Mitochondrial Myopathies - Part 1 Flashcards
What kind of DNA is the mitochondrial genome and where is it located?
- Mitochondrial genome is double stranded DNA circular molecules that sit within the mitochondria.
Outline the differences between the nuclear DNA and the mitochondrial DNA.
1) . Nuclear DNA:
- 3.2 billion base pairs.
- Approximately 20,000 genes.
- Exons (1.5%) and Introns (25%) of the genome.
- Average gene 9 exons (145bp) and Introns (3365bp)
- Range from 1 exon (0.57kb) in the IFNA6 gene to 79 exons (2090kb) in the DMD gene.
2) . mtDNA:
- Single circular molecule - multiple copies of this molecule however.
- 16,569 base pairs.
- 37 genes.
- No introns.
Describe the mitochondrial genome.
- Exclusively of maternal origin.
- Circular double stranded DNA molecule.
- 16,569bp long.
- Slightly different genetic code to nuclear DNA.
- Contains 37 genes.
- 13 encode components of the respiratory complex - most have many more subunits encoded by nuclear DNA than mtDNA however.
- 22 encode mitochondrial tRNAs.
- 2 encode mitochondrial rRNAs.
- All genes lack introns.
What are the features of mitochondrial inheritance?
- Both sexes equally affected.
- Only maternal transmission of disease - female who is effected could potentially transmit the disorder to all of her children.
- Mainly effects mitochondrial rich tissue that are heavily dependant on oxidative phosphorylation for energy production (skeletal and cardiac muscle / CNS / inner ear / kidney / pancreas).
- Heteroplasmy and Homoplasmy - if you have a mitochondrial mutation it will probably be present in only some of the copies of mitochondrial DNA. May have a mix of normal and mutant mtDNA (heteroplasmy) or a situation where every single mtDNA copy carries the mutation (homoplasmy). Homoplasmy is less common but you do get it in LHON.
- Mitochondrial bottleneck - primary oocyte has about 100,000 mtDNA molecules but when the secondary oocytes are produced not all the mtDNA molecules are replicated. Only about 100 mtDNA molecules are selected for replication. This is called the mitochondrial bottleneck. For some reason the mitochondrial DNA molecules containing the mutations are the ones that get selected for replication when forming the secondary oocytes. What can happen then is you can have this switch within a single generation from a mother whose eggs contain very few mutant mitochondrial DNA molecules to a child that has very high mitochondrial DNA mutation load.
- Threshold effect - it is possible that although you might select out mutated mtDNA it is also possible that some normal mtDNA may be selected out. Therefore, some of the offspring of a mother carrying the mutation may have very few of the mutant mtDNA molecules and thus a very low mutation load. You may also get some offspring that have a very high mutation load and anything in between these two extremes. As long as you have a mutation load that lies below a certain threshold you will not develop clinical symptoms of mitochondrial disease. As soon as you cross this threshold you start to get symptoms. Once you cross the threshold every small percentage increase beyond the threshold value will make your symptoms far far worse.
What do we mean by heteroplasmy and homoplasmy when talking about mitochondrial diseases?
Heteroplasmy and Homoplasmy - if you have a mitochondrial mutation it will probably be present in only some of the copies of mitochondrial DNA. May have a mix of normal and mutant mtDNA (heteroplasmy) or a situation where every single mtDNA copy carries the mutation (homoplasmy). Homoplasmy is less common but you do get it in LHON.
What does the term ‘mitochondrial bottleneck’ mean?
Mitochondrial bottleneck - primary oocyte has about 100,000 mtDNA molecules but when the secondary oocytes are produced not all the mtDNA molecules are replicated. Only about 100 mtDNA molecules are selected for replication. This is called the mitochondrial bottleneck. For some reason the mitochondrial DNA molecules containing the mutations are the ones that get selected for replication when forming the secondary oocytes. What can happen then is you can have this switch within a single generation from a mother whose eggs contain very few mutant mitochondrial DNA molecules to a child that has very high mitochondrial DNA mutation load.
What is meant by the ‘threshold effect’ with regards to mitochondrial diseases?
Threshold effect - it is possible that although you might select out mutated mtDNA it is also possible that some normal mtDNA may be selected out. Therefore, some of the offspring of a mother carrying the mutation may have very few of the mutant mtDNA molecules and thus a very low mutation load. You may also get some offspring that have a very high mutation load and anything in between these two extremes. As long as you have a mutation load that lies below a certain threshold you will not develop clinical symptoms of mitochondrial disease. As soon as you cross this threshold you start to get symptoms. Once you cross the threshold every small percentage increase beyond the threshold value will make your symptoms far far worse.
What is the incidence of pathogenic mitochondrial DNA mutations?
1 in 250 people carry pathogenic mitochondrial DNA mutations.
What is the birth prevalence of mitochondrial disease?
The birth prevalence of mitochondrial disease is approximately 1 in 5000.
Describe the epidemiology of mitochondrial disease.
- 1 in 250 people carry pathogenic mitochondrial DNA mutations.
- Birth prevalence is approximately 1 in 5000.
- Presentation slightly different in adults to children.
- Can present at any age. Encephalo-myopathic disease is common in children. Myopathic or cerebellar disease is common in adults.
- Another rule of thumb - if you see a child with a mitochondrial disorder it is very likely that it is caused by mutations in nuclear genes. If you see an adult with a mitochondrial disease it is more likely to be caused by mutations in the mtDNA.
Give a general overview of which tissues / organs mitochondr ial disorders usually affect.
General - Intrauterine Growth Restriction (IUGR), Failure to thrive (FTT), Short Stature.
CNS - Acute encephalopathy, seizures, hypotonia or hypertonia, dystonia, cerebellar ataxia, migrane, stroke-like episodes, developmental delay, regression.
Ear/Eye - Sensorineural hearing loss (SNHL), auditory neuropathy, ptosis, PEO, cataract, optic atrophy, pigmentary retinopathy.
Neuromuscular - Muscle weakness, exercise intolerance, rhabdomyolysis, axonal or demyelinating peripheral neuropathy.
Cardiac - Hypertrophic Cardiomyopathy (HCM) or Dilated Cardiomyopathy (DCM), non-compaction of myocardium, conduction defects.
Endocrine - Diabetes mellitus, GH deficiency, adrenal insufficiency, hypothyroidism, hypoparathyroidism.
Renal - Proximal tubulopathy, steroid resistant nephrotic syndrome with focal glomerulosclerosis.
Blood - Sideroblastic anaemia, neutropenia, thrombocytopenia, pancytopenia.
What you would see is a very well delineated syndrome, or with some combination of features from each of these tissues or groups which together should make you think about mitochondrial disease as a possibility.
What are the basic investigations that you might want to do when you think that a patient might have an underlying mitochondrial disorder?
- Full Blood Count which might show pancytopenia.
- Plasma glucose / lactate / amino acids / acylcarnitine profile / thyroid function tests / Liver Function Tests / Urea and Creatinine / Biotinidase.
- Urine organic acids.
- Faecal elastase.
- CSF protein / lactate / amino acids / pterins / neurotransmitters.
- ECG.
- Echocardiogram.
- Eye examination.
- Audiological assessment.
- EMG and nerve conduction studies.
- MRI brain and MRS.
- Muscle biopsy (most important diagnostic tool for diagnosing mitochondrial disease).
What might you expect to see on an MRI/MRS test for a mitochondrial dystrophy?
MRI:
- Cerebellar cortical atrophy.
- Cerebellar atrophy.
- Calcification in basal ganglia.
- Leukodystrophy.
- T2 hyperintensities in basal ganglia and brain stem.
- Stroke-like lesions.
MRS:
- Lactate doublet at 1.3 ppm.
What is one of the key diagnostic tools for mitochondrial disease?
Muscle Immunohistochemistry:
3 stain types used in muscle biopsy for diagnosing mitochondrial disease.
1) . Gomori Trichrome stain - will see ragged red fibres, essentially they are red because there is sub-sarcolemma collection of mitochondria due to excessive replication of mitochondria.
2) . COX stain - get uniform brown staining of muscle fibres which could be dark or light. Sometimes will see fibres that don’t stain at all and these are COX negative fibres. If you see more than about 1-2% of COX negative fibres then you are probably looking at mitochondrial disease.
3) . SDH stain - the SDH stain will show deep blue staining in the same fibres that are COX negative on COX staining. Excessive staining of abnormal muscle fibres.
Can also do electron microscopy which may show you abnormal mitochondria.
Outline specialised / targeted investigations that may be used for the investigation of mitochondrial diseases.
- Plasma / urine thymidine and deoxyuridine levels together with platelet or leukocyte thymidine phosphorylase assay are diagnostic tests for MNGIE.
- Peripheral blood mononuclear cell Coenzyme Q10 assay may be used if you are specifically thinking that there may be a Q10 deficiency which can present as cerebellar disease.
- You can quantify cardiolipin in blood or in cultured skin fibroblasts doing something called the MLCL/TLCL ratios - would get more MLCL compared to TLCL if you had Barth syndrome. This is a very specific test for Barth syndrome.
- Can check the activity of pyruvate dehydrogenase (PDH) if you are thinking of PDH deficiency.
- Can also assay each of the mitochondrial respiratory complexes.
