Basophilic peripheral nerve inclusions in patient with L144F SOD1 amyotrophic lateral sclerosis

Introduction. Histopathological findings of various inclusions were reported
 in the central nervous system of amyotrophic lateral sclerosis (ALS)
 patients, but not in the peripheral nerves. Case report. Our patient was a
 66-year-old man with lower limb weakness later involvement of upper limbs,
 and loss of sphincter control. Neurological examination showed affection of
 both upper and lower motor neurons. He had paresthesia of the left side of
 his body and socks-distribution numbness. Histopathology of the sural nerve
 and genetic report showed basophilic periodic acid-Schiff (PAS)-positive
 intra-axonal inclusions and heterozygous L144F mutation in the exon 5 of the
 SOD1 gene. Conclusion. It seems that presence of the basophilic peripheral
 nerve inclusions may suggest diagnosis of SOD1 positive ALS.


Introduction
Around 5-10% of patients with amyotrophic lateral sclerosis (ALS) have a genetic basis of the disease. Mutations in SOD1 gene are one of the most common causes of ALS, accounting for around 23% of familial ALS and about 7% of apparently ALS in all studies. Over 185 disease-associated variations in SOD1 have been identified so far 1 .
From 5% to 10% of ALS individuals have at least one additional family member affected by ALS (so called familial ALS -fALS), but the majority of ALS cases (90-95%) occur in people with no prior family history (so called sporadic ALS -sALS). 2 5 . In a patient with the D90A SOD1 mutation, sural nerve biopsy showed loss of large myelinated fibers, and degenerative changes of myelinated and unmyelinated fibers 6 .
To the best of our knowledge, we present the unique case of apparently SALS patient with the L144F mutation in the SOD1 gene who had intra-axonal basophilic periodic acid-Schiff (PAS)-positive inclusions.

Case report
A 66 year old male presented with five years history of progressive weakness with atrophy of distal muscles, especiallly of both legs, as well as a slight weakness and distal muscle atrophy in both arms, rare fasciculations and loss of sphincter control. On admission, he was not able to walk.
Neurological examination of the patient revealed bilateral distal muscle atrophy and rare fasciculations with global mild weakness in the upper limbs. In the lower limbs, patient had bilateral distal muscle atrophy, and provoked fasciculations with severe global weakness. The tendon reflexes were brisk, except ankle jerks which were diminished, and presence of bilateral Babinski sign were noticed. Standing and walking were impossible. He had a socks-distribution paresthesia. Urinary catheter was placed due to incontinence. ALSFRS-r was 38/48. The mini-mental status examination (MMSE) score was normal. Electromyography showed neurogenic changes with actual denervation activity at rest in cervical and lumbosacral regions. Nerve conduction study showed axonal sensory and motor neuropathy, which was more pronounced in the lower limbs.
Laboratory and CSF studies were normal. Magnetic resonance imaging of the brain and spinal cord were unremarkable. Conclusion was that patient fulfilled El Escorial revised criteria for probable ALS, with atypical clinical features such as axonal neuropathy and bladder dysfunction. In the next three months, nerve and muscle biopsy, and genetic examinations were preformed. The patient died five months later at age of 67, due to the cardiopulmonary arrest.
Biopsy of the right gastrocnemius muscle revealed a neurogenic lesion, rare and atrophic end-stage muscle fibers and dominant presence of fatty and connective tissue.
Sural nerve biopsy showed the following: fatty and connective tissue infiltration ( Fig 1A); intense reduction of small and large myelinated fibers in number; secondary remyelination and regeneration ( Figure 1B), together with axonal degeneration (Fig 1C). Basophilic PAS positive inclusions, 14-79 μm in diameter, which were o served intraaxonally in several fascicles. These inclusions were PAS positive (Figs 1D and 1E). The inclusions were described as polyglucosan body-like alterations. Genetic analysis showed pathogenic heterozygous missense L144F mutation in the exon 5 of the SOD1 gene, which led to the final diagnosis of genetic ALS.

Discussion
We presented a patient with the L144F mutation of the SOD1 gene and atypical APBD may be diagnosed incorrectly, based only on the presence of polyglucosan bodies which is a non-specific finding 7 . We registered reduction of small and large myelinated fibers in number, axonal degeneration, as well as remyelination and regeneration that have been previously described in ALS patients 5,6 with PAS-positive basophilic inclusions in the sural nerve specimens that had not been described in SOD1 patients so far. PAS-positive inclusions resembled findings in APBD, so only genetic analyses were able to distinguish between ABPD and ALS 9 . Based on the clinical presentation, needle electromyography and even nerve biopsy, it is not always possible to distinguish with certainty between APBD and L144F SOD1 ALS, so it is necessary to perform genetic analyses that will confirm APBD or ALS.
It is well known that neuronal inclusions containing aggregated SOD1 are pathological hallmark of SOD1 ALS seen both in patients and in transgenic animal models overexpressing mutant SOD1 protein. Some SOD1 ALS patients, including those with L144F mutation, have larger skein-like and Lewy body-like inclusions, in addition to smaller missfolding SOD1 inclusions 10 . We were not able to perform additional enzymatic and genetic analyses for adult polyglucosan body disease, as well as imunohystohemical studies covering epitopes across the entire SOD1 protein to see if the inclusions seen in our patients contain SOD1 aggregates.

Conclusion
Basophilic peripheral nerve inclusions may be associated with SOD1 positive ALS.