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Mitochondria are cell organelles that produce energy. These little power plants transform the energy content of the our food into a form our tissues can use. This process uses up most of the inhaled oxygen. Both inhaled oxygen and nutrients end up eventually into mitochondria where they are transformed into a form of chemical energy that a cell can use, called ATP. In addition of producing ATP, mitochondria also take part in other cellular functions, e.g. in storing calcium, heme synthesis and in synthesizing steroid hormones.
Mitochondrial diseases are a heterogeneous group of probably over hundred different diseases. A common feature for them is some kind of mitochondrial dysfunction. It is believed that the symptoms are caused by defects in energy metabolism. The ost severe mitochondrial diseases manifestdirectly after birth and may lead to the death of an infant. Other mitochondrial diseases have their onset during childhood, teenage, early orlate adulthood, and the mildest forms of the disorders may not affect the patients expected life span. Every age group and almost every medical field has its own mitochondrial diseases. Mitochondrial diseases most often affect muscle, heart and/or brain, and in early childhood also the liver . Some hearing defects, especially in combination with maternal inherited diabetes, are also typical for mitochondrial diseases.
Genes control the function of mitochondria, just as they control all cellular activity. Most of the mitochondrial proteins are produced by nuclear genes. Mitochondria contain also their own genes, called mtDNA, that all are necessary for the energy metabolism of tissues and organs.
Defects in mtDNA affect the energy production in tissues. There are several kinds of known mtDNA defects: large deletions and single amino acid defects. For example the disease called MELAS is caused by a mtDNA defect. It is a tiny mistake in the gene; approximately of the same magnitude as one letter misspelled on 5 pages of text. Yet this is enough to disturb the energy metabolism: mitochondria can’t process all the energy from food to a form that cells can use, which then leads to constant shortage of energy in the tissues. This is why e.g. MELAS mutation can cause skeletal and/or cardiac muscleweakness, diabetes, deafness or symptoms in the brain.
There are up to 100 000 identical copies of mtDNA in every single egg cell. Children inherit their mtDNA from the maternal gamete in conception. So unlike nuclearDNA , originating equally from the mother and father, mtDNA is inherited maternally. How exactly mtDNA defects are passed on to the next generation depends on the quality of the defect, as described below:
Point mutations, i.e. mutations of one nucleotide (mutations associated with e.g. MELAS, MERRF, NARP diseases)
If some of the mtDNA copies in an egg cell carry a gene defect, it is passed on to the offspring. If a mother’s ovum has mtDNA with e.g. MELAS-mutation, then her children will almost always inherit it. If all the mtDNA copies in an egg cell carry a MELAS-mutation, the lack of energy in that cell is so severe that it couldn’t be fertilized. Only mothers pass on mtDNA point mutations to the next generation, and thereforea father carrying a point mutation do not pass on the disease to his children. Not all of the children whose mother carries an mtDNA point mutation get ill. If they have inherited just a few copies of mtDNA with MELAS-defect, they may never display any symptoms.
Large single mtDNA deletions (e.g. mitochondrial muscle disease, some mitochondrial eye muscle weaknesses, Pearson’s disorder, Kearns-Sayre disorder). Large single mtDNA deletions are not usually inherited. They are most likely formed during early stages of foetal development, and do not originate from been the mother. Therefore, it is unlikely that a mother who has a child with a large single mtDNA deletion would get another child with the same defect. Most likely a woman with a large single mtDNA deletion does not pass this defect to her children.
Not at all. Only point mutations in mtDNA (see above) are inherited from a mother to most of her children. A large number of genes in the cell nucleus affect the function of mitochondria, and these genes are inherited from both parents. In that case the disease may be dominant (offspring have a 50% chance to inherit the defective gene from a sick parent) or recessive (the disease manifests only, if the child inherits a defective gene from both parents. The risk is 25%if both parents are healthy).
If a person carries a defect in a protein that maintains mtDNA, mistakes in mtDNA may accumulate with age. These proteins are all coded by genes in the cell nucleus and thus inherited 50% from the mother and 50% from the father. Defects in genes encoding proteins called Polymerase gamma and Twinkle are the most common causes of accumulating mtDNA defects. Polymerase gamma (POLG) and Twinkle replicate and repair mtDNA in cell division. If POLG or Twinkle does nott function properly, it disturbs the replication and repair of mtDNA, which eventually damages mtDNA. Typical consequences of POLG and Twinkle defects are numerous large partial mtDNA deletions, so called multiple mtDNA deletions. These deletions underlie for example the inherited Progressive External Ophthalmoplegia (PEO-disease) and MIRAS (MItochondrial Recessive Ataxia Syndrome, a cerebellar disorder that also causes dysfunction of the peripheral sensory nervous system). If a patient’s muscle biopsy shows multiple mtDNA deletions or lowered amount of mtDNA, usually the POLG-gene is the first gene investigated. Yet a finding of multiple mtDNA deletions is not alone enough for a diagnosis of a mitochondrial disease, since some accumulation of deletions is a usual finding in normalaging tissues.
First of all the doctor may suspect a mitochondrial disease based on the patients symptoms. If the patient displays symptoms in the brain, a brain MRI can be a useful examination, since mitochondrial diseases often display typical changes that are visible in MRI.
Laboratory tests: Measuring the use of energy and the amount of by- and end products of energy metabolism can give information about how well mitochondria are working. If the patient’s tissues use oxygen and produce carbon dioxide effectively, the mitochondria are working effectively (since the end products of energy metabolism are carbon dioxide and water). In case of a disturbance in cellular energy metabolism, the process may produce an overload of lactic acid. This happens if the cells switch to an alternative, non-oxidative and non-mitochondrial way of producing energy, which is also less efficient in ATP production. If mitochondrial energy metabolism does not function properly, amount of lactic acid in tissues and blood will rise. Amount of lactic acid can be measured from blood or cerebrospinal fluid. If lactic acid concentration is increased it points to a mitochondrial disease, but on the other hand not all mitochondrial diseases increase the amount of lactic acid. Sometimes brain symptoms cause only local increase in lactic acid, which can be seen in a spectroscopy-examination that can be done during MRI. Increase of lactic acid can also be followed by exercise test by bicycle ergometry, during and after whichthe amount of lactic acid is measured. Simultaneous measurement of oxygen consumption and carbon dioxide production by spiroergometry can suggest mitochondrial dysfunction. If mitochondria do not function with full capacity, the patient will use less oxygen and exhale less carbon dioxide. If muscle symptoms are evident,the levels of creatine kinase (CK) may increase in blood. Usually CK does not elevate to thousands of units, except in the case of severe children’s dystrophic myopathies, such as thymidine kinase 2 defect.
Muscle biopsy: Examination of patient’s muscle by analyzing a biopsy sample is an important when doctors suspect a mitochondrial disease. The biopsy is usually taken from the quadriceps (musculus vastus lateralis) or deltoid muscle (musculus deltoideus). For adults, the operation is done by a surgeon, with local anaesthesia, or by a neurologist (needle biopsy). Children are usually in general anesthesia during the procedure. The size of the biopsy sample varies from half a little finger nail to approx 1 x 3 cm, or sometimes a smaller sample that can be taken with a biopsy needle, is enough.Muscles are different with different levels of energy metabolism, so not all muscles can be used as biopsy sites. There are different tests that can be made from the biopsy a) Functional tests of mitochondria b) mtDNA tests c) microscopic study of the tissue.
Functional tests of mitochondria. Mitochondria are gently separated from the fresh biopsy sample. The aim is to keep the mitochondria functional even after the separation. The functional testing should be done within 4 hours after taking the biopsy to obtain reliable results. The use of oxygen and/or production of ATP in mitochondria is measured by giving them different nutrients that they can use for producing ATP while using oxygen. In addition, activity levels of respiratory chain enzymes are measured. For example the MELAS-disease tends to cause lack of the enzyme complex I of the respiratory chain, which often can be seen with this test. However, sometimes mitochondrial disorders do not affect the enzyme activities in skeletal muscle.
mtDNA tests. Mitochondrial DNA can be studied from DNA that has been isolatedfrom the biopsy sample. Some diseases (e.g. PEO, decreased amount of mtDNA) typically display clear mtDNA defects in the muscle, but these defects can be completely absent (PEO, large deletions of mtDNA) or be found in smaller amounts (MELAS, point mutations of mtDNA) in the blood cells. In these cases DNA tests from muscle biopsy are the only way to verify the gene defect. Some centres use cells collected from urine for testing mtDNA mutations. Also the amount of mtDNA can be studied.
Microscopic study of the tissue. A part of the biopsy is frozen and stained in a pathology laboratory for analysis of the amount and function of mitochondria. The preparations are investigated under a microscope. So called COX-negative fibres (see picture) are a sign of lowered activity of cytochrome c oxidase , a key enzyme of the mitochondrial respiratory chain. COX negative fibers in children are always an indication of mitochondrial disorder, but in adults their finding is suggestive for the disorder, but not specific. . If the tissue is examined with an electron microscope, large amounts of mitochondria ofabnormal size and shape can be found (see picture).
On the left is a cross-section of a skeletal muscle sample. The fibres shown in blue are COX-negative, which means that they suffer from lowered activity of COX , have mitochondrial dysfunction, and also have a compensatory increase of mitochondria. (photo by A.Paetau)
On the right an electron microscopy image of enlarged mitochondria with abnormal “parking lot” inclusions.
If a patient has symptoms that fit a mitochondrial disease andmtDNA mutation is found, that mutation is the cause of the disease. MtDNA defects can be found from a healthy human usually only in the case that a relative has been diagnosed with a mitochondrial disease with maternally inherited mtDNA point mutation, for example MELAS. The finding of an mtDNA mutation in a patient concerns a wide range of people, in addition to the patient, who may also carry the mutation: all his/her maternal relatives, own siblings, and in the case of a female patient her children, mother, siblings and siblings’ children.
There are hundreds, even thousands of copies of mtDNA in a single cell, so a small amount of mutant mtDNA does not yet disturb energy metabolism. Crucial for disease manifestation is the amount of mutant mtDNA. It all comes down to the egg cell the person originated from: how large amount of the mtDNA of that specific ovum was mutated. For example a patient’s muscle sample may show that 70% of mtDNA carries a MELAS mutation, and 30% of mtDNA is normal. The amount of mutant mtDNA in a tissue is often defines the severity of the disorder . However, the symptoms may manifest late in life, be mild, or never manifest, at all if the amount of mutant mtDNA is small. Therefore, it is possible to be a carrier of MELAS mutation without knowing it, or it affecting health. A family may have subjects with mitochondrial diseases of varyingseverity, as well ashealthy maternal relatives, who carry just a small amount of the MELAS mutation. However, healthy female carriers may pass the mutant mtDNA to their children.
The specific gene defect and disease in question determines whether a relative is in risk of carrying the gene defect, and whether it has potential to affect the subject’s life. A key person giving information is the physician who is taking care of the patient with a mitochondrial disease.
In the case of a mtDNA point mutation, inherited from amother to her children (e.g. MELAS), finding of the mutation in a patient may directly concern a large number of maternal relatives, as they are potential carriers of the mutant mtDNA as well. In this case thephysician should direct healthy relatives and the patient to a expert physician in medical genetics.
In the case of a single large deletion of mtDNA, the genetic defect is usually not inherited, and thus the finding does not affect the patient’s relatives.
In the case of a nuclear gene defect, a specialist in medical genetics can give advice on how the finding affects the patient, the relatives and how this affects family planning.
In Finland, appointments with medical geneticists can be organized by admission at University Central Hospitals, some central hospitals and in Family Federation (Väestöliitto).
Patients with a mitochondrial disease are usually recommended to use cholesterol medication that affects only the uptake of cholesterol, and does not affect mitochondrial function. Statins suppress the function of HMG-coA reductase, an enzyme that is connected to production of cholesterol. This enzyme is also active in synthesizing the mitochondrial enzyme ubiquinon, a part of the mitochondrial respiratory chain, which is often malfunctioning in mitochondrial patients. Since statins affect the synthesis of ubiquinon, they are usually not recommended as cholesterol medication for patients with mitochondrial disease.
The treatment of mitochondrial diseases is always planned individually, and pros have to be weighted against cons when choosing medication. If statins are chosen as medication, regular follow-ups with blood tests are important, with special emphasis on CK-levels.
We originally did our mouse studies with NR (nicotinamide riboside) and it did improve metabolism and also rescue mitochondrial disease signs in these animals. However, NR was not used for humans at the time when we started the trial, and even now, there are no studies in mitochondrial patients with NR. The effects of the two vitamin B3 forms (NR and niacin) are partially different, and for safety reasons, we chose niacin for our study - it has been used for 50 years for patients with high blood lipids. NR is unstable as a compound, and our experience is that even from solid suppliers, the NR content of products vary.
All in all, it may be that NR and niacin turn out to have similar effects for mitochondrial diseases. However, because they affect the NAD+ pathways somewhat differently, we don’t have yet evidence of NR in humans. We showed that niacin is a strong NAD-booster in humans. NR’s safety and effects in mitochondrial disease (and healthy individuals) remain to be studied.
The Finnish Association for Mitochondrial Patients and Families (Mitokondrioyhdistys ry) was founded in March 2018.
Chair: Kim Storås
phone: 0500 853 001