Endocrine Journal 1997, 44(6), 805-809

A Case of Mi

Acidosis and

Mellitus and

tochondrial Encephalomyopathy, lactic

Stroke-like Episodes Associated With D Hypothalamo-Pituitary Dysfunction

iabetes

TAi TosHI JOKO, KEN-IcHI IWASHIGE, TosHIHIKO HASHIMOTO, YASUHIRO ONO, KUNIHIsA KOBAYASHI, NAOTAKA SEKIGUCHI, TATSUYA KUROKI, TosHIHIKO YANASE, RYoIcHI TAKAYANAGI, FUMIo UMEDA, AND HAJIME NAWATA

Third Department of Internal Medicine, Faculty of Medicine, Kyushu University, Fukuoka 812-82, Japan

Abstract. A 45-year-old woman with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS) had muscular atrophy, severe cerebral and cerebellar atrophy, and cardiac hypertrophy. She also had diabetes mellitus treated with insulin, and sensorineural hearing loss. Ragged-red fibers were observed on muscle biopsy and an adenine to guanine transition mutation at

position 3243 of her mitochondrial DNA was confirmed. Further investigations revealed that she also had hypothalamo-pituitary dysfunction. It appears that diabetes mellitus, hypothalamo-pituitary dysfunction, and the other abnormalities are all associated with mitochondrial dysfunction in this

patient.

Key words: Mitochondrial dysfunction, Endocrine abnormalities, Position 3243 mitochondrial mutation, MELAS, Diabetes mellitus

(Endocrine Journal 44: 805-809,1997)

MITOCHONDRIAL encephalomyopathies are increasingly recognized as a group of multi-system

disorders, often associated with ragged red fibers on muscle biopsy and ultrastructural abnormalities

of the mitochondria. These diseases have been

grouped into syndromes such as mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), myoclonus epilepsy with

ragged-red fibers (MERRF), Kearns-Sayre syndrome (KSS), and chronic progressive external

ophthalmoplegia (CPEO). MELAS patients are usually normal at birth and during the first year of

life, but then show stunted growth, episodic

Received: March 12, 1997

Accepted: August 25, 1997 Correspondence to: Dr. Toshihiko HASHIMOTO, Third Department of Internal Medicine, Faculty of Medicine, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-82, Japan

vomiting, seizures and recurrent cerebral insults

resembling strokes that cause hemiparesis, hemianopia or cortical blindness [1]. The MELAS

syndrome has been shown to be associated with an adenine to guanine transition mutation at

nucleotide 3243 in the dihydrouridine loop of mitochondrial tRNALeu(UUR) [2]. Recent reports have suggested that this mitochondrial gene

mutation is also associated with diabetes and

sensorineural hearing loss [3-6]. Furthermore, the reported endocrinological abnormalities are not

limited to diabetes but also include hypogonadism

[7] as well as hypoparathyroidism [8]. Here we report a case of MELAS associated with both diabetes and hypothalamo-pituitary dysfunction.

Case Report

A 45-year-old woman had a history of normal

806 JOKO et al.

birth and development except for short stature. Her first menstruation was observed at the age of 14 years, and she subsequently married and delivered a child at the age of 27. She was diagnosed as having diabetes at the age of 28 and started insulin therapy 1 year later. Sensorineural hearing loss and rapid, brief myoclonic jerks appeared at the age of 35 and 45 years, respectively. One year before admission her menstruation became irregular and was not observed during admission (3 months). She was admitted to our hospital in April, 1995 with disturbance of consciousness . Physical examination revealed her to be

emaciated, with a weight of 27 kg and a height of 147 cm. Skeletal abnormalities were absent. Her intelligence was below normal limits, with an IQ of 48. Funduscopic examination showed diabetic retinopathy. An ejection systolic murmur (Lev II/ VI) was audible. Muscle atrophy and weakness were generalized, but were more severe in the

proximal limb muscles. Rapid, brief myoclonic jerks were noted and her gait was ataxic. Examination of the cranial nerves showed sensorineural hearing loss. The deep tendon reflexes were decreased and vibration sense was impaired. We informed the patient and her family of the necessity of the following examinations including muscle biopsy and they consented to our

performance of these examinations. Routine laboratory tests on admission are shown in Table 1. Plasma glucose (253 mg/dl) and hemoglobin Aic (8.6%) levels were high. The C-

peptide immunoreactivity (CPR) response to glucagon stimulation was decreased and urinary 24-h excretion of CPR was noticeably reduced. Serum lactate and pyruvate levels were moderately increased, and the former was noticeably increased in the cerebrospinal fluid (CSF). Echocardiography indicated left ventricular hypertrophy. Magnetic resonance imaging demonstrated severe cerebral and cerebellar atrophy with dilatation of the lateral and fourth ventricles. The pituitary gland seemed normal on this imaging study. Light microscopy of a biopsy from the left biceps muscle showed some "ragged red fibers" on modified Gomori trichrome staining (Fig. 1). DNA isolated from peripheral leukocytes and

skeletal muscle was analyzed for the point mutation associated with MELAS. A 322 base pair (bp)

fragment encompassing the mutation site at nucleotide 3243 was therefore amplified by the

polymerase chain reaction (PCR) and digested with Apa I. The forward primer was 5'GGACAAG-

ADAAATAAGGCCT3', and reverse primer was

5'AAGGGTTGTAGTAG000GTA3'. PCR reaction was performed in a final volume of 50 pd containing

25 pmol sense and antisense primers, 0.1 µg DNA, 7.5 pmol of each deoxyribonucleoside triphosphate,

10 mM Tris-HC1 (pH 8.3), 50 mM KCI, 1.5 mM MgC12, 1.25 U Taq polymerase. The reaction

Table 1. Laboratory findin gs on ad mission

Fig. 1. Ragged red fibers in a muscle biopsy specimen

stained by the modified Gomori trichrome method.

Arrow shows ragged-red fibers.

MELAS AND ENDOCRINE ABNORMALITIES 807

conditions were: initial denaturation for 3 min at 94 °C then 33 cycles of denaturation for 30 sec,

annealing at 55 °C for 20 sec and extension at 72 °C for 40 sec

, with final extension at 72 °C for 1 min. The PCR product of our patient was cleaved by Apa I to give three fragments (Fig. 2), suggesting

the presence of an adenine to guanine nucleotide 3243 in the dihydrouridine loop of mitochondrial tRNALeu(UUR) These findings were compatible with

a diagnosis of MELAS. Hormonal studies showed low basal levels of

TSH, FSH, LH, GH and somatomedin C, normal basal levels of ACTH, and high basal levels of

cortisol and PRL (Table 1). Moreover, as shown in Fig. 3, there were abnormal responses of pituitary

hormones to hypothalamic releasing factors, i.e., no responses of ACTH to corticotropin-releasing

factor (CRF), TSH to TRH, or LH to LH-RH, as well as a delayed response of FSH to LH-RH, and

an increased response of GH to GH-releasing factor

(GRF) and PRL to TRH. These data indicated that hypothalamo-pituitary function was abnormal in

our patient.

Discussion

Mitochondrial encephalomyopathy is

characteristic of mitochondrial disorders. Many studies have demonstrated that MELAS patients are also prone to develop diabetes mellitus, and

insulin secretion is reported to be decreased in these

patients [6]. Current evidence indicates that

pancreatic B cells secrete insulin in response to a rise in the ATP level caused by glucose oxidation.

Our patient was diagnosed as having diabetes in her twenties and started insulin therapy about 1

year later. Her insulin response to glucose stimulation was noticeably decreased, indicating

that energy production in the pancreatic B cells

was impaired. Although we did not analyze mitochondrial DNA from pancreatic B cells, an

abnormality of the B cell mitochondria is suspected to underlie such decreased energy production.

She also had other endocrinological abnormalities related to the hypothalamo-pituitary

axis. To our knowledge, there have been 9 patients

Fig. 2. Agarose gel electrophoresis of PCR products

digested with Apa I. Fragments of mitochondrial

DNA (322 bp) encompassing position 3243 obtained from the patients and normal control were

amplified by the PCR, digested with Apa I, and analyzed by agarose gel electrophoresis. lane 1,

leukocyte from the present case; lane 2, skeletal

muscle from the present case; lane 3, positive control from the known patient with a mutation

from adenine to guanine at position 3243; lane 4, normal control.

Fig. 3. Responses of pituitary hormones to their respective releasing factors. Shaded areas show the normal

range. Doses of the releasing factors used in these

studies were as follows: CRF,100 jig; GRH, 50 µg; TRH, 500 µg; LH-RH,100 µg. Plasma level of ACTH

and serum levels of GH, TSH, PRL, FSH and LH were measured by solid phase radioimmunoassay.

808 JOKO et al.

reported who had hypothalamo-pituitary hypofunction associated with mitochondrial

encephalomyopathy (Table 2) [9-15]. The features of these patients were: short stature (130-140 cm), female predominance, and failure of GH, LH and

FSH secretion which frequently occurs in pan-hypopituitarism. The short stature of these patients was probably due to GH deficiency. Hypothalamo-

pituitary hypofunction can be seen in all of the mitochondrial encephalomyopathies, including

KSS, CPEO, MERRF and MELAS. Since hormone secretion is an energy dependent process, a defect of mitochondrial oxidative phosphorylation may

lead to the failure of hormone secretion.

Alternatively, abnormalities in the mitochondria of the blood vessels may cause ischemic damage to

the endocrine organs. Indeed, Hasegawa et al. have reported abnormal mitochondria in the smooth muscle cells of small arteries obtained from a

MELAS patient [16]. In our patient, imaging studies excluded tumors of the pituitary gland or hypothalamus, suggesting that hypothalamo-

pituitary dysfunction was associated with mitochondrial abnormalities affecting her endocrine

organs, but the precise mechanism of such dysfunction in the mitochondrial encephalo-

myopathies remains to be clarified.

Table 2. Cases of mytochondrial encephalomyopathy associated with hypoth

mo-pituitary dysfunction

ala-

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