MELAS point mutation with unusual clinical presentation

MELAS point mutation with unusual clinical presentation

Neuromusc.Disord.,Vol.3. No. 3, pp. 191 193.1993 Printed in Great Britain 0960-8966/93 $6.00 + 0.00 1~,1993 PergamonPressLtd MELAS POINT MUTATION WI...

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Neuromusc.Disord.,Vol.3. No. 3, pp. 191 193.1993 Printed in Great Britain

0960-8966/93 $6.00 + 0.00 1~,1993 PergamonPressLtd

MELAS POINT MUTATION WITH UNUSUAL CLINICAL PRESENTATION ALAN L. SHANSKE,* SARA SHANSKE,$ GABRIELLA SILVESTRI,~ KURENAI TANJI,t DAVID WERTHEIM* and STANLEY LIPPER* *Department of Pediatrics, Queens Hospital Center Affiliation, Albert Einstein College of Medicine, Jamaica, NY 11432; tH. Houston Merritt Clinical Research, Center for Muscular Dystrophy and Related Disorders, and Department of Neurology, College of Physicians and Surgeons, Columbia University, New York, U.S.A.

(Received 26 October 1992; revised31 December 1992; accepted 28 January 1993)

Abstract--Mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) is a multisystemic mitochondrial disorder (Pavlakis et al. Advances in Contemporary Neurology. Philadelphia: Davis, 1988: 95-133) and most patients with the typical MELAS phenotype have a point mutation in mitochondrial DNA, an A to G transition at nucleotide 3243 (Goto et al. Nature 1990; 348: 651-653; Koboyashi et al. Bioehem Biophys Res Commun 1990; 173: 816-822; Ciafaloni et al. Ann Neurol 1992; 31: 391-398). A 9-yr-old boy presenting with chronic asthma and depression was found to have abnormal mitochondria, partial defects of respiratory chain enzymes, and the MELAS point mutation. Key words: MELAS, mtDNA mutation, encephalomyopathy.

revealed severe bifrontal atrophy. Worsening tachycardia and widened pulse pressure were noted the next day. An E K G showed ST/segment depression in leads II, III and AVF consistent with inferior wall ischemia. The patient could not be resuscitated after a cardio-pulmonary arrest on day 5. A muscle biopsy and fibroblasts culture were obtained 4 h post-mortem.


A 9-yr-old African-American male with chronic asthma, gastritis and depression was admitted with a 1 day history of blood-tinged vomitus, shortness of breath and lethargy. At the time of his admission, he had been residing in a chronic care facility for 4 months because of his severe asthma. No muscle weakness had been noted prior to admission to our institution. He was the product of a 28 weeks gestation born to unrelated parents who had lost four midtrimester pregnancies. At the time of his admissfon, he was alert and oriented but had generalized muscle wasting and hyporeflexia. He exhibited Kussmaul respirations, scattered rhonchi subcostal retractions and was tachycardic. Laboratory investigation revealed a metabolic acidosis with a venous pH of 7.21, a bicarbonate of 7, an anion gap of 33, and a lactic acid of 18.9 (normal range 14). A review of multiple past admissions showed the same pattern of acidbase disturbance with the highest bicarbonate after correction of 12. He failed to respond to vigorous fluid, bicarbonate and asthma treatment and on day 3 developed an ileus which was felt to be secondary to his acidosis. On day 4 he developed his first and only tonic-clonic seizure. A CAT scan


Biochemical assays of mitochondrial enzymes in muscle homogenates, D N A extraction and PCR amplification were as previously described [1]. RESULTS

Muscle biopsy

Gomori trichrome showed the presence of ragged-red fibers (RRF), succinate dehydrogenase stain showed increased enzymatic activity, and electron microscopy showed a proliferation of mitochondria with abnormal structure. Biochemistry

Biochemical analysis of mitochondrial enzymes 191


A . L . SHANSKE el al.





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Fig. I. (A) M a p of P C R amplified region o f m t D N A . The b o u n d a r i e s of the P C R fragment are denoted by the p r i m e r termini (P). F r a g m e n t distances are in bp. The t R N A ~c~muR~gene (hatched box) in p a t i e n t s c o n t a i n s an extra H a e III (H) site (bold). In n o r m a l individuals, digestion of the P C R p r o d u c t with H a e I l l generates three fragments (169, 37 a n d 32 bp). In patients, there are two a d d i t i o n a l f r a g m e n t s (97 and 72 bp). (B) A u t o r a d i o g r a m of digestion profile of P C R f r a g m e n t s digested with H a e IIl and electrophoresed t h r o u g h a 12% a c r y l a m i d e gel. N - normal; P = patient. Table 1. Activities of mitochondrial enzymes in muscle homogenate of patient and controls Enzyme Cytochrome c oxidase Succinate-cyt c reductase NADH-cyt c reductase Citrate synthase NADH-dehydrogenase Succinate dehydrogenase

Activity Patient 1.39 0.20 0.66 44.43 19.30 1.09

Controls 2.80 = 0.52 (71) 0.70 = 0.23 (69j 1.02 = 0.38 (78) 9.88 ± 2.55 (73) 35.48 ± 7.07 (76) 1.00 5:0.53 (781

Activity expressed as micromoles substrate utilized rain ~ g ', Controls: mean 5: S.D. Number of controls in parentheses.

in muscle homogenate showed partial defects of cytochrome c oxidase, succinate cytochrome c reductase and NADH-dehydrogenase (Table 1). We feel that these biochemical data are meaningful as this autopsy material was obtained only 4 h post-mortem and mitochondrial enzyme activities remain fairly stable under these conditions. In addition, succinate dehydrogenase, the only respiratory chain enzyme without mtDNA-encoded subunits, had normal activity and citrate synthase, a matrix enzyme reflecting mitochondrial volume, was increased, indicating mitochondrial proliferation. Death

Molecular genetics DNA isolated from muscle of the patient was amplified using primers encompassing the gene for tRNA LeuluuR1 of mitochondrial DNA (Fig. I A). Digestion with the restriction endonuclease HAE III yielded fragments of 97 and 72 bp, which are diagnostic for the A to G transition at nt 3243 found in patients with MELAS (Fig. 1B). Quantitation of the autoradiogram showed that the patient had 83% mutant genomes in muscle. Similar analysis showed that the patient had 52% mutant genomes in fibroblasts and his mother had 8% in blood. DISCUSSION

The MELAS syndrome was first described in 1984 [2] and subsequent reports [3, 4] suggested that this may be a familial disease with maternal inheritance. It is one of several disorders affecting mitochondrial function which have diverse clinical manifestations and which are characterized pathologically by RRF on muscle biopsy. These

MELAS Point Mutation

disorders also include M E R R F (myoclonus epilepsy with RRF), the Kearns-Sayre syndrome (KSS), and progressive external ophthalmoplegia (PEO). In each syndrome, a prominent symptom such as myoclonic epilepsy in M E R F F , ophthalmoplegia in KSS and PEO, and stroke in MELAS, dominates the clinical presentation. Additional features in patients with MELAS include seizures, short stature, recurrent headaches and vomiting, as well as altered mental status [5]. In 1990, an A to G transition at nucleotide 3243 in the tRNA Leu~UtJRl gene of mitochondrial DNA (mt DNA) was described in patients with MELAS, and has now been confirmed in almost all cases with the typical MELAS phenotype [1, 6, 7]. More recently, a T to C transition at position 3271, in the same tRNA LeulUuR~gene was reported in three patients with MELAS [8]. With definition of this disease at the molecular level, its full clinical extent can now be better elucidated. We found the A to G mutation at position 3243 of mtDNA in a 9-yr-old boy with a mitochondrial myopathy who did not have the clinical features associated with the MELAS syndrome. Our patient had severe chronic asthma and depression, and lacked stroke-like episodes and seizures (except for a single near terminal paroxysm). This mtDNA mutation may, therefore, be more common than patients with the typical MELAS phenotype would suggest, and should be looked for in patients with muscular or encephalopathic abnormalities. In fact, molecular genetic analysis of maternal relatives of patients with MELAS has shown that this mutation is often present in individuals who have not had stroke-like episodes, some of whom are asymptomatic and others having only some features of the disease, such as headaches or short stature [1, 9]. In addition, the same mutation has been found in patients presenting with PEO [6, 10, 11], and in one patient with generalized myopathic


weakness alone [11]. In keeping with the maternal inheritance of mtDNA alterations, the mutation was also present in the mother of our patient. She was clinically asymptomatic but had four spontaneous abortions. This mtDNA point mutation, may, therefore, be associated with fetal wastage. REFERENCES

1. Ciafaloni E, Ricci E, Shanske S, et al. MELAS: clinical features, biochemistry, and molecular genetics. Ann Neurol 1992; 31: 391-398. 2. Pavlakis S G, Phillips P C, DiMauro S, DeVivo D C, Rowland L P. Mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes. Ann Neurol 1984; 16: 481-488. 3. Driscoll P F, Larsen P P, Gruber A B. MELAS syndrome involving mother and two children. Arch Neurol 1987; 44: 971-973. 4. Montagna P, Gallassi R, Medori R, et al. MELAS syndrome: characteristic migrainous and epileptic features and maternal transmission. Neurology 1988; 38: 751-754. 5. Pavlakis S G, Rowland L P, DeVivo D L, et al. Mitochondrial myopathies and encephalomyopathies. In: Plum F, ed. Advances in Contemporary Neurology. Philadelphia: Davis, 1988: 95-133. 6. Goto Y-l, Nonaka I, Horai S. A mutation in the tRNA Le°~uuk~ gene associated with the MELAS subgroup of mitochondrial encephalomyopathies. Nature 1990; 348: 651-653. 7. Koboyashi Y, Momo M Y, Tominaga K, et al. A point mutation in the mitochondrial tRNA t*u~UUR~gene in MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes). Biochem Biophys Res Commun 1990; 173: 816-822. 8. Goto Y-I, Nonaka I, Horai S. A new mtDNA mutation associated with mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). Biochem Biophys Aeta 1991; 1097: 238-240. 9. Inui K, Fukushima H, Tsukamoto H, et al. Mitochondrial encephalomyopathies with the mutation of the mitochondrial tRNA L~u~oUR~gene. J Pediatr 1992; 120: 62-66. 10. Ciacci F, Moraes C T, Silvestri G, Shanske S, Schon E A. The "MELAS-3,243" mutation in mt DNA is found in many patients with progressive external ophthalmoplegia (PEO). Neurology 1992; 42:417 (abstract). 11 Hammans S R, Sweeney M G, Brockington M, Morgan-Hughes J A, Harding A E. Mitochondrial encephalopathies: molecular genetic diagnosis from blood samples. Lancet 1991; 337:1311-1313.