice, at least partially by the inhibition of JNK activation. According to our findings, NMN could be a new target for disease-modifying treatments of AD.

References

  1. Kapogiannis, D. and M.P. Mattson, Disrupted energy metabolism and neuronal circuit dysfunction in cognitive impairment and Alzheimer’s disease. Lancet Neurol, 2011. 10(2): p. 187-98.
  2. Gong, Y., et al., Alzheimer’s disease-affected brain: presence of oligomeric A beta ligands (ADDLs) suggests a molecular basis for reversible memory loss. Proc Natl Acad Sci U S A, 2003. 100(18): p. 10417-22.
  3. Hardy, J. and D.J. Selkoe, The amyloid hypothesis of Alzheimer’s disease: progress and problems on the road to therapeutics. Science, 2002. 297(5580): p. 353-6.
  4. Lesne, S., et al., A specific amyloid-beta protein assembly in the brain impairs memory. Nature, 2006. 440(7082): p. 352-7.
  5. Tayeb, H.O., et al., Pharmacotherapies for Alzheimer’s disease: beyond cholinesterase inhibitors. Pharmacol Ther, 2012. 134(1): p. 8-25.
  6. Imai, S. and L. Guarente, NAD+ and sirtuins in aging and disease. Trends Cell Biol, 2014. 24(8): p. 464-71.
  7. Long, A.N., et al., Effect of nicotinamide mononucleotide on brain mitochondrial respiratory deficits in an Alzheimer’s disease-relevant murine model. BMC Neurol, 2015. 15: p. 19.
  8. Kristian, T., et al., Mitochondrial dysfunction and nicotinamide dinucleotide catabolism as mechanisms of cell death and promising targets for neuroprotection. J Neurosci Res, 2011. 89(12): p. 1946-55.
  9. Owens, K., et al., Mitochondrial dysfunction and NAD(+) metabolism alterations in the pathophysiology of acute brain injury. Transl Stroke Res, 2013. 4(6): p. 618-34.
  10. Liu, D., M. Pitta, and M.P. Mattson, Preventing NAD(+) depletion protects neurons against excitotoxicity: bioenergetic effects of mild mitochondrial uncoupling and caloric restriction. Ann N Y Acad Sci, 2008. 1147: p. 275-82.
  11. Yamamoto, T., et al., Nicotinamide mononucleotide, an intermediate of NAD+ synthesis, protects the heart from ischemia and reperfusion. PLoS One, 2014. 9(6): p. e98972.
  12. Imai, S., Dissecting systemic control of metabolism and aging in the NAD World: the importance of SIRT1 and NAMPT-mediated NAD biosynthesis. FEBS Lett, 2011. 585(11): p. 1657-62.
  13. Mehan, S., et al., JNK: a stress-activated protein kinase therapeutic strategies and involvement in Alzheimer’s and various neurodegenerative abnormalities. J Mol Neurosci, 2011. 43(3): p. 376-90.
  14. Yarza, R., et al., c-Jun N-terminal Kinase (JNK) Signaling as a Therapeutic Target for Alzheimer’s Disease. Front Pharmacol, 2015. 6: p. 321.
  15. Kim, D., et al., SIRT1 deacetylase protects against neurodegeneration in models for Alzheimer’s disease and amyotrophic lateral sclerosis. EMBO J, 2007. 26(13): p. 3169- 79.
  16. Zhang, W., et al., Multiple inflammatory pathways are involved in the development and progression of cognitive deficits in APPswe/PS1dE9 mice. Neurobiol Aging, 2012. 33(11): p. 2661-77.
  1. Ishrat, T., et al., Amelioration of cognitive deficits and neurodegeneration by curcumin in rat model of sporadic dementia of Alzheimer’s type (SDAT). Eur Neuropsychopharmacol, 2009. 19(9): p. 636-47.
  2. Li, J.G., et al., Homocysteine exacerbates beta-amyloid pathology, tau pathology, and cognitive deficit in a mouse model of Alzheimer disease with plaques and tangles. Ann Neurol, 2014. 75(6): p. 851-63.
  3. Chu, J. and D. Pratico, Involvement of 5-lipoxygenase activating protein in the amyloidotic phenotype of an Alzheimer’s disease mouse model. J Neuroinflammation, 2012. 9: p. 127.
  4. Vukic, V., et al., Expression of inflammatory genes induced by beta-amyloid peptides in human brain endothelial cells and in Alzheimer’s brain is mediated by the JNK-AP1 signaling pathway. Neurobiol Dis, 2009. 34(1): p. 95-106.
  5. Querfurth, H.W. and F.M. LaFerla, Alzheimer’s disease. N Engl J Med, 2010. 362(4): p. 329-44.
  6. Wang, X., et al., Exendin-4 antagonizes Abeta1-42-induced suppression of long-term potentiation by regulating intracellular calcium homeostasis in rat hippocampal neurons. Brain Res, 2015. 1627: p. 101-8.
  7. Braithwaite, S.P., et al., Inhibition of c-Jun kinase provides neuroprotection in a model of Alzheimer’s disease. Neurobiol Dis, 2010. 39(3): p. 311-7.
  8. Sclip, A., et al., c-Jun N-terminal kinase regulates soluble Abeta oligomers and cognitive impairment in AD mouse model. J Biol Chem, 2011. 286(51): p. 43871-80.
  9. Thakur, A., et al., c-Jun phosphorylation in Alzheimer disease. J Neurosci Res, 2007. 85(8): p. 1668-73.
  10. Guglielmotto, M., et al., Amyloid-beta(4)(2) activates the expression of BACE1 through the JNK pathway. J Alzheimers Dis, 2011. 27(4): p. 871-83.
  11. Rahman, M., et al., Intraperitoneal injection of JNK-specific inhibitor SP600125 inhibits the expression of presenilin-1 and Notch signaling in mouse brain without induction of apoptosis. Brain Res, 2012. 1448: p. 117-28.
  12. Colombo, A., et al., JNK regulates APP cleavage and degradation in a model of Alzheimer’s disease. Neurobiol Dis, 2009. 33(3): p. 518-25.
  13. Yoon, S.O., et al., JNK3 perpetuates metabolic stress induced by Abeta peptides. Neuron, 2012. 75(5): p. 824-37.
  14. Godoy, J.A., et al., Signaling pathway cross talk in Alzheimer’s disease. Cell Commun Signal, 2014. 12: p. 23.
  15. Killick, R., et al., Clusterin regulates beta-amyloid toxicity via Dickkopf-1-driven induction of the wnt-PCP-JNK pathway. Mol Psychiatry, 2014. 19(1): p. 88-98.
  16. Hong, H.S., et al., Interferon gamma stimulates beta-secretase expression and sAPPbeta production in astrocytes. Biochem Biophys Res Commun, 2003. 307(4): p. 922-7.
  17. Liao, Y.F., et al., Tumor necrosis factor-alpha, interleukin-1beta, and interferon-gammastimulate gamma-secretase-mediated cleavage of amyloid precursor protein through aJNK-dependent MAPK pathway. J Biol Chem, 2004. 279(47): p. 49523-32.
  18. Morales, I., et al., Neuroinflammation in the pathogenesis of Alzheimer’s disease. A rational framework for the search of novel therapeutic approaches. Front Cell Neurosci,2014. 8: p. 112.
  19. Kim, S.H., C.J. Smith, and L.J. Van Eldik, Importance of MAPK pathways for microglialpro-inflammatory cytokine IL-1 beta production. Neurobiol Aging, 2004. 25(4): p. 431-9.
  1. Waetzig, V., et al., c-Jun N-terminal kinases (JNKs) mediate pro-inflammatory actions of microglia. Glia, 2005. 50(3): p. 235-46.
  2. Marcello, E., et al., Synaptic dysfunction in Alzheimer’s disease. Adv Exp Med Biol, 2012. 970: p. 573-601.
  3. El-Husseini, A.E., et al., PSD-95 involvement in maturation of excitatory synapses. Science, 2000. 290(5495): p. 1364-8.
  4. Janz, R., et al., Essential roles in synaptic plasticity for synaptogyrin I and synaptophysin I. Neuron, 1999. 24(3): p. 687-700.
  5. Costello, D.A. and C.E. Herron, The role of c-Jun N-terminal kinase in the A beta- mediated impairment of LTP and regulation of synaptic transmission in the hippocampus. Neuropharmacology, 2004. 46(5): p. 655-62.
  6. Sclip, A., et al., c-Jun N-terminal kinase has a key role in Alzheimer disease synaptic dysfunction in vivo. Cell Death Dis, 2014. 5: p. e1019

Can NMN really reverse Aging? (backup)

As we age, our levels of the Co-enzyme Nicotinamide Adenine Dinucleotide NAD+ drop significantly in multiple organs in mice and humans  (5,8,10).

NAD+ decrease is described as a trigger in age-associated decline(23), and perhaps even the key factor in why we age (5).

In 2013, research published by Dr David Sinclair demonstranted that short term supplementation with Nicotinamide MonoNucleotide (NMN) reversed many aspects of aging, making the cells of old mice resemble those of much younger mice, and greatly improving their health (8).

 

The quotes below are directly from that research:

NMN was able to mitigate most age-associated physiological declines in mice”

“treatment of old mice with NMN reversed all of these biochemical aspects of aging”

Since that landmark 2013 study, dozens of others have been published investigating the efficacy of supplementation with NMN in treatment and prevention of a wide range of disease including cancer, cardiovascular disease, diabetes, Alzheimers, Parkinsons, and more (5,6,7,9,10,11,13,14,15,16).

According to Dr Sinclair:

enhancing NAD+ biosynthesis by using NAD+ intermediates, such as NMN and NR, is expected to ameliorate age-associated physiological decline

WHAT IS NAD+

NR benefits chartNAD+ is a key co-enzyme that the mitochondria in every cell of our bodies depend on to fuel all basic functions. (3,4)

NAD+ play a key role in communicating between our cells nucleus and the Mitochondria that power all activity in our cells (5,6,7)

NAD+ LEVELS DECREASE WITH AGE

NAD+ levels decreaseAs we age, our bodies produce less NAD+ and the communication between the Mitochondria and cell nucleus is impaired. (5,8,10).

Over time,  decreasing NAD+ impairs the cell’s ability to make energy, which leads to aging and disease (8, 5) and perhaps even the key factor in why we age (5).

NAD+ METABOLISM IN HUMANS


NAD+ can be synthesized in humans from several different molecules (precursors), thru 2 distinct pathways:
De Novo Pathway

  • Tryptophan
  • Nicotinic Acid (NA)

Salvage Pathway

  • NAM – Nicotinamide
  • NR – Nicotinamide Riboside
  • NMN – Nicotinamide MonoNucleotide

The NAD+ supply is constantly being consumed and replenished through the Salvage Pathway, with approximately 3g of NAM metabolized to NMN and then to NAD 2-4 times per day (14).

  • The salvage pathway sustains 85% or more of our NAD+ (14)
  • Nampt is the rate-limiting step in the salvage process (97).
  • As we age, Nampt enzyme activity is lower, resulting in less NAM recycling, less NAD+, more disease and aging (97,101).

ALL PRECURSORS BOOST NAD+ SIGNIFICANTLY IN LIVER

NAM, NA, NMN, NR, and Tryptophan ALL elevate levels of NAD+ significantly in the liver, which has many benefits for metabolic health.

This chart from the Trammell thesis shows the impact on liver NAD+ for mice given NR, NAM, and NA by oral gavage 0.25, 1, 2, 4, 6, 8 and 12 hours before testing.

Charts showing NMN impact on NAD+ levels in the liver are below.

* Note:  These charts are somewhat deceptive. It shows NAM (green bar) elevated NAD+ nearly as much as NR (black bar)

However if they used equal mg of each supplement, which is how people actually purchase and use them, it would show NA about equal with NR and NAM far effective than NR at elevating NAD+ in the liver.

Mice in these experiments didn’t receive equal WEIGHTS of each precursor. Instead researchers chose to use quantity of molecules, which makes NR look “better” by comparison.

In this case, “185 mg kg−1 of NR or the mole equivalent doses of Nam and NA”(16).

Molecular weight for NR is 255 grams, NAM is 122 grams, and NA 123 grams.  So this chart used a ratio of  255 grams of NR to 122 and 123 grams of NAM and NA.

NMN

  • “NMN makes its way through the liver, into muscle, and is metabolized to NAD+ in 30 minutes” (R)

NR

  • Is much slower, taking 8 hours to reach peak NAD+ in humans (R)

NAM

  • Has very similar NAD+ profile to NR, taking 8 hours to reach peak NAD+ in humans (R)
  • Has been shown to increase NAD+ level in liver (47%), but was weaker in kidney (2%), heart (20%), blood (43%) or lungs (17%) (R)

NA (Niacin)

  • Elevates NAD+ to peak levels in liver in 15 minutes (R)
  • raised NAD+ in liver (47%), and impressively raised kidney (88%), heart (62%), blood (43%) and lungs (11%) (R)

TRYPTOPHAN

  • In the liver  tryptophan is the preferable substrate for NAD+ production (R)
  • Administration of tryptophan, NA, or NAM to rats showed that tryptophan resulted in the highest hepatic NAD+ concentrations(R)

ONLY NMN BYPASSES THE NAMPT BOTTLENECK IN TISSUES THROUGHOUT THE BODY

Restoring NAD+ to youthful levels in ALL CELLS throughout the body is the goal.

However, many tissues cannot utilize NAD+ directly from the blood as NAD+ cannot readily pass through the cellular membrane.

Muscle tissue, for example, depends on cells internal recycling of NAD+ through the salvage pathway which is controlled by Nampt.

To restore depleted NAD+ levels in such cells, a precursor must:

  • Be available in the bloodstream
  • Once inside a cell, be able to bypass the Nampt bottleneck

NA and Tryptophan
NA and Tryptophan act through the De Novo pathway, which supplies a small percentage of our NAD+, primarily in the liver

NAM
NAM is abundant in the blood and easily carried into such cells throughout the body, but  depends on Nampt, which is the rate limiting enzyme in the salvage pathway.

NR
When taken orally as a supplement, most NR does not make it through the digestive system intact, but is broken down to NAM (97,98,99).

For more info on how NR is converted to NAM in the body.

NR can bypass the Nampt bottleneck, but is not normally available in the bloodstream

After oral NMN supplementation, levels of NMN in the bloodstream are quickly elevated and remain high longer than NAM, NA, or NR (18,22,97,98,99)

Oral NMN supplements:

  • Make their way intact thru the digestive system (22)
  • Quickly elevates levels of NMN in the bloodstream for use throughout the body (22)
  • Quickly elevates levels of  NMN in tissues throughout the body (22)
  • Quickly raises levels of NAD+ in blood, liver and tissues  through the body (22,23)
  • Remain elevated longer than NAM, NA, or NR (18)

Only NMN is readily available in the bloodstream to all tissues, and bypasses the Nampt bottleneck in the Salvage pathway

ORAL NMN IS READILY AVAILABLE THROUGHOUT THE BODY

The chart at right shows levels of a double labeled NAD+ (C13-d-nad+) in liver and soleus muscle at 10 and 30 minutes after oral administration of double labeled NMN.

This clearly shows that NMN makes it way through the liver intact, through the bloodstream, into muscle, and is metabolized to NAD+ in 30 minutes (22) .

This quote below is directly from that study.

Orally administered NMN is quickly absorbed, efficiently transported into blood circulation, and immediately converted to NAD+in major metabolic tissues (22).

NMN QUICKLY RAISES NAD+ IN LIVER AND BLOOD

mouse-single-dose
In this 2016 study, mice were given a single dose of  NMN in water.

NMN  levels in blood showed it is quickly absorbed from the gut into blood circulation within 2–3 min and then cleared from blood circulation into tissues within 15 min

 

 

 

NMN INCREASES NAD+ and SIRT1 DRAMATICALLY IN ORGANS

In this 2017 study, NMN supplementation for 4 days significantly elevated NAD+ and SIRT1, which protected the mice from Kidney damage.

NAD+ and SIRT1 levels were HIGHER in OLD Mice than in YOUNG Mice that did not receive NMN.

LONG TERM SUPPLEMENTATION WITH NMN

mouse-long-term-research

In a long-term experiment documented in the 2016 study (22) , mice were given 2 different doses of NMN over 12 months.

Testing revealed that NMN  prevents some aspects of  physiological decline in mice, noting these changes:

  • Decreased body weight and fat
  • Increased lean muscle mass
  • Increased energy and mobility
  • Improved visual acuity
  • Improved bone density
  • Is well-tolerated with no obvious bad side effects
  • Increased oxygen consumption and respiratory capacity
  • Improved insulin sensitivity and blood plasma lipid profile

Here are some quotes from  the  study:

NMN suppressed age-associated body weight gain, enhanced energy metabolism, promoted physical activity, improved insulin sensitivity and plasma lipid profile, and ameliorated eye function and other pathophysiologies

NMN-administered mice switched their main energy source from glucose to fatty acids

These results strongly suggest that NMN has significant preventive effects against age-associated impairment in energy metabolism

NMN effectively mitigates age-associated physiological decline in mice


LOWER FAT AND INCREASED LEAN MUSCLE MASS

Researchers found that NMN administration suppressed body weight gain by 4% and 9% in the 100 and 300 mg/kg/day groups.

Analyses of  blood chemistry panels and urine did not detect any sign of toxicity from NMN.

Although health span was clearly improved, there was no difference in maximum lifespan observed.

These results suggest that NMN administration can significantly suppress body weight gain without side effects

INCREASED OXYGEN CONSUMPTION AND RESPIRATORY CAPACITY
screen-shot-2016-11-04-at-2-22-48-pm

Oxygen consumption significantly increased in both 100 and 300 mg/kg/day groups during both light and dark periods (Figure 3A).

Energy expenditure also showed significant increases  (Figure 3B).

Respiratory quotient significantly decreased in both groups during both light and dark periods (Figure 3C),

This suggests that NMN-administered mice switched their main energy source from glucose to fatty acids.

The mice that had been receiving NMN for 12 months exhibited energy levels, food and water consumption equivalent to the mice in the control group that were 6 months younger.

NMN administration has significant preventive effects against age associated physical impairment

HUMAN STUDIES – LONG TERM SUPPLEMENTATION WITH NMN

The first clinical trial of NMN in humans is currently underway by an international collaborative team between Keio University School of Medicine in Tokyo and Washington University School of Medicine (33).

Participants are 50 healthy women between 55 and 70 years of age with slightly high blood glucose,BMI and triglyceride levels.

Using a dose of 2 capsules of 125mg NMN per day over a period of 8 weeks, researchers are testing for:

  • change in insulin sensitivity
  • change in beta-cell function
  • works to control blood sugar
  • blood vessels dilate
  • effects of NMN on blood lipids
  • effects of NMN on body fat
  • markers of cardiovascular and metabolic health

According to the study:

“Data from studies conducted in rodents have shown that NMN supplementation has beneficial effects on cardiovascular and metabolic health, but this has not yet been studied in people”

Testing of metabolic parameter will continue for 2 years after supplementation has ended, so final results will not be published for some time yet, but preliminary results are expected to be announced in early 2018.

FOODS THAT CONTAIN NMN

NMN is found in many food sources such as edamame, broccoli, cucumber,cabbage, avocado, tomato, beef and shrimp.

As such, it is likely free from serious side effects in humans, and has been available for purchase commercially for over 2 years.

DOSAGE

In the long term (12 month) 2016 mouse study (22), both 100 and 300mg/kg per day improved oxygen consumption, energy expenditure, and physical activity more.

According to the FDA guidelines, an equivalent  would be about 560 mg for a 150lb human.

It should be noted that NMN administration did not generate any obvious toxicity, serious side effects, or increased mortality rate throughout the 12-month-long intervention period, suggesting the long-term safety of NMN.

The current Human study uses a dosage of 2 capsules of 125 mg, which seems to be the most commonly used dosage.

SUMMMARY

NAD+ levels decrease throughout the body as we age, contributing to disease and aging.

Restoring NAD+ levels can ameliorate many age released health issues.

All the NAD+ precursors are effective at raising NAD+ levels in the liver.

Raising NAD+ in the liver has many benefits, but is not effective in tissues and organs that cannot access NAD+ directly from the bloodstream and so depend on internal cellular NAD+ recycling.

For these tissues, utilizing each cells internal Salvage Pathway is necessary to restore NAD+ levels.

NR is not stable in the body and not normally found in the bloodstream, so is not readily available as NR to many tissues. Once metabolized to NAD+ it cannot enter cells. If metabolized to NAM it cannot bypass the Nampt bottleneck.

NMN is the only precursor that is stable and available to cells through the bloodstream, and can bypass the Nampt bottleneck to quickly restore NAD+ throughout the body.

NMN IS THE WHOLE BODY NAD+ BOOSTER

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OTHER RESEARCH WITH NMN

Aging

Head to Head Comparison of Short-Term Treatment with the NAD(+) Precursor Nicotinamide Mononucleotide (NMN) and 6 Weeks of Exercise in Obese Female Mice (Uddin, 2016)

NAD(+) levels were increased significantly both in muscle and liver by NMN
NMN-supplementation can induce similar reversal of the glucose intolerance
NMN intervention is likely to be increased catabolism of fats
NMN-supplementation does mimic exercise

DNA Damage

A conserved NAD+ binding pocket that regulates protein-protein interactions during aging (Sinclair, 2017)

This study showed supplementation with NMN was able to repair the DNA in cells damaged by radiation.

the cells of old mice were indistinguishable from young mice after just one week of treatment.”


Diabetes & Metabolic disease

Nicotinamide Mononucleotide, a Key NAD+ Intermediate, Treats the Pathophysiology of Diet- and Age-Induced Diabetes in Mice (Yoshino, 2011)

NMN was immediately utilized and converted to NAD+ within 15 min, resulting in significant increases in NAD+ levels over 60 min

administering NMN, a key NAD+ intermediate, can be an effective intervention to treat the pathophysiology of diet- and age-induced T2D

Surprisingly, just one dose of NMN normalized impaired glucose tolerance

Declining NAD+ Induces a Pseudohypoxic State Disrupting Nuclear-Mitochondrial Communication during Aging (Gomes, Sinclair,2013)

raising NAD+ levels in old mice restores mitochondrial function to that of a young mouse

treatment of old mice with NMN reversed all of these biochemical aspects of aging

restore the mitochondrial homeostasis and key biochemical markers of muscle health in a 22-month-old mouse to levels similar to a 6-month-old mouse

CardioVascular Disease

Nicotinamide mononucleotide, an intermediate of NAD+ synthesis, protects the heart from ischemia and repercussion (Yamamoto, 2014)

NMN significantly increased the level of NAD+ in the heart

NMN protected the heart from I/R injury

Nicotinamide mononucleotide supplementation reverses vascular dysfunction and oxidative stress with aging in mice (de Picciotto, 2016)

NMN reduces vascular oxidative stress
NMN treatment normalizes aortic stiffness in old mice
NMN represents a novel strategy for combating arterial aging

Short-term administration of Nicotinamide Mononucleotide preserves cardiac mitochondrial homeostasis and prevents heart failure (Zhang, 2017)

NMN can reduce myocardial inflammation

NMN thus can cut off the initial inflammatory signal, leading to reduced myocardial inflammation

Nicotinamide mononucleotide requires SIRT3 to improve cardiac function and bioenergetics in a Friedreich’s ataxia cardiomyopathy model

Remarkably, NMN administered to FXN-KO mice restores cardiac function to near-normal levels.

restoration of cardiac function and energy metabolism upon NMN supplementation
remarkable decrease in whole-body EE and cardiac energy wasting

Neurological Injury

Nicotinamide mononucleotide attenuates brain injury after intracerebral hemorrhage by activating Nrf2/HO-1 signaling pathway (Wei, 2017)

NMN treats brain injury in ICH by suppressing neuroinflammation/oxidative stress

NMN treatment protects against cICH-induced acute brain injury
NMN treatment reduces brain cell death and oxidative stress
These results further support the neuroprotection of NMN/NAD+

Alzheimers

Effect of nicotinamide mononucleotide on brain mitochondrial respiratory deficits in an Alzheimer’s disease-relevant murine model (Long, 2015)

We now demonstrate that mitochondrial respiratory function was restored

Nicotinamide mononucleotide protects against β-amyloid oligomer-induced cognitive impairment and neuronal death (Wang, 2016)

NMN could restore cognition in AD model rats.
The beneficial effect of NMN is produced by ameliorating neuron survival, improving energy metabolism and reducing ROS accumulation.
These results suggest that NMN may become a promising therapeutic drug for AD

Nicotinamide mononucleotide inhibits JNK activation to reverse Alzheimer disease(Yao, 2017)

NMN Treatment Rescues Cognitive impairments
NMN Treatment Improves Inflammatory Responses

Kidney Disease
Nicotinamide Mononucleotide, an NAD+ Precursor, Rescues Age-Associated Susceptibility to AKI in a Sirtuin 1-Dependent Manner (Guan, 2017)

Supplementation with NMN restored kidney SIRT1 and NAD+ content in 20-month-old mice and protected both young and old mice from acute kidney injury.

References:

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