Three unrelated and non-consanguineous patients with clinical features of MM were evaluated (Table 1). All the patients experienced early adulthood onset of slowly progressive muscle weakness, which preferentially involved muscles in the lower extremities. Patients 1 and 2 had a history of steroid medication under the presumptive diagnosis of polymyositis according to the muscle biopsies and both showed transient responses. Patient 3 suffered from diabetes and showed the EMG findings of superimposed peripheral polyneuropathy.

The genomic DNA was extracted from the peripheral blood leukocytes using a Wizard Genomic DNA Purification kit according to the manufacturer's instructions (Promega, Madison, WI, U.S.A.). All the coding exons as well as the flanking introns of the DYSF gene were amplified using the primer sets designed by the authors (primer sets available on request). A polymerase chain reaction was performed with a thermal cycler (model 9700, Applied Biosystems, Foster City, CA, U.S.A.) using the following conditions: 32 cycles of denaturation at 94℃ for 30 sec, annealing at 60℃ for 30 sec, and extension at 72℃ for 30 sec. After the amplicon (5 µL) treatment with 10 U shrimp alkaline phosphatase and 2 U exonuclease I (USB Corp., Cleveland, OH, U.S.A.), direct sequencing was performed with the BigDye Terminator Cycle Sequencing Ready Reaction kit (Applied Biosystems) using a ABI Prism 3,100 genetic analyzer (Applied Biosystems). All the novel mutations were confirmed by sequencing 100 control chromosomes.

Patient 1 was a 45 yr-old male patient who had presented with progressive weakness in both lower extremities for the previous 17 yr. He noticed difficulty in running and atrophy of the lower leg muscles at 27 yr of age. The muscle weakness and atrophy progressed. A neurological examination showed bilateral motor weakness and symmetric atrophy of the entire lower extremity muscles, which were more severe on the distal part than on the proximal part. Marked atrophy of the lower leg muscles with relatively spared pelvic girdle and upper thigh muscles resulted in a stalk-leg appearance. He required a cane for level walking and also complained of decreased pinch strength. He had been diagnosed of having a polymyositis at the age of 28 yr as a result of a muscle biopsy of the medial gastrocnemius showing myopathic changes with inflammatory cell infiltration. Immunohistochemical stain was not performed on the biopsy specimen. His 43 yr-old younger sister was affected in the same way in her early thirties and found it difficult to walk up and downhills. Since then, the muscle weakness worsened so that walking was still possible but only for short distances using a cane. She also had difficulty in holding her baby due to extended weakness of her upper extremity muscles. Standing on their tip-toe or heels were impossible in both patients. They could stand from sitting position only with the aid of their hands.

Patient 2 was a 30 yr-old female who presented with difficulty in stair walking and pain in her low back and knee. Difficulty in climbing stairs developed at 26 yr of age and this difficulty in walking gradually worsened so that she could not climb stairs without the support of her upper extremities. A neurological examination showed atrophy of both calf muscles, decreased strength of the lower extremity muscles, and an absent tendon reflex. The Gowers' sign was positive. She could not stand on her tip-toe, but standing on her heels was possible. The EMG revealed the myopathic changes in gastrocnemius, rectus femoris, and biceps brachii muscles. A muscle biopsy without immunohistochemial stain from the biceps brachii suggested a diagnosis of inflammatory myopathy. Magnetic resonance imaging of her lower leg disclosed symmetric fatty degeneration and decreased muscle volume of the posterior compartment muscles in both lower legs. The anterior and lateral compartment muscles had either a normal or a relatively normal signal intensity and morphology. The patient denied a family history of neuromuscular disease.

Patient 3 was a 37-yr-old man with quadriparesis who first became aware of periodic weakness in his lower extremities at the age of 18 yr. He noticed difficulties in standing and climbing stairs at the age of 21 and could not walk by himself by the age of 33. An examination at the age of 37 showed incomplete quadriparesis and he could walk with a four-point crutch. His older sister developed lower extremity weakness and atrophy at the age of 18 yr and could not walk by the age of 36.

Direct sequencing analysis identified five heterozygous mutations in the DYSF gene including three novel mutations (Fig. 1). Patient 1 and his sister had compound heterozygous mutations of a novel C to T transition (c.2494C>T) in exon 24 resulting in a nonsense mutation (Gln832X) along with a previously reported missense mutation (c.2997G>T; Trp999Cys). The Gln832X mutation was also found in the patient 2 with a novel missense mutation (c.2974C>T; Trp992Arg). Patient 3 had two mutations including a novel A to T transversion (c.3307A>T) in exon 30, which resulted in a nonsense mutation (Lys1103X), and a previously reported insertion mutation (c.4200dupC) in exon 39 resulting in a frame shift mutation (Ile1401HisfsX8). Three novel mutations (Gln832X, Trp992Arg, and Lys1103X) were not detected in 100 control chromosomes.