MELAS Case Study: A 17-Year-Old's Journey

• Mitochondrial Encephalomyopathy, Lactic Acidosis, and Strokelike Episodes (MELAS) is a mitochondrial disease
• Frank J. Castora is the author of the case report

Case Report:

• A 17-year-old Caucasian female was referred to a neurologist due to severe headache and repeated vomiting over 3 days
• The patient experienced weakness in arms and legs, fatigue after minor exercise, migraine headaches since childhood with auditory and visual hallucinations lately, slurred speech, and slight right-side paralysis (hemiparesis)
• The patient's birth history was normal, 3850 gm weight after unremarkable pregnancy, labor, and delivery, but motor and mental development milestones were abnormal
• The patient was of short stature (<25th percentile) and low weight (<15th percentile), identified with learning impairment since age 8
• Physical examination revealed the patient was febrile at 38.4°C, with elevated serum lactate (24.6 mg/dL) and pyruvate (3.8 mg/dL) concentrations
• Normal values are 5-18 mg/dL and 0.55-1.0 mg/dL, respectively
• The patient's lactate:pyruvate ratio was elevated at 34:1, normal range 10:1 to 20:1
• Analysis of lumbar cerebrospinal fluid (CSF) showed high protein at 0.66 mg/dL
• Normal range 0.18-0.58 mg/dL, and increased lactate at 3.7 mmol/L
• normal is < 2.8 mmol/L
• Increased pyruvate at 284 µmol/L
• normal is 8–150 µmol/L
• Electroencephalogram indicated conduction slowing in the right hemisphere
• CT scan showed hypodensity in the right temporoparietal and occipital cortices
• Audiometric exam revealed decreased perception of high tones and moderate bilateral sensorineural hearing loss
• Several maternal relatives had hearing loss
• Figure 8-1 indicates ptosis of the left eye, bilateral external ophthalmoplegia, and proximal muscle weakness
• MRI revealed signal intensity and diffusion coefficient changes compatible with acute ischemic infarct and cerebral atrophy
• Lactic acidosis, bilateral hearing loss, progressive muscle weakness, and strokelike episodes indicate a mitochondrial disorder
• DNA from the patient, her mother, grandmother, mother's brother, and younger symptomless sister tested positive for a mutation at nucleotide position 3243 of the mitochondrial DNA (mtDNA) genome

Diagnosis:

• Mitochondrial encephalomyopathy, lactic acidosis, and strokelike episodes (MELAS) is a progressive neurodegenerative disorder with high morbidity and mortality
• Early death is common, ranging from 10 to 35 years, although patients in their 50s have been documented
• Most deaths are due to medical complications, some due to status epilepticus
• Typical presentation includes mitochondrial encephalomyopathy
• Some patients experience neurosensory hearing loss and diabetes mellitus
• Multiple organ systems may be involved
• Physical examination often includes short stature, muscle weakness, and psychiatric problems
• Cardiomyopathy with signs of congestive heart failure can be observed

Strokelike Episodes:

• Hallmark feature of MELAS
• Presentation often occurs at age 4-15 years with the first stroke-like episode
• Episodes initially manifest as vomiting and headache, lasting up to several days
• Seizures and visual abnormalities are followed by hemiplegia
• Seizure types may be tonic-clonic or myoclonic
• Patients may experience migraine headaches
• Some patients may experience hearing loss, which may accompany diabetes
• Palpitations and shortness of breath may be present due to cardiac conduction abnormalities like Wolff-Parkinson-White syndrome
• Acute onset of abdominal pain may reflect pancreatitis, ischemic colitis, and intestinal obstruction
• Numbness, tingling, and pain in extremities can be due to peripheral neuropathy
• Some patients may have Leigh syndrome
• A CT scan or MRI of the brain reveals a lucency consistent with infarction following a strokelike episode
• Cerebral atrophy and calcifications may be observed later
• Serial MRI studies demonstrate lesion resolution
• Electroencephalogram is performed when seizures are a concern
• Mental deterioration usually progresses after repeated episodic attacks
• Psychiatric abnormalities may accompany episodes of stroke
• Encephalopathy may lead to dementia, eventually dying
• High mortality can be caused by cardiac involvement, hypertrophic cardiomyopathy and conduction abnormalities

Diagnosis:

• Combined clinical, biochemical, and morphological information is used to establish a diagnosis
• A three-tiered testing approach is undertaken
• Tier 1: Noninvasive screening, including basic glucose and electrolyte analysis, blood counts, screening for metabolic disorders, measuring blood and urine ammonia, ketones, and lactic acid levels
• Tier 2: Blood and CSF lactate and pyruvate levels, lactate/pyruvate ratio, measuring amino acids in blood, urine, CSF and organic acids in urine and CSF, and ketones and free and total carnitine in blood in blood and urine
• Tier 3: Repeat testing under different conditions (e.g., after a fast), perform skin or muscle biopsy, mtDNA testing
• Lactic acidosis is a very important feature
• Arterial lactate and pyruvate are high, and CSF lactate also may be high
• Unlike tissue-injury lactic acidosis where an increased ratio is coincident with decreased O2 saturation, the increased lactate/pyruvate ratio of MELAS patients is observed at normal O2

Imaging:

• PET studies may reveal a reduced cerebral metabolic rate for oxygen utilization
• SPECT studies can ascertain strokes in individuals with MELAS, tracer accumulates in the parieto-occipital region, can delineate the extent of the lesion, used to monitor the evolution of the disease

Genetic Testing:

• mtDNA isolated from blood cells can be tested for any known mutations associated with the suspected disorder
• Many disease-causing mutations are not detectable in mtDNA isolated from blood cells

Biopsy:

• Includes repeat testing for some of the same compounds and skin and muscle biopsy
• Material from the skin or muscle biopsy must be appropriately prepared
• Biopsy material is used to obtain biochemical, morphological, and genetic information to establish a diagnosis
• Enzyme histochemical staining of muscle fibers can identify abnormal levels of respiratory Complexes II, IV, and V
• Specific immunohistochemical stains can be used to evaluate specific respiratory chain subunits
• Patients with MELAS often demonstrate significantly increased staining for succinate dehydrogenase and reduced staining for cytochrome c oxidase
• Fresh or even frozen muscle can be used to determine electron transport chain (ETC) activity by spectrophotometric assays

Molecular Basis:

• Molecular analysis of mtDNA from muscle biopsies often provides a definitive diagnosis of MELAS
• Individuals with severe clinical manifestations generally have greater than 80% mutant mtDNA in postmitotic tissues such as muscle
• In ~80% of MELAS cases, the mutation is an A→G base substitution at nucleotide position 3243
• 7.5%Possesses a T→C transition at nucleotide position 3271
• At least eight additional point mutations and one four-base pair deletion mutation also have been described.
• Mutations in the nuclear genes that specify mitochondrial proteins can lead to mitochondrial disorders that obey Mendelian genetics

Biochemical Aspects:

• Mitochondrial myopathies involve multiple organ systems, display a wide variety of symptoms
• Central to the utilization of fuel molecules (carbohydrates, protein, and fats) is the catabolism of these large macromolecules into smaller molecules that serve as substrates for the TCA cycle
• The mitochondrion is the site of fatty acid oxidation, the tricarboxylic acid (TCA) cycle, electron transport, and amino acid metabolism
• Carbohydrates and fats are metabolized to acetyl-CoA, which condenses with oxaloacetate to form citrate
• Protein degradation also results in the generation of acetyl-CoA and in amino acids used for several of the TCA cycle intermediates
• The TCA cycle also provides numerous intermediates for anabolic reactions

Electron Transport Chain (ETC):

• The oxidation of fuel molecules results in the release of carbon dioxide and the formation of the reduced electron carriers NADH and FADH2
• Electrons carried by NADH and FADH2 are transferred to the electron transport chain (ETC)
• Electrons enter the ETC at respiratory Complexes I and II
• The electrons from NADH enter at respiratory Complex I (RC I, NADH dehydrogenase) with the oxidation of NADH to NAD+
• Electrons carried by FADH2 are transferred to RC II (succinate dehydrogenase) as the FADH2 is oxidized to FAD
• Electrons from RC I and II are transferred to the quinone form of coenzyme Q (CoQ), which delivers them to RC III (UQ-cytochrome c reductase)
• Cytochrome c then accepts the electrons from RC III, and reduced cytochrome c delivers the electrons to RC IV, cytochrome c oxidase
• Electrons are then used by RC IV to reduce molecular oxygen to water
• At RC I, III, and IV, the transfer of electrons is accompanied by the pumping of H+ in the mitochondrial matrix across

Oxidative Phosphorylation (OXPHOS):

• Approximately 1000 proteins comprise the mitochondrion, the majority are encoded on genes located on nuclear DNA
• Mitochondria-encoded proteins include 7 subunits of NADH-dehydrogenase (RC I), 1 subunit of RC III, 3 subunits of cytochrome c oxidase (RC IV), and 2 subunits of the ATP synthase (RC V)
• In addition to these 13 protein-coding genes, the mtDNA encodes 22 mitochondrial transfer ribonucleic acids (tRNAs) and two ribosomal RNA (rRNA) molecules

NADH & Lactate Regulation:

• Disruption in the flow of electrons through the ETC can lead to an increase in NADH
• The increase in NADH will shut down the TCA cycle
• The buildup of NADH will thus result in depletion of NAD+ stores
• NADH is used to reduce pyruvate to lactate
• Failure of electrons to move through the ETC increases reactive oxygen species (ROS), such as superoxide anion (O2¯)
• Elevated levels of lactate causes a lower pH in the cell
• ROS cause damage to nearby proteins, membranes, and nucleic acids

Genetic Considerations:

• The brain and CNS have the highest demand of all the cells in the body for OXPHOS
• Cardiac and skeletal muscle have the next highest need for OXPHOS
• the endocrine system and skin have lesser requirements for OXPHOS
• Deleterious mutations in mtDNA can cause mitochondrial dysfunction, with the earliest manifestations seen in neurological and muscular abnormalities
• The high error rate and lack of efficient DNA repair systems can lead to a high mutation rate for mtDNA

Genetic Mutations:

• MELAS is a mitochondrial genetic disease
• At least 10 different causative point mutations have been described in mtDNA
• The most common mutation is an A → G transition at nucleotide pair (np) 3243 (A3243G)
• 7.5% Have a heteroplasmic T → C point mutation at np 3271 (T3271C)
• Since both normal and abnormal mitochondria may be present in tissues
• Only offspring of affected mothers are themselves affected; none of the affected males pass on their mitochondrial disease
• Maternal inheritance is one aspect of mitochondrial inheritance

Genetics:

• There are three other aspects of mitochondrial inheritance

• Replicative segregation leading to heteroplasmy (the presence of both mutant and wild-type mtDNA molecules within a cell or tissue).

• During cell division, mitochondria distribute to daughter cells according to their position in the cell during cytokinesis

• If mutant mitochondria are concentrated in a particular area of the cytoplasm, then these mutant mitochondria may locate as a group in one of the daughter cells

• The threshold effect describing the different degrees of dependency of various tissues on OXPHOS

• High error rate and lack of efficient DNA repair systems, leading to a high mutation rate for mtDNA

• Presence of this most abundant MELAS-associated mutation, the A3243G mutation, can be quickly determined using PCR amplification of mtDNA followed by restriction enzyme Apa I

• The sample from the proband had the highest proportion of mutant mtDNA, 71%

Mutations:

• Mutations in tRNALeu (UUR) affect protein synthesis in mitochondria
• The MELAS linked tRNA mutation causes aminoacylation deficiency and a defect in translation initiation
• Mutations is for a signal for terminating through the two rRNA genes

Therapy:

• Although there is no cure palliative treatments can alleviate symptoms
• Modifying the diet, adding vitamins and supplements, and minimizing or avoiding external stresses are important for the patient
• It is important for the MELAS patient to maintain a good nutritional status, and to avoid fasting

Metabolic Therapies

• The goal of the metabolic therapies is to increase the production of ATP and interrupt the progression of this mitochondrial disorder
• Metabolic therapies used for the management of MELAS include: CoQ10, idebenone, dichloroacetate (DCA), carnitine, menadione, phylloquinone, ascorbate, riboflavin, nicotinamide, succinate, and creatine monohydrate
• A lack of long-term follow-up has hampered evaluation
• CoQ10 is a naturally occurring substance that stabilizes respiratory complexes located in the mitochondrial inner membrane, scavenges free radicals and acts as a potent antioxidant

Common Interventions:

• Diet supplements like carnitine may stimulate the uptake of long-chain fatty acids into affected cells Supplementing the diet with carnitine may stimulate the uptake of long-chain fatty acids into affected cells
• Fatty acid oxidation generates acetyl-CoA, which produce NADH and FADH2 to feed into the ETC
• Succinate is an intermediate formed during the metabolism of acetyl-CoA
• Creatine monohydrate increases muscle strength in high-intensity anaerobic & aerobic activities
• This is related to increased intracellular creatine/phosphocreatine, maintaining cellular ATP & stabilizing the mitochondrial permeability transition pore

Medications

• Menadione is a form of Vitamin K3
• Phylloquinone is a form of Vitamin K1
• and Ascorbate are forms of Vitamin C, all donating electrons to cytochrome c
• B-blockers, calcium channel blockers, digoxin, and amiodarone can be used to control cardiac conduction abnormalities (arrhythmias)
• A pacemaker may be inserted to combat heart failure
• Seizures can be managed with anti-epileptic medications such as carbamazepine
• The common antiepileptic drug valproic acid is contraindicated because it depletes the body of carnitine, which may exacerbate symptoms.

Exercise:

• Patients can have moderate treadmill training may leading to the improvement of aerobic capacity and a reduction in resting and postexercise lactate levels
• With concentric exercise training (lifting a weight), a remarkable increase report-edly occurs in the ratio of wild-type to mutant mtDNAs and in the proportion of muscle fibers with normal respiratory chain activity
• If conditions such as cardiomyopathy are present, then exercise should be restricted
• However strenuous exercise may lead to rhabdomyolysis