Mito Flashcards
what is a primary mitochondrial disease?
Disease due to a mtDNA mutation- affects the mitochondrial respiratory chain and OXPHOS (Oxidative phosphorylation)
what is the pathogenesis of mitochondrial disease?
Disease is due to- low energy production- ROS generation - reactive oxygen species- lactic acidosis
what is the incidence and age of onset of mitochondrial disease?
Mitochondrial diseases can be present at birth, butcan also occur at any age.Incidence:12.5 per 100,000 in adults4.7 per 100,000 in children
What is the phenotype of mitochondrial disease?
can affect any tissuemost affected are those with the highest energy demand e.g. brain and muscleCNSencephalopathyseizuresdementiastroke-like episodeataxiadepressionEYEopthalmoplegiacataractsptsosisoptic atrophyCARDIAChypertrophic cardiomyopathydilated cardiomyopathyGIdysphagiapsuedo obstructionconstipationhepatic failureENDOCRINEdiabeteshypothyrpidismgonadal failurePNSmyopathyneuropathyHEARINGsensironeural deafness
describe the mitochondrial genome
replicates independently of nDNAcircular- 16.6Kb long and codes 37 genes- 13 mitochondrial peptides, 22tRNAs and 2rRNAsno introns and so splicing is not a mechanism of disease.maternally transmittedeach cell has 100-100,000’s of copeis of the MT genomecan be heterplasmic or homoplasmicThe level of mutant mtDA can vary within individuals from the same family and between tissues in the same individual due to the mitochondrial bottleneck where WT and Mut mt is randonly distributed to daughter cells - this can be uneven resultin in cell lines with higher or lower levels of heteroplasmy
why may a mt variant not be detected in blood?
some variants are lost from blood due to rapid mitotic division.e.g. m.3243A>G- most common mtDNA mut and found in MELAS and MIDD- this makes genetic testing difficult and need to carefully consider the choice of tissue
what is the function of the mitochondria?
Produces energy for the cell via the oxidative phopshorylation pathway (OXPHOS)also involved in ca2+ signalling, cellular metabolism, haem and steroid synthesis
why is there a high mutation rate in mitochondria?
NAME?
what is OXPHOS?
the OXPHOS pathway transfers electrons through e- transport carriers to create an electrochemical gradient across the MM which is used to drive the production of ATP from ADP
What is the heteroplasmy threshold for mitochondrial disease.
For mtDNA variants to manifest with a phenotype there is a threshold level of mutant that needs to be present~40-60% for mtDNA deletions~90% for point mutations in mtDNA tRNAs
what are the 3 mitochondrial deletion diseases and there characteristics?
- de novo- heteroplasmic- several genes deletedPearsons- anemia, pancytopenia, lacic acidosis, pancreatic failure (onset in infancy)Kearns-Sayre- myopathy deafness, opthalmoplegia, cardiomyopathy (adult onset)CPEO- opthalmoplegia, ptosis, imparied eye movement
what are secondary mitochondrial disease?
due to nDNA defects and involves genes that have a direct effect on the function of the mitchondria
Give an example of nDNA mutation affecting mtDNA maintenance and expression
direct effects e.g. POLG, POL2 (specific mtDNA polymerase and TWNKindirect effect e.g. SLC25A4 and RRM2B- affect nucleoside transport, synthesis and salvage- mtDNA needs a supply of dNTPs for replication, im mitotically active tissue this is provided by import from the cytoplasm. In non-mitotically active tissue the dNTPS are obtained from the salvage pathway- these differences result in tissue specific phenotype
Give an example of nDNA mutation resulting in mt dysfunction without affecting mtDNA maintenance or function?
Mutations in the assembly factors of the respiratory chain e.g SURF mutations in MELASmutations in genes involved in mt dynamics e.g. fusion and fission which in turn perturbs the number and distribution of mitochondriae.g. MFN2 in CMT (fusion)OPA1 in optic atrophy (fission)
what are the mtDNA point mutation diseases?
maternally inherited (except AID- sporadic)heteroplasmicmost common mut is m.3243A>G in MEALS and MIIDMELAS- myopahty, encephalopathy, acidosis and stroke like episodesMERRF- myoclonic epilepsy and ragged red fibresNARP- neurogenic weakness, ataxia and retinitis pigmentosaMIDD- maternally inherited diabetes and deafnessAID - amnio glycoside induced deafnessLHON- leber hereditary optic neuropathy
How is mt disease associated with neurodegenerative disorders?
Parkinsons- mtDNA deletions are observed in neurons of parkinsons patients and mutations in genes involved in mitochondrial function are mutated in early onset familal parkinsonsalzheimers - respiratory chain dysfunction due to mtDNA mut found in neurons of patients
How can mt be used as a cancer biomarker?
WARBURG EFFECTtumours preferentially use glycolysis to produce ATP and injury of the respiratory chain is a key event in carcinogenesismt genome could be used as an early biomarker of cancer as mutation does not appear to be restricted to certain cancer types.
What approach is used for diagnosis of mitochondrial disease?
considers phenotype and family history and often requires several approaches including biochemistry, immunohistochemistry, nuclear and mitochondrial DNA testing
what are the requirements and usage of histochemistry in mitochondrial disease diagnosis?
Needs muscle biopsycan stain for ragged red fibres using a gromi trichome stain (sub sarcolemma collection of MT)can test for the activity of specific mitochondrial enzymes- SDH (succinate dehydrogenase) loss is indicative of a complex II deficiency- COX (oxidase) subunitss encoded by both nuclear and mitchondrial DNANormal IHC does not exclude a diagnosis and can see age related mitchondrial defects so need to consider this in older patients
what are the requirements and usage of biochemistry in mitochondrial disease diagnosis?
requires muscle biopsy- want a sample enriched for mtcan measure rates of flux, substrate oxidation and AT|P generation and can measure the level of activity of each XOPHOS substrate seperatelyNeed a lot of sample (50-100mg) may not detect subtle OXPHOS deficiencies especially when mosaic and only a few muscle fibres are affected
How are mt DNA rearrangements detected?
NAME?
How are mt point mutations detected?
common mt mutations can be detected by sequencing- technique need to be sensitive to detect low heteroplasmy (mosaicism)
What is mutation searching for mitchondrial disease?
real time PCR can be used to detect mt copy number- an increase in mtDNA CN suggests that the moelcualr effect is likely to be located in the mitochondrial genome, hence whole mtDNA genome sequencing may be consideredneed to consider heteroplasmy and use a technique that is sensitive to low level mutation85-90% of patients with a suspected primary mitochondrial disease do not have a pathogenic mutation detected in the mdt genome highlighting the importance of considering an nDNA mutation- screen genes involved in OXPHOS or mt genome maintenanceNGS is proving pivital in mt disease diagnosis and >250 nDNA genes have now been shown to cause mitochondrial disease
What are the considerations for mt disease testing (sample type)?
Muscle is best but may not be practical or desired by the patient- however failure make a diagnosis in blood does not mean the patient does not have mt disease, just that the mutation is not detectable in blood- many labs include caveat on their reports to this affect and request a muscle sample if a diagnosis is still strongly suspected.Urine can be used for some mutations e.g. m.3243A>G in MELAS as levels in urine correspond with the level in blood.
