33 Natural Ways to Improve Mitochondrial Function

A Combination of Nutriments Improves Mitochondrial Biogenesis and Function in Skeletal Muscle of Type 2 Diabetic Goto–Kakizaki Rats

33 Natural Ways to Improve Mitochondrial Function

Recent evidence indicates that insulin resistance in skeletal muscle may be related to reduce mitochondrial number and oxidation capacity.

However, it is not known whether increasing mitochondrial number and function improves insulin resistance.

In the present study, we investigated the effects of a combination of nutrients on insulin resistance and mitochondrial biogenesis/function in skeletal muscle of type 2 diabetic Goto–Kakizaki rats.

We demonstrated that defect of glucose and lipid metabolism is associated with low mitochondrial content and reduced mitochondrial enzyme activity in skeletal muscle of the diabetic Goto-Kakizaki rats.

The treatment of combination of R-α-lipoic acid, acetyl-L-carnitine, nicotinamide, and biotin effectively improved glucose tolerance, decreased the basal insulin secretion and the level of circulating free fatty acid (FFA), and prevented the reduction of mitochondrial biogenesis in skeletal muscle.

The nutrients treatment also significantly increased mRNA levels of genes involved in lipid metabolism, including peroxisome proliferator–activated receptor-α (Pparα), peroxisome proliferator–activated receptor-δ (Pparδ), and carnitine palmitoyl transferase-1 (Mcpt-1) and activity of mitochondrial complex I and II in skeletal muscle.

All of these effects of mitochondrial nutrients are comparable to that of the antidiabetic drug, pioglitazone. In addition, the treatment with nutrients, un pioglitazone, did not cause body weight gain.

These data suggest that a combination of mitochondrial targeting nutrients may improve skeletal mitochondrial dysfunction and exert hypoglycemic effects, without causing weight gain.

Citation: Shen W, Hao J, Tian C, Ren J, Yang L, Li X, et al. (2008) A Combination of Nutriments Improves Mitochondrial Biogenesis and Function in Skeletal Muscle of Type 2 Diabetic Goto–Kakizaki Rats. PLoS ONE 3(6): e2328. https://doi.org/10.1371/journal.pone.0002328

Editor: Don Husereau, Canadian Agency for Drugs and Technologies in Health, Canada

Received: November 1, 2007; Accepted: April 26, 2008; Published: June 4, 2008

Copyright: © 2008 Shen et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Funding: This study was supported by the Pujiang Talent Award (05PJ14104) and the Diabetes Research Grant from the Science and Technology Commission of Shanghai Municipality (04dz14007), and the grant by the Chinese Academy of Sciences. The sponsors has no role in the design and conduct of the study, in the collection, analysis, and interpretation of the data, and in the preparation, review, or approval of the manuscript.

Competing interests: Patent has been filed on combination of mitochondrial nutrients metioned in this paper: Chinese patent application No 200610162888.4. The authors have declared that no competing interests exist.

Increasing evidence suggests that mitochondrial dysfunction due to oxidative damage is a major contributor to aging, degenerative diseases such as cancer, and metabolic syndrome, such as obesity and type 2 diabetes [1], [2], [3].

Skeletal muscle insulin resistance may play an important role in the pathogenesis of the metabolic syndrome and 2 type diabetes [4]. Recent studies reported insulin resistance is associated with impaired skeletal muscle oxidation capacity and reduced mitochondrial number and function [5], [6].

Therefore, protecting mitochondria from oxidative damage to improve mitochondrial function in skeletal muscle seems a possible strategy to prevent and treat diseases associated with mitochondrial dysfunction [7], [8], [9], 10.

For example, rosiglitazone improves the suppression of adipose mitochondrial biogenesis in db/db and high fat diet-fed mice [11].

Pioglitazone reduces hyperglycemia, hyperlipidemia, and hyperinsulinemia in male fatty rats [12], improves mitochondrial function and stimulates mitochondrial biogenesis in human adipocyte/tissue in vitro [13], [14] or human neuron- cells [15].

Metformin delays the manifestation of diabetes and vascular dysfunction and reduces mitochondrial oxidative stress in Goto-Kakizaki (GK) rats [16]. However, the use of any pharmacological therapy for type 2 diabetes, such as thiazolidinedione, insulin, metformin, and other oral hypoglycemic agents or combination therapy with or without insulin appears to be associated with an increased risk of heart failure and body weight gain [17]. Therefore, effective treatments without apparent side effects are greatly needed for preventing and treating diabetes and other metabolic syndromes.

We have defined a group of mitochondrial targeting antioxidants/metabolites as mitochondrial nutrients [8], [9], [18], i.e.

, nutrients which improve mitochondrial function and protect mitochondria from oxidative damage, including those that can 1) inhibit or prevent oxidant production in mitochondria; 2) scavenge and inactivate free radicals and reactive oxygen species; 3) repair mitochondrial damage and enhance antioxidant defenses by stimulating mitochondrial biogenesis and inducing phase-2 enzymes; and 4) act as cofactors/substrates to protect mitochondrial enzymes and/or stimulate enzyme activity. One good example of mitochondrial nutrients is R-α-lipoic acid (LA) [9], [19], [20], [21].

More recently, we have examined the effect of LA and acetyl-L-carnitine (ALC), as well as their combination, on mitochondrial biogenesis in 3T3-L1 adipocyte.

We found that treatment with a combination of LA and ALC significantly improved mitochondrial function and increased mitochondrial biogenesis related transcription factors while the treatments with only LA and ALC alone at the same concentrations showed little effect [22].

From these results, we have concluded that the combination of mitochondrial targeting nutrients may complementarily promote mitochondrial synthesis and adipocyte metabolism and possibly, thiazolidinedione drugs, prevent and treat insulin resistance in type 2 diabetes.

In the present study, we investigated the effects of a combination of LA and ALC, with two other mitochondrial nutrients, nicotinamide and biotin, on improving glucose tolerance, insulin release, fatty acid metabolism, and mitochondrial biogenesis and function in the spontaneous diabetic GK rats.

The metabolic characteristics of GK rats are summarized in Table 1. After the 12-week administration, there was no difference in body weight between the untreated and the nutrients-treated GK rats, however, the body weight in the pioglitazone-treated group was significantly higher than that of other groups.

The pioglitazone-induced gain of body weight is consistent with a previous report [23]. The pioglitazone treatment tended to increase food intake (measured for individual rats in g/kg/24 h) and significantly reduced the levels of triglyceride and total cholesterol in blood.

The nutrients-treatment did not affect fasting glucose and triglyceride levels, but significantly reduced fasting plasma insulin (p

Source: https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0002328

Mitochondrial Dysfunction as a Key Event during Aging: From Synaptic Failure to Memory Loss

33 Natural Ways to Improve Mitochondrial Function

Open access peer-reviewed chapter

By Claudia Jara, Angie K. Torres, Margrethe A. Olesen and Cheril Tapia-Rojas

Submitted: March 21st 2019Reviewed: July 8th 2019Published: August 7th 2019

DOI: 10.5772/intechopen.88445

Mitochondria are important cellular organelles with key regulatory functions in energy production, oxidative balance, and calcium homeostasis. This is especially important in the brain, since neurons require a large number of functional mitochondria to supply their high energy requirement, mainly for synaptic processes.

A decrease in the activity and quality of mitochondria in the brain, particularly in the hippocampus, is associated with normal aging and a large number of neurodegenerative diseases compromising memory function.

Although synaptic and cognitive dysfunction is multifactorial, growing evidence demonstrates that mitochondria play a key role in these processes and suggests that maintaining mitochondrial function could prevent these age-dependent alterations.

In this chapter, we will discuss the hippocampal mitochondrial dysfunction present in aging and how these defects promote age-associated synaptic damage and cognitive impairment. We will summarize evidence that shows how neurodegeneration can be accelerated or attenuated during aging by modulating mitochondrial function.

  • aging
  • mitochondria
  • oxidative stress
  • synapses
  • memory

Aging is an extensively studied process, identifying a growing interest in how and why cognitive processes are affected from a neurobiological approach [1].

Aging is a multifactorial biological process, characterized by deterioration of physiological and cellular functions including brain function [2], where age is the main risk factor for the development of pathologies such as cancer, diabetes, cardiovascular disorders, and neurodegenerative diseases [3].

Cognitive deterioration occurs during aging, where reasoning, attention, and memory, among other processes, decrease gradually with the age [4].

Cellular senescence and alterations to mitochondria and in proteolytic systems are considered hallmarks of aging [3], where one of the most studied is the mitochondria [5]. In fact, mitochondrial dysfunction has been directly associated with the aging phenotype and the majority of diseases that lead to cognitive damage.

Over the last decades, a great interest has arisen regarding mitochondrial structure and function due to its relation with the aging brain [5]. Mitochondria are organelles essential for energy production, whose size is usually 0.5–1 μm, composed by two membranes, forming the intermembranous space and the mitochondrial matrix [6].

The outer membrane contains many copies of the transport protein porin (or voltage-dependent anion-selective channels (VDAC)), which allows the passage of molecules with a maximum weight of 5 KDalton (KDa), and the inner membrane forms numerous invaginations, tubular structures, called cristae [6].

Mitochondria are capable of remodeling their architecture through fission and fusion processes, allowing morphological adaptation to different situations [6]. Fission is essential for mitochondrial duplication and is necessary for mitophagy, allowing dysfunctional mitochondrial sections to be recycled.

Fusion allows mitochondria to interconnect, allowing damaged mitochondria to maintain their function. However, fission-fusion processes are interrupted during aging, generating damaged mitochondria [7].

Mitochondria have a small circular genome called mtDNA, which encodes 22 tRNAs, 2 mitochondrial rRNAs, and 13 subunits of the electron transport chain (ETC) [8]. mtDNA can be damaged by exposure to reactive oxygen species (ROS), chemical carcinogens, and ionizing radiation affecting the mitochondrial function; changes are also observed during aging [9].

The internal mitochondrial membrane contains the ETC, responsible for generating ATP. ETC is formed by five protein complexes; complex I (NADH dehydrogenase) receives electrons of NADH which pass through the ETC via oxidation-reduction reactions forming an electrochemical gradient that allows the formation of ATP.

In addition, FADH2 donates its electrons to complex II (succinate dehydrogenase) performing the same action for ATP generation but at lower production levels [10]. As a secondary product, the ETC forms ROS, specifically by complexes I and III, but its production is controlled by antioxidant enzymes [11].

Therefore, in normal conditions ROS production is moderate, providing certain physiological roles [11]; however, during aging ROS accumulation causes biological damage known as “oxidative stress” [12].

In the past, mitochondria have always been highlighted for its role in ATP production; however, another key function is to maintain intracellular calcium homeostasis [13].

The outer mitochondrial membrane is permeable to ions and ~5 KDa metabolites because its lipid bilayer has transmembrane proteins that form the mitochondrial permeability transition pore (mPTP). mPTP opening and closing dynamics regulates the concentration of calcium [13].

However, in conditions of high calcium concentrations, permanent mPTP opening generates massive transport of ions and small molecules

Source: https://www.intechopen.com/online-first/mitochondrial-dysfunction-as-a-key-event-during-aging-from-synaptic-failure-to-memory-loss

Mitochondrial Dysfunction and Diabetes: Is Mitochondrial Transfer a Friend or Foe?

33 Natural Ways to Improve Mitochondrial Function

Open AccessReview

byMagdalene K Montgomery

Department of Physiology, School of Biomedical Sciences, University of Melbourne, Melbourne 3010, Australia

Biology 2019, 8(2), 33; https://doi.org/10.3390/biology8020033

Received: 9 November 2018 / Revised: 21 November 2018 / Accepted: 20 December 2018 / Published: 11 May 2019

View Full-TextDownload PDF Obesity, insulin resistance and type 2 diabetes are accompanied by a variety of systemic and tissue-specific metabolic defects, including inflammation, oxidative and endoplasmic reticulum stress, lipotoxicity, and mitochondrial dysfunction. Over the past 30 years, association studies and genetic manipulations, as well as lifestyle and pharmacological invention studies, have reported contrasting findings on the presence or physiological importance of mitochondrial dysfunction in the context of obesity and insulin resistance. It is still unclear if targeting mitochondrial function is a feasible therapeutic approach for the treatment of insulin resistance and glucose homeostasis. Interestingly, recent studies suggest that intact mitochondria, mitochondrial DNA, or other mitochondrial factors (proteins, lipids, miRNA) are found in the circulation, and that metabolic tissues secrete exosomes containing mitochondrial cargo. While this phenomenon has been investigated primarily in the context of cancer and a variety of inflammatory states, little is known about the importance of exosomal mitochondrial transfer in obesity and diabetes. We will discuss recent evidence suggesting that (1) tissues with mitochondrial dysfunction shed their mitochondria within exosomes, and that these exosomes impair the recipient’s cell metabolic status, and that on the other hand, (2) physiologically healthy tissues can shed mitochondria to improve the metabolic status of recipient cells. In this context the determination of whether mitochondrial transfer in obesity and diabetes is a friend or foe requires further studies.View Full-Text

Keywords: mitochondrial dysfunction; insulin resistance; type 2 diabetes; mitochondrial transfer; exosomes mitochondrial dysfunction; insulin resistance; type 2 diabetes; mitochondrial transfer; exosomes

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Montgomery MK. Mitochondrial Dysfunction and Diabetes: Is Mitochondrial Transfer a Friend or Foe? Biology. 2019; 8(2):33.

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Calcium, ATP, and ROS: a mitochondrial love-hate triangle | American Journal of Physiology-Cell Physiology

33 Natural Ways to Improve Mitochondrial Function

Mitochondrial oxidative phosphorylation (ox-phos) is the major ATP synthetic pathway in eukaryotes.

In this process, electrons liberated from reducing substrates are delivered to O2 via a chain of respiratory H+ pumps.

These pumps (complexes I-IV) establish a H+ gradient across the inner mitochondrial membrane, and the electrochemical energy of this gradient is then used to drive ATP synthesis by complex V (ATP synthase) (136).

Chemically, the stepwise reduction of O2 (O2 → O2−· → H2O2 → OH· → H2O) proceeds via several reactive oxygen species (ROS).

These ROS can damage cellular components such as proteins, lipids, and DNA (70), but recent evidence also highlights a specific role in redox cell signaling for mitochondrial ROS (55, 203).

In the fine balancing act of aerobic metabolism, mitochondrial ox-phos accomplishes the reduction of O2 to H2O while maximizing ATP synthesis and maintaining ROS production to only the amounts required for microdomain cell signaling (19, 87).

In addition to ATP synthesis, mitochondria are the site of other important metabolic reactions, including steroid hormone and porphyrin synthesis, the urea cycle, lipid metabolism, and interconversion of amino acids (39, 141). Mitochondria also play central roles in xenobiotic metabolism, glucose sensing/insulin regulation (113), and cellular Ca2+ homeostasis (65, 66), which affects numerous other cell signaling pathways.

Despite these critical metabolic roles of mitochondria, classic “mitochondriology” was considered a mature field as recently as 1990.

However, several important observations have fueled a renaissance in mitochondrial research, including 1) mitochondrial ROS are not just damaging by-products of respiration, but important for cell signaling (19, 23); 2) mitochondrial release of factors such as cytochrome c is an important step in programmed cell death (100, 110, 112); 3) nitric oxide (NO·) is an potent regulator of mitochondrial function (19, 23, 34); 4) mitochondrial morphology is far from static, with the organelles being subject to fission, fusion, and intracellular movement on a rapid timescale (95, 218); and 5) mitochondria actively orchestrate the spatiotemporal profiles of intracellular Ca2+, under both physiological and pathological conditions (65, 66). Together these observations suggest an extensive regulatory role for mitochondria in both normal and pathological cell function.

The interplay between the conventional and novel roles of mitochondria has received little consideration, and an examination of recent mitochondrial science reveals several incompatibilities with classic bioenergetic viewpoints. An example is the requirement of ATP for apoptosis (137).

How does the cell maintain ATP synthesis in the face of mitochondrial disassembly that occurs during apoptosis? Another example, which is the focus of this review, is the role of Ca2+ in regulating organelle function and dysfunction.

How can Ca2+, a physiological stimulus for ATP synthesis (5, 72, 118), become a pathological stimulus for ROS generation, cytochrome c release, and apoptosis? As will be discussed extensively, this apparent mitochondrial Ca2+ paradox revolves around a “two-hit” hypothesis (Fig.

1) in which a concurrent pathological stimulus can turn Ca2+ from a physiological to a pathological effector.

Fig. 1.Two-hit hypothesis for mitochondrial Ca2+ in physiology and pathology. Under physiological conditions, Ca2+ is beneficial for mitochondrial function.

However, in the presence of an overriding pathological stimulus, Ca2+ is detrimental. Similarly, Ca2+ can potentiate a subthreshold pathological stimulus, resulting in pathogenic consequences. See text for full explanation.

[Ca2+]m, mitochondrial matrix Ca2+ concentration; ROS, reactive oxygen species.


Most of the mitochondrial effects of Ca2+ require its entry across the double membrane into the matrix.

Although the mitochondrial outer membrane was thought to be permeable to Ca2+, recent studies suggest that the outer membrane voltage-dependent anion channel (VDAC) is a ruthenium red (RuRed)-sensitive Ca2+ channel and thus serves to regulate Ca2+ entry to mitochondrial intermembrane space (60). Furthermore, transport across the inner membrane is highly regulated (for review see Ref. 65).

Figure 2 outlines the major mechanisms for mitochondrial Ca2+ transport, with Ca2+ uptake achieved primarily via the mitochondrial Ca2+ uniporter (MCU). Uptake is driven by the membrane potential (Δψm), and therefore the net movement of charge due to Ca2+ uptake consumes Δψm.

A recent patch-clamp study suggests the that MCU is a highly selective (Kd < 2 nM) Ca2+ channel (99), but attempts to define its molecular nature have been largely unsuccessful. The channel is known mostly for its pharmacological sensitivity to RuRed (127), and a colorless component of RuRed (Ru360) is the active MCU-binding agent (156, 216).

Saris et al. (165) identified a 40-kDa glycoprotein of the intermembrane space as an MCU regulatory component, although the transmembrane component of the MCU has been more difficult to isolate, with limited reports of such an entity (124).

Interestingly, reverse MCU transport (Ca2+ export) was shown to be regulated by Ca2+ binding to the outer surface of the inner membrane (86) and was also linked to a soluble intermembrane space component.

Fig. 2.Pathways of mitochondrial Ca2+ uptake and export. The respiratory chain is shown with (left to right) complexes I, III, IV, and V. The outer mitochondrial membrane and complex II are omitted for clarity.

Where possible, known 3-dimensional (3D) structures obtained from the Protein Data Bank ( http://www.rcsb.org/pdb) are shown.

UP, Ca2+ uniporter; RaM, rapid-mode Ca2+ uptake; RyR, ryanodine receptor; PTP, permeability transition pore; Δψm, membrane potential.

Das et al. (41) showed that a complex of Ca2+-polyphosphate and β-hydroxybutanoate can form a Ca2+ channel indistinguishable from that in Escherichia coli, raising the possibility that the MCU (by virtue of mitochondrial/bacterial relationships) may be a nonproteinaceous entity.

However, the second-order Ca2+ transport kinetics of the MCU suggest a more complex structure with separate activation and transport sites (169, 206). From a physiological perspective, a role was recently demonstrated for p38 MAP kinase in regulating RuRed-sensitive Ca2+ transport (126).

Clearly, identification of the molecular nature of the MCU will aid greatly in understanding the physiological and pathological regulation of mitochondrial Ca2+ uptake.

Two additional mechanisms of Ca2+ entry into mitochondria have also been identified. The first, called “rapid-mode” uptake (RaM), occurs on a millisecond timescale and allows fast changes in mitochondrial matrix Ca2+ concentration ([Ca2+]m) to mirror changes in the cytosol ([Ca2+]c) (186).

Second, we have found (11) that ryanodine receptor isoform (RyR)1 is localized to the inner membrane of mitochondria in excitable cells and have termed this channel “mRyR.” Kinetic analysis of the MCU predicts a tetrameric structure RyR, which exists as a tetramer of ∼500-kDa subunits (17).

Together, mRyR and RaM are thought to underlie the phenomenon of excitation-metabolism coupling, in which [Ca2+]c-induced contraction is matched by [Ca2+]m stimulation of ox-phos (see below).

A fast response of [Ca2+]m to [Ca2+]c requires rapid Ca2+ efflux from the mitochondrial matrix, and several mechanisms exist for this purpose (65). Primarily, Ca2+ efflux is achieved by exchange for Na+, which is in turn pumped the matrix in exchange for protons (Fig. 2).

Thus both Ca2+ uptake and efflux from mitochondria consume Δψm and are therefore reliant on H+ pumping by the respiratory chain to maintain this driving force. In addition to these pathways of Ca2+ efflux, an additional mechanism exists in the form of the permeability transition (PT) pore (10).

The PT pore is assembled from a group of preexisting proteins in the mitochondrial inner and outer membranes (38), with Ca2+ binding sites on the matrix side of the inner membrane believed to regulate pore activity. Normally, “flickering” of the PT pore between open and closed states serves to release Ca2+ from the matrix (84, 141, 205).

However, prolonged PT pore opening due to [Ca2+]m overload can result in pathological consequences (38).


The primary role of mitochondrial Ca2+ is the stimulation of ox-phos (5, 40, 72, 118, 123). As shown in Fig.

3, this occurs at many levels, including allosteric activation of pyruvate dehydrogenase, isocitrate dehydrogenase, and α-ketoglutarate dehydrogenase (118), as well as stimulation of the ATP synthase (complex V) (40), α-glycerophosphate dehydrogenase (211), and the adenine nucleotide translocase (ANT) (123). Overall the effect of elevated [Ca2+]m is the coordinated upregulation of the entire ox-phos machinery, resulting in faster respiratory chain activity and higher ATP output. Thus mitochondrial ATP output can be changed to meet the cellular ATP demand. An example of this is β-adrenergic stimulation in cardiomyocytes signaling the demand for increased contractility. The concomitant upregulation of ox-phos via [Ca2+]m elevation provides the ATP needed for increased contractile force.

Fig. 3.Ca2+ activation of the TCA cycle and oxidative phosphorylation. Thin arrows represent metabolic pathways/reactions; thick arrows represent actions of Ca2+. The outer membrane is omitted for clarity.

Where possible, known 3D structures obtained from the Protein Data Bank are shown. For α-glycerophosphate (α-GP) dehydrogenase (α-GPDH), the cytosolic isoform structure is shown.

Succ, succinate; α-KG, α-ketoglutarate; Isocit, isocitrate; Cit, citrate; OAA, oxaloacetate; Mal, malate; Fum, fumarate; Ac-CoA, acetyl coenzyme A; Pyr, pyruvate; PDH, pyruvate dehydrogenase; Acon, aconitase; CS, citrate synthase; MDH, malate dehydrogenase; ICDH, isocitrate dehydrogenase; α-KGDH, α-ketoglutarate dehydrogenase; DHAP, dihydroxyacetone phosphate; CxI–V, complexes I–V; SDH, succinate dehydrogenase.

Many other mitochondrial functions are also regulated by Ca2+.

For example, Ca2+ activation of N-acetylglutamine synthetase generates N-acetylglutamine (92), a potent allosteric activator of carbamoyl-phosphate synthetase, the rate-limiting enzyme in the urea cycle (119).

In addition, Ca2+- and diacylglycerol-sensitive protein kinase (PKC) isoforms and calmodulin have been reported in mitochondria, although their precise targets within the organelle are less well understood (54, 160).

Overall, it appears that Ca2+ is a global positive effector of mitochondrial function, and thus any perturbation in mitochondrial or cytosolic Ca2+ homeostasis will have profound implications for cell function, for example, at the level of ATP synthesis. Also, it cannot be ignored that Ca2+, particularly at the high concentrations experienced in pathology, appears to have several negative effects on mitochondrial function, as discussed in the following sections.


In contrast to the beneficial effects of Ca2+, the PT pore embodies the pathological effects of Ca2+ on mitochondria. The PT pore, as described nearly 25 years ago by Haworth and Hunter (81–83), is an assembly of preexisting proteins of the inner and outer mitochondrial membranes into a large conductance channel permeable to solutes of

Source: https://journals.physiology.org/doi/10.1152/ajpcell.00139.2004

9 Ways to Boost Your Mitochondria, Increase Energy and Enhance Longevity

33 Natural Ways to Improve Mitochondrial Function

When patients tell me they’re feeling wiped out, exhausted and dragging through their days, often it’s because they’re not treating their mitochondria right. Most people don’t realize it but mitochondria play a massive role in their energy levels, how well their metabolism functions and even how much brain fog they deal with every day.

In a nutshell, your mitochondria are the trillions of microscopic energy factories that power your body, turning the food you eat and the air you breathe into the energy that powers the biochemical reactions in your cells. That energy is used for everything from flexing muscles to making essential enzymes and hormones.

So, if you’re pummeling your mitochondria daily with a litany of bad habits – crappy food, poor sleep, high stress levels and an office-chair-to-couch-potato lifestyle – you won’t have enough energy on hand to power your day.

When we’re young, we have plenty of mitochondria. But, as with every other system in the body, over time, our mitochondria decline in both size and number – and with it, much of the energy we once took for granted.

In fact, researchers now think mitochondrial decline is one of the primary drivers of aging – of feeling tired and looking tired – and a major culprit behind the diseases of the brain and the cardiovascular system that impact so many people as they age.

But the good news is that we do have some control over how fast or slow we age – and a lot of it comes down to how well we treat our mitochondria.

Not surprisingly, many of the healthy habits I encourage everyone to adopt to sustain health are, at the microscopic level, great for your mitochondria too.

So, to support and manage your mighty ‘mitos’ I recommend the following mitochondria-boosting moves:

1) Don’t eat crap — particularly the stuff that spikes blood sugar.

Say ‘no’ to sugar, refined grains (think flour), even the whole grains people think of as healthy, all of which spike blood sugar and contribute to unwanted pounds and body fat – which, in turn, promotes mitochondria-crushing inflammation. In addition, try to keep your diet ‘clean’ by avoiding processed foods, pesticide-laden produce, and factory-farmed foods meats.

2) Feed your mitochondria well

While the mitochondria can use either fatty acids or carbohydrates to create the ATP needed to produce energy, doing so with fat is loads more efficient and creates fewer free radical byproducts. Keeping your carb intake low so your mitochondria will burn fat for energy, will help keep you trim to boot.

In addition, look for foods packed with vitamins, phytonutrients and antioxidants. On your plate, that means goodies high-quality, pasture-raised animals, wild-caught fish, (preferably) organic veggies, avocados, extra virgin olive oil, nuts and seeds, and some low-sugar fruits.

You can’t go wrong loading up on leafy greens and cruciferous veggies, cauliflower and Brussels sprouts.

3) Get into intermittent fasting.

To boost mitochondrial function and longevity, practicing intermittent fasting a few days a week is a great ‘bio hack’ that supports mitochondria health by reducing mitochondrial free radical production.

How to do it:  Compress your ‘eating window’ from the typical 12 -16 hour graze-all-day routine down to 8-hours. You’ll eat dinner earlier and breakfast later – and give your body many more non-eating hours.

4) Keep moving!

Need another reason to exercise? Your mitochondria love it.

Get into a high-intensity interval training (HIIT) groove that will boost mitochondrial production, and minimize the risk of overtraining and mitochondrial damage.

A regular HITT routine will build up muscular endurance as well as the number and size of the mitochondria that power those muscles, so don’t just sit there – move for your mitochondria!

5) Add meditation and massage to your routine.

Recent research suggests that meditation and other relaxation-based techniques can reduce oxidative stress – and that means, as time passes, less damage to the mitochondria. And don’t forget to relax your body too. One recent study showed that massage could give the body’s production of mitochondria a boost.

6) Focus on good quality sleep.

Sleep protects your brain by clearing out neural waste products that build up daily and, the research suggests, preserves the mitochondria as well. Think of sleep as your brain’s time to ‘take out the garbage,’ so don’t cut corners on this mitochondria-protecting activity.

7) Soak up some sun.

It turns out that exposing your body to sensible amounts of sunlight (turning pink means you’ve overdone it) is nature’s way boosting mitochondria production, so don’t be afraid to catch a few rays every now and then.

8) Expose yourself to the cold.

Another way to trigger new mitochondrial production is to expose yourself to quick bursts of cold temperatures, be it in the great outdoors for 20 -30 seconds a shot, or in the shower. Doing so will, in essence, trick your body into survival mode, and kick mitochondria production into high gear.

9) Supplement your mitochondrial health.

While nothing encourages the mitochondria to thrive more than a clean, healthy diet, you can give your ‘mitos’ a targeted boost with some carefully chosen nutrient supplements. The right combo of diet and supplements translates to enhanced mental sharpness, fewer body aches and pains and protection against the most common and dreaded diseases of aging.

There’s no better mind-body therapy than that! The supplements I recommend to support mitochondrial health include: Nicotinamide Riboside, Alpha Lipoic Acid; Glutathione; CoQ10; the B vitamins; Magnesium; Fish or krill oil; L Carnitine; and PQQ, a relatively new one on the research radar and it looks not only to reduce oxidative damage but to stimulate new mitochondrial growth.

Source: https://drfranklipman.com/2019/04/22/9-ways-to-boost-your-mitochondria-increase-energy-and-enhance-longevity/

Delivery of exogenous mitochondria via centrifugation enhances cellular metabolic function

33 Natural Ways to Improve Mitochondrial Function

Mitochondria are essential organelles involved in the maintenance of cell growth and function, and have been investigated as therapeutic targets in various diseases.

Recent studies have demonstrated that direct mitochondrial transfer can restore cellular functions of cells with inherited or acquired mitochondrial dysfunction. However, previous mitochondrial transfer methods are inefficient and time-consuming.

Here, we developed a simple and easy mitochondrial transfer protocol using centrifugation, which can be applied to any cell type.

By our simple centrifugation method, we found that the isolated mitochondria could be successfully transferred into target cells, including mitochondrial DNA-deleted Rho0 cells and dexamethasone-treated atrophic muscle cells.

We found that mitochondrial transfer normalised ATP production, mitochondrial membrane potential, mitochondrial reactive oxygen species level, and the oxygen consumption rate of the target cells. Furthermore, delivery of intact mitochondria blocked the AMPK/FoxO3/Atrogene pathway underlying muscle atrophy in atrophic muscle cells. Taken together, this simple and rapid mitochondrial transfer method can be used to treat mitochondrial dysfunction-related diseases.

Mitochondria are powerful and dynamic organelles responsible for essential cell functions, including energy metabolism, generation of free radicals, maintenance of calcium homeostasis, cell survival and death.

Mitochondrial dysfunction is being recognized as being involved with many serious health problems such as aging1, cancer2, metabolic disorders3 and neurodegenerative diseases4. Muscle disorders such as muscle atrophy, degeneration and myopathy are also caused by mitochondrial malfunction5,6.

Abnormal activities of enzymes of the mitochondrial respiratory chain and mitochondrial DNA (mtDNA) deletions have been observed in aged skeletal muscles7. These mtDNA mutations cause cellular dysfunction and lead to loss of muscle mass and strength.

Oxidative damage resulting from errors in mtDNA replication and the repair system are thought to be at the root cause of these diseases8. Although mitochondrial dysfunction and muscle disorders are closely related, the detailed underlying mechanisms remain enigmatic.

Diverse mechanisms lead to mitochondrial dysfunction, including changes in the nuclear or mitochondrial genome, environmental insults or alterations in homeostasis9.

Accumulation of dysfunctional mitochondria (>70–80%) upon exposure to intracellular or extracellular stress leads to oxidative stress, and in turn, affects intracellular signalling and gene expression6,10. Under severe oxidative stress, ATP is depleted, which prevents controlled apoptotic death and instead causes necrosis11.

A recent study indicates that increased production of mitochondrial reactive oxygen species (mROS) is a major contributor to mitochondrial damage and dysfunction associated with prolonged skeletal muscle inactivity6. In addition, increased mitochondrial fragmentation caused by mROS production results in cellular energy stress (e.

g., a low ATP level) and activation of the AMPK-FoxO3 signalling pathway, which induces expression of atrophy-related genes, protein breakdown and ultimately muscle atrophy5,6,12. Collectively, these results indicate that modulation of mROS production plays a major role in the prevention of muscle atrophy.

Although recent studies provide direct evidence linking mitochondrial signalling with muscle atrophy, no mitochondria-targeted therapy to ameliorate muscle atrophy has been developed to date.

Existing mitochondria-targeted therapeutic strategies can be categorised as follows: 1) repair via scavenging of mROS, 2) reprogramming via stimulation of the mitochondrial regulatory program and 3) replacement via transfer of healthy exogenous mitochondria13.

However, since modulation of mitochondrial function via repair and reprogramming can’t overcome genetic defects, replacement of damaged mitochondria represents an attractive option14.

In this regard, recent studies have shown that the healthy or modified mitochondria can be delivered to damaged cells, restoring cellular function and treating the disease15,16,17,18,19,20.

There have also been reports of direct delivery of healthy mitochondria to specific cells in vitro21,22,23. However, these methods have limitations in terms of efficiency and require cell cultivation process for mitochondrial delivery.

In this study, we developed a simple method to transfer mitochondria into cells by first mixing them together followed by centrifugation. This method makes mitochondrial delivery possible into any cell type, and no additional incubation is required. The transfer efficiency remained high irrespective of the amounts of mitochondria used.

We also evaluated the effects of mitochondrial transfer on cells with induced mitochondrial dysfunction by treatment with oligomycin24 and ethidium bromide (EtBr) (Rho0 cells)25.

Finally, we compared changes in mitochondrial metabolic function and the signalling pathway from dexamethasone (Dexa)-induced atrophic L6 muscle cells receiving intact mitochondria and damaged mitochondria.

We first isolated mitochondria from human umbilical cord-derived mesenchymal stem cells (UC-MSCs) by differential centrifugation (Fig. S1A). Mitochondrial viability, mitochondrial purity, and mitochondrial function were then comprehensively analysed.

To assess mitochondrial viability, we used the MitoTracker Red CMXRos (CMXRos) probe that stains mitochondria and its accumulation is dependent on the mitochondrial membrane potential (MMP). In other words, this probe allows for identification of viable, respiration competent mitochondria26.

Also, the identity of endogenous mitochondria was confirmed by counterstaining with MitoTracker Green (MTG). As shown in Fig. S1B–D, isolated mitochondria were clearly stained both with MTG and CMXRos, indicating that isolated mitochondria from UC-MSCs maintained their membrane potential and were viable.

The purity of isolated mitochondria was determined by assessing the presence of functional mitochondrial markers [cytochrome C oxidase (COX IV) and cytochrome c] and absence of nuclear markers [proliferating cell nuclear antigen (PCNA) and β-actin].

Western blots against COX IV and cytochrome c confirmed the presence of mitochondrial proteins in the isolated mitochondria, while PCNA and β-actin proteins were absent, confirming the high purity of the isolated mitochondria (Fig. S1E). Also, Fig. S1F shows mitochondria with reticulated morphology by electron microscopy. Finally, the ATP content of various amounts of mitochondria (0.05, 0.5 and 5 μg) increased proportionately thus confirming their functionality (Fig. S1G).

Validation of mitochondrial transfer

In order to effect transfer of isolated mitochondria into target cells, we elected on using a quick and simple centrifugation method. Figure 1A shows our experimental scheme for mitochondrial transfer and further application.

Isolated intact mitochondria can be efficiently transferred into prepared recipient cells by centrifugation at 1,500 × g for 5 min.

This condition was established through preliminary experiments assessing transfer efficiency over time and centrifugal force (Fig. S2A).

Figure 1

Confocal microscopic analysis of target cells following mitochondrial transfer. (A) Experimental scheme for mitochondrial transfer and further application. The picture was drawn by us.

(B) Representative images of UC-MSCs co-stained with fluorescent mitochondrial dyes (MitoTracker Green and MitoTracker Red CMXRos) at 24 h after mitochondrial transfer in the before mitochondrial transfer (upper panels) and after mitochondrial transfer (lower panels).

Green: endogenous mitochondria of UC-MSCs (recipient cells), red: transferred mitochondria isolated from UC-MSCs, yellow: merged mitochondria. (CE) Three confocal sections are shown in Z-stack overlay mode.

Transferred mitochondria (red) within UC-MSCs were detected in the orthogonal view (upper panels; Z) and the corresponding signal profile (lower panels; S) together with endogenous mitochondria (green). Results are from the centre of the mitochondrial network of UC-MSCs (D) and 2 μm below (C) and 2 μm above (E) it. Z: Z stack image-ortho analysis, S: signal profile of each section. Scale bar, 50 μm.

We confirmed the presence of the transferred mitochondria by confocal microscopy. As shown in Fig. 1B, exogenous mitochondria stained with CMXRos were mixed with UC-MSCs whose endogenous mitochondria were stained with MTG, and then immediately subjected to centrifugation.

As expected, exogenous mitochondria were transferred into UC-MSCs (Fig. 1B) by simple centrifugation. Transferred exogenous mitochondria (red) co-localised with endogenous mitochondria (green) from UC-MSCs, indicating movement of exogenous mitochondria inside the cells as evidenced by the merged yellow staining.

To further assess the localisation of internalised mitochondria, we generated a z-stack of confocal images of UC-MSCs having received exogenous mitochondria stained with CMXRos (Fig. 1C,D and E).

Our results clearly show that the presence of transferred mitochondria within recipient cells, as evidenced by the merged yellow staining, confirming that exogenous mitochondria can be efficiently transferred into recipient cells by centrifugation.

Quantification of mitochondrial transfer

To quantify the efficiency of mitochondrial transfer, flow cytometry (FACS) and PCR analyses were performed. Mixtures of UC-MSCs with exogenous mitochondria were subjected to centrifugation and then to FACS.

The results of FACS analysis showed that the uptake of exogenous mitochondria increased with the amount of mitochondria used; 33.1 ± 0.8%, 77.1 ± 1.1% and 92.7 ± 5.9% of cell exhibited green fluorescence following transfer of 0.05, 0.5 and 5 μg of mitochondria into UC-MSCs, respectively (Fig. 2A).

To further support the efficiency of our centrifugation transfer method, mitochondria (0.05 μg of protein) were passively transferred into UC-MSCs by co-incubation at 37 °C without centrifugation (Fig. S2B).

Different concentrations of Pluronic F-68 (PF-68), which increases the fluidity of cell membranes27, were used during passive transfer. Nevertheless, the passive transfer efficiency of mitochondria was incomparable to that of the centrifugation method (Fig. S2B).

Additionally, we tested the effect of UC-MSCs pretreatment with PF-68 prior to mitochondrial transfer by centrifugation. In contrast with the passive transfer results, increases in transfer efficiency were observed along with membrane permeability modulation of recipient cells by pre-treatment with PF-68 prior to centrifugation.

Especially, the highest transfer efficiency was obtained after pretreatment with 20 mg/mL PF-68 for 2 hours, compared to centrifugation alone (90.7 ± 8.8% vs. 34.3 ± 1.8%) (Fig. S2C). Taken together, our results suggest that mitochondrial transfer by centrifugation is very effective. It also shows that controlling membrane permeability enhances transfer efficiency.

Figure 2

Effect of mitochondrial dose on mitochondrial delivery efficiency. (A) Flow cytometric analysis of MitoTracker Green fluorescence in UC-MSCs at 24 h after transfer of various amounts of mitochondria (expressed as μg of protein).

(B) qPCR analysis of human mtDNA (h-mtDNA), rat-mtDNA and rat-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in rat L6 muscle cells after transfer of mitochondria isolated from UC-MSCs.

(C) Real-time PCR analysis of the mean h-mtDNA copy number of mitochondria isolated from UC-MSCs for transfer (prepared mitochondria in bar definition), rat L6 cells after mitochondrial transfer via centrifugal force (centrifugation) and rat L6 cells after 1 day of co-culture (co-culture). Relative mtDNA copy numbers normalised to the GAPDH level are shown. All values are mean ± SEM. N = 3, *P 

Source: https://www.nature.com/articles/s41598-018-21539-y