Treatment of Fluoroquinolone Associated Disability - the pathobiochemical implications
Krzysztof Michalak 1,2, Aleksandra Sobolewska-Włodarczyk 3, Marcin
Włodarczyk 3, Justyna Sobolewska 4, Piotr Woźniak 4, Bogusław Sobolewski 4
Correspondence author: olasobolewska1@poczta.onet.pl
Re-written (less scientifically) for the benefit of those suffering from QTS/FQAD (see abstract) and other interested lay-persons by Fluoroquinolone Toxicity Support UK. With apologies to the original authors and those who find some of the science in our text possibly inaccurate in detail. Our aim is for as many people as possible to be able to read and understand how painful and damaging the after effects from these drugs can be. The original paper can be viewed here:-
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5632915/
Numbers appearing thus [1] indicate reference papers found at the end of this study.
Letters & symbols appearing thus (FQAD) are explained at first use.
Abstract
In recent years a significant medical and social problem has arisen where patients, after being prescribed the fluoroquinolone class of antimicrobials, suffer for many years from tiredness, concentration problems, neuropathies, tendinopathies and other symptoms. These long-term effects have been referred to as Fluoroquinolone Associated Disability (FQAD) or Quinolone Toxicity Syndrome (QTS).
Detailed knowledge about the molecular activity of Fluoroquinolones (FQs) in the cells remains unclear in many aspects, while the 'effective' treatment of the resulting chronic state remains difficult and not at all effective. This paper reviews the biochemical properties of FQs that can cause changes in tissues and organs, hints at directions for further research and reviews the existing research concerning the proposed treatment of patients.
Based on the analysis of existing literature, seven main directions of possible effective treatment of FQAD are proposed:-
(A) reduction of the oxidative stress
(B) restoring reduced effectiveness of the mitochondrial membrane
(C) supplementation of metals that are chelated (“hijacked”) by FQs
(D) stimulating mitochondrial proliferation
(E) removing FQs permanently accumulated in the cells (if this happens)
(F) regulating disturbed gene expression
(G) regulating disturbed enzyme activity
1. Long Term Adverse Reactions caused by Fluoroquinolones
Fluoroquinolones (FQ) belong to the broad-spectrum antibiotic group, meaning they are effective against both gram-negative and gram-positive bacteria. There are many FQs but the most frequently prescribed are ciprofloxacin, levofloxacin, moxifloxacin, norfloxacin and ofloxacin.
FQs destroy bacteria by preventing the bacterial DNA from duplicating. They do this by inhibiting two essential enzymes called topoisomerase II and gyrase. For the last three decades FQs have played an important role in the treatment of serious bacterial infections, especially hospital-acquired infections. However, due to the possibility of serious side-effects and also the UK's ongoing Antimicrobial Stewardship Campaign, these drugs are not currently first-line medicines and their use has become more restricted. FQs should be reserved for those who do not have alternative treatment options.
In 2016 the US Food and Drug Administration (FDA) upgraded existing “black box” warnings for FQs, acknowledging that they are associated with disabling and potentially permanent serious side effects. These side effects can involve the disruption of tendons, joints, muscles, nerves and central nervous system and can also initiate type 2 diabetes. Due to the increasing number of reports about FQ toxicity and long-term complications, the FDA has introduced significant restrictions on their use in recent years, particularly in children and in people aged 65 years and over.
1.1 Tendon rupture
FQs are associated with an increased risk of tendinitis and tendon rupture. This risk is further increased in those over 60, in kidney, heart and lung transplant recipients, and those also receiving steroid therapy [1-4]). High levels of physical activity may also increase risk, and for this reason some sports medicine specialists have advised avoidance of FQs for athletes.
1.2. Nervous system disturbances
Taking FQs is associated with neurotoxicity [5-8] of which the main symptoms that are linked to FQ treatment include insomnia, restlessness and, rarely, seizure, convulsions and psychosis [9-11]. Many reports point to chronic persistent peripheral neuropathy being generated by FQs [12-18] and one study in 2001 showed a possible association between FQs and severe, long-term adverse effects involving the peripheral nervous system as well as other organ systems [19].
1.3 Cardiotoxicity
A study has shown that FQs prolong the heart's QT interval ('heartbeat'). In some cases, this can be a life threatening condition because a prolonged QT interval can lead to torsades de pointes, a specific type of abnormal heart rhythm that can lead to sudden cardiac death [20].
1.4. Hepatotoxicity, nephrotoxicity - poisoning of the liver and kidneys
Other adverse reactions generated by FQs include hepatotoxicity [21] and nephrotoxicity [22]. A significant study in 2015 reported persistent and delayed multi-symptom serious side-effects apparently triggered by FQ use causing severe problems and disability in previously vigorous and healthy individuals. This study described patients who developed new-onset symptoms both during and also in the weeks following FQ use. The study also showed cognitive, psychiatric, peripheral nervous, gastrointestinal and endocrine issues. [23]
1.5. Diabetes mellitus
A study about FQ use and development of type 2 diabetes mellitus (T2DM) hypothesized that FQs predispose an individual to develop diabetes. It also shows a strong correlation between the increase in FQ use in the U.S. between 1990-2012 and the increase in type 2 diabetes in subsequent years which suggests a large part of type 2 diabetes may possibly be generated by FQ exposure. [24]
1.6. Fluoroquinolone Associated Disability (FQAD)
In 2016 a clinical investigation of a newly identified adverse drug reaction, termed FQ-Associated Disability (FQAD) was conducted. It proved that severe toxicities which develop when cancer patients receive supportive care drugs such as FQs are important, despite being difficult to understand, detect, and to communicate to clinicians. These findings supported the recommendations of the FDA’s advisory committee (see above). Revision of FQ-product labels should be considered to include prominent descriptions of the newly identified FQAD. [8]
1.7. Conclusions
Patients with CNS problems (e.g. epilepsy or arteriosclerosis), prolonged QT interval, elderly persons, individuals who are also using corticosteroids and individuals who have chronic renal diseases should not be treated with FQs. FQs are already contraindicated in children because they cause destruction of the immature joint cartilage in animals, thus their use in paediatrics is restricted to life-threatening infections.
2. Oxidative stress
One of the main effects of FQs in cells is the generation of oxidative stress. Very briefly, each cell in the body contains hundreds or thousands (depending on the type of cell) of mitochondria which together 'generate' energy, or adenosine triphosphate (ATP), from 'food supplies' transported into the cell via the electron transport chain. Mitochondria are the source of the body's power and the total electron current in all human mitochondria can be easily estimated to be about 70 amperes. When something in the generation process goes wrong (such as FQs causing blocks or leakages in the transport system as discussed later) one side effect is the creation of reactive oxygen species which then generate oxidative stress.
Studies [25, 26] have shown the relationship between oxidative stress and percentages of electron leakage (i.e. loss of power). These are normally estimated to be 0.1-0.5% during rest [27-29] and 0.01-0.03% during exercise [30, 31]. Anything disturbing the precise mechanism of electron flow through the electron transport chain nearly always causes more leakage and the percentages can increase up to possibly 10% (about 7 amperes). The majority of toxins (including FQs) that find their way to the transport chain can cause this leak. Increased oxidative stress is one of the limitations of mental or physical exercise, but the extra increase caused by leakage during rest will further reduce the body's capacity for exercise – meaning the patient will feel very tired during even the smallest amounts of physical or mental exercise.
Further studies [30, 32, 33] have looked more closely at cellular metabolism and conclude that this area is of great importance to FQ sufferers as their energy production pathway is strongly disturbed. Two dangerous reactive oxygen species (free radicals) are created during oxidative stress, one is hydrogen peroxide and the other is hydroxyl radical. Overproduction of the latter can lead to cell death and too much free radical damage is thought to lead to ageing and many cancers.
2.1. The role of Mitochondrial Permeability Transition Pores (PTP) in the regulation of energy production
Studies [34, 35] are concerned with the mitochondrial membrane, or rather the mitochondrial Permeability Transition Pore (PTP) which is precisely regulated by many factors. It is essential to the proper functioning of a cell and therefore to the process of energy creation within that cell. If the regulatory cycle is not balanced the outcome can be either cell death or a new 'equilibrium' which might be far from ideal where energy production is very poorly controlled. We will see later that FQs do have an influence on the detailed regulation of PTP and this is a very urgent topic for further research. Interestingly, this influence on PTP regulation means that FQs are also used to treat cancers [36-41]
2.2. How the cell adapts to oxidative stress state
Note: The metabolic rate is the speed at which chemical reactions take place to allow the mitochondria to release energy from food. It varies due to several factors e.g. age, gender, fitness and genetic traits.
A cell's increased oxidative stress state is first characterised by raised hydrogen peroxide levels. Normally, hydrogen peroxide levels inform the cell and nucleus about mitochondrial energy production because it is connected to the metabolic rate of the cell. In the case of disturbed metabolic regulation, however (e.g. due to mitochondrial toxins such as FQs), the increased oxidative stress and disrupted PTP cause energy production to be reduced. The main process that uses energy in the cell is the Na/K (sodium/potassium) pump which is actually the most fundamental process of life because it consumes up to 50% of the generated energy. The Na/K pump is responsible for everything being transported into and out of the cell. If there's not enough energy created to maintain the balance things start to go wrong including the creation of oxidative stress and thus hydrogen peroxide. We have already seen how this reduces the amount of energy available, thus a vicious cycle begins. The cell enters a state which can be called “permanent stress adaptation” and the effect of this on the patient is a feeling of “lack of energy”. Many regulatory processes of oxidative stress adaptation take place, however, and this important process is just one example.
3. Molecular mechanisms of FQ toxicity
It is very important to understand the consequences of oxidative stress generation by FQs because the molecular mechanisms involved may help to determine a possible effective treatment. Possibly thousands of people are waiting for FQ toxicity mechanisms to be understood and for treatment methods to follow [112].
Fluoroquinolones also have epigenetic effects which are highly important. Epigenetics is the study of changes in the genes that, while not involving changes to the underlying DNA sequence, can be passed on to daughter cells. Epigenetic change is a regular and natural occurrence, but it can also be influenced by several factors including age, the environment, lifestyle and disease. Epigenetic modifications can be simply the way in which cells end up as skin cells, liver cells, brain cells, etc. or they can have more damaging effects that can result in diseases like cancer. These epigenetic effects depend on various factors, one of which is reactive oxygen species [42].
There is a great similarity between bacterial DNA (targeted by FQs, see 1. above) and mammalian (i.e. human) mitochondrial DNA. Some authors point to this as indicating FQs can have a direct effect on mitochondrial DNA leading to disturbed mitochondrial regeneration and division [43, 44]. Also at the molecular level is the cytoskeleton, the complex network responsible for each cell's shape and movement. It is connected to energy dissipation and organization in mitochondria [46-49]. Changes in the cytoskeleton have been observed after FQ treatment [45].
3.1. Chelation
Note: a cofactor is a non-protein chemical compound or metallic ion that is required for an enzyme's biological activity to happen. Cofactors can be considered "helper molecules" that assist in biochemical transformations.
Chelation is a type of bonding of molecules to metal ions, and FQs are very good at this. Studies have shown that FQs bond with aluminium, iron, copper, zinc, manganese, calcium and magnesium [50,51]. Unfortunately, the research did not include selenium which is very important in a cell as the co-factor of glutathione (a vital anti-oxidant) which removes hydrogen peroxide (see section 2.2 above), thus this is an important topic for further research.
A further study (52) analysed the ability of different FQs to form complexes (or bind) with various metal ions and human serum albumin (the most abundant protein in human blood plasma). The results found that approximately 50% - 73% of metal ions were bound by FQs in most cases. However, the study also indicates that the resulting complexes can cause a significant reduction in the antimicrobial action of FQs.
Other studies [53, 54] point to the ability of FQs to create complexes with different intracellular molecules (which control proliferation, differentiation and survival of cells for example). One other important consideration that was demonstrated is that the crystal forms of FQs are very stable (i.e. resistant to being broken down or changed) meaning they are hard to eliminate [55].
All these features strongly support the thesis that FQs can survive in the cell for a long time. However, the question remains unclear as to what extent this phenomenon actually takes place and whether it contributes to the chronic symptoms of FQAD.
Metal ion chelation seems to be the most fundamental feature of FQs, which probably leads to all other observed toxic effects. Their antibacterial effect is connected with chelating magnesium which disturbs the gyrase and topoisomerase interaction with bacterial DNA (the bactericidal property of FQs). However, the magnesium bond is weaker than the iron bond [50] which has been studied in its own right [56], with results pointing to the high ability of FQs to absorb iron. This reduces the activity of three enzymes which are essential for collagen building but need iron as co-factor. The inhibition of these enzymes leads to significant changes in the mechanical properties of the collagen, leading to tears, ruptures and weak repairs, i.e. FQ-induced tendinopathy [51, 52, 53].
Iron is also essential for the cytochromes, which are proteins that have important roles in oxidative processes and the electron transfer chain during cell metabolism and cellular respiration. They need iron (as a co-factor) but we don't know how their role is affected if the amount of available iron is reduced due to FQ chelation. It's possible that reduced availability of iron in a cell leads to inhibition of the electron transport chain, electron leakage and oxidative stress as discussed above. Copper is another important cytochrome co-factor, while many enzymes require calcium as a co-factor, and zinc is needed by about 300 enzymes. Zinc is the most abundant metal in the brain and an essential component of numerous proteins involved in biological defence mechanisms against oxidative stress. The depletion of zinc may enhance DNA damage by impairing its repair mechanisms [57].
The effects of magnesium chelation are already well known, especially with respect to cartilage damage. Magnesium is another important intracellular ion and is co-factor of about 300 enzymes. It has been demonstrated that an individual dose of ofloxacin has identical effects on cartilage as a magnesium deficient diet [58], suggesting that quinolone-induced arthropathy (joint disease) is probably caused by a reduction of available magnesium in cartilage [58].
Similar results were observed during cultivation of cartilage cells (called chondrocytes) in a magnesium-free medium [45, 59-61]. Supplementation of magnesium alongside the FQ treatment restored the cartilage damage to some degree [60, 61] however it did not restore the reduced cell division [61]. This suggests that there are other reasons why FQs reduce cell division [44]. Magnesium deficiency in immature dogs induced similar clinical symptoms as FQ treatment [62, 63]. Dietary magnesium is stated to reduce intestinal FQ absorption [64-70] and it's interesting to note that information leaflets for FQs warn that their effects will be reduced if medicines or supplements containing calcium, magnesium, aluminium or iron are taken at the same time. It is also hypothesized that FQs create an intracellular magnesium deficit (too little magnesium within a cell) that can lead to insulin resistance and type 2 diabetes [24].
Summing up, the number of enzymes with reduced activity due to their ion-cofactor chelation is probably long and thus an important topic for further research. The long-term effects of multiple ion chelations by FQs is actually a more important problem as the presented research is only concerned with what is taking place during FQ treatment. It does not describe the chronic problems and damage after the treatment ends. The full outcome of persistent ion chelation and the degree to which it takes place in FQ patients must be analysed and reported.
3.2. Oxidative stress generated by FQs
We have already discussed oxidative stress and indeed many papers study the ability of FQs to generate oxidative stress in the cells. The generation of this stressed state differs in detail for each of the FQs depending on their individual ability to chelate metal ions and also on their possible different abilities to change the activity of enzymes in an ion-independent manner. Studies suggest that the ability of the various FQs to change the activity of different antioxidative enzymes can vary significantly [71,72].
While a lot of literature looks at FQ induced oxidative stress, one paper [73] has measured levels of the redox status change (oxidation) that occurs with different FQs at various concentrations both below and above the therapeutic ones. The observations showed that all FQs showed moderate cytotoxicity (cell poisoning) on tendon cells after 24 hours and more severe, significant toxicity after 72 hours. Also, a rapid increase in toxicity takes place after a given concentration of FQs is reached which is only slightly higher than the therapeutic one. The substantial increase in reactive oxygen species (free radical) production which can lead to serious consequences begins at concentrations approximately 10x higher than the therapeutic one.
Assuming that some people already have compromised mitochondria or otherwise increased free radicals (e.g. as presented in [21]), this toxicity limit may occur at even lower therapeutic concentrations. The paper referenced above [73] is also in agreement with clinical observations that FQAD is observed especially in patients who were treated with higher FQ doses or for a longer period, or were given several FQ courses in a relatively short period of time.
Some papers point to detailed FQ effects on different enzymes, for example, it was noted in one study that increased free radicals led to disruption of critical mitochondrial enzymes and gene regulation [21]. This paper also found that FQ induced oxidative stress generation may take place at lower FQ concentrations in people who have an existing problem with oxidative stress. The most important reasons for this seems to be other toxins already contributing to this problem, trace element deficiencies, or genetic mutations.
Many studies [75-77] demonstrate other effects along with the increased oxidative stress state after FQ treatment, however changes in enzyme activity differ between experiments, tissues and the individual FQs, suggesting there is variability in the common outcomes. What seems to be important is the reduction in antioxidant activity which is the first line barrier against free radical damage and apoptosis (cell death). New experiments must estimate these enzyme changes in detail, while more attention must also be paid to the FQ-induced disruption of the antioxidant processes, both in the cell itself and also in the mitochondria.
3.3. Reduction of mitochondrial membrane potential by FQs
The mitochondria, as we have seen, are responsible for generating the body's power using food molecules as their fuel. These molecules have to be transported through the double-walled membrane and, here, the word 'potential' refers to the difference in voltage found on either side of the membrane. One of the symptoms of FQ-damaged cells is the reduction of the mitochondrial membrane potential [37, 38, 77-79], however, the detailed mechanism of this phenomenon remains unknown. It's possible that it stems from factors that regulate mitochondrial Permeability Transition Pores (PTP) opening (see 2.1 above), which can induce the PTP to create large holes thus causing leakage [80, 81]. One such factor would be oxidative stress.
It's worth noting that the decrease in membrane potential is characteristic of the oxidative stress state, while an increase is characteristic for some types of cancers [32]. This observation explains the ability of FQs to treat cancers [36-41] and they are commonly found as part of a chemotherapy cocktail.
FQs are also known to have a tendency to induce epilepsy (epileptogenic activity) in some people which is possibly connected to the PTP. One of the proteins which can support PTP opening is called Translocator Protein* (TSPO) which is also known as a benzodiazepine receptor. TSPO, located on the surface of mitochondria, may activate PTP opening causing membrane potential reduction and leading to apoptosis [80, 81]. Some authors suggest that the epileptogenic activity of FQs is because they have GABA-like structures (similar to benzodiazepines) which may allow them to act as GABA antagonists (i.e. produce stimulant and convulsive effects) [82, 83]. Since TSPO is also a benzodiazepine receptor, similar interaction possibly also takes place between FQs and TSPO leading to opening PTP.
(* called translator protein in original)
The whole problem of the PTP opening is very complicated, however, and involves many other factors related to energy production and the causes of cell death. Broad reviews about regulation of PTP and the main PTP protein (VDAC) are presented in [34,35], while the subject of mitochondrial uncoupling proteins regulation, which may provide another possible way to help FQAD patients, is discussed in [84].
3.4. Does Fenton Reaction take place in FQAD patients?
Fenton Reaction (FR) is important because, if it does occur in FQ patients, it creates free radicals which contribute to oxidative stress. A simple overview of Fenton Reaction is that available metal ions such as iron and copper react with hydrogen peroxide to create the highly dangerous hydroxyl radical which we have already seen (in section 2) leads to cell death.
There is no evidence to prove such a reaction takes place in FQAD patients, however four theories could be put forward for further research:
(a) Risk of FR is higher due to increased concentration of superoxide and hydrogen peroxide;
(b) Reduced membrane potential can affect levels of iron in the cell;
(c) Risk of FR is higher due to ability of FQ to chelate iron, creating complexes that generate hydroxyl radical;
(d) Risk of FR is higher due to upregulation of iron-transporting protein which increases iron concentration in the cell. Similar upregulation has been observed in bacteria [85, 86], however bacterial gene regulation cannot be directly compared to the mammalian one [87, 88].
3.5. Changes in gene expression and enzyme activities after FQ treatment
Many papers point to other effects of FQ toxicity, e.g.:
1. Changes in RNA expression effects in tendon tissue of FQ treated rats including production of significantly less fibrocartilage and poorly organized collagen compared with control animals [89].
2. Changes in gene expression of ciprofloxacin treated prostate cancer cells compared to healthy control cells, where the treated cells showed induced apoptosis (cell death) and a significant increase in some factors involved in the PTP opening state [41].
3. As iron chelators, various FQs inhibit essential enzymes that require iron as co-factor leading to epigenetic effects (i.e. changes in gene expression that do not involve changes to the underlying DNA sequence) which may explain FQ-induced nephrotoxicity and tendinopathy [56].
4. Like all drugs and toxins FQs undergo biotransformation in the liver (at the cytochrome P-450 site) to make them less toxic, but it's possible they create their own delayed toxicity by actually inhibiting the detoxification ability of P-450 [90, 91] plus [92, 93, 94, 95].
3.6. FQs and HIF-1α
Hypoxia is a condition where the tissues are not oxygenated adequately, usually due to an insufficient concentration of oxygen in the blood, and can either affect the whole body or be localised. HIF-1α is a protein which activates over 40 genes in response to hypoxia. The natural function of the HIF-1α system is to change the metabolism of the cell into the anaerobic pathway (i.e. not requiring oxygen) in order to protect the cell against oxidative stress from hydrogen and to produce lactate instead. One study discovered that the HIF-1α protein was eliminated by FQ treatment [56] and the resulting lack of this 'safety valve' in FQ patients may cause a shift of excess amounts of hydrogen into the electron transport chain, causing an electron leak [96].
Assuming that this phenomenon is important in FQ-treated patients, then inhibitors of glycolysis and lipolysis (i.e. the breakdown of sugar and lipids, or fats, respectively) could possibly act to reduce oxidative stress. One natural glycolysis inhibitor in the diet is citric acid which could be supplemented. On the other hand, vinegar (acetic acid) perhaps should be contra-indicated for FQAD patients because it promotes glyco- and lipolysis, both of which might contribute to too much 'hydrogen pressure' and oxidative stress [111].
4. Therapeutic conclusions
The treatment of FQAD, especially that lasting for many years, is a very difficult therapeutic problem. The effectiveness of different therapies carried out on patients is rather low. A large number of patients suffer from tendinopathies, neuropathies, chronic tiredness and need a lot of sleep, sometimes more than 12 hours a day. Understanding all the molecular mechanisms of FQ activity in the cell is an urgent aim for current science in order to find methods for helping these people.
The main question that arises is why, in many cases, do FQAD symptoms last for many years, sometimes after just a standard 5-day treatment. Three reasons can be taken into consideration:
A. Long lasting oxidative stress destroys mitochondrial DNA (mtDNA) resulting in newly synthesized proteins which create cytochrome complexes with disturbed structures leading to permanent electron leakage and even more oxidative stress.
B. The complexes FQs make with proteins and cations (ions) are so stabile (unbreakable) that they exist in the cells for many years, continuing to disturb energy production and epigenetics.
C. Epigenetic changes in gene regulation become persistent many years after FQ treatment even when there is no FQ in the cell.
The answer to which of these three possible reasons contribute to the chronicity (long-term effects) of FQAD symptoms is of high importance with respect to the problem of finding an effective treatment. Research that can answer these questions must be performed as quickly as possible.
In the case of destroyed mtDNA treatment is difficult and it must focus on the stimulation of mitochondrial replication. The destroyed and damaged mitochondria must be removed and the less damaged must replicate in order to replace the removed ones and reduce the total electron leakage (see section 2). After many replications the most healthy mitochondria would dominate the cell although the final effect would depend on the state of the most healthy mitochondrion in each cell.
The second possibility is to increase the ratio of cell exchange (renewal) in a given tissue. The cells with more destroyed mitochondria must be shifted to apoptosis (death) while more healthy cells must replace them. This process, however, cannot take place in the Central Nervous System and muscles because cell exchange is close to zero in these tissues. Also, the exchange rate of collagen cells is very low, causing tendon regeneration to be a difficult and long-lasting problem. (HIF-1α elimination may also contribute to the collagen problem [56]).
If new research confirms the existence of FQs in the cells and mitochondria, in amounts which make their permanent interactions with proteins and cations possible many years after FQ use, the research must also focus on methods of how to remove the FQs from strong protein and cation complexes. The simplest way seems to be the application of increased doses of metal positive ions of iron, copper, manganese, zinc and magnesium, which are all natural FQ-competitors for the protein binding sites. It should be pointed out that metal ions enter the cell to some degree due to the negative membrane potential of the cell and then enter the mitochondria due to the negative value of mitochondrial potential. If the membrane voltage decreases, the lowered potential (characteristic of oxidative stress) may not be able to pull the ions into the cell or mitochondrion up to the required concentration.
There is a strong relationship between the actual membrane potential and ability of each cell or mitochondrion to attract and absorb the ions. The question is, to what extent is the transport of ions into a cell disrupted in FQAD patients? Does lower ion concentration only depend on reduced membrane potential or also on the membrane transport being blocked by FQs joining to the metal binding sites of transport proteins? Every metal ion requires a separate analysis.
The third possible reason for the permanent FQAD state is the ongoing disorder in gene expression caused by some positive loop regulations (or “vicious circles”). For example, reduced iron levels disturb cell metabolism which leads to reduced iron absorption in the cell. Many such loops are possible which could cause the chronic state of the patient despite the lack of any FQs in the cell. These must be recognized in order to find methods to restore the normal regulatory state. This case is the most hopeful for patients because, while mtDNA damage is very difficult to treat effectively, regulation of gene expression is difficult but possible.
Until detailed knowledge concerning FQ toxicity is recognised, the following directions in supporting FQAD patients are proposed according to the known and probable mechanisms of FQ toxicity:
A. reduction of oxidative stress;
B. restoring reduced mitochondrial membrane potential;
C. supplementation of uni- and bivalent cations (metals) that are chelated by FQs;
D. supporting mitochondrial replication in the cell;
E. removing FQs permanently accumulated in the cells (if this phenomenon takes place);
F. regulating disturbed epigenetics and enzyme activities.
A. Reduction of oxidative stress. Assuming that hydrogen peroxide is not effectively removed from the cell after FQ treatment, serious consequences may occur as outlined above. Detailed comparisons of the different FQs with respect to oxidative stress is urgent to determine which FQs are safer to use and which ones are more dangerous. There are thousands of natural substances which possess an antioxidant capacity and which are able to reduce free radicals leaked from the electron transport chain.
The antioxidants which easily enter the mitochondria are the most interesting ones. The mitochondria-targeted antioxidant MitoQ protects against FQ-induced oxidative stress and mitochondrial membrane damage in human Achilles tendon cells. In cells treated with MitoQ the oxidative stress was lower and mitochondrial membrane potential was maintained [79]. Other studies respectively report oxidative damage to collagen was prevented by co-administration of N-acetylcysteine [97]; and similar effects with resveratrol [98]; Vitamin E reduces the consequences of FQ-induced damage [99]; while Vitamin C has been shown to possess the ability to protect mice against lethal gamma photon irradiation (a strong source of oxidative stress) [100].
Trace elements zinc, copper, selenium, iron and manganese are co-factors of important antioxidative enzymes, and selenium supplementation is reported to partially restore oxidative stress and sperm damage in FQ-treated cells [101]. Manganese seems to be very important because it is a co-factor of a mitochondrial enzyme known as SOD2 which protects mtDNA against free radical damage.
This all means that the amount of trace elements in the cell must be satisfactory. Detailed research is required regarding iron and copper in order to ensure their supplementation does not cause Fenton reaction to occur. Also important is that citrate (e.g. citric acid) and other glycolysis inhibitors may reduce the ‘hydrogen pressure’ thus reducing leakage and oxidative stress while acetic acid (e.g. vinegar) could contribute to possible overdosed 'hydrogen pressure', and thus increase leakage and oxidative stress. [111]*
B. Restoring reduced mitochondrial potential. This may be one of several important steps in achieving the proper regulatory balance in FQ-patients, however it is not a trivial task. Reducing oxidative stress may contribute to restoring mitochondrial membrane potential, however the reasons for mitochondrial Permeability Transition Pores (PTP) opening are complex and require advanced research. The points of interest may be re-activating the HIF-1α system (see 3.6 above), reducing intracellular and intramitochondrial calcium concentrations, restoring membrane potential and restoring intracellular magnesium, all of which contribute to PTP closing. Cyclosporine A (an immuno-suppressant medication and natural product) and Metformin (a diabetic drug) are thought to be able to close PTPs [102, 103] and to protect against oxidative stress [102, 104-106], so may be interesting substances for possible FQAD treatment.
C. Supplementation of metal ions that are chelated by FQs. In addition to the points raised in A., the roles of magnesium and potassium must be considered. Both ions have much higher concentrations within cells (intracellular) than outside cells (extracellular). Potassium is probably removed from the cell in the oxidative stress state while magnesium, as we have seen, is strongly chelated by FQs. Supplementation must take into consideration the regulatory effect of the kidneys, which removes excessive amounts of both via the urine. Thus small but frequent doses are recommended in order to keep slightly higher concentrations of both ions in the blood plasma which will facilitate access across the cell membranes and into the cells.
D. Supporting mitochondrial replication in the cell. Supporting mitochondrial exchange (removal of destroyed mitochondria and replication of the healthier ones) is the only answer in the case of irreversible mtDNA damage. A substance that is claimed to possess the ability to promote the mitochondrial biogenesis (production) is pyrroloquinoline quinone (PQQ) [107, 108]. This substance is also stated to be protective against oxidative stress [109].
E. Removing FQs permanently accumulated in the cells. The problem of FQ accumulation is only hypothesised in the available research, therefore it is urgent to establish if this phenomenon really does take place and to what extent. Removing accumulated FQs may take place in two ways: by cytochromes P-450 (the body's de-tox proteins, see 3.5.4 above) and by other processes which can remove the molecule from the cell. Re-activating the reduced ability of cytochrome P-450 detoxification may be an important point in FQAD patients. On the other hand, ozonation (the treatment of a substance e.g. water with ozone) has been described to be an effective method for removing a first generation FQ (flumequine) from liquid water [110]. Thus, ozone therapy should be examined as a method of FQ degradation in the body.
F. Regulating disturbed epigenetics and enzyme activities. All the factors presented above contribute to disturbed gene expression which can then contribute to a vicious circle of degradation causing a new regulatory balance which is very far from the optimal one. If this is the main reason for the chronic state of FQAD patients, then there is a very good chance of finding methods for quick and effective treatment of this state. However, the problem is one of high complexity.
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