Abstracts of the Fifth Annual Meeting of the Italian Association for the Study of the Peripheral Nervous System (ASNP).

Journal of the Peripheral Nervous System 20(Supplement):S1–S31 (2015)

PROCEEDINGS

Abstracts of the Fifth Annual Meeting of the Associazione Italiana per lo studio del Sistema Nervoso Periferico (ASNP) April 9–11, 2015 Torino, Italy President: Leonardo Lopiano Scientific Committee: Guido Cavaletti, Emilia Bellone, Chiara Briani, Marinella Carpo, Dario Cocito, Gianmaria Fabrizi, Fiore Manganelli, Luca Padua, Davide Pareyson, Stefano Previtali, Angelo Schenone Organizing Committee: Dario Cocito, Palma Ciaramitaro, Aristide Merola, Erdita Peci Organizing Secretariat: the office www.theoffice.it; www.asnp.net CROSS-SECTIONAL STUDY OF A POPULATION OF FABRY DISEASE PATIENTS: PILOT STUDY FOR A FABRY-RELATED NEUROPATHY REGISTRY

showed in 3 patients a mild reduction of Sensory Action Potential (SAP) amplitude only for sensory nerves in lower limbs; all these patients were classified as having CRF. Only 2 (14.3%) patients reported a DN4 score more than 0; median DN4 scale value was 0 (range: 0–1). Performing QST, supra-threshold stimulus for cold induced pain was 10.8∘ C (range: 2.6–19.5∘ C) and through PI-NRS cold-induced pain was rated 5 (range: 2–9) at left hand and 4 (range: 2–10) at right hand. The study population does not show a significant burden of neuropathy at TNSc and NCS evaluation, apart from patients previously diagnosed CRF. Similarly, DN4 score does not show significant persistent neuropathic pain. On QST cold is able to induce mild pain. This study is in the pilot phase for the creation of a registry; the feasibility of the project has been verified. We now aim to enlarge the study population to comprehend also pediatric patients, in particular males, since it is known from the literature that young males report neuropathic pain more frequently. Small fiber neuropathy detection will be implemented, possibly with skin biopsy, to verify if there could be a surrogate endpoint for ERT efficacy and/or a marker of organ damage.

Alberti P1 , Pieruzzi F2 , Salerno F2 , Torti G2 , Santoro P3 , Cavaletti G1 . 1 Department of Surgery and Translational Medicine; 2 Department of Health Sciences, University of Milano-Bicocca, Clinical Nephrology and San Gerardo Hospital, Monza, Italy; 3 Neurology O.U., San Gerardo Hospital, Monza, Italy. Fabry’s disease is a rare X-linked lysosomal storage disorder caused by deficiency of 𝛼-galactosidase A, resulting in deposition of glycosphingolipids, particularly globotriaosilceramide (Gb3), in different tissues and organs (peripheral and central nervous system, skin, eyes, heart and kidneys). Gb3 accumulation occurs preferentially within the vascular endothelium and smooth muscle cells leading to progressive vessel occlusion, ischemia and organ dysfunction. Neurological manifestations include cerebrovascular accidents, neuropsychological alterations and small fiber neuropathy. Outcome measures to detect neuropathy were obtained for all subjects as follows: neurological examination formalised through TNSc, Nerve Conduction Studies (NCS) in upper and lower limbs, pain assessment through DN4, Quantitative Sensory Testing (QST). Fourteen patients were enrolled, 78.6% female (n = 11). Median age was 51 years (range: 27–72). Nine (64.3%) patients were given Enzyme Replacement Therapy (ERT) at the time of evaluation; 7 patients (50%) were classified as having Chronic Renal Failure (CRF), of which 2 had grade I CRF, 4 had grade II CRF and 1 had grade IV. Neurological assessment formalised through TNSc showed a score more than 0 in 4 (28.6%) patients; overall, the median TNSc score was of 0 (range: 0–5). NCS © 2015 Peripheral Nerve Society

CHARCOT-MARIE-TOOTH DISEASE TYPE 1A WITH RESPIRATORY INSUFFICIENCY: CASE REPORT AND REVIEW OF THE LITERATURE Allegri I1,3 , Aiello M2 , Gemignani F1,3 . 1 UO di Neurologia, AOU Parma; 2 UO di Pneumologia, AOU Parma; 3 Polo Neurologico Interaziendale, Sezione Malattie Neuromuscolari, Italy. Charcot-Marie-Tooth disease type 1A (CMT1A) presents usually with mild to moderate clinical manifestations and, although phenotypic variability has been recognized, only

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their o-diphenolic structure, OO derived polyphenols possess higher antioxidant activity than other phenols. Among them, oleuropein and its derivatives such as hydroxytyrosol and tyrosol, exhibit potent scavenger activity of superoxides and hydroxyl radicals, with anti-thrombotic, anti-atherogenic and anti-inflammatory activities. In addition to OO, the most useful preparations are extracts from leaves and vegetation water. We tested the potential neuroprotective activity of a phenol-enriched extract from olive mill waste waters (OMP). Our study compared the effects of orally delivered OMP (50 mg/kg, hydroxytyrosol equivalent, five times/week) on CDDP (2 mg/kg/d i.p. twice weekly for 4 weeks) neurotoxicity in vivo. We investigated if OMP could prevent CINP (prevention protocol) or rescue CINP (therapeutic protocol). CDDP administered to Sprague-Dawley rats significantly impaired their growth rate (P < 0.05), enhanced hind paw thermal response latencies (P < 0.01), decreased mechanical sensitivity (P < 0.001) and slowed sensory and motor nerve conduction velocity (P < 0.001). OMP delivered according to preventive protocol showed protective effects, significantly improving all studied parameters, and these beneficial effects lasted for 4 weeks after treatments discontinuation. In parallel, we assessed platinum concentration, in plasma and nervous tissues obtained from rats sacrificed after four weeks of OMP administration. We found that OMP treatment significantly decreased platinum concentration, in plasma and nervous tissues. OMP administration after the last injection of CDDP (therapeutic protocol) also induced a significant recovery of thermal and mechanical sensitivities with partial effect on conduction velocities. OMP administration also improved all altered clinical parameters examined, in particular plasma high-density lipoproteins (HDL), creatinine and urea, indicating further possible beneficial effects. In conclusion, the possible protective activity of OMP CINP may stem from chelating activity of polyphenols on transitional metals. This finding suggests that OMP may interfere with in vivo platinum antitumor activity. Therefore, OMP warrants further investigations before being used in patients following antineoplastic treatment with CDDP. Moreover, focused studies should be designed to figure out whether its chelating activity may have a role to limit CDDP coasting phenomenon.

a few patients may develop an incapacitating disease. In particular, respiratory insufficiency, which is not uncommon in other CMT genotypes, has been reported only in a few CMT1A patients, and is not considered a feature of the disease. A 73-year-old woman had a diagnosis of CMT1A at age 66, when she presented with a 20-year history of trips and falls, with increasing frequency. At that time she had mild to moderate distal wasting, severe weakness of foot and toe dorsiflexion (MRC 3) and mild weakness of plantar flexion and of the intrinsic hand muscles, absent deep tendon reflexes at lower limbs, impairment of pinprick and vibration sensation in the feet, and markedly stepping gait. Motor nerve conduction velocity in the median nerve was 32 m/s, whereas motor action potentials in the peroneal nerve and sensory action potentials in the sural and radial nerve could not be obtained. DNA testing confirmed a chromosome 17p11.2 duplication. When re-evaluated at age 73, she complained of increasing exertional dyspnoea since 1 year. Weakness of the distal muscles of the lower limbs had progressed to MRC 2, and she walked with bilateral ankle-foot orthoses and with the aid of a cane. She had tachypnea on minimal effort and when lying flat, and showed paradoxical movement of the anterior abdominal wall on respiration. Restrictive dysfunction was seen on spirometry, and treatment with Bilevel Positive Airway Pressure (B-PAP) was started. Clinical features of CMT1A were outlined in several large series, which did not mention respiratory problems. However, diaphragmatic weakness was reported in 2 severely affected patients out of 57 CMT1A Brazilian cases by Marques et al. (2005), and in 3 of 61 by Thomas et al. (1997): also these latter patients had advanced forms, with fecal incontinence in 2 and dysarthria and dysphagia in one. Considering that CMT1A is a length-dependent neuropathy, involvement of the diaphragm is not unexpected in view of the length of the phrenic nerve, thus subclinical or oligosymptomatic dysfunction of the respiratory muscles in CMT1A might be more common than usually thought. This deserves to be further investigated prospectively, as it could be relevant in clinical practice.

CISPLATIN-INDUCED PERIPHERAL NEUROPATHY IN RATS IS PROTECTED BY POLYPHENOL RICH EXTRACT FROM OLIVE OIL ADMINISTRATION

COMPARISON OF CLINICAL AND ELECTROPHYSIOLOGICAL RESPONSE TO LENALIDOMIDE OR STEM CELL TRANSPLANTATION IN PATIENTS WITH POEMS

Bianchi R1 , Porretta-Serapiglia C1 , Taiana MM1 , Lombardi R1 , Sassone J1 , Oggioni N2 , Canta A2 , Meregalli C2 , Cavaletti G2 , Lauria G1 . 1 3rd Neurology Unit, Carlo Besta Foundation IRCCS National Neurological Institute, Milan, Italy; 2 Experimental Neurology Unit, Department of Surgery and Translational Medicine, University of Milan “Bicocca”, Monza, Italy.

Bianco M1 , Terenghi F1 , Gallia F1 , Nozza A2 , Scarale A1 , Fayoumi MZ1 , Di Pietro D1 , Nobile-Orazio E1 . 1 2∘ Neurology, Department of Medical Biotechnology and Translational Medicine, Milan University, 2 Department of Medical Oncology and Haematology, Humanitas Clinical and Research Centre, Rozzano, Milan, Italy.

Cisplatin (CDDP) is an antineoplastic drug that causes, as a side effect, peripheral neuropathy in a high percentage of patients. The availability of neuroprotective compound able to prevent and, possibly, reverse chemotherapy-induced peripheral neurotoxicity (CINP) would increase patients’ compliance and be a major advance in treatment. Olive oil (OO) and some extracts or residual components from its production process are sources of healthy unsaturated fatty acids, antioxidants, as phenol compounds, vitamin E and carotenes. Because of

POEMS (polyneuropathy, organomegaly, endocrinopathy, M protein and skin changes) syndrome is an unusual multisystemic disease with neurological disability due to a severe disabling polyneuropathy, with high mortality by multiorgan failure. Electrophysiological features of POEMS neuropathy are characterized by axonal loss and demyelinating

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with a more frequent and severe injury in the lower limbs and in the intermediate nerve segments than in the distal portions. Hematopoietic stem cell transplantation (HSCT) is considered the treatment of choice for POEMS while lenalidomide is the most promising therapy for patients not eligible for HSCT. The objective of this study was to assess the long-term effects on clinical and neurophysiologic parameters in patients with POEMS treated with lenalidomide or HSCT. The clinical and neurophysiologic data were reviewed in 15 POEMS patients treated with HSCT (n: 6) or lenalidomide (n: 9). The MRC sumscore on 16 muscles and nerve conduction studies were assessed before (T0) and after 1 (T1) and 2 years (T2) of treatment and the differences were compared using ANOVA. Combining the two groups of patients, there was a significant improvement after treatment in the mean MRC sumscore (T0 = 65 ± 15; T1 = 69 ± 11; T2 = 71 ± 8; p = 0.000) and in the ulnar mean distal motor latency (DML) (T0 = 3.8 ± 1.2 msec; T1 = 3.3 ± 0.4 msec; T2 = 3.0 ± 0.4 msec; p = 0.02), distal compound muscle action potential (CMAP) amplitude (T0 = 6.8 ± 2.8 mV; T1 = 7.2 ± 3.2 mV; T2 = 7.4 ± 2.9 mV; p = 0.0000) and motor conduction velocity (MCV) (T0 = 37.4 ± 10.7 m/sec; T1 = 43 ± 9 m/sec; T2 = 47.9 ± 9.3 m/sec; p = 0.0003). The difference was also significant when we separately analyzed patients treated with lenalidomide and HSCT while there was no difference in the clinical and neurophysiologic data between the two groups. Treatment with HSCT and lenalidomide significantly but similarly improved clinical and neurophysiologic parameters in patients with POEMS syndrome.

diffuse distal and symmetric neurogenic chronic changes with total denervation at tibialis anterior, gastrocnemius and extensor digitorum brevis, bilaterally. Family history was negative for neurological diseases. The patient was born to healthy non consanguineous parents, and has a 42-year-old healthy sister. By using a custom Ion Ampliseq panel and the Ion Torrent NGS platform, we simultaneously analyzed 33 genes associated with hereditary axonal motor neuropathies and dHMNs (AARS, DNM2, GJB1, IGHMBP2, MFN2, RAB7A, TRPV4, ATP7A, DYNC1H1, HINT1, KIF1B, MPZ, REEP1, VAPB, BSCL2, FGD4, HSPB1, LMNA, NEFL, SETX, YARS, DCTN1, GARS, HSPB3, LRSAM1, PDK3, SLC5A7, DHTKD1, GDAP1, HSPB8, MED25, PLEKHG5, TFG). A new HINT1 homozygous mutation (NM_005340:c.148_149delAC; NP_005331:p.H51Ffs*18) was identified in the exon 2 and confirmed by Sanger sequencing. The mutation leads to a premature stop codon with the loss of critical protein domains and its pathogenicity was supported by in silico predictions. Direct sequencing was performed also in the proband’s parents and healthy sister who were all heterozygous for the mutation. Considering that neuromyotonia may occur at later stages of the disease and that about 20–30% of HINT1-mutated cases show peripheral neuropathy without neuromyotonia, HINT1 mutations should be considered (and screened for) in patients with an autosomal recessive form of distal-motor neuropathy or axonal neuropathy.

MONOMELIC LUMBOSACRAL PLEXOPATHY: A FOCAL VARIANT OF CIDP? NOVEL LOSS-OF-FUNCTION MUTATION OF THE HINT1 GENE IN A PATIENT WITH DISTAL MOTOR AXONAL NEUROPATHY WITHOUT NEUROMYOTONIA

Bono V, Gallia F, Di Pietro D, Scarale A, Nobile-Orazio E. 2∘ Neurology, Department of Medical Biotechnology and Translational Medicine, Milan University, Humanitas Clinical and Research Centre, Rozzano, Milan, Italy.

Boaretto F1 , Cacciavillani M2 , Mostacciuolo ML1 , Spalletta A1 , Vazza G, Briani C3 . 1 Department of Biology, University of Padova; 2 CEMES, Data Medica Group, EMG Lab, Padova; 3 Department of Neurosciences, DNS, University of Padova, Italy.

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is a sensory and motor neuropathy characterized by progressive or recurrent symmetric proximal and distal weakness and sensory dysfunction, developing over at least 2 months. Many atypical clinical variants have been described, including Lewis-Sumner syndrome, distal acquired demyelinating symmetric and purely motor or sensory neuropathy. It also can present with weakness and/or sensory symptoms localized to one or both upper limbs. This variant is known as focal CIDP. A 78-year-old man presented with a history of acute pain and progressive weakness of the right leg leading to complete distal paralysis. Lumbar spine MRI only showed slight disc protrusion L5–S1. Pain improved with therapy for neuropathic pain but strength remained stable. Three months later pain relapsed on the right lower limb with concomitant worsening of weakness that also involved proximal muscles. He needed two crutches to walk. Neurological examination showed severe proximal and distal weakness of the right lower limb and touch dysesthesia below the knee. Tendon reflexes were absent in the right leg, normal in the other limbs. Electrophysiological studies were consistent with right lumbosacral plexopathy with normal findings in the arms and minimal contralateral abnormalities. Cerebrospinal fluid (CSF) study showed increased

Mutations in the HINT1 gene, encoding the histidine triad nucleotide-binding protein 1, have been recently shown to be associated with hereditary axonal neuropathy with or without neuromyotonia and also with distal hereditary motor neuropathies (dHMNs). Here we describe a patient with distal motor axonal neuropathy without neuromyotonia due to a new mutation of the HINT1 gene. A 39-year-old man came to our attention after progressive lower limbs weakness with onset in early childhood. At the age of 12 he had undergone tendon release to improve gait, with little benefit. Subsequently distal upper limbs impairment arose with impaired dexterity (difficulty in opening bottles, lacing shoes) and progressively worsened. When we first saw the patient, he presented with bilateral stepping gait, severe lower limbs distal atrophy and loss of strength. Upper limbs showed diffuse atrophy, more severe at intrinsic muscles of the hands, hypothenar and thenar eminences, and severe weakness. Deep tendon reflexes were reduced at upper limbs, abolished at lower limbs. Neurophysiologic studies revealed signs of motor axonal polyneuropathy. Needle EMG showed

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NEUROPATHY IN POEMS SYNDROME: THE ROLE OF NEW DRUGS IN A MONOCENTRIC COHORT OF PATIENTS

proteins (112 mg/dL) with normal cell count. Sural nerve biopsy revealed a markedly reduced density of myelinated fibers, onion bulb formations, clusters of regenerating fibers and perivascular and endo/epineurial inflammatory cells. MRI of the lumbosacral plexus was normal. An IgM lambda monoclonal gammopathy of undetermined significance (1519 mg/dL) was present. The patient did not improve with intravenous prednisolone while treatment with IVIg was followed by improvement of pain and strength that further improved with subsequent courses of IVIg. He became able to walk a few steps without crutches. This patient had a motor and sensory lumbosacral plexopathy restricted to one limb. The recurrent clinical course, the increased concentration of CSF proteins, the presence of inflammatory cells and demyelinating features in sural nerve biopsy and the consistent improvement after immune therapy support the diagnosis of focal CIDP despite the absence of demyelinating features on nerve conduction studies. Since focal CIDP has been previously reported only on upper limb, this would represent a peculiar case of focal CIDP affecting the lower limb.

Campagnolo M1 , Lessi F2 , Dalla Torre C1 , Lucchetta M1 , Adami F2 , Briani C1 . Departments of 1 Neuroscience, DNS, and 2 Medicine, Hematology Unit, University of Padova, Padova, Italy. POEMS syndrome is a rare multisystemic disease due to a plasma cell disorder, whose pathogenic mechanisms are not fully defined. Therapeutic options are those effective in multiple myeloma, but univocal recommendations are still lacking. We describe our experience regarding the clinical efficacy of two different drugs, Lenalidomide and Bortezomib, in patients with POEMS syndrome. Six patients (4 M, 2 F, mean age 59, range 45–73) were considered. Five patients underwent for at least one year Len-Dex protocol (Lenalidomide 25 mg/day for 21 every 28 days and Dexamethasone 40 mg weekly), whereas the remaining patient, with localized osteosclerotic myeloma, was treated with radiation. The clinical response was scored with Overall Neuropathy Limitations Scale (ONLS), MRC Score and INCAT Sensory Sum Score. VEGF levels were assessed by enzyme-linked immunoadsorbent assay (ELISA) kit (Quantikine; R&D Systems). After the 6th cycle of lenalidomide, four patients showed improvement of at least one point in ONLS score and MRC Sum Score that persisted stable after the 12th cycle. ONLS score and MRC Score improved in the remaining two patients after the 12th cycle. Four patients also presented a reduction of the INCAT Sensory Sum Score after the 6th cycle; among them, only one showed further improvement after the 12th cycle. Of the remaining patients, one patient worsened and the other one remained unchanged. Both these patients had to withdraw the treatment in the following months due to disease progression, and were treated according to BorD and CyBorD protocol (Cyclophosphamide 300 mg/m2 , Bortezomib 1.5 mg/m2 , Dexamethasone 40 mg weekly), respectively. One patient is currently undergoing treatment every 28 days, while the other one, after 5 cycles with monthly administration, is now being treated every 3 months. After one year of therapy, both patients disclosed neurological improvement. After the 12th cycle of Len-Dex, VEGF serum levels decreased in all patients, from a mean initial value of 1947.2 pg/mL to a mean value of 651.75 pg/mL. Both patients who shifted to Bortezomib-based therapy showed a sharp increase of VEGF levels before the new treatment. After initial Bortezomib administration, sVEGF levels fell quickly to normal ranges. In our experience, Lenalidomide is an effective and safe therapeutic option in patients with POEMS syndrome; nevertheless, due to disease progression, two patients needed to be shifted to Bortezomib. Neither of these drugs, potentially neurotoxic, seemed to have worsened the neurological status in our patients. The ongoing follow-up study will help gain further data.

HNPP: NOT ONLY ENTRAPMENT SITES. NERVE ULTRASOUND FINDINGS IN A GUITAR PLAYER Cacciavillani M1 , Briani C2 . 1 CEMES, Data Medica Group, EMG Lab, Padova; 2 Dept. of Neurosciences, DNS, University of Padova, Padova, Italy. Hereditary neuropathy with liability to pressure palsies (HNPP) is an autosomal-dominant neuropathy characterized by focal painless pressure neuropathies at common entrapment sites. HNPP is associated with a 1.5-Mb deletion (rarely a point mutation) of the gene for PMP22. Neurophysiological data show a sensorimotor demyelinating polyneuropathy with conduction abnormalities preferentially localized at common entrapment sites. The diagnosis is easy if family history is present and accurate neurophysiological evaluation is performed. The diagnosis may, however, be challenging in the presence of poor clinical history or atypical presentation. Recently, nerve Ultrasound (US) has increased the diagnostic power showing the presence of nerve abnormalities in multiple entrapment sites, often prompting molecular analysis. We describe curious US findings in a young amateur guitar player, who eventually turned out to be affected with HNPP. A 21-year-old man complained of left ulnar nerve weakness and sensory loss after playing guitar for several hours. Neurological evaluation revealed weakness at left abductor digiti minimi (3/5 MRC) and hypoesthesia at IV and V digits. Neurophysiology showed conduction abnormalities mostly at common entrapment sites, where US also evidenced increased nerve cross-sectional-area (CSA), as already described in HNPP. Interestingly, the palmar cutaneous branch of left median nerve, which had undergone repetitive stress while playing guitar, appeared significantly enlarged (CSA 3.28 mm2 , normal values ≤ 1 mm2 ). Despite negative family history, molecular testing confirmed the clinical, neurophysiological and ultrasound suspicion of HNPP.

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and Maternal-Infantile Sciences and CEBR, University of Genova; 2 Istituto Italiano di Tecnologia (IIT), Drug Discovery and Development Department, Genova; 3 Department of Informatics, Bioengineering, Robotics and System Engineering (DIBRIS), University of Genova, Italy.

WORK-RELATED NEURALGIC AMYOTROPHY OF PRONATOR TERES Capoccitti G1 , Filippou G1 , Vegni V2 , Franci L1 , Insana L1 , Giannini F1 . 1 Dip. Scienze Mediche, Chirurgiche e Neuroscienze, Università degli Studi di Siena, Siena, 2 Servizio di Radiologia, Clinica “Rugani”, Siena, Italy.

Active myelination requires high demand of lipid biosynthesis in Schwann cell processes, and the disruption of lipid biosynthesis may result in abnormal myelin and PNS pathology. In experimental models of CMT1A, we found a down-regulation of genes primarily related to lipid metabolism, a striking and early reduction of sphingomyelin content and a significant up-regulation of acid sphingomyelinase activity, which mirror the dysmyelinating phenotype observed in this disease. Interestingly, the analysis of purified myelin from sciatic nerves showed that the decrease of sphingomyelin in affected nerves is not merely due to the lack of myelin but also to an altered arrangement of these lipids in the myelin sheath. This prompted us to perform a sphingolipid-targeted analysis by high resolution mass spectrometry (LC MS/MS) in sciatic nerves and purified myelin of CMT1A rats to clarify the critical step causing the significant reduction of sphingomyelin. Interestingly, we found an overall derangement of the sphingolipid pathway since the very early precursor synthesis. Therefore, we performed LC MS/MS complete lipid profiling by shotgun analysis. In this case data were acquired over a broad mass range and analyzed altogether at the same time to capture the general picture of the metabolic profile of the samples and to identify metabolic differences between the CMT1A and control groups. Together with sphingolipids deficit, we found a striking impairment of phospholipids species. Moreover, CMT1A rat cerebrospinal fluid lipid profiling revealed a remarkable decrease in myelin glycosphingolipids content that might suggest their use as biomarkers. As sphingo- and phospho-lipids pathways are pharmacologically targetable, we treated CMT1A DRG cultures with compounds able to restore the lipid homeostasis in Schwann cells and improve in vitro myelination. Our preliminary data focused on PIP3 and L-serine as the most promising drugs to ameliorate the CMT1A phenotype. We are confident that improving our knowledge on the lipid pathways altered in CMT1A may contribute to the development of pharmacological approaches aiming at the preservation of the myelin membrane.

Pronator teres (PT) syndrome is a compression neuropathy of the median nerve (MN) at the elbow. It is very rare compared to compression at the wrist (carpal tunnel syndrome) and usually caused by compression in its natural path between the two heads of PT mainly due to muscle hypertrophy, but several other causes are described. Patients typically complain of aching discomfort and weakness in the proximal forearm, and paresthesias in the distal territory of MN. EMG abnormalities in muscles innervated by MN main trunk are common. Despite the syndrome name, PT is involved only in 1/3 of cases, mainly due to nervous branch originating distally to compression site. A 27-year-old male, working as a waiter, reported a 10-month history of mild pain in the proximal aspect of his left forearm, lasting two weeks and followed by increasing focal atrophy in the proximal third of its volar aspect. On examination pronation was weak (4/5 MRC scale) and all other proximal and distal muscles had normal strength. Upper arm tendon reflexes were normal and no sensory loss was detected. NCS studies of the main trunk and anterior interosseous branch of MN were normal. CMAP recorded from PT showed severe amplitude reduction and normal median MCV between axilla and elbow. EMG of PT showed 5/5 fibrillations/positive waves and single unit pattern at maximal voluntary recruitment. EMG of all distal muscles supplied by MN were normal. Ultrasonographic study confirmed severe atrophy of PT and integrity of the main trunk of MN. MRI scan of left elbow showed diffuse hyperintense signal and atrophic involution of PT, edema of soft tissues in the anterior space of the elbow (between lacertus fibrosus and ulnar head of PT) and tendinopathy of common flexor. MN path was not recognizable. In about half of the cases, humeral and ulnar head of PT are independently or together supplied from a single branch arising from the main trunk of MN between 4.5 cm above and 3.5 cm below the interepicondylar line (Gunther 1992, Dogan 2010). Clinical, imaging and electrophysiological features in the present case could be explained by isolated compression of a proximal PT nervous branch by lacertus fibrosus edema sparing the main trunk of MN. Occupational history suggests chronic exogenous compression during working activity as the main pathogenic mechanism. To our knowledge, only two cases of isolated PT neuralgic amyotrophy have been reported, both without pathogenic hypothesis (England 1987).

EFFECTS OF ETHOXYQUIN ON PLATINUM DRUGS-INDUCED PERIPHERAL NEUROTOXICITY Carozzi VA1 , Meregalli C1 , Oggioni N1 , Zhou J2 , Reed N2 , Ruifa M2 , Hoke A2 , Marmiroli P1 , Cavaletti G1 . 1 Department of Surgery and Translational Medicine, University of Milan-Bicocca, Monza (MB), Italy, 2 Departments of Neurology and Neuroscience, Johns Hopkins School of Medicine, Baltimore, MD, USA.

SPHINGO- AND PHOSPHO-LIPIDS METABOLISM IMPAIRMENT AS A POTENTIAL THERAPEUTIC TARGET IN CHARCOT-MARIE-TOOTH TYPE 1A (CMT1A) NEUROPATHY

Ethoxyquin (EQ) is a synthetic antioxidant FDA-approved for use in animal food in order to protect against lipid peroxidation and stabilize fat soluble vitamins. Even if an antimutagenic effect was observed in rodents, its primary use is still as a supplement in animal food. Since oxidative stress is

Capodivento G1 , Visigalli D1 , Basit A2 , Pastore VP3 , Armirotti A2 , Schenone A1 , Nobbio L1 . 1 Department of Neurosciences, Rehabilitation, Ophthalmology Genetics

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cytoskeleton of Schwann cells. DG and DG-interacting proteins have been implicated in the pathogenesis of a wide array of peripheral neuropathies. Merosin-deficient congenital muscular dystrophy results from insufficient expression of laminin-2 (DG’s main ECM ligand), whereas in Charcot-Marie-Tooth 4F periaxin is mutated. Furthermore, DG is a host receptor for Mycobacterium leprae infection and the DG-cleaving enzymes MMP2/9 have been found to be elevated in inflammatory neuropathies. In this study, we found that the genetic deletion of DG in murine Schwann cells (DG Floxed/Floxed//PoCre mice, DGKO mice) leads to progressive demyelination associated with endoneurial edema and inflammatory infiltrates. This phenotype is strictly limited to motor nerves and spontaneously arises at 6 months with a chronic persistence at 8, 10 and 12 months. The demyelination is associated with inflammatory infiltrates mainly composed by macrophages. An increase in proinflammatory cytokines expression has been also detected. According to histological data showing demyelination, a significant slowing of motor nerve conduction is evident at the neurophysiological analysis. Even though further studies are needed to dissect the molecular mechanisms underlying demyelination and inflammation and the selective involvement of motor nerves, the DG deficient mice could provide a useful animal model to better understand the pathogenesis of human neuropathies sustained by altered expression of proteins interacting with DG in a genetic or acquired fashion.

one of the main causes of platinum drugs-induced peripheral neurotoxicity, EQ should be a potential neuroprotective drug against platinum-based chemotherapy. We tested the potential neuroprotective as well as non-interfering properties of EQ in in vitro and in vivo settings investigating the involved mechanistic pathways. In vitro studies demonstrated that EQ was able to restore cisplatin (CIS)-induced ATP deprivation in 50B11 DRG neuronal cells as well as prevent axonal degeneration in primary DRG neurons. These data were corroborated by in vivo experiments when EQ was co-administered with CIS in rats: CIS-induced functional abnormalities in nerves, DRG neuronal atrophy and neuropathic pain were significantly prevented by EQ. Moreover, in vitro and in vivo data support that EQ did not interfere with CIS antitumoral properties; EQ, added together with CIS in PA-1 ovarian cancer cells, had no effect on cancer cell death. Similarly, in vivo studies demonstrated that EQ did not prevent CIS’s ability to reduce tumor growth in PA-1 tumor bearing mice. These results were validated by dot blot analysis that revealed the EQ ability to reduce DNA-platinum adducts in DRG and sciatic nerve but not in PA-1 ovarian cancer cells. Furthermore, EQ reduced the levels of heat shock protein 90 (Hsp90) and two client proteins SF3B2 and ataxin-2. Reducing levels of SF3B2 using RNAi in tissue-cultured neurons was effective against neurotoxicity of CIS and suggested that EQ effect is mediated by these proteins. By contrast, both in vitro and in vivo data revealed that EQ was less effective in protecting against oxaliplatin (OHP)-induced peripheral neurotoxicity: EQ, if added to 50B11 neuronal cells was not able to counteract OHP-induced ATP deprivation. Moreover, EQ co-administered with OHP in human colorectal cancer-bearing mice was not really effective in protecting against OHP-induced functional nerve damage and neuropathic pain. It, however, did not interfere with OHP ability to slow tumor growth. These results support the non-interfering effect of EQ on platinum drugs antineoplastic action. However, it is effectively neuroprotective against CIS, but not OHP-induced peripheral neurotoxicity. This differential effect should be justified by the activation of different molecular pathways and allows future deeper investigations. This work was partially supported by Cariplo Foundation.

PERIPHERAL NEUROTOXICITY INDUCED BY CISPLATIN: A POSSIBLE ROLE OF OCT2 GENE Chiorazzi A, Ceresa C, Canta A, Pozzi E, Cavaletti G, Marmiroli P. Department of Surgery and Translational Medicine, Experimental Neurology Unit, University of Milano-Bioccca, Monza (MB), Italy. Cisplatin (CDDP) is an antineoplastic drug used in clinical practice for the treatment of several tumors such as lung and ovarian cancer. Its use is limited by the development of severe nephrotoxicity and peripheral neurotoxicity characterized by sensory alteration. It was shown that the organic cation transporter-2 (OCT2) is able to mediate cellular transport of cisplatin and that this transporter is expressed in kidneys and in dorsal root ganglia (DRG) of mice. There is some evidence that mutations in OCT2 gene are protective against cisplatin nephrotoxicity but its role as a possible target of specific organ protection is under investigation. To investigate if OCT2 plays an important role in the development of peripheral neurotoxicity induced by cisplatin, we performed an animal model in which we used C57BL/6 mice wild-type for this gene (WT) and C57BL/6 mice knock-out for OCT2 gene (OCT2-/-). We treated all the animals intraperitoneally with cisplatin at the dose of 4 mg/kg twice a week for 4 weeks; during the experiment the body weight changes and the general clinical signs of the animals were monitored once a week. At the end of the study, to analyze the development of peripheral neurotoxicity, caudal and digital nerve conduction velocity (NCV) were performed. At the sacrifice, caudal and sciatic nerves were collected for histopathological analysis, DRG were collected for morphological and

DELETION OF PNS DYSTROGLYCAN RECAPITULATES ASPECTS OF MOTOR INFLAMMATORY NEUROPATHIES Cerri F1 , Lopez ID1 , Bianchi F2 , Riva N1 , Feltri L3 , Comi G1 , Del Carro U2 , Quattrini A1 . 1 Department of Neurology, Experimental Neuropathology Unit, INSPE, Division of Neuroscience, San Raffaele Scientific Institute, Milan, Italy; 2 Department of Neurology, Neurophysiology Unit, INSPE, Division of Neuroscience, San Raffaele Scientific Institute, Milan, Italy; 3 Departments of Neurology and Biochemistry, Hunter James Kelly Research Institute, University at Buffalo, Buffalo, NY, USA. Dystroglycan (DG) is a ubiquitous membrane-associated dimeric protein that connects the cytoskeleton to the basal lamina. In peripheral nerves, DG is mainly expressed by Schwann cells. DG provides mechanical and functional connection between the extracellular matrix (ECM) and the

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morphometrical analysis and skin biopsies were collected to study the intra-epidermal nerve fiber density. The NCV measured in mice knock-out for OCT2 was less impaired versus WT animals and the morphological and morphometrical analysis showed an increase in somatic and nuclear area of DRG neurons, indicating that the expression of OCT2 could be critical for the development of peripheral neurotoxicity. These results evidenced that OCT2 seems to be an ideal target for the establishment of protective therapies aimed to specifically reduce cisplatin side effects and increase the quality of life of the patients. Partially granted by “Fondazione banca del Monte di Lombardia”.

values have been calculated: accuracy, sensitivity, specificity, VVP, VVN, LR+, LR-. Moreover, a multiple regression model has been adopted, with the electrophysiological markers (CMAP amplitude, SAP amplitude, RVM increase, reinnervation MUPs) as predictive variables. 307 consecutive patients (232 m, 75 f; mean age 47.64; range 14–94 yrs; SD 17.16), with 444 TN: 113 brachial plexopathies, 75 radiculopathies, 252 mononeuropathies and 4 lumbosacral plexopathies. The SAP amplitude increase turned out to have a highly specific prognostic role for clinical improvement with a high prognostic accuracy (90% in brachial plexopathy, 60% in nerves), sensibility and specificity (100%). The multiple regression model suggests that in all patients and in the NS group the clinical improvement is mostly related with SAP amplitude increase.

SAP AMPLITUDE IS PREDICTIVE FOR THE CLINICAL OUTCOME IN TRAUMATIC NEUROPATHIES EFFICACY OF 5% LIDOCAINE MEDICATED PLASTER IN LOCALIZED PERIPHERAL NEUROPATHIC PAIN IN ADULTS

Ciaramitaro P1 , Mondelli M3 , Rota E4 , Romano M5 , Logullo F6 , Battiston B7 , Sard A8 , Faccani G2 , Cocito D9 . 1 Clinical Neurophysiology, 2 Neurosurgery Division, CTO, Neuroscience Department, AOU Città della Salute e della Scienza di Torino, Torino; 3 EMG Service, Siena; 4 UOC Neurologia, Ospedale Guglielmo da Saliceto, Piacenza; 5 UOC di Neurologia, AO Ospedali Riuniti Villa Sofia-Cervello, Palermo; 6 Clinica di Neurologia, Ancona; 7 Divisione di Traumatologia e 8 Chirurgia della Mano, CTO, AOU Città della Salute e della Scienza di Torino, Torino; 9 Clinical Neurophysiology, Neuroscience Department, AOU Città della Salute e della Scienza di Torino, Torino, Italy.

Ciaramitaro P1 , Stella M3 , Risso D3 , Germano P3 , Faccani G2 , Cocito D4 . 1 Clinical Neurophysiology, 2 Neurosurgery Division, CTO, Neuroscience Department, AO Città della Salute e della Scienza di Torino, Italy; 3 Department of Plastic Surgery, Burn Center, CTO, AO Città della Salute e della Scienza di Torino; 4 Clinical Neurophysiology, Neuroscience Department, AO Città della Salute e della Scienza di Torino, Italy. Localized peripheral neuropathic pain (NP) may be defined as “a type of neuropathic pain characterized by consistent and circumscribed area(s) of maximum pain”. 5% lidocaine medicated plaster (LMP) is a topical peripheral noninvasive analgesic approved (FDA) as the first line of medication for treating allodynia generated by post-herpetic neuralgia; LMP is recommended as the first line of treatment for localized peripheral NP in US, Europe, and Latin America. According to recent reports, LMP is an effective, safe, and comfortable therapeutic option in patients with localized peripheral NP secondary to traumatic peripheral neuropathies. The objectives of this study are: 1) to evaluate the efficacy of LMP in localized peripheral NP secondary to post-burn plastic surgery; 2) to confirm safety, tolerability and absence of side effects in a short term treatment in adults. In a prospective study we enrolled consecutive patients with painful post-burn scars and localized NP treated with 5% lidocaine medicated plaster-Versatis®. Demographic variables, the size of painful area and pain intensity (measured using VAS) were recorded. The possible neuropathic origin of this pain was defined on the basis of a DN4 questionnaire score ≥4. Inclusion criteria: age 18–75 years; DN4 ≥ 4; VAS ≥ 4; localized peripheral pain (painful area < 70 cm2 ). Exclusion criteria: polyneuropathy, major depressive disorders, treatment with other analgesic drugs. The mean duration of pain before starting treatment with LMP was calculated. NRS was measured before starting treatment (T0), after 1 (T1) and 3 (T2) months. VAS at T0 and T2, PGIC and functional recovery at T2 were performed to evaluate the efficacy of the treatment. Pain reduction ≥50%, percentage of functional recovery and of PGIC ≤ 2 was calculated at T2 to evaluate the efficacy. Twelve patients were included (5 males, 7 females; age 30–64, mean 52.4 ± 9.6 SD). Localized neuropathic pain

The increasing number of high-speed motor vehicle accidents results in more frequent traumatic neuropathies (TN), notably of brachial plexus, with residual limitation in hand function, which may significantly impair the working ability and the quality of life. Less evidence is available regarding the natural history and the neurophysiological time course of TN, both in the absence and in the presence of a surgical repair. In order to improve the clinical management of TN patients, the availability of neurophysiological markers able to predict the functional recovery would be extremely useful. The aim of the study is to evaluate the prognostic value of electrodiagnostic studies (EDX) and to identify the neurophysiological markers predictive of clinical recovery in a large sample of TN patients. In a longitudinal, multicentric study, we enrolled all consecutive patients with a clinical diagnosis of TN, referred to the Centers included in the Italian Network for Traumatic Neuropathies in the period between January 2010 and December 2012. All the patients were divided into three groups based on the type of intervention (Surgery-S or No Surgery-NS) and the correspondent timing for clinical and electrophysiological follow up. EDX study including nerve conduction study (NCS) and electromyographic examination (EMG) were performed according to a standard protocol shared among the participant Centres. The clinical evaluation in the follow-up was carried out by using MRC and the modified Rankin Scale (mRS), both in the pre-surgical and in post-surgical phase. Statistical analysis has been carried out on the patients who have had at least one clinical (mRS) and one EDX follow-up to calculate the prognostic accuracy of each EDX marker in predicting the surgical/non surgical recovery. For each EDX outcome measure the following

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affected upper (9 patients) and lower (3 patients) limbs. The mean duration of pain before starting treatment with LMP was 3.2–14.8 months; DN4 score was 6.7 ± 1.2. 8/12 (67%) of patients used LMP as monotherapy. Functional recovery after treatment was observed in 68% of patients; pain and painful area reduction, respectively, in 68% and in 92%. None of the patients reported adverse local or systemic reactions to the use of LMP. This study suggests LMP-Versatis® efficacy in short term treatment of localized peripheral NP secondary to post-burn plastic surgery, reducing both pain intensity and size of the painful area; moreover, our results confirm safety, tolerability and absence of side effects.

PROLONGED PHONE-CALL POSTURE AS RISK FACTOR FOR DEVELOPING ULNAR NERVE ENTRAPMENT AT ELBOW: A DYNAMIC NEUROPHYSIOLOGICAL STUDY Coraci D1,2 , Padua L2,3 , Erra C3 , Doneddu P4 , Granata G3 , Rossini PM3 . 1 Board of Physical Medicine and Rehabilitation, Department of Orthopedic Science, “Sapienza” University, Rome, Italy; 2 Department of Rehabilitation, Don Gnocchi ONLUS Foundation, Milan, Italy; 3 Institute of Neurology, Università Cattolica del Sacro Cuore, Rome, Italy; 4 Department of Clinical and Experimental Medicine, University of Sassari, Sassari, Italy. Ulnar neuropathy at the elbow (UNE) is the second most common entrapment syndrome and its occurrence appears to be increasing. Since it is often related to dynamic processes involving surrounding structures, postures and work-hobby activities may have a role in its origin and progression. The time spent on mobile phones has drastically increased in the last decades leading to prolonged phone posture (PPP) and consequently to an increased time spent with flexed elbow. We aimed to assess the effect of PPP both in patients with symptoms of UNE and in symptom-free subjects. Patients with pure sensory symptoms of UNE and negative neurophysiological tests (min-UNE) and symptoms-free subjects were enrolled. We evaluated ulnar motor nerve conduction velocity across the elbow at baseline and after 6, 9, 12, 15, 18 minutes of PPP in both groups. Thirty-eight symptom-free subjects and thirty-eight patients were enrolled. Globally 121 ulnar nerves from 76 subjects were studied. Moreover, a pilot ultrasonographic study of ulnar nerve was started in these patients. Conduction velocity of ulnar nerve across the elbow significantly changed over PPP time both in the control group and, to a greater extent, in patients with min-UNE. Conduction velocity during PPP showed different evolution between the two groups. The changes became significantly different after 6 minutes of PPP and progressively increased, with the greatest difference at 15 minutes. In conclusion, PPP causes a modification of ulnar nerve function as expressed by slowing of motor nerve conduction velocity, which is greater in patients with min-UNE although it also occurs in symptom-free subjects. Excessive PPP should be avoided in patients with UNE-symptoms.

PURINERGIC MODULATION AND TWO PHOTON IN VIVO MICROSCOPY OF SPINAL NEUROGLIAL MALADAPTIVE PLASTICITY FOLLOWING PERIPHERAL NERVE INJURY Cirillo G1 , Papa M2 , Todisco V1 , Tedeschi G1 . 1 Department of Medical, Surgical, Neurological, Metabolic and Aging Sciences, Second University of Naples, Italy; 2 Department of Public Clinic and Preventive Medicine, Institute of Human Anatomy, Second University of Naples, Italy. Modulation of spinal reactive gliosis following peripheral nerve injury (PNI) is a promising strategy to restore synaptic homeostasis. Oxidized ATP (OxATP), a nonselective antagonist of purinergic P2X receptors, was found to recover the neuropathic behavior (allodynia and hyperalgesia) following PNI. We investigated the role of intraperitoneal (i.p.) OxATP treatment in restoring the expression of neuronal and glial markers in the mouse spinal cord following sciatic spared nerve injury (SNI). Using in vivo two-photon microscopy, we imaged spinal astrocytes using the red fluorescent dye sulphorhodamine 101 (SR101) and astrocytic as neuronal Ca2+ levels with Oregon-Green BAPTA-1 (OGB), at rest and upon right hind paw electrical stimulation in sham, SNI, and OxATP-treated mice. Neuropathic behavior was investigated by von Frey and thermal plantar test. The expression of glial (GFAP and Iba1) and GABAergic markers (vGAT and GAD65/67) as also glial (GLT1 and GLAST) and neuronal glutamate transporters (EAAC1 and vGLUT1) have been evaluated. In SNI mice, we found (i) increased glial response, (ii) decreased glial amino acid transporters, and (iii) increased expression of neuronal amino acid transporters; (iv) in vivo analysis of spinal neurons and astrocytes showed a persistent increase of Ca2+ levels. OxATP administration reduced glial activation, modulated the expression of glial and neuronal glutamate/GABA transporters, restored neuronal and astrocytic Ca2+ levels, and prevented neuropathic behavior. In vitro studies, moreover, validated that OxATP (i) reduced levels of reactive oxygen species (ROS), (ii) reduced astrocytic proliferation, and (iii) increased vGLUT expression. All together, these data support the correlation between reactive gliosis and perturbation of the spinal synaptic homeostasis and the role played by the purinergic system in modulating spinal plasticity following PNI. Moreover, in vivo two-photon microscopy allowed real time morpho-functional imaging of the neuro-glial network in the living central nervous system.

AN OBSERVATIONAL STUDY ON TAFAMIDIS FOR TRANSTHYRETIN-RELATED FAMILIAL AMYLOID POLYNEUROPATHY IN ITALY Cortese A1 , Russo M2 , Obici L3 , Cavallaro T4 , Fabrizi GM4 , Manganelli F5 , Santoro L5 , Luigetti M6 , Sabatelli M6 , Schenone A7 , Grandis M7 , Mauro A8 , Pradotto LG8 , Mazzeo A2 , Gentile L2 , Piscosquito G9 , Calabrese D9 , Vita G2 , Merlini G3 , Pareyson D9 . 1 C. Mondino National Neurological Institute, Pavia; 2 Department of Neurosciences, University of Messina, and Clinical Centre NEMO SUD, Fondazione Aurora Onlus, Messina; 3 Amyloidosis Research and Treatment Center, Scientific Institute Policlinico San Matteo and Department of Molecular Medicine, University

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of Pavia, Pavia; 4 Department of Neurological, Neuropsychological, Morphological and Motor Sciences, University of Verona, Verona; 5 Department of Neurosciences, Reproductive and Odontostomatological Sciences, University Federico II of Naples, Naples; 6 Catholic University Department of Neurosciences, Rome; 7 Department of Neurology, Ophthalmology and Genetics, University of Genoa, Genoa; 8 Division of Neurology and Neurorehabilitation, Ospedale San Giuseppe, Istituto Auxologico Italiano, IRCCS, Piancavallo (Verbania); 9 IRCCS Foundation, C. Besta Neurological Institute, Milan, Italy.

EPIDERMAL NERVE FIBER LOSS IN AMYOTROPHIC LATERAL SCLEROSIS Dalla Bella E, Lombardi R, Porretta-Serapiglia C, Dacci P, Cazzato D, Sassone J, Lauria G. 3rd Neurology Unit, Carlo Besta Foundation IRCCS National Neurological Institute, Milan, Italy. Amyotrophic lateral sclerosis (ALS) is dominated by upper and lower motor neuron pathway degeneration and electrodiagnostic findings require normal sensory nerve conduction. Nevertheless, sensory abnormalities, including pain, have been reported in anecdotal ALS patients, suggesting that the disease may affect also the somatosensory system. Most recently, small nerve fiber loss has been described by skin biopsy and corneal confocal microscopy in two separate small groups of ALS patients. We performed a systematic study of the somatosensory pathway in a large cohort of familial and sporadic ALS patients to correlate genotype, phenotype and disease duration to somatosensory evoked potentials, sensory nerve conduction study and skin biopsy findings. Sixty patients with laboratory-supported probable, probable or definite ALS according to El Escorial revised criteria were consecutively recruited. Patients with any risk factor for peripheral neuropathy were excluded. Patients were divided according to the following phenotypes: spinal (25), bulbar (10), pyramidal (6), flail forms (7), FOSMN (4). Ten patients carried pathogenic mutations in known ALS causative genes (4 in SOD1, 1 in VCP, 5 in C9orf72). Eight patients complained of positive sensory symptoms, such as paresthesia, hyperesthesia, and dysesthesia at distal extremities or more diffuse. Neurophysiological studies revealed mild impairment of sensory nerve action potential amplitude of sural nerve in only 5 patients. Skin biopsy showed reduced intraepidermal nerve fiber density (IENFD) at the distal leg in all except 5 patients (2 flail-arm, 1 bulbar and 2 spinal onset). Small diameter nerve fiber loss seems to be part of the neurodegenerative process in ALS, suggesting that the spectrum of neuronal cell involvement is larger than formerly supposed.

Tafamidis meglumine is a transthyretin (TTR) stabilizer able to prevent mutated TTR tetramer dissociation into toxic misfolded amyloidogenic monomers in vitro and has been licensed in Europe and Japan for the treatment of TTR-familial amyloid polyneuropathy (TTR-FAP). In four previous randomized placebo-controlled or open-label trials it reduced TTR-FAP progression particularly in early-onset early-stage Met30 TTR-FAP patients, while the results on late-onset forms are less satisfactory. We performed a multicenter observational study following with a homogenous protocol a series of TTR-FAP patients who have been prescribed tafamidis by their treating physicians according to the existing national regulations. Main inclusion criteria: adults with symptomatic TTR-FAP, prescribed to receive tafamidis 20 mg/day, written informed consent. Patients’ standardized assessment at baseline and after 6–12–18–24 months included Neuropathy Impairment Score (NIS), NIS-Lower Limb (NIS-LL), Kumamoto Scale, Charcot-Marie-Tooth Neuropathy Score, FAP Stage, Polyneuropathy Disability (PND) score, Modified Body Mass Index (mBMI), NT-proBNP, echocardiography for inter-ventricular septum (IVS) thickness, ECG, Adverse Events (AEs). Responders were defined as patients with NIS-LL progression 6, in 2 severe NRS > 8). Two patients developed tactile hypoesthesia in the same areas, the oldest one (55 years) also a complete anhidrosis, tactile and pinprick hypoesthesia and ataxic gait. Skin biopsies showed severe epidermal denervation in anhidrotic areas and reduced intraepidermal nerve fiber (IENF) density in hyperhidrotic areas and at lower limbs. SSR were absent or reduced, warm thermoalgesic thresholds were increased, Ewing’s tests were normal. H-reflex was absent at rest in all. In 4 patients NCS showed reduced sensory action potential amplitudes with patchy pattern and SEPs were consistent with sensitive ganglionopathy. Our data suggest that Ross syndrome is a degenerative disorder characterized by progressive multifocal involvement of different fiber populations, starting from autonomic to sensory fibers, related to the disease duration. Neuropathic pain, tactile hypoesthesia and sensory ataxia represent the wide spectrum of a syndrome due to the progressive involvement of dorsal root neurons in a generalized still unknown injury.

evaluation, sensory profile assessment by quantitative sensory testing (QST) battery, nerve conduction studies (NCS) and skin biopsy at distal leg (DL) and proximal thigh (Pth). Twenty-five patients were enrolled (13 women, 12 men), 23 on ERT for more than 5 years, 2 untreated. Twelve of 25 patients complained of chronic pain. None had bone crisis in the last year. All of them complained of painful sensation suggestive of neuropathic pain with proximal patchy distribution, in 6 with severe pain paroxysms. Pinprick hypoesthesia was found in 9 and thermal hypoesthesia in 17. The sensory profile at QST battery showed a sort of stereotypical pattern with high cold thresholds with errata sensation (burning instead of cold), presence of paradoxical heat sensation (PHS) and mechanic hypoesthesia MDT, while only in 3 presence of pressure pain hyperalgesia (PPT). Skin biopsy showed epidermal denervation in 19 patients, in 12 with a non length-dependent pattern. In 10 of them we found only skin denervation at proximal site compared with 96 age- and sex-matched controls. The conventional nerve conduction studies showed only mild L5 radiculopathy in 2 and Carpal Tunnel Syndrome in 1. The presence and distribution of the sensory symptoms and signs and skin denervation in most of GD1 patients is consistent with small fibre neuronopathy, in some of them subclinical and probably not responsive to ERT.

PRIMARY BILIARY CIRRHOSIS: AN UNUSUAL CAUSE OF SENSORY AXONAL NEUROPATHY Di Pietro DA, Nobile-Orazio E. 2nd Neurology, Department of Medical Biotechnology and Translational Medicine, Humanitas Clinical and Research Center, Milan University, Rozzano, Milan, Italy. Acquired chronic sensory neuropathies are frequent in middle and late adulthood and represent a common cause for referral to neurologist. These forms of neuropathies can be challenging for physicians since, once known etiologies are excluded, still 10 to 35% of cases remain idiopathic (Wolfe et al., 1999). In a long-term follow-up study, Jann (2001) reported that repetition of common laboratory investigations do not add information to the basal evaluation, and response to symptomatic therapy is often not satisfactory (Pasnoor, 2013). This suggests that new serologic markers and associations are needed in view of possible etiological treatment. We describe 2 patients with chronic sensory neuropathy associated with primary biliary cirrhosis (PBC). Patient 1 was a 59-year-old woman presenting with tingling in both hands and electric shock-like paraesthesias at feet. Neurological examination showed sensory loss for all modalities with a length-dependent fashion. Nerve conduction studies (NCS) and sural biopsy were consistent with a chronic sensory axonopathy. Laboratory examinations revealed mild colestatic index elevation and anti-mitochondrial (AMA-M2) antibodies positivity. Symptoms improved after 3 months of oral prednisone. Patient 2 was a 51-year-old woman presenting with a 2-year history of tingling at right foot extended after months to the contralateral foot and to both hands. NCS were consistent with a purely sensory axonal neuropathy and laboratory findings revealed marginally increased levels of alkaline phosphatase. ANA (1:320) and AMA-M2 positivity led to

SMALL NERVE FIBRE IMPAIRMENT AND NEUROPATHIC PAIN IN TYPE 1 GAUCHER DISEASE Devigili G1 , Lettieri C1 , Rinaldo S1 , Sechi A2 , Deroma L2 , Ciana G2 , Dardis A2 , Bembi B2 , Eleopra R1 . 1 Neurological Unit, Neuroscience Department, 2 Regional Coordinator Centre for Rare Diseases, Academic Hospital of Udine, Italy. Type 1 Gaucher (GD1) disease is a lysosomal storage disorder usually differentiated from types II and III Gaucher disease by the absence of nervous system involvement. However, recently, an increasing number of reports described neurological manifestation including subclinic large fibre neuropathy. Pain is one of the more disabling symptoms in GD1, often persisting despite enzyme replacement therapy (ERT). It is currently considered as nociceptive pain secondary to bone involvement but it is described even in its absence without explanation. The aim of study was to investigate the presence and characteristics of pain in a cohort of GD1 patients and to assess peripheral nerve involvement. Patients with diagnosis of GD1 were enrolled during 1 year. Patients with comorbidity for peripheral neuropathy were excluded. Pain intensity and qualities were recorded. Then patients underwent pain questionnaires (Douleur Neuropathique en 4 questionnaire, DN4) and Neuropathic Pain Symptom Inventory (NPSI), clinical and neuroalgological

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common entrapment sites (right ulnar nerve at the elbow and fibular nerve at caput fibulae). The patient was treated with ivIg with clinical benefit. The increased duration of SAP is dependent on distal temporal dispersion and may be considered a sign of focal distal demyelination in sensory nerves. In conclusion, the duration of the distal SAPs may be considered an early and specific electrophysiological feature of distal demyelination in sensory nerves, and may be relevant in patients with acute/chronic dysimmune neuropathies with prevailing sensory deficits.

the diagnosis of PBC. Symptoms remained stable after one year of follow-up. Three cases of pure sensory neuropathy associated with PBC have been previously described (Charron, 1980; Illa, 1989; Toyooka, 2010). Clinical presentation is homogeneous, with no patient reporting symptoms of PBC before the onset of neuropathy and with a usually marginal liver impairment at diagnosis. Retrospective analysis of our 91 patients with sensory neuropathy evaluated from 2004 to 2014 showed that 31% of them had a cryptogenetic neuropathy and that 7.1% of them had PBC. This prevalence might be, however, underestimated since AMA antibodies were not always evaluated in the presence of mild and asymptomatic cholestasis. Testing for AMA-M2 may be therefore useful in patients with idiopathic sensory neuropathy. The pathogenic role of these antibodies in the neuropathy remains however unclear.

IMPLEMENTING A NEXT-GENERATION-SEQUENCING (NGS) PANEL FOR CHARCOT-MARIE-TOOTH DISEASE TYPE 2 (CMT2) Ferrarini M, Testi S, Taioli F, Cabrini I, Fabrizi GM. Department of Neurological and Movement Sciences, University of Verona, University Hospital G.B. Rossi, Verona, Italy.

INCREASE OF THE DISTAL SENSORY ACTION POTENTIAL DURATION AS A SPECIFIC FEATURE IN ACUTE INFLAMMATORY DEMYELINATING POLYNEUROPATHY: A CASE REPORT

CMT2 is an autosomal dominant (rarely recessive) motor-sensory neuropathy with high locus/allelic heterogeneity and incomplete clinical and neurophysiological penetrance. More than forty genes have been discovered. Diagnostic molecular yield does not exceed 25% of patients/families even in more selected series (Rossor AM et al., Nature Rev. 2013). By AmplySeqTM (Life Technologies) we designed a first-level custom panel including fourteen more frequent CMT2 genes: MFN2, MPZ , GJB1, GARS, BSCL2, TRPV4, NEFL, HSPB1, HSPB8, GDAP1, RAB7A, DNM2, HINT1, REEP1. The NGS procedure was based on two-tube multiplex amplification of DNA pools and semiconductor sequencing with the Ion Torrent Personal Genome Machine (PGM). Our custom panel allowed a theoretical coverage of 97.72%. Sequencing raw data were analyzed by Ion Reporter Suite program. Sensitivity and specificity of NGS was analyzed in a cohort of 25 CMT2 probands with mutations in MFN2 (which is the more common CMT2 gene though associated with 10–15% of cases only) detected previously by DHPLC analysis and Sanger sequencing; in the reference cohort 5 MFN2 mutations were clearly pathogenic, 4 variations were clearly polymorphic, 7 mutations had an unclear role (rare variants or mutations in singleton case without co-segregation study available). By analyzing 11 patients for a 316-chip, the uniformity coverage was greater than 91% with an average base depth of 1100 reads; the target base-coverage was 81.65% at 500X. NGS detected all variations previously identified by conventional techniques. Furthermore, in two probands with variations with unclear role (MFN2 p.D77N and p.V705I), NGS identified two alternative mutations: a pathogenic BSCL2 p.N88S (a well-characterized pathogenic mutation, associated with dHMN and pyramidal features in three members of the examined pedigree) and TRPV4 p.Y491X (a 70-year-old man with mild-to-moderate intermediate CMT). The preliminary study disclosed a 100% sensitivity of this custom panel PGM-NGS. NGS represents a promising approach in molecular screening of CMT2. As a corollary, the report emphasizes that reports of MFN2 mutations should be critically evaluated.

Donato F, Basaldella F, Zuco C, Moretto G, Ottaviani S, Bovi T, Romito S Squintani G. Neurology Unit, Neuroscience Department, Azienda Ospedaliera Universitaria Integrata, Verona, Italy. The diagnosis of acute inflammatory demyelinating polyneuropathy (AIDP) is based on a combination of specific clinical, electrophysiological and cerebrospinal fluid (CSF) data. Demyelination is the main electrophysiological feature of this subtype of Guillain-Barré syndrome (GBS). Increased duration of distal compound muscle action potentials (cMAPs) has recently been proposed to be an index of demyelination in distal nerve segments of chronic inflammatory demyelinating polyneuropathy (CIDP). However, the contribution of sensory neurography has not been well characterized and an increased duration of distal sensory action potentials (SAPs) is not taken into account in the diagnostic criteria of AIDP or CIDP. We describe a case of AIDP with predominant sensory clinical and electrophysiological impairment. A 26-year-old woman came to our attention for oral and upper limbs paresthesias occurring ten days after flu-like symptoms. The clinical picture progressively worsened in one week and tingling appeared in lower limbs. A first electrophysiological test revealed an increased distal motor latency of cMAPs obtained stimulating the left median, ulnar nerves, and fibular nerves bilaterally, associated with increased duration of cMAPs from right fibular nerve, and interestingly an increased duration (total and negative) of SAPs of median and ulnar nerve bilaterally. For this purpose, we collected 20 recordings of SAPs from median, ulnar, radial and sural nerves as age-matched controls. At admission, two weeks after symptom onset, neurological examination revealed a diffuse hypopallesthesia and lower limbs areflexia. No motor deficits were detected. CSF examination revealed a mild hyperproteinorrachia (0.74 g/l, normal value C p.M115T, NDRG1: c.442C > T p.R148X, AARS: c.986G > A p.R329H and MPZ :c.233C > T p.S78L. These variants were verified by Sanger sequencing and by familial segregation in other affected family members. Validation by Sanger sequencing and by family segregation of other “weaker mutation” is still ongoing, leading to speculate a possible increase of the final diagnostic rate. This Inherited Peripheral Neuropathies Panel, although ongoing, allowed us to define a molecular diagnosis for patients still lacking a genetic definition, identifying mutations in genes not routinely screened in our laboratory practice. Our results confirm that gene panels will became the best approach to test for mutations in genetic heterogeneous diseases, leading to a faster analysis of a broad spectrum of genes.

PERIPHERAL NEUROPATHIES SECONDARY TO TREATMENT WITH IMMUNOMODULATING MABS TARGETING IMMUNE CHECKPOINTS: AN EMERGING CHALLENGE? Giannini F1 , Calabrò L2 , Danielli R2 , Alberici A3 , Insana L1 , Gasparotti R4 , Cerase A1 , Capoccitti G1 , Di Giacomo AM2 , Maio M2 .1 Dipartimento Scienze Mediche, Chirurgiche e Neuroscienze, Università di Siena, Siena; 2 U.O.C. Immunoterapia Oncologica, Azienda Ospedaliera Universitaria Senese, Siena; 3 Clinica Neurologica, Università di Brescia, Brescia; 4 U.O.C. Neuroradiologia, Università di Brescia, Brescia, Italy.

SACRAL NERVE NEUROMODULATION FOR FECAL INCONTINENCE IN A PATIENT WITH ANTIPHOSPHOLIPID SYNDROME-RELATED AUTONOMIC NEUROPATHY Giorli E1,2,5 , Franciotta D3 , Serventi A4 , Binda GA4 , Canepa G4 , Siciliano G2 , Giannini F5 , Mannironi A1 , Schenone A6 , Benedetti L1 . 1 Department of Neurology, S. Andrea Hospital, La Spezia; 2 Department of Clinical and Experimental Medicine, Neurological Clinic, University of Pisa, Pisa; 3 Laboratory of Neuroimmunology, IRCCS, “C. Mondino” National Neurological Institute, University of Pavia, Pavia; 4 Department of Abdominal Surgery - Galliera Hospital – Genova; 5 Department of Medical, Surgical

Immune-checkpoints represent new targets for cancer immunotherapy. The use of mAbs blocking CTLA4, PD-1 receptors or its ligand PD-L1, induces activation of T cells increasing reaction to cancer neoantigens. However several immune-related adverse events (irAEs) due to the unopposed T-cell activation have been reported, the most

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and Neurological Sciences, University of Siena, “Le Scotte” Hospital, Siena; 6 Department of Neuroscience, Rehabilitation, Ophthalmology, Genetics, Maternal and Child Health (DiNOGMI), University of Genova, Genova, Italy.

Torino, Department of Mechanical and Aerospace Engineering, Politecnico of Torino, Torino; 4 Nanostructured Interfaces and Surfaces, Department of Chemistry, University of Torino, Torino; 5 CNR-IPCF UOS, Pisa, Italy.

Originally used for urinary incontinence, sacral nerve neurostimulation (SNS) has been increasingly viewed as an effective treatment for fecal incontinence over the last years, especially when other conventional therapies result ineffective. In patients with fecal incontinence following sacral spinal cord injury, inflammatory demyelinating syndromes of the CNS, and entrapment or traumatic pudendal neuropathies, sacral neurostimulators can significantly improve continence and quality of life. To our knowledge, SNS has never been exploited in autoimmune autonomic neuropathies. We describe a patient with Hughes syndrome who developed an autonomic neuropathy-related fecal incontinence that was successfully treated with SNS. A 54-year-old woman with antiphospholipid syndrome presented with constipation and fecal incontinence. Electroneurography showed decreased amplitudes of the motor responses on pudendal nerves bilaterally, and increased latencies to the left side. Sensory evoked potentials of pudendal nerves, stimulated on the front (vaginal) and on the back side (anal), showed increases in response latency of P37, with poor definition of somatosensory potentials. Pudendal motor evoked potentials, after brain stimulation and anal recording, showed decreased amplitudes of the motor responses, but central motor conduction time normal values. Electromyography of the striated muscle of the anal sphincter, using bilateral recording at rest, at maximum effort, and at the anal sphincter response to the Valsalva maneuver showed important denervation and muscular sphincteric depletion (likely due to axonal injury), and very poor reflex responses. Anorectal manometry and defecography showed ineffectiveness in the evacuation of the contrast medium due to ineffective pushing of the abdominal press and paradoxical contraction of pubo-rectalis muscles. A sacral neurostimulator was implanted, which led to an immediate benefit on the symptoms. Indeed, the Cleveland Clinic Continence (CCC) score and Fecal Incontinence Quality of Life (FIQOL) Scale, administered before and after implantation, showed an improvement of 50% in bowel function with a CCC score of 13 before insertion and a CCC score of 7 after a follow-up of 36 months, and a significant improvement in all the four FIQOL scales. In conclusion, our case is the first one reporting the effectiveness of SNS for fecal incontinence following autoimmune neuropathy with autonomic manifestations, thus suggesting that SNS is an advantageous therapeutic option in these disorders.

Fibrous substrates functioning as temporary extracellular matrices (ECM) can be prepared easily by electrospinning, yielding fibrous matrices suitable as internal fillers for nerve guidance channels. Several parameters can be adjusted in order to produce random or aligned fibers. The aim of this study was to understand the influence of electrospun nano-fibre alignment on Schwann cell (SC) adhesion, morphology and proliferation and sensory neuron-like cells differentiation. Since SC and axonal regrowth processes play an important role in peripheral nerve regeneration, we used RT4-D6P2T cell line, primary SC cultures and neuron-like 50B11 cell line to perform in vitro tests. Particularly, we performed adhesion, proliferation and vitality assays on SC and neurite growth evaluation on 50B11 differentiated cells. When seeded on aligned fibers RT4-D6P2T and SC showed a lower adhesion than under control conditions or random fibers. Both RT4-D6P2T and primary SC displayed high actin cytoskeleton organization and many focal adhesion points under all conditions tested. Actin cytoskeleton staining showed that both RT4-D6P2T and primary SC cultured under aligned fibers displayed elongated actin fibers in comparison to control condition and random fibers. Both MTT and proliferation assay showed that RT4-D6P2T and primary SC cultures had lower proliferation rates when seeded on aligned fibres than under control conditions or random fibres. The alignment of gelatin electrospun fibres did not affect the number of adherent 50B11 neuron-like cells. Immunostaining against 𝛽-tubulin shows that 50B11 cell maintaining their capability to differentiate on gelatin electrospun fibres. Confocal images shows that aligned fibre resulted in neurites alignment which is parallel to the fibres direction. Neurites growth on random fibres is similar to control conditions. These data suggest that the topography of electrospun gelatin fibers can be adjusted to modulate SC and axon organization and that aligned nano-fibres are promising fillers for the design of devices for peripheral nerve repair.

NETWORK TOPOLOGY OF NAV1.7 MUTATIONS IN SODIUM CHANNEL-RELATED PAINFUL DISORDERS Kapetis D1 , Yang Y2,3,4 , Galbardi B1 , Szklarczyk R5 , Faber CG6 , Gerrits M6 , Merkies ISJ6,7 , Dib–Hajj SD2,3,4 , Tononi P8 , Sassone J8 , Mantegazza M9 , Waxman SG2,3,4 , Lauria G8 . 1 Bioinformatics Unit and 8 Neuroalgology Unit, IRCCS Foundation “Carlo Besta” Neurological Institute, Milan, Italy; 2 Department of Neurology and 3 Center for Neuroscience and Regeneration Research, Yale University School of Medicine, New Haven, CT 06515, and 4 Neurorehabilitation Research Center, Veterans Affairs Medical Center, West Haven, CT 06515, USA; 5 Department of Clinical Genetics, Maastricht University Medical Center, Maastricht, the Netherlands; 6 Department of Neurology, Maastricht University Medical Center, Maastricht, the Netherlands; 7 Department of Neurology, Spaarne Hospital, Hoofddorp, the Netherlands;

THE INFLUENCE OF ELECTROSPUN FIBRE ALIGNMENT ON SCHWANN CELL BEHAVIOUR AND AXONAL OUTGROWTH Gnavi S1,2 , Fornasari BE1,2 , Tonda-Turo C3 , Laurano R3 , Zanetti M4 , Ciardelli G3,5 , Geuna S1,2 . 1 Department of Clinical and Biological Sciences, University of Torino, Orbassano; 2 Neuroscience Institute of the Cavalieri-Ottolenghi Foundation, University of Torino, Orbassano; 3 Politecnico di

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9 Institute of Molecular and Cellular Pharmacology, CNRS & University of Nice-Sophia Antipolis, France.

to subjects’ condition (diagnosis, side, and time of biopsy). Findings were referred to gender- and age-adjusted normative values. Forty patients and 17 healthy subjects performed bilateral skin biopsies; 15 SFN patients and 8 healthy subjects underwent follow-up skin biopsies. Sural and dorsal nerve action potential conduction study was normal in all subjects. Interside IENF density did not differ both in SFN patients (median 2.45 ± 1.45 SD right; 2.2 ± 1.32 left) and healthy subjects (median 6.3 ± 2.81 right; 6.2 ± 2.3 SD left). The right-to-left correlation coefficients were excellent (Pearson 0.95 in SFN and 0.97 in healthy subjects). The analysis of IENF density at 20-day follow-up biopsy showed no difference between sides in both the groups, and yielded excellent correlation coefficients. The diagnosis of SFN can be reliably ascertained by unilateral skin biopsy at the distal site of the leg, and IENF density is not expected to vary within 3 weeks.

Voltage-gated sodium channels (NaV) regulate ion flux and generate electrical signals in excitable cells by opening and closing the pore gates. Gain-of-function mutations in NaV1.7 that change the biophysical properties of the channel and increase the excitability of dorsal root ganglion (DRG) nociceptors have been found in patients with inherited erythromelalgia (IEM), paroxysmal extreme pain disorder (PEPD) and painful small fibre neuropathy (SFN). The variations in the interatomic interactions caused by these mutations and the rearrangement of connectivity within the channel are largely unexplored. To address these issues, we used the graph theory approach to analyse several topological parameters, such as betweenness and closeness centrality, clustering coefficients, and eccentricity in pathogenic NaV1.7 mutations identified in patients with IEM PEPD, and SFN. Findings were compared with mutations not causing biophysical abnormalities (nABN) in the NaV1.7 channel and single nucleotide polymorphisms (SNPs) between human and homologous NaV1.7 genes. Here we demonstrate that in pathogenic NaV1.7 mutations are characterized with significant variation of betweenness centrality (ΔBct ). Conversely, nABN mutations and SNPs did not show any significant association with ΔBct value. In addition, we demonstrated that in pathogenic NaV1.7 mutations, polar and charged residues with high ΔBct values may play a key role in altering the normal network connectivity of the channel facilitating long-range interdomain interactions. Our novel observations, based on the analysis of the topological features of the protein, deepen our understanding of how changes induced by pathogenic single amino acid substitutions found in patients with painful conditions can modulate the interatomic network connectivity of the NaV1.7 channel. These findings also provide a novel approach to predict the biophysical effects of NaV1.7 mutations, thus increasing the cost-effectiveness of cell electrophysiological assays and contributing to the assessment of the pathogenicity of gene variants.

AN OLD PROCEDURE DRESSED WITH A NEW PURPOSE: SKIN BIOPSY TO DETECT AMYLOIDOSIS Lopez ID1 , Cerri F1 , Scarlato M1 , Riva N1 , Marcatti M2 , Fazio R1 , Quattrini A1 . 1 Department of Neurology, Institute of Experimental Neurology (INSPE) and Division of Neuroscience, San Raffaele Scientific Institute, Milan; 2 Hematology and Bone Marrow Transplantation Unit, San Raffaele Scientific Institute, Milan, Italy. Amyloidosis is a systemic disease, that may be acquired or hereditary, in which deposits of abnormally folded proteins share distinctive staining properties and fibrillar ultrastructural appearance. Amyloid can be found in a variety of tissues, ultimately leading to their progressive dysfunction. With recent advances in the treatment of systemic amyloidosis, an early diagnosis of the disease is mandatory. Thus far, no biochemical markers are available and the diagnosis is strictly based on the histological evidence of amyloid deposits in tissues. Biopsy of a clinically affected organ (liver, heart, kidneys, muscle or nerve) is the most sensitive method, however, such a biopsy is invasive and carries the risk of many complications. For this reason, since the early 1970s subcutaneous fat pad biopsy, obtained via fine-needle aspiration, being safe, cheap, and rapid, has been introduced as a screening test for the detection of amyloidosis, while rectal mucosa and labial salivary glands are less frequently the target of biopsy. At this time, Congo red staining continues to be the gold standard to detect amyloid fibrils both in bright field and under polarized light. Skin biopsy, a safe, easy and out-patient procedure with minimal complications, is commonly performed in cutaneous amyloidoses and systemic amyloidoses with cutaneous involvement in order to determine the abnormal protein deposits in the dermis. On the basis that amyloidosis is a systemic disease and that amyloid fibrils could eventually gather in every organ, we postulated skin biopsy as a plausible tool to detect amyloid. To our knowledge, no data are reported in the literature on the potential usefulness of this method either in the acquired or hereditary form of amyloidosis. We analyzed 12 patients (2 affected by familiar amyloidosis, 4 presenting with an acquired form and 6 patients, used as control, 4 affected by small fibers neuropathy and 2 healthy control). Each patient

SIDE AND TIME VARIABILITY OF INTRAEPIDERMAL NERVE FIBER DENSITY Lauria G, Dacci P, Lombardi R, Cazzato D, PorrettaSerapiglia C, Taiana M, Sassone J, Dalla Bella E, Rinaldo S, Lettieri C, Eleopra R, Devigili G. 3rd Neurology Unit and Skin Biopsy, Peripheral Neuropathy and Neuropathic Pain Clinic, IRCCS Foundation “Carlo Besta” Neurological Institute, Milan, Italy; Neurological Unit, University-Hospital “S. Maria della Misericordia”, Udine, Italy. The objective of this study is to assess the right-to-left and short-term variability of intraepidermal nerve fiber (IENF) density at the distal site of the leg. Patients with possible or probable small fiber neuropathy (SFN) and healthy volunteers underwent skin biopsies at the right and left distal leg. A subgroup of subjects underwent follow-up biopsies 20 days later. Biopsies were immunostained by polyclonal anti-protein gene product 9.5 antibodies and intraepidermal nerve fiber (IENF) density was quantified in non-consecutive sections following published guidelines by operators blinded

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underwent a punch skin biopsy in a proximal and distal site in the leg; subsequently a Congo red staining and the intraepidermal nerve fibers count was performed. Our preliminary analysis showed that the skin biopsy is easy, fast and well tolerated by the patients as well as the fat pad, and the two, stained with Congo red, detected patients with amyloid fibrils with a complete overlap. However, more patients will be required to determine accurate specificity and sensitivity of skin biopsy in amyloidosis and statistically compare it to the most common biopsy sites.

ADMISSION NEUROPHYSIOLOGICAL ABNORMALITIES IN GUILLAN-BARRÉ SYNDROME: A SINGLE-CENTER EXPERIENCE Luigetti M, Servidei S, Modoni A, Rossini PM, Lo Monaco M, Sabatelli M. Department of Geriatrics, Neurosciences & Orthopedics, Institute of Neurology, Catholic University of the Sacred Heart, Rome, Italy. Although patients with Guillain-Barré syndrome (GBS) are often hospitalized a few days after symptoms onset, nerve conduction study (NCS) abnormalities in early phases of the disease are not well characterized. Our aim was to report early neurophysiological abnormalities from a cohort of GBS patients. In this single-center study, we retrospectively reviewed the NCS data of 71 consecutive GBS patients in whom neurophysiology was performed within two weeks after disease onset. We further divided our cohort into three sub-groups according to the interval between disease onset and NCS (≤4 days; 5–7 days; 8–14 days). A great proportion of patients (37%) with an early NCS ( ≤ 4 days) showed normal neurophysiological results. The most altered parameters were F waves with a proportional increase, according with days after onset. Conduction blocks were observed preferentially in upper limbs, in about one-third of cases. This study confirms that NCS may be normal in the early phases of GBS and suggests to perform an extensive neurophysiological evaluation in these patients. Our results may help clinicians in the interpretation of NCS in early-onset GBS.

SEARCH FOR A ROLE OF COMORBIDITIES IN VARIABILITY TO IVIG THERAPEUTIC RESPONSE IN CIDP Lucchesi C1 , Schirinzi E1 , Piazza S1 , Buda G2 , Tavoni A3 , Siciliano G1 . 1 Department of Clinical and Experimental Medicine, Clinic of Neurology, University of Pisa, Pisa; 2 Department of Clinical and Experimental Medicine, Hematology Unit, University of Pisa, Pisa; 3 Department of Clinical and Experimental Medicine, Clinical Immunology Unit, University of Pisa, Italy. Among the clinical and laboratory factors that more influence the classic presentation of CIDP, the occurrence of possible comorbidities appears to play a relevant role also toward its therapeutic response to available treatment. The aim of this study was to analyse clinical variability of a cohort of CIDP patients affected by a concurrent disease of different origin but targeting the peripheral nervous system. We report our experience in a series of 9 patients with a CIDP diagnosis (EFNS/PNS criteria) treated with IVIG and different comorbid/coexisting conditions. In particular, in our case series CIDP was associated with the following disorders mainly of autoimmune origin: myeloid dysplasia, thyroid-related ocular myopathy, multiple sclerosis and ocular myopathy, macular oedema, popliteal deep venous thrombosis, monoclonal IgG gammopathy; a patient showed a complex picture of systemic autoimmune disorders; in another patient CIDP was associated with Charcot-Marie-Tooth type 1A and in the last one history of successfully surgically treated kidney cancer was present. All patients were treated with IVIG at standard doses (0.4 mg/kg/daily for five consecutive days) with variable frequency of administration. The patient with CIDP and ocular myositis showed a clear improvement in CIDP symptoms as well as in diplopia, related to ocular muscle involvement. A clinical response was not obtained in the patient with CIDP and multiple sclerosis, actually performing plasmapheresis, azathioprine and glatiramer acetate, with partial benefit on both conditions. In the patient with CIDP and a complex autoimmune picture, IVIG treatment was discontinued for intolerability. The remaining patients showed a clear benefit on CIDP related symptoms, although at variable rate of IVIG delivery, and a good profile of tolerability. Taken together, data from the present case series suggest a highly heterogeneous profile of clinical response to IVIG treatment in CIDP associated with concomitant disorders. To better understand the mechanisms which can be associated with the inflammatory demyelinating process of CIDP can be useful also in terms of identifying parameters predictive of therapeutic response in individual patients affected by this disease.

LIVEDO AND MULTIPLE MONONEUROPATHY Lupidi F1 , Gelardi C2 , Danieli MG2 , Logullo F1 . 1 Section of Neurology, Ospedali Riuniti, Torrette di Ancona; 2 Section of Clinical Medicine, Ospedali Riuniti, Torrette di Ancona, Italy. Livedo is a rare, usually chronic, dermatological vasculopathy characterized by skin purplish or bluish-red lesions, primary affecting the lower extremities, due to cutaneous venous plexus dilatation or de-oxygenation. We can distinguish livedo reticularis, characterized by regular unbroken circles forming a netlike pattern, and livedo racemosa, characterized by irregular broken circles. It is usually a benign and self-limited isolated condition, although it may reflect a more complex underlying disease, mainly a vasculitis or a thrombo-occlusive vasculopathy due to hypercoagulable states. Livedo may be rarely associated with a polyneuropathy, a multiple mononeuropathy or a ganglionopathy. We describe four cases of livedo associated with an axonal sensory-motor multiple mononeuropathy which underline different clinical conditions (Sneddon’s syndrome linked to antiphospholipid syndrome, thrombotic microangiopathy related to thrombophilia, cutaneous polyarteritis nodosa, non systemic vasculitic neuropathy). All the patients were women younger than 50 years presenting with a cutaneous livedo pattern. A skin biopsy was performed in all patients; a sural nerve biopsy was also performed in three patients. In all patients lesions started to improve after 1 to 3 months of therapy (ASA, prednisone) and completely

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resolved in about 6 months. In the fourth subject, despite the complete healing of cutaneous ulcers, there was a progressive neurological deterioration, documented by clinical and serial EMG-ENG examinations. The patient was then treated with oral cyclophosphamide for 6 months with sustaining functional and clinical remission, followed by a maintenance therapy with oral methyl-prednisolone and ASA. The other patients are now in remission with low dose oral PDN and ASA.

higher sensitivity and uniformity of the probe-based vs. the amplicon-based strategy (98% vs. 78% of target region with ≥20X coverage). Moreover, the uniformity of the probe-based approach allowed quantitative CNV analysis for the detection of large deletions. In conclusion, this approach allowed us to increase by ≥10-fold the number of genes analyzed, significantly expanding the diagnostic yield. Our data confirm the usefulness and time/cost effectiveness of NGS-based gene panels for the genetic diagnosis of hereditary neuropathies. Supported by Italian Ministry of Health grant to FT.

A NGS TARGETED-RESEQUENCING APPROACH FOR THE CHALLENGING GENETIC DIAGNOSIS OF INHERITED PERIPHERAL NEUROPATHIES

IS THE RESPONSIVENESS TO IVIG OR CORTICOSTEROIDS PREDICTIVE FOR TREATMENT DEPENDENCE IN CIDP?

Magri S1 , Di Bella D1 , Saveri P2 , Baratta S1 , Milani M1 , Piscosquito G2 , Moroni I3 , Pareyson D2 , Taroni F1 . 1 Unit of Genetics of Neurodegenerative and Metabolic Disease, 2 Clinic of Central and Peripheral Degenerative Neuropathies Unit, 3 Child Neurology Unit, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milan, Italy.

Manganelli F, Topa A, Iodice R, Pisciotta C, Santoro L. Department of Neurosciences, Reproductive Sciences and Odontostomatology, University Federico II of Naples, Naples, Italy. The objective of this study was to assess whether the responsiveness to treatment with intravenous immunoglobulin (IVIG) or corticosteroids (CS) may be a predictive factor for treatment dependence in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). We retrospectively evaluated therapeutic features in 35 CIDP patient responders to IVIG or CS. IVIG were used as first-line treatment in 20 patients and CS in 15 patients. The number of patients who failed to respond to the first-line therapy was higher in CS group (8/15; 53.3%) compared to IVIG group (1/20; 5%). Overall, 27 patients (77.2%) were responsive to IVIG and 8 patients (22.8%) to CS. Twenty-six (74.3%) out of 35 patients needed maintenance therapy (treatment-dependent group). Nine patients were OFF-treatment at least since 1 year. In the treatment-dependent group the proportion of patients with IVIG, as the effective treatment (23/26; 88.5%), was higher than that of patients with CS as the effective treatment (3/26; 11.5%). Interestingly, all the 8 patients who failed to respond to CS as first-line treatment became treatment-dependent; conversely, the only patient who was unresponsive to IVIG as first-line treatment, was in complete remission. CIDP patients seem to be more frequently responsive to IVIG. The responsiveness to IVIG together with the resistance to CS treatment seems to be a predictive factor for treatment dependence in CIDP.

More than 70 genes are known to cause hereditary peripheral neuropathies, which include: demyelinating (CMT1-4) and axonal (CMT2) Charcot-Marie-Tooth disease, distal hereditary motor neuropathies (HMN), and hereditary sensory neuropathies (HSN). These disorders represent a phenotypic continuum and show significant genetic overlap. Therefore, efficient genetic diagnosis would require a comprehensive and systematic approach such as that provided by NGS technology. We developed and validated customized gene panels with two different enrichment strategies in order to find the best compromise between sensitivity, specificity, uniformity, and cost: 1) an amplicon-based enrichment strategy using TruSeqCustomAmplicon technique [panel 1, CMT1-4 (37 genes) + HSN (17 genes); panel 2, CMT2 (29 genes) + HMN (13 genes)]; 2) a probe-based enrichment strategy using Nextera Rapid Capture technique (one single panel containing 95 genes for CMT1-4, CMT2, HMN, and HSN). We analyzed 141 patients previously screened for the CMT1A deletion/duplication and for ∼3–5 genes selected according to the phenotype. We identified 300–600 variants per patient, which were reduced to ≤10 by filtering. All variants were subsequently imported into a local database created specifically for the purpose of: 1) filtering identified variants according to quality criteria, functional consequence, and frequency in variant databases; 2) annotating the frequency of polymorphic variants in the Italian population; 3) collecting data to allow an easy and quick query also by other laboratories. The molecular cause of the disease was identified in 43/141 patients (30.5%) in the following genes: SH3TC2 (n = 10), MPZ (n = 7), GJB1 (n = 5), GDAP1 (n = 4), MFN2 (n = 2), FIG4 (n = 2), HINT1 (n = 2), DHTKD1 (n = 1), EGR2 (n = 1), GNB4 (n = 1), HARS (n = 1), HSPB1 (n = 1), HSPB8 (n = 1), IGHMBP2 (n = 1), MTMR2 (n = 1), NDRG1 (n = 1, non-Gypsy), SPTLC1 (n = 1), TRIM2 (n = 1). In addition, uncertain variants and/or one heterozygous mutation in recessive genes were identified in most of the remaining patients. In these cases, further analyses are required to validate variants and to assess the possible presence of large in/dels. Coverage analysis revealed a

BRENTUXIMAB VEDOTIN: A MICROTUBULE APOLLYON Mariotto S, Ferrari S, Cavallaro T, Monaco S. Department of Neurological and Movement Sciences, University of Verona University Hospital G.B. Rossi Verona, Italy. Some patients with advanced-stage Hodgkin lymphoma (HL) and anaplastic large cell lymphoma (ALCL) are refractory to or relapse after frontline, second-line chemotherapy and autologous stem cell transplant. Brentuximab vedotin (BV) is an anti-CD30-drug conjugate that has shown efficacy in these patients. The most relevant adverse effects of BV include peripheral neuropathy (PN). Despite being a major limiting factor of prolonged therapy, the mechanism involved in the neurotoxicity has not been characterized.

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We describe clinical and neurophysiological data of two patients affected by HL and treated with BV and pathological examination of sural nerve biopsy of one case. We also report their follow-up. Clinical and neurophysiological data show that BV related neuropathy can be a reversible process. A significant cytoskeleton derangement with microtubule loss has been observed at nerve biopsy suggesting that the main causative agent of PN should be the antimicrotubule drug monomethyl-auristatin that blocks the polymerization of tubulin. The analysis and characterization of neurological damage in these patients is of fundamental importance. If the toxicity of BV become acceptable this treatment will become an intriguing option and may be extend to other hematological malignancies.

distribution and concentration of the different gangliosides. Further studies are required to better understand various BBE phenotypes and to give them the right positioning in the MFS/BBE/GBS spectrum. CHARCOT-MARIE-TOOTH SUBTYPES IN A COHORT OF ITALIAN PATIENTS ENROLLED AT MESSINA NEUROMUSCULAR CENTRE SINCE 1994 TO 2014 Mazzeo A1 , Stancanelli C1,2 , Alfonzo A1 , Gentile L1 , Russo M1 , Fabrizi GM3 , Taioli F3 , Ferrarini M3, Toscano A1 , Vita G1 . 1 Department of Neurosciences, University of Messina; 2 Biomedical Department of Internal and Specialistic Medicine, University of Palermo; 3 Department of Neurological and Movement Sciences, University of Verona, Italy. The Centre for Neuromuscular Diseases of the University of Messina is a tertiary center for neuromuscular disorders in South Italy and a Regional Reference Centre for Rare Diseases. We analyzed the percentage of Charcot-Marie-Tooth (CMT) patients assessed at our Center in the last twenty years, with the aim to evaluate how many patients received a genetic diagnosis and the distribution of the genetic subtypes. This was a retrospective study. We analyzed the clinical records from 823 patients with anamnestic and/or clinical suspect of CMT seen since 1994 to 2014. The distribution of CMT subtypes and pathogenic genetic mutations was determined. 310/823 (37.6%) received a genetic diagnosis. The most common subtypes found were CMT1A/PMP22 duplication (51.6%), HNPP/PMP22 deletion (16.1%), CMT1B/MPZ mutation (10.6%), CMTX/GJB1 mutation (8.3%), CMT2F/HSP27 mutation (5.1%). Other mutations included PMP22 point mutation (n. 7 pts), MFN2 mutation (n. 5 pts), GDAP1 mutation (n. 4 pts), BSCL2 mutation (n. 3 pts), GARS mutation (n. 2 pts), TRPV4 mutation (n. 2 pts), LITAF mutation (n. 1 pt) and NEFL mutation (n. 1 pt). Our findings confirm the heterogeneity of CMT. The percentage of genetic confirmation is slightly smaller than the ones previously reported. One explanation might be that the time period we considered is twenty years, with a limited number of known genes in the first decade. The subtypes distribution showed a major prevalence of CMT2F/HSP27 mutation compared to previous reports, with atypical phenotypic features in some individuals.

BICKERSTAFF ENCEPHALITIS: REPORT OF TWO ATYPICAL CASES WITH NEGATIVE ANTI GQ1B ANTIBODIES Mataluni G1 , Studer V1 , Terracciano C2 , Rocchi C1 , Marfia GA1 . 1 Unit of Disimmune Neuropathies - Institute of Neurology, Department of Systems Medicine, University of Tor Vergata, Rome; 2 Department of Clinical Biochemistry and Molecular Biology, University of Tor Vergata, Rome, Italy. Bickerstaff brainstem encephalitis (BBE) is conventionally defined as a syndrome presenting progressive, external ophthalmoplegia, ataxia and disturbance of consciousness or hyperreflexia as its major manifestations, and has subsequently been associated with anti-GQ1b antibodies. Although the typical clinical features of BBE are well recognized, some cases with serological and clinical atypical presentations have been described. Here we report two cases of BBE characterized by negative anti-GQ1b antibodies: in the first patient we observed high serum titer of IgG and IgM GM1 whereas no anti ganglioside antibodies were demonstrated in the second. Fever and diarrhea episode occurred 10 days earlier in these patients: IgG and IgA for Campylobacter jejuni were detected only in the patient who carried anti GM1 antibodies. Both of them displayed disturbance of consciousness, diplopia, slurring of speech, dysphagia and limb ataxia. A minor motor deficit of the left limbs was observed in the first patient, a more severe sensory-motor deficit in the left limbs instead occurred in the second one. In the first patient multiple supra and infra-tentorial T2 hyperintensity were detected in brain MRI and CSF examination demonstrated high protein level and mononuclear pleocytosis. CSF and brain MRI showed no abnormalities in the second patient. Investigation for bacterial infection and neurotropic viruses were negative in both. Nerve conduction studies performed longitudinally within 1 month were unremarkable in these patients. They both significantly recovered within five days of plasma exchange. The clinical, serological and instrumental features of these 2 cases are different from the classical symptoms described in the Miller-Fisher syndrome (MFS)/BBE/Guillain-Barré syndrome (GBS) spectrum. The cause for this variability could be due in the existence of still unknown neural antigens target of molecular mimicry; likewise host factors such as different antibody accessibility to the blood-brain and blood-nerve barrier as well as the

RITUXIMAB IN PATIENTS WITH REFRACTORY CHRONIC INFLAMMATORY DEMYELINATING POLYRADICULONEUROPATHY NOT ASSOCIATED WITH HAEMATOLOGICAL DISEASES Medici D1,3 , Ferrante T1,3 , Manneschi L1,3 , Pedà G1,3 , Montanari E1,3 , Pavesi G2,3 , Gemignani F2,3 . 1 UO Neurologia, AUSL Parma; 2 UO Neurologia, AOU Parma; 3 Polo Neurologico Interaziendale, Sezione Malattie Neuromuscolari, Italy. In occasional reports, rituximab proved to be efficacious in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) refractory to first-line conventional immunosuppressive therapies, especially in the presence of co-occurring autoimmune and haematological diseases. We report our

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motor and sensory nerves of the four limbs. Routine blood tests, including HbA1c and oral glucose tolerance test, were normal, except for MGUS IgG-lambda. Anti-MAG antibodies and DNA analysis for PMP22 gene were negative. Because his symptoms were mild, a decision was made not to initiate any treatment. By age 60 he noted some difficulty walking caused by right leg weakness, which gradually progressed over one year. At the age of 61 years, examination showed absent deep tendon reflexes, and distal weakness of the lower limbs mainly on the right, with some proximal impairment. Electroneurography showed overall worsening, more marked for conduction velocity and motor action potential amplitude of the right peroneal nerve. Cerebrospinal fluid examination showed a protein level of 54.4 mg/dl (normal, 15–50). He was started on plasma exchange and prednisone 25 mg, with mild improvement and stabilization at follow-up. Our observation, as well as a few other reports, shows that DADS may progress to classic CIDP, thus it is unlikely to be a separate disorder. However, DADS might serve as a useful clinical designation for the characterization of the disorder as a regional variant of CIDP, as its atypical presentation can be diagnostically perplexing, and it might require a somewhat different, non-aggressive, therapeutic approach.

experience in the treatment with rituximab of patients with CIDP not associated with systemic diseases. During the period 2010–2014, four patients with “pure” CIDP (3 women, 1 man, age 39–76 years) were treated with rituximab after unsatisfactory responses or intolerance to first-line therapies. Rituximab dosage was 375 mg per sqm IV, weekly for four consecutive weeks. Disease course was relapsing-remitting in 3 patients and chronic progressive in 1, with a disease duration of 5–10 years pre-rituximab. Patients who improved by at least one point in lower limb score of Overall Neuropathy Limitation Scales (ONLS) were considered as responders. Patients treated with rituximab for neuropathy associated with anti-MAG antibodies were not included. Three of four patients responded to rituximab, with discontinuation of previous therapies, whereas a 73-year-old woman with a 7-year history of chronic progressive CIDP continued to worsen. Improvement occurred 4–6 months after rituximab was started. Responders maintained the improvement during the follow-up period (22–50 months). Previous reports of small series or single cases tend to show that about half of patients with refractory CIDP can be responsive to rituximab, but this is mainly seen in patients with associated haematological disease. It can be supposed that B cells play a predominant role in the pathogenesis of a CIDP subtype responder to rituximab, as this is a monoclonal antibody against CD20+ B lymphocytes. Our data, although anecdotal, confirm that rituximab may represent a treatment option in a subset of refractory CIDP patients, and controlled trials may be warranted. Predictors of response to rituximab have not been identified, and our cases show that the treatment can be effective also in patients with longstanding disease.

SUBCUTANEOUS IMMUNOGLOBULIN IN CIDP AND MMN: A DIFFERENT LONG-TERM CLINICAL RESPONSE? Merola A1 , Cocito D1 , Romagnolo A1 , Peci E1 , Toscano A2 , Mazzeo A2 , Gentile L2 , Russo M2 , Fazio R3 , Filosto M4 , Siciliano G5 , Schirinzi E5 , Lopiano L1 . 1 Department of Neuroscience, University of Turin; 2 Department of Neuroscience, Psychiatry and Anesthesiology, A.O.U. Policlinico “G. Martino” Messina; 3 Department of Neurology, IRCCS San Raffaele Milano; 4 Neurological Clinic - Neuromuscular Diseases and Neuropathies Section, University and A.O. “Spedali Civili” of Brescia; 5 Department of Clinical and Experimental Medicine, University of Pisa, Italy.

DISTAL ACQUIRED DEMYELINATING SYMMETRIC POLYNEUROPATHY (DADS) WITH IGG-LAMBDA MGUS PROGRESSING TO CLASSIC CHRONIC INFLAMMATORY DEMYELINATING POLYRADICULONEUROPATHY Medici D1,3 , Ferrante T1,3 , Pavesi G2,3 , Gemignani F2,3 , Montanari E1,3 . 1 UO di Neurologia della AUSL di Parma; 2 UO di Neurologia della AOU di Parma; 3 Polo Neurologico Interaziendale, Sezione Malattie Neuromuscolari, Italy.

Several studies confirmed the short-term clinical efficacy of subcutaneous immunoglobulin in chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) and multifocal motor neuropathy (MMN), suggesting a similar effectiveness than intravenous immunoglobulin, with the possible advantages of stable plasmatic concentration, lower patient’s emotional distress during infusions and independence from the hospital care. The main objective of this study is to assess the long-term efficacy of subcutaneous immunoglobulin in dysimmune peripheral neuropathies, evaluating the two-year follow-up data of 45 CIDP and 20 MMN patients switched from intravenous to subcutaneous immunoglobulins. Adherence to therapy was the primary outcome measure, while quality of life and clinical predictors of long-term disability progression were analyzed as secondary outcomes. The overall adherence to therapy was 92.2% at 12 months, 87.3% at 18 months and 77.5% at 24 months, with some differences between MMN and CIDP subjects: 15% of MMN returned to the intravenous therapy and 25% required an increase of subcutaneous immunoglobulin dose or an implementation with intravenous infusions

Distal acquired demyelinating symmetric polyneuropathy (DADS) represents an uncommon phenotype of inflammatory neuropathy, which has been proposed either as a regional variant, or as a distinct entity from classic chronic inflammatory demyelinating polyneuropathy (CIDP). It is characterized by a strictly length-dependent pattern, usually with mainly or pure sensory manifestations, and antimyelin-associated glycoprotein (anti-MAG) antibodies are present in the majority of these patients. We report a patient who presented with DADS, later evolving to classic CIDP. This 63-year-old man presented at age 58 with a 6-month history of non-painful dysesthesias in the feet; he also complained of intermittent positional numbness of his hands since two years. He had bilateral pes cavus, normal motor strength and deep tendon reflexes, slightly reduced thermal sensation in the fingertips and vibration perception in the legs. Electroneurography showed mainly demyelinating features, with uniformly slowed conduction velocities in the

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for inadequate therapy efficacy. On the contrary, only 13.3% of CIDP returned to intravenous therapy or required an increase of dose. Quality of life improved in almost all subjects and only two patients reported significant cutaneous side effects, which required the suspension of therapy. Altogether these findings confirm the safety and tolerability of subcutaneous immunoglobulin in dysimmune peripheral neuropathies, but highlight a possible differential efficacy between CIDP and MMN.

CI, 1.3–4.8) in the moderate/severe stage. This study confirms that manual work is an important risk factor for CTS, independently of personal anthropometric risk factors. The association between manual work and CTS severity tends to increase from mild to severe stage of electrophysiological scale. This kind of trend does not seem to be confirmed in the case of subjective symptoms as evaluated by the clinical scale.

EARLY MORPHOLOGICAL ABNORMALITIES OF MYELINATED FIBERS IN A RAT MODEL OF CMT 1A.

CARPAL TUNNEL SYNDROME SEVERITY AND MANUAL WORK: A CASE-CONTROL STUDY

Nolano M1 , Nobbio L2 , Provitera V1 , Pisciotta C3 , Caporaso G1 , Stancanelli A1 , Visigalli D2 , Capodivento G2 , Manganelli F3 , Schenone A2 , Santoro L3 . 1 S. Maugeri Foundation IRCCS, Medical Center of Telese Terme (BN); 2 Department of Neurosciences, Rehabilitation Ophthalmology, Genetics and Maternal-Infantile Sciences (DINOGMI), University of Genoa; 3 Department of Neurosciences, Reproductive Sciences and Odontostomatology, University “Federico II” of Naples, Italy.

Mondelli M1 , Curti S2 , Farioli A2 , Aretini A1 , Ginanneschi F3 , Greco G4 , Argentino A2 , Salce C2 , Mattioli S2 . 1 EMG Service, Local Health Unit 7, Siena, 2 Dpt. Medical and Surgical Sciences, University of Bologna, 3 Dpt. Neurological, Neurosurgical and Behavioural Sciences, University of Siena, 4 EMG Service, Local Health Unit 7, “Nottola” Hospital, Montepulciano (Siena), Italy. Carpal tunnel syndrome (CTS) is a socially relevant condition. This case-control study aims to investigate the association between CTS severity and manual work considering personal anthropometric risk factors as well. We prospectively and consecutively enrolled one CTS case for two controls regardless of age and gender who were admitted to the same three outpatient electromyography labs. CTS diagnosis was made according to clinical findings and distal conduction delay of the median nerve. Case and controls with diabetes, connective and thyroid diseases, renal failure, polyneuropathy, cervical radiculopathy, wrist trauma were excluded, as well as, women in pregnancy or lactation. CTS cases were grouped in two classes of progressive clinical and electrophysiological severity according to two validated five-stage scales: mild (stages I/II) and moderate/severe (stages III-IV-V). Anthropometric measures and occupational history were collected for both cases and controls. Job titles were coded according to the International Standard Classification of Occupations (ISCO 88) by two occupational physicians who were blind to case/control status. Job titles were grouped in two main occupational categories: manual workers and non-manual workers. To assess the association between CTS severity and manual work, multinomial logistic regression models (adjusted for age, sex, wrist-palm ratio and waist-stature ratio) were performed. Relative risk ratios (RRR) and 95% confidence intervals (95% CI) were calculated taking controls as reference category. This case-control study enrolled 370 cases and 747 controls. After the exclusion of retired subjects and housewives, we included 224 cases (age 47.6 ± 12.0 yrs; 154 females, 70 males) and 500 controls (age 42.8 ± 11.6 yrs; 299 females, 201 males) in the main analysis. These subjects were grouped in 496 manual workers and 228 non-manual workers. For manual workers in respect to non-manual workers, the RRR for the electrophysiological severity scale were 2.6 (95%CI 1.4–4.7) for the mild class and 3.3 (95%CI 2.0–5.6) for the moderate/severe class. Regarding the clinical severity scale, the RRR for manual workers compared to non-manual workers were 3.3 (95% CI, 2.0–5.4) in the mild stage and 2.5 (95%

Abnormalities of internodal and nodal length associated with aspects of axonal degeneration have been described in skin samples of Charcot-Marie-Tooth type 1A (CMT1A) patients. It is not clear how early such abnormalities occur and if they undergo changes with aging. In order to elucidate these aspects, we assessed myelinated fiber density and morphometric features in skin samples and in the sciatic nerve of hemizygous CMT1A transgenic rats (Sereda et al., 1996) either in an asymptomatic stage and in a late disease course (22 days and 1 year of age). Specimens were obtained from anterior and posterior foot pads, tail and sciatic nerve of CMT1A rats and wild-type littermates. They were fixed in Zamboni solution, cryoprotected in 20% sucrose in PBS, cut in 50-micron-thick sections and processed with indirect immunofluorescence technique. A panel of antibodies to stain myelin, axon and node-paranode was used. Quantitative assessment of myelinated fibers and measurements of caliber, G-ratio, internodal and nodal length were performed on digital confocal images. We observed in both skin and nerve sections of young and old CMT rats, in addition to nerve fiber loss, several and frequent morphological abnormalities of myelinated fibers, such as caliber irregularities, thinning, myelin fragmentation, shortened internode, enlargement of the nodal-paranodal region. Such abnormalities were rarely observed in controls and were easier to detect on skin samples where myelinated fibers from nerve fascicles separate to reach their terminal territories in dermal papillae. Interestingly, the loss of nerve fibers associated at severe aspects of dysmyelination was very marked in the homozygous rat. Measurements on dermal myelinated fibers showed values of internode length and caliber with a large variability but significantly lower than in controls while nodal length values were significantly higher.

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OPEN QUESTIONS IN CHARCOT-MARIE-TOOTH DISEASE (CMT). A WEB-BASED QUESTIONNAIRE STUDY LINKED TO THE NATIONAL CMT REGISTRY

THE NEUREGULIN1/ERBB SYSTEM, A POTENTIAL TOOL TO IMPROVE PERIPHERAL NERVE REGENERATION Pascal D1 , Tos P2 , Ronchi G1,3 , Raimondo S1,3 , Gambarotta G1,4 , Gnavi S1,3 , Geuna S1,3 . 1 Department of Clinical and Biological Sciences, University of Torino; 2 Reconstructive Microsurgery, Orthopedics Department, Trauma Center Hospital (CTO), University of Torino; 3 Neuroscience Institute Cavalieri Ottolenghi (NICO), Torino; 4 Neuroscience Institute of Torino (NIT), Interdepartmental Centre of Advanced Studies in Neuroscience, University of Torino, Italy.

Pareyson D, Schenone A, Fabrizi GM, Santoro L, Vita G, Quattrone A, Padua L, Previtali SC, Allegri I, Filippini G, Calabrese D; for the Italian CMT Network. Milan, Genoa, Verona, Parma, Rome, Naples, Catanzaro, Messina – Italy. A Charcot-Marie-Tooth disease (CMT) National Registry has started (http:www.registronmd.it) with collection of clinical/genetic information (minimal dataset) and reporting of clinical scales. There is conflicting evidence from the CMT literature about five important issues: disease course and complications during pregnancy; use, efficacy and tolerability of orthotics and assistive devices; outcome of surgery for skeletal deformities; safety of anesthesia; occurrence of sleep disorders. The objective of this study was to collect information and provide answers to these five issues in CMT by using a web-based questionnaire panel linked to the registry. We collected information about these unresolved questions through self-reported questionnaires administered to the Registry population, which is well-characterized and fairly representative. Some already validated questionnaires and other ad hoc developed ones are used. We have developed four novel ad hoc questionnaires following common rules of items generation and reduction by experts in the different fields and by patients. A questionnaire on pregnancy and delivery in CMT has been developed through two focus groups with patients and a psychologist, with the supervision of a gynecologist. Three ad hoc questionnaires on orthotics and assistive devices, surgery for skeletal deformities, and safety of anesthesia have been designed with the contribution of neurologists, methodologists, orthopedists, physiatrists, anesthesiologists, and patients. The four ad hoc questionnaires have been tested on 21 patients and then formally finalized. The list of questionnaires has been completed by adding existing validated scales: QUEST (Quebec User Evaluation of Satisfaction with Assistive Technologies) on use of foot orthotics, FFI (Foot Function Index), ESS (Epworth Sleepiness Scale) and PSQI (Pittsburgh Sleep Quality Index) for sleep, m-FIS (modified-Impact Fatigue Scale), HDAS (Hospital Anxiety and Depression Scale). A website linked to the Registry has just been opened and finalized where all the 10 questionnaires can be filled online by the CMT subjects adhering to the Registry and by controls. Controls fill the questionnaires for pregnancy, anesthesia, and sleep, and are recruited among friends and unaffected relatives of CMT participants, matched for age and sex. We have finalized a series of questionnaires which are now on the website ready to be filled by the CMT patients participating in the Registry and by controls. We aim at collecting information from 500 CMT subjects and from at least 100 healthy controls. We will gain important information to give advice about: pregnancy, orthotics, surgery, anesthesia, sleep and fatigue in CMT. Funded by a Telethon-UILDM grant (GUP13006).

Peripheral nerve trauma or injuries may lead to sensory or motor function deficits if not properly treated. For this reason, in the last years, nerve repair surgical techniques and the regenerative properties of the peripheral nervous system were the focus of many studies. With the aim of improving peripheral nerve regeneration, surgical approaches have matched with new biomedical strategies. In the field of tissue engineering, gene therapy has interesting perspectives. This approach consists of the over expression of specific therapeutic protein and study their regenerative effects on the injured nerve. Among the different trophic factors that regulate the nerve regeneration process, the Neuregulin1/ErbB system plays an important role both in myelination and re-myelination processes. In this study we explored the possibility to manipulate the system, in vitro and in vivo, in order to increase Schwann cell migration and axon survival, thus improve injured peripheral nerve regeneration. To interfere with the Neuregulin1/ErbB system soluble ecto-ErbB4, a protein fragment endogenously released by cells expressing the cleavable isoform of the NRG1 receptor ErbB4, was expressed in vitro in Neuregulin1 expressing glial cells. A strong increase in cell motility was observed. Experiments suggest that activation of a back signaling, mediated by the transmembrane Neuregulin1 isoform, plays a crucial role in the migratory activity induced by fragment expression. Contrary to what happens in neurons, in which back signaling promotes AKT phosphorylation, in glial cells the activated pathway stimulates ERK phosphorylation. Indicating the ability of this protein to activate different pathways depending on the cells type. In the second part of the study we manipulated in vivo the Neuregulin1/ErbB system through local delivery of soluble ecto-ErbB4 by gene transfer in the muscle-vein-combined nerve guide. Data show that fragment delivery has a positive effect on fiber maturation. These results indicate that manipulation of the NRG1/ErbB system by gene therapy could be a promising tool to improve peripheral nerve regeneration. However, to obtain stronger positive effects, a more finely tuned release of the ecto-ErbB4 fragment might be required.

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was unremarkable, but her non-affected parents were consanguineous. She had delayed motor development with gait difficulties due to foot deformities and muscle weakness. At age 5 years, Nerve Conduction Study (NCS) showed markedly decreased velocities (ubiquitously C and p.F491LfsX32 variants were novel. We found SH3TC2 mutations in 50% of this highly selected series, indicating that this phenotype is strongly suggestive of CMT4C. Indeed, scoliosis and/or cranial nerve involvement should strongly address the clinician to perform SH3TC2 gene analysis. Interestingly, in the remaining 9 probands no SH3TC2 mutation was found, indicating that other genes may be causative of this peculiar phenotype. We confirm that the neuropathy of CMT4C is moderate to severe. All are loss-of-function mutations. In our series, the most frequent mutations were p.R954X and p.R1109X (both already reported), the former found in several families of different ethnic origin and the latter mainly in Gypsy families (Claramunt at al., Clin Genet 2007). Moreover, there are possible founder effects, with the splice-site mutation (c.805+2 T > C) carried by apparently unrelated patients from Calabria in Southern Italy, and another mutation (p.Q892X) in different patients from Albania.

of the following iminosugars: Swainsonine, Kifunensine, Castanospermine, N-Butylnojirimycine (NB-DNJ), Deoxymannojirimycine (DMJ), then we tested the effect of those drugs on HeLa cells and Schwannoma rat cells expressing wild type P0 or P0K109N, evaluating the trafficking and the intercellular adhesion as outcome measures. Adhesion test on HeLa cells expressing P0K109N treated with NB-DNJ showed a significant improvement (p < 0.001). Furthermore, western blot analyses demonstrated the ability of NB-DNJ to reduce the molecular weight of the mutant hyperglycosylated protein. In conclusion, different clinical phenotypes of CMT1B are the result of the disruption of a variety of cellular and molecular pathways, including mistrafficking and gain of glycosylation, so that therapy for this disease may need to be equally as complex and individualized as the pathways effected by the expression of mutant forms of P0.

A CASE OF CMTX5 CAUSED BY A MISSENSE MUTATION IN THE PRPS1 GENE ON XQ22.3 AND AXONAL ATROPHY IN THE SURAL NERVE BIOPSY Prada V, Ursino G, Geroldi A, Gotta F, Gemelli C, Emilia B, Grandis M, Mandich P and Schenone A. Department of Neuroscience, Rehabilitation, Ophthalmology, Genetics and Child/Maternal Sciences, (DINOGMI), University of Genova, Genova, Italy. Charcot-Marie-Tooth (CMT) neuropathy represents a clinically and genetically heterogeneous group of hereditary peripheral neuropathies characterized by chronic motor and sensory impairment. Mutations in up to 70 genes may cause CMT. Approximately 10%–20% of CMT is inherited in an X-linked manner (CMTX). Normally CMTX is due to mutations in the GJB1 gene (CMTX1). Recently, the PRSP1 gene has been identified as a rare cause of CMTX (CMTX5). Here, we report a case of CMTX5 caused by a missense mutation at conserved amino acids in the PRPS1 gene on Xq22.3 (c.344T > C;p.M115T). LITAF, MFN2, SH3TC2, dup/del 17p11, GJB1, EGR2, MPZ , GDAP1, SOX10, CX26 mutation were excluded. The disease is inherited in a X-linked recessive manner (mother and the sister of the patient show hearing loss, pes cavus and mild peripheral neuropathy). Disease onset was early in infancy, with distal weakness and sensory impairment, hearing loss and visual abnormalities. From age 4 he used ankle foot orthosis and hearing aid. The neurophysiological findings showed diffuse slowing of conduction velocity, mainly in the lower limbs, which worsened over time to diffuse marked reduction of motor and sensory action potentials and slowing of conduction velocity. Sural nerve biopsy performed when the patient was 6 yrs old showed a normal density of myelinated fibers (MFs), some axons with increased myelin thickness, rare demyelinated axons and onion bulbs. We present a case of CMTX5, in which, the neuropathological study, differently form the sural nerve biopsy in a previously reported case, shows normal density of MFs with clear signs of severe axonal atrophy and only mild demyelination.

THERAPEUTIC OPTIONS FOR MPZ MUTATION CAUSING MISGLYCOSYLATION Prada V1 , Callegari I3 , Salerno A1 , Passalacqua M2 , Tonetti M2 , Schenone A1 , Grandis M1 . 1 Department of Neuroscience, Ophthalmology, Rehabilitation, Genetics and Child/Maternal Sciences, DINOGMI, University of Genova, Genova; 2 Department of Experimental Medicine, DIMES, University of Genova, Genova; 3 Neurology Department, IRCCS National Neurological Institute C. Mondino, University of Pavia, Pavia, Italy. The increasing interest around the role of glycosylation in disease has uncovered the existence of nearly 30 genetic disorders affecting N−linked and O−linked pathways. Moreover, mutations introducing new or removing constitutive N−linkage sites are more frequent than expected. We have recently identified different mutations in the Myelin Protein Zero (MPZ ) gene, encoding P0, causing a gain or loss of glycosylation. In particular we proved that four mutations (N93S, T95A, T95M and T95K) cause a loss-of-glycosylation, while other mutations (D32N, D80N, D89N and K109N) create a new glycosylation consensus sequence. Iminosugars, a class of drugs interfering with the N-glycosylation pathway, are an intriguing group of molecules that might complement mutations causing misglycosylation. Therefore, we investigated the effect of selected iminosugars on transfected cells expressing the fusion protein P0K109N-EGFP corresponding to a hyperglycosylated P0 mutant. We preliminary selected, in HeLa and Schwannoma cells, the sub-toxic concentration

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site (lateral distal lower leg or medial posterior mid-calf) and participating laboratory as possible influential variables. Age, sex and biopsy site showed an independent linear correlation with IENF density. For each decade, the 5th quantile IENF cut-off showed a 0.54 fibers/mm decrease, while females exhibited a 1.0 fibers/mm cut-off greater than males. Compared to the lateral distal lower leg, biopsies from the calf showed a 3.4 fibers/mm lower 5th percentile cut-off, documenting a variation linked by site. We did not find significant differences in terms of IENF density between Caucasian and non-Caucasian subjects although our cohort was not suitable to draw definitive conclusions on this issue. In conclusion, we presented for the first time an age and gender adjusted normative dataset for intra-epidermal nerve fiber density at distal leg obtained with indirect immunofluorescence by sharing data from four experienced laboratories worldwide. This dataset can be used as reference for laboratories processing skin biopsies with this technique.

GENOMIC APPROACH FOR INHERITED MOTOR AND CMT2 NEUROPATHIES: A COLLABORATIVE STUDY Previtali SC1 , Fabrizi GM2 , Schenone A3 , Pareyson D4 , Manganelli F8 , Bellone E3 , Lunetta C6 , Penco S5 , Mandich P3 , Taroni F4 , Corbo M7 , Magri S4 , Ferrarini M2 , Piscosquito G4 , Scarlato M1 , Riva N1 , Viganò F1 Carrera P1 , Pisciotta C8 , Santoro L8 , Ferrari M1 , Bucci G1 , Lazarevic D1 , Cittaro D1 , Comi G1 , Casari G1 , Bolino A1 . 1 IRCCS San Raffaele Scientific Institute, Milano; 2 University of Verona; 3 University of Genova; 4 IRCCS Foundation, “C. Besta” Neurological Institute; 5 Niguarda Ca’ Granda Hospital, Milano; 6 CENTRO CLINICO NEMO, Milano; 7 Casa di Cura del Policlinico, Milano; 8 University Federico II of Naples, Naples, Italy. Patients with hereditary motor neuropathy (HMN) may clinically and genetically overlap with axonal Charcot-MarieTooth (CMT2) neuropathies and motor neuron disorders (MND). Recent evidence shows that the same gene can be mutated in all these diseases, suggesting that they represent a unique spectrum of disorders. There is currently no effective treatment for all HMN/CMT2/MND, and responsible genes are still mostly unknown. Thus, the identification of new molecules and signaling pathways remains a major challenge to understand their pathogenesis and to develop prospective therapies. We have designed a collaborative study aimed at the identification of new genes responsible for HMN/CMT2/MND by using an integrated approach based on NGS panel and exome sequencing. Candidate genes will undergo validation studies including expression and functional analyses. As a preliminary result, we have identified a new candidate gene for HMN/CMT2 neuropathy in a consanguineous family.

TRAUMATIC LESION OF RADIAL NERVE: ELECTROPHYSIOLOGICAL AND ULTRASOUND EVALUATION Romano M1 , Cosentino G2 , Gagliardo A3 , Cottone S1 , Brighina F2 , Fierro B2 . 1 UOC di Neurologia A.O.O.R Villa Sofia Cervello Palermo; 2 Dipartimento di Biomedicina sperimentale e Neuroscienze cliniche (BioNeC), Università di Palermo; 3 U.O. Neurofisiopatologia, accreditata con SSN, Palermo, Italy. A 27-year-old man was accidentally shot during hunting. The dorsal portion of the right arm was involved. Soon he presented with motor weakness in the right hand. The neurological examination revealed right hand drop, with deep tendon reflexes present. Neurophysiology one month after the incident showed: absence of cMAP in the right radial nerve (recorded at the common extensor digitorum muscle). Normal sensory conduction of the radial nerve; normal sensory-motor conduction of all the other nerves. EMG: active muscle denervation of the common extensor digitorum and partially of muscle brachioradialis. Ultrasound examination showed multifocal damage of the radial nerve, in particular in the posterior interosseous branch. A neuroma was also observed.

IMMUNOFLUORESCENT INTRAEPIDERMAL NERVE FIBER NORMAL DENSITY AT DISTAL LEG: A MULTICENTER STUDY Provitera V1 , Gibbons CH2 , Wendelchafer-Crabb G3 , Donadio V4 , Vitale DF1 , Stancanelli A1 , Caporaso G1 , Liguori R, Wang N2 , Santoro L5 , Kennedy WR3 , Nolano M1 . 1 “S. Maugeri” Foundation IRCCS, Medical Center of Telese Terme (BN), Italy; 2 Department of Neurology, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA; 3 Department of Neurology, School of Medicine, University of Minnesota, Minneapolis, MN, USA; 4 IRCCS Institute of Neurological Sciences, University of Bologna, Italy; 5 Department of Neurosciences, Reproductive and Odontostomatological Sciences, University “Federico II” of Naples, Italy.

NRG1/ERBB SYSTEM REGULATION AFTER PERIPHERAL NERVE INJURY Ronchi G1,2 , Viano N1 , Ceccopieri S1 , Perroteau I1 , Geuna S1,2 , Gambarotta G1 . 1 Dept. of Clinical and Biological Sciences, University of Turin, Orbassano (TO); 2 Neuroscience Institute of the “Cavalieri Ottolenghi” Foundation (NICO), University of Turin, Orbassano (TO), Italy.

The objective of the study was to create an age and gender adjusted normative dataset for intra-epidermal nerve fiber (IENF) density at distal leg in skin samples processed with immunofluorescence. We collected from four experienced laboratories worldwide IENF density data of 528 healthy individuals. In all laboratories skin samples were collected, processed and analyzed according to standard procedures. We employed quantile regression analysis to tailor the fit of the 5th percentile as the normal cut-off value and to test and measure the effect of age, sex, body mass index, race, biopsy

One of the most important trophic factors involved in Schwann cell activity is Neuregulin1 (NRG1), which is involved in survival, proliferation, differentiation, migration and myelinating activity of glial cells during development. It is well known that Schwann cells play a key role in peripheral nerve regeneration and that NRG1 is also required for axon

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(AMSAN). Serology for HIV, EBV, CMV, mumps and Borrelia was negative, as was CSF polymerase chain reaction assay (PCR) for Herpes virus, Borrelia, Mycoplasma Pneumoniae, Cryptococcus and Mycobacterium tubercolosis. Positivity for TOSV IgM antibodies was found on both CSF and serum; the patient remembered being recently exposed to mosquitoes. He was treated with plasma-exchange with complete clinical recovery and was discharged home. In the June 2013, the patient complained of relapsing numbness and tingling paraesthesias, along with episodic weakness, at lower limbs. The nerve conduction studies confirmed the presence of the AMSAN, but some demyelinating features were also detected. The lumbar puncture was repeated; the serology for Borrelia, Rickettia Conori, West Nile, Mosaico Sandy Fever was negative, but positivity for TOSV IgG antibodies was found on both CSF and serum. The patient was started on steroid treatment, which was slowly tapered, as he was prone to complain of a clinical worsening if the dosage was decreased too quickly. This is the first case, to the best of our knowledge, of a TOSV infection associated to a peripheral neuropathy. Remarkably, the TOSV infection was able to clinically and electrophysiologically mimic a GBS syndrome. Hence, we suggest that TOSV infection should be taken into account in the diagnostic work up of GBS-like polyneuropathy in the summer months, especially in geographic areas where mosquitos are abundant for climatic reasons.

regeneration and remyelination after nerve injury in adulthood. NRG1 is a very large gene that codes for several different isoforms, arising by alternative splicing and/or use of different promoters; NRG1 exerts its effect by binding the ErbB2/ErbB3 heterodimer receptor expressed by Schwann cells. In this study we focused our attention on the changes of the NRG1/ErbB system during nerve degeneration and regeneration. In order to clarify the relationship between the expression of the NRG1/ErbB system and the different phases of nerve degeneration and regeneration, three types of experimental lesions were carried out: (I) axonotmesis (crush lesion), that is a complete interruption of axon continuity without discontinuity of the nerve, characterized by fast nerve regeneration; (II) neurotmesis, that is a complete transection of the nerve, followed by end-to-end repair, characterized by slow nerve regeneration; (III) neurotmesis without nerve repair, characterized by chronic nerve degeneration. mRNA and proteins were extracted from the distal region of adult rat median nerves at different time points after the injury. Quantitative real-time-PCR and western blot analysis for the different NRG1 isoforms, the NRG1 tyrosine kinase receptors (ErbB1-ErbB2-ErbB3) and the myelin basic protein (MBP) were performed. In parallel, to better understand and explain mRNA and protein expression results, nerve morphology ultrastructure changes were also investigated by electron microscopy analysis. Results showed intense and significant changes in gene and protein expression, especially shortly after injury. mRNA and protein expression is modulated with different time courses following the different phases of nerve degeneration and regeneration. This analysis shows that the different NRG1 isoforms and ErbB receptors display a highly specific expression patterns, suggesting that they play important roles after peripheral nerve injury.

PICC INSERTION IMPROVES NUTRITIONAL STATUS, AUTONOMIC SYMPTOMS AND QOL IN PATIENTS WITH TTR-FAP Russo M1 , Vita GL1 , Stancanelli C2,3 , Vita G1,3 , Mazzeo A3 , Messina S1,3 . 1 Centro Clinico Nemo Sud, University of Messina; 2 Biomedical Department of Internal and Specialistic Medicine, University of Palermo; 3 Department of Neuroscience, University of Messina, Italy.

GUILLAIN-BARRÉ SYNDROME-LIKE ONSET OF ACUTE POLYNEUROPATHY ASSOCIATED WITH TOSCANA VIRUS INFECTION

Familial amyloidotic polyneuropathy (TTR-FAP) usually presents itself as a progressive sensorimotor polyneuropathy with severe autonomic dysfunction and cardiomyopathy. Autonomic symptoms, such as gastrointestinal dysfunction, orthostatic hypotension, sexual and urinary dysfunction, may be the first signs of the disease. Loss of more than 10% of body weight can be also an early manifestation of TTR-FAP. Cachexia is regularly present after few years from the clinical onset, due to gastrointestinal dysautonomia, muscle atrophy, and infection. In late stage of disease (stage 3) patients become bedridden and exposed to bedsores, venous thrombosis, and pulmonary embolism. How malnutrition can affect survival and quality of life in TTR-FAP is not well described. However, a recent study on a cohort of AL amyloidosis patients shows how malnutrition affects the prognosis independently of hematologic response to treatments. Herein we describe the good clinical outcome in two patients affected by TTR-FAP in which a satisfactory nutritional status has been achieved after a peripherally inserted central catheter (PICC) was placed. The two patients (a 52-year-old female and a 63-year-old-male, carrying Ala49 and Gln89 mutation, respectively) underwent a series of regular follow-up

Rota E1 , Morelli N1 , Immovilli P1 , De Mitri P1 , Ratti G2 , Guidetti D1 . 1 Department of Neurology, Guglielmo da Saliceto Hospital, Piacenza; 2 Department of Infectivology, Guglielmo da Saliceto Hospital, Piacenza, Italy. Numerous disorders mimicking Guillain-Barré syndrome (GBS) have been reported in the literature. Toscana virus (TOSV) is an arthropod-borne emerging pathogen in the Mediterranean area, mainly in the summer. Despite the increasing evidence of a role played by TOSV in CNS infections, it remains a neglected agent and no cases of polyneuropathy associated to TOSV infection have been reported to date. Here we describe a 40-year-old male patient with no antecedent disease at anamnesis, admitted to the hospital in September 2012 for acute facial weakness, associated to numbness paraesthesias at lower and upper limbs. The neurological examination revealed facial diplegia and reduced tendon reflexes. In the suspicion of a GBS syndrome, the patient underwent a lumbar puncture, which detected increased protein levels (242 mg/dl), and the presence of 8 lymphocytes/microl. The nerve conduction studies documented an acute motor and sensory axonal neuropathy

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before and after the PICC insertion including: modified body mass index (mBMI), an autonomic symptoms questionnaire, 24-hour blood pressure holter and Norfolk Quality of Life Questionnaire-Diabetic Neuropathy (QOL-DN). Our findings showed an improvement of the general clinical status, gain of weight and mBMI, reduced orthostatic intolerance and ameliorated quality of life. Our experience suggests that PICC has to be considered in TTR-FAP patients, when severe malabsorption and diarrhea make unuseful all dietary adjustments causing malnutrition and dehydration.

NEUROIMAGING OF SCIATIC INTRANEURAL PERINEURIOMA Salvalaggio A1 , Cacciavillani M2 , Gasparotti R3 , Ferraresi S4 , Briani C1 . 1 Dept. of Neurosciences, DNS, University of Padova, Padova; 2 CEMES-EMG Lab, Data Medica Group, Padova; 3 Dept. of Medical and Surgical Specialties, Radiological Sciences and Public Health, University of Brescia, Brescia; 4 Rovigo Hospital, Neurosurgery, Rovigo, Italy. Intraneural perineurioma is a benign peripheral nerve sheath tumor of perineurial cell origin. We describe the clinical, neurophysiological, Ultrasound and 3D magnetic resonance (MR) Neurography findings of a perineurioma involving the sciatic nerve. An 8-year-old girl, with an unremarkable medical history, presented three years ago with gait impairment initially ascribed to the fact that her right leg was 1 cm shorter than the left one. Neurological examination revealed loss of strength at the plantar and dorsal flexion of the right foot and distal hypotrophy of right lower limb; there was no impairment of touch sensation or pain. A neurophysiological/EMG examination in March 2014 revealed denervation of biceps femoris (long and short head), tibialis anterior, extensor digitorum longus, gastrocnemius muscles of the right leg. Nerve ultrasounds revealed an increased cross sectional area (CSA) of the sciatic nerve, more noticeable in the middle third of the thigh (CSA 34 mm2 on the right leg, 14 mm2 on the left one). A perineurioma was suspected and a 3D MR Neurography performed. A fusiform enlargement of the right sciatic nerve, from the great ischiatic foramen (CSA 31mm2 at the right leg, 15 mm2 at the left leg) to the popliteal fossa (52 mm2 vs, 11 mm2 ) was demonstrated. The fusiform enlargement was 20 cm long and showed contrast enhancement. In December 2014 the patient underwent surgical tendon transposition; since the diagnosis was clear, no histological examination was performed. In conclusion, the present case confirms the role of ultrasound, as a precious complementary examination to the EMG, very useful to localize the lesion along the course of the involved nerve. 3D MR neurography confirmed the suspect of perineurioma, thus allowing to preserving the nerve from surgery in an already severely affected child.

3D MR NEUROGRAPHY OF AXILLARY NERVE INJURY: CASE REPORT Salvalaggio A1 , Cacciavillani M2 , Baldo M3 , Briani C1 , Gasparotti R4 . 1 Dept. of Neurosciences, Sciences NPSRR, University of Padova, Padova; 2 CEMES, Data Medica Group, EMG Lab, Padova; 3 Villa Ferri Medica, centro di fisiatria e riabilitazione, Padova; 4 Dept. of Medical and Surgical Specialties, Radiological Sciences and Public Health, University of Brescia, Brescia, Italy. A 26-year-old male player on the rugby national team, with an unremarkable past medical history, in October 2013 had a trauma to his right shoulder during a match. He immediately felt an electric shock-like sensation in his right shoulder radiating to the arm. Soon after he presented with loss of strength of the right deltoid muscle. In December 2013 an electromyography (EMG) demonstrated a complete denervation of the deltoid muscle likely secondary to axillary nerve injury. Being a part of the rugby national team, the prognosis of the axillary nerve damage (axonotmesis or a neurotmesis) was crucial for the ongoing championship. An EMG response would have been possible only after 3 months from the trauma. Nerve ultrasound was little useful given the axillary nerve course and surgery inspection had also been considered. MR imaging of the brachial plexus and of the right shoulder was performed in January 2014, using 3D MR neurography sequences and multiplanar reconstructions. High-resolution 3D MR neurography showed integrity of the brachial plexus and a fusiform enlargement of the right axillary nerve suggesting a posttraumatic neuroma, with nerve continuity on either side. Surgery was therefore excluded. In February 2014 an EMG examination still revealed a complete deltoid denervation. In April 2014 (six months after the trauma) an EMG examination revealed reinnervation potentials even before clinical evidence of improvement, then progressively improved. In October 2014 the patient returned to play in the rugby league. The novel MRI reconstruction technique that was applied was able to differentiate axonotmesis from neurotmesis avoiding a surgery inspection. This MR technique is suited for the examination of nerves that run deeply (and so not suitable for US evaluation) and, in the present case, provided evidence on prognosis earlier than EMG.

EFFICACY OF CYCLOSPORINE AND PREDNISONE COMBINATION THERAPY IN A-CIDP ASSOCIATED WITH MINIMAL CHANGE NEPHROTIC SYNDROME Schirinzi E1,2 , Caponnetto C3 , Garnero M3 , Novi G3 , Schenone A3 , Siciliano G2 , Mannironi A1 , Benedetti L1 . 1 Department of Neurology, S. Andrea Hospital, La Spezia; 2 Department of Clinical and Experimental Medicine, Neurological Clinic, University of Pisa, Pisa; 3 Department of Neuroscience, Rehabilitation, Ophthalmology, Genetics, Maternal and Child Health (DINOGMI), University of Genoa, Italy. The simultaneous occurrence of GBS and nephrotic syndrome has previously been reported. However A-CIDP and nephrotic syndrome remained rarely reported. A 78-year-old woman complained of paresthesias at hands and feet and progressive fatigability to both legs two weeks after an upper respiratory tract infection. Neurological examination showed

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diaphragmatic ultrasounds 2–4 months postoperatively). 11 subjects showed unilateral or bilateral postsurgical phrenic nerve palsy, 5 of which were permanent, with electrophysiological demonstration of prolongation of diaphragmatic action potential (DAP) latency and/or fall in DAP amplitude. A consensual diaphragmatic hypomotility or immobility was found by echography in all cases except one. In particular, a striking correlation seemed to be found between the amplitude of the DAP and both muscular motility at ultrasound and ventilatory dysfunction. Between the 6 infants with transient phrenic neuropathy, 2 showed a recovery at T2, 3 recovered at T3 and 1 at T4. We found that both neurophysiology and echography represent important tools in the respiratory follow-up after heart surgery in children, and considerations regarding the timing of the possible recovery of a phrenic neuropathy are discussed.

loss of strength, decreased pinprick sensation, and tendon areflexia at four limbs. Neurophysiological studies showed slowed velocity of motor nerves, with conduction blocks in left peroneal and ulnar nerve, and non-recordable sural nerve conduction velocity. Cerebrospinal fluid examination showed normal cell count, hyper-proteinorrachia, and identical cerebrospinal fluid and serum oligoclonal bands. The patient, with a diagnosis of GBS, was treated with intravenous immunoglobulin (IVIg) (0.4 g/Kg/day for 5 days). Neurological symptoms transiently improved, but after three weeks a second IVIg treatment was performed due to a clinical worsening. The patient didn’t improve and a further progressive clinical worsening was recorded. After eight weeks from the onset of symptoms the neurological examination revealed a severe flaccid paresis at four limbs. The diagnosis was changed in A-CIDP. As we were going to begin plasma exchange an extensive laboratory examination was performed. A severe proteinuria (34 g/L) with hypoalbuminemia and hypoproteinemia discovered a nephrotic syndrome. So we decided to not perform plasma exchange and we began high-dose intravenous methylprednisolone. Renal biopsy showed a minimal change nephrotic syndrome (MCNS). Both neuropathy and proteinuria improved with prednisone (1 mg/Kg/day) combined with cyclosporine (2–3 mg/Kg/day). She remained free of proteinuria and clinically stable during three months of follow-up. A-CIDP and MCNS responded to immunosuppressive therapy suggesting a common pathogenesis of the two conditions.

CLINICAL FEATURES AND DIAGNOSIS OF AUTONOMIC FAILURE IN FAMILIAL AMYLOIDOTIC POLYNEUROPATHY (FAP) Stancanelli C1 , Iodice V2 , Mazzeo A1 , Vita G1 , Mathias CJ2 . 1 Department of Neurosciences, University of Messina, Italy; 2 Autonomic Unit, National Hospital for Neurology and Neurosurgery, Institute of Neurology, University College London, UK. Amyloidosis is a group of diseases characterized by tissue deposition of insoluble proteins and fibril aggregates. Mutations on the transthyretin (TTR) gene are the most common forms of familial amyloidotic polyneuropahty (FAP). Other genes include apolipoprotein A1, gelsolin or alpha fibrinogen. Progressive autonomic failure is a common clinical feature in patients with FAP. The aim of this study was to characterize the autonomic cardiovascular function in FAP patients. Retrospective study of a cohort of 113 patients (100 symptomatic and 13 asymptomatic) with confirmed diagnosis of FAP. Clinical history and examination, and autonomic functional tests results were collected. The patients underwent head-up tilt (HUT), standing test, Valsalva maneuver, pressure stimuli and deep breathing, liquid meal challenge, modified exercise tests and supine and tilted cathecholamine. Autonomic dysfunction is strongly present in FAP (only 12/100 symptomatic patients had normal autonomic functional tests. Widespread cardiovascular autonomic failure was found in 19% of the patients). Parasympathetic dysfunction is predominant in FAP (detected in 69% of symptomatic patients and asymptomatic patients). Orthostatic hypotension (OH) was found in 34% of the symptomatic patients. Postprandial and exercise induced OH was found in 10% of FAP patients who did not present OH on standard HUT or standing tests. We have characterized the autonomic dysfunction in a large cohort of patients with FAP using an extensive battery of autonomic cardiovascular testing. Abnormalities of the HR variability (using the deep breathing test and the Valsalva maneuver) represent early sign of autonomic impairment in FAP. In addition, the study highlighted the key role of additional cardiovascular test, including liquid meal challenge and modified exercise tests to assess the autonomic dysfunction profile in FAP.

PHRENIC NERVE PALSY IN PEDIATRIC HEART SURGERY: THE NEUROPHYSIOLOGICAL CONTRIBUTION IN FOLLOW-UP AND DECISION MAKING Segatti A1 , Zanelli S2 , Santuz PA2 , Biban P2 , Luciani GB3 , Romito S1 . 1 Neurology Unit, Azienda Ospedaliera Universitaria Integrata, Verona; 2 Pediatric Intensive Care Unit, Azienda Ospedaliera Universitaria Integrata, Verona; 3 Department of Surgery, Pediatric Cardiac Surgery Unit, University of Verona, Azienda Ospedaliera Universitaria Integrata, Verona, Italy. Phrenic nerve palsy after cardiac surgery in infants is a well-known undesirable event. The resulting dyaphragmatic dysfunction can be transient or permanent, and lead to failure of weaning from ventilatory support and other respiratory severe complications. Diaphragmatic palsy can be resolved with plication, but the timing for the procedure is still debated, just for the possible transient nature of the phrenic nerve damage. Moreover, information on long-term outcome in plicated children, in terms of growing and ventilatory function, are still lacking. The aims of this study were to demonstrate the possibility of a temporary phrenic nerve paralysis, more prolonged than reported in previous works, and to compare the electroneurographic findings with those of ultrasound. 28 infants, younger than one year, affected by different kinds of heart congenital malformations, were studied both by phrenic nerve conduction study (pnNCV) and by diaphragmatic ultrasonography, before the surgical correction (T0) and periodically in the postoperative period (T1 within 72 hours, T2 after 7–10 days, T3 after 14-21 days, T4 after 21–28 days; two babies underwent pnNCV and

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Mutations in Cu/Zn SOD1 cause familial amyotrophic lateral sclerosis (ALS), a degenerative disease characterized by loss of lower and upper motor neurons in the motor cortex, brain stem and spinal cord. Although ALS course reflects the loss of motor neurons, a number of reports have described sensory abnormalities in patients. Moreover, loss of small nerve fiber was demonstrated in patients by skin biopsy and confocal corneal microscopy, although the underlying molecular mechanisms have never been investigated. We aimed at addressing this issue by analyzing SOD1 levels and neurite stress features in dorsal root ganglion (DRG) neurons from two SOD1G93A mouse models. Our results demonstrated that DRG neurons express SOD1 and that SOD1G93A protein significantly accumulates in DRG neurons from presymptomatic and symptomatic SOD1G93A mice compared to wild-type (SOD1WT). We also found that SOD1G93A expression decreased neurite length and promoted the transcription of the early axonal injury sensor ATF3. To test the hypothesis that mutant SOD1 differentially affects neuronal sub-types, we analyzed neurite length and ATF3 labeling in in vitro small (30μm) DRG neurons from SOD1G93A and non-transgenic littermates. We demonstrated a specific neurite length decrease and ATF3 expression increase in peripherin and IB4-positive small DRG neurons, whereas no change was found in NF200-positive large DRG neurons. The specific impairment of small DRG neurons explained the loss of epidermal nerve fiber density we found in SOD1G93A mice. Ongoing study will assess the molecular mechanisms underlying these findings.

ROLE OF RAB7 IN CHARCOT-MARIE-TOOTH TYPE 2B DISEASE Stasi M1 , Bramato R1 , Manganelli F2 , Nolano M3 , Cogli L1 , Santoro L2 and Bucci C1 . 1 Dept. Biological and Environmental Sciences and Technologies, University of Salento, Lecce; 2 Dept. Neurological Sciences, University of Naples “Federico II”, Naples; 3 Salvatore Maugeri Foundation IRCCS, Telese Terme, Benevento, Italy. Charcot-Marie-Tooth type 2B disease is a peripheral ulcero-mutilating neuropathy caused by five missense mutations in the rab7 gene. This rare neuropathy is characterized by prominent sensory loss, distal muscle atrophy and high frequency of ulcers and infections that often lead to toe and foot amputations. Rab7 is a small GTPase controlling transport to late endosomes and lysosomes. Although the biochemical properties of the CMT2B-causing Rab7 mutant proteins have been investigated, it is not yet clear how mutations in a ubiquitous protein like Rab7 cause a peripheral neuropathy. Others and we have demonstrated that Rab7 has multiple functions in the cell and specific functions in neurons. Indeed, Rab7 is involved in phagocytosis, in autophagy and in mitophagy but also in neurotrophin trafficking and signaling, regulating neurite outgrowth and neuronal migration. Furthermore, Rab7 interacts with RILP (Rab Interacting Lysosomal Protein), which has been demonstrated to be down-regulated in sural nerve biopsy of a CMT2B patient. In addition, Rab7 interacts with vimentin and peripherin, two intermediate filament proteins. Thus, Rab7 mutations could impact on several cellular processes affecting in particular peripheral neurons. For instance, injury to the peripheral nervous system triggers an immediate damage-response mechanism leading to extensive axonal regeneration. This damage-response mechanism recapitulates the developmental stages required for neuronal differentiation and therefore axonal regeneration after injury depends on neurotrophin trafficking and signaling, efficient autophagy and mitophagy, and on vimentin and peripherin functions. All these processes seem to be controlled by Rab7 and thus CMT2B-causing Rab7 mutants could perturb one or more of these events in peripheral neurons. Using transient transfection we demonstrated that CMT2B-causing Rab7 mutants exhibit altered interaction with vimentin and peripherin affecting their assembly, inhibit neurite outgrowth and perturb the authophagic flux. Recently, we isolated dermal fibroblast from a CMT2B patient carrying the V162M mutation and we are confirming and extending some of these findings. Future studies on relevant cells (DRG neurons) will be necessary to establish if and how these processes are altered and to pinpoint target molecules for future therapeutical approaches.

RECURRENCE OF APPARENTLY REMITTED DIFFUSE LARGE B-CELL LYMPHOMA PRESENTING WITH ISOLATED THIRD NERVE PALSY Terenghi F1 , Balducci C1 , Crocchiolo R2 , Morabito L2 , Nobile-Orazio E1 . 1 2∘ Neurology, Department of Medical Biotechnology and Translational Medicine, Milan University, 2 Department of Medical Oncology and Haematology, both at Humanitas Clinical and Research Centre, Rozzano, Milan, Italy. Neurolymphomatosis (NL) is a rare neurologic manifestation of non-Hodgkin lymphoma (NHL) and leukemia. The diagnosis is often elusive with available diagnostic procedures. We report a 40-year-old man with a two-year history of diffuse large B-cell lymphoma (DLBCL, III2B) with abdomen localization treated with six R-CHOP cycles. Two recurrences of B-cell lymphoma were treated with radiotherapy and second line chemotherapy with two DHAP cycles. After 1-year with negative radiologic follow up, allogenic stem cell transplantation was planned. At this time he reported episodic diplopia in the evening, lasting few minutes and associated with cervical pain. At that time neurological examination was normal. Cerebrospinal fluid (CSF) cells were increased (83 cell/mm3 , 88% leukocytes, with normal lymphocytes subpopulation on flow cytometry), proteins were 139 mg/dl, and glucose 42 mg/dl. Brain MRI with gadolinium showed an enhancement of III, IV and V bilateral cranial nerves without parenchymal changing whereas cervical and dorsal spine

SMALL DRG NEURONS ARE SPECIFICALLY AFFECTED IN MOUSE MODELS OF AMYOTROPHIC LATERAL SCLEROSIS Taiana MM, Sassone J, Lombardi R, Porretta-Serapiglia C, Cazzato D, Bianchi R, Lauria G. Neuroalgology and Headache Unit, IRCCS Foundation “Carlo Besta” Neurological Institute, Milan, Italy.

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plantar surface on the midfoot (right foot: 10.6 ± 9.3 mm2 vs. 22.2 ± 8.4 mm2 , p < 0.001; left foot: 13.1 ± 8.6 mm2 vs. 21.8 ± 8.6 mm2 , p = 0.003) with respect to the controls. At stabilometric analysis CMT patients had a higher velocity of CoP (24.7 ± 10.8 mm/s vs. 15.3 ± 4.8 mm/s, p = 0.001) and a greater length of the sway path (1123.9 ± 505 mm vs. 742.4 ± 281.8 mm; p = 0.005) than controls. These parameters were not influenced by eye closure. The velocity of CoP correlated with clinical motor disability as assessed by CMTES (p = 0.03) and with weakness of dorsi-flexor muscles assessed by MRC (p < 0.05) and dynamometer (p = 0.03). No correlation was found with either the other clinical features, including the vibration, or the static baropodometric findings. Static baropodometric findings were consistent with foot deformity (pes cavus). Stabilometric data showed an altered balance in CMT1A patients during upright standing. The imbalance in our CMT patients seems to be related to the weakness of the dorsiflexor muscles rather than proprioception impairment or foot deformity.

MRI were normal. After six days neurological examination revealed divergent strabismus on the right side in association with absent ocular movements and ptosis on the left side. CSF studies showed 21 cell/mm3 (92% leukocytes and normal flow cytometry), 180 mg/dl proteins and 26 mg/dl glucose. Laboratory and radiologic findings ruled out infections (tuberculosis, viral) and inflammatory (sarcoidosis) disease. A subsequent brain gadolinium MRI after seven days showed leptomeningeal involvement. The patients was treated with high dose intravenous methylprednisolone but progressively worsened with headache, nausea, cognitive impairment and lower limb hypotonic and areflexic palsy. Spinal gadolinium MRI showed multiple dorsal and lumbar meningeal involvement. DLBCL is the most common form of lymphoma with secondary CNS recurrence that usually occurs early, during the initial course of therapy. Isolated oculomotor nerve palsy is rare. In our patient unilateral isolated oculomotor palsy was the initial manifestation of lymphoma recurrence. CSF findings with elevated protein, low glucose levels and high CSF cells and MRI abnormalities supported the diagnosis of NL despite the normality of CSF flow cytometry. This diagnosis should be always suspected in a patient with remission lymphoma and new neurological symptoms.

RITUXIMAB IN REFRACTORY CHRONIC INFLAMMATORY DEMYELINATING POLYRADICULONEUROPATHY Velardo D, Bianchi F, Mauri E, Comi G, Fazio R. Department of Neurology, Institute of Experimental Neurology, Division of Neuroscience, San Raffaele Scientific Institute, Vita-Salute San Raffaele University, Milan, Italy.

THE SWING OF CMT1A PATIENTS Tozza S1 , Pisciotta C1 , Aceto G1 , Bruzzese D2 , Santoro L1 , Manganelli F1 . 1 Department of Neurosciences, Reproductive Sciences and Odontostomatology, University Federico II of Naples, Naples; 2 Department of Preventive Medical Sciences, University Federico II of Naples, Naples, Italy.

Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is a progressive or relapsing-remitting disease causing muscle weakness and loss of sensitivity, probably due to an autoimmune process. First-line therapies include corticosteroids, immunoglobulin and plasma-exchange, but about 25-30% of patients do not have a good response to these treatments, so neurologists are searching for new effective drugs. In clinical practice CIDP patients were treated with different immunosuppressive drugs, including cyclophosphamide, cyclosporine, mycophenolate mofetil, interferon alpha and peripheral blood stem cell transplantation; more recently, therapies using monoclonal antibodies, such as rituximab, offered the best promise for treatment of CIDP, even if needing further data. Here we report four cases of CIDP patients who were non-responders to first-line treatments (corticosteroids or immunoglobulin) and other immunosuppressive agents, whose disease improved after treatment with rituximab. The patients had different ages, ranging from 16 to 79, and different disease stages at the beginning of treatment, only one of them showing IgM kappa monoclonal gammopathy. All patients received a first course of rituximab infusion (two 1000 mg intravenous infusions separated by two weeks) and a different number of further treatment cycles (rituximab 1000 mg) every 6 months; they all showed increased muscle strength and improvement in quality of life and are now undergoing clinical and electrophysiological follow-up. Controlled trials are needed to better selection of patients and to evaluate short- and long-term efficacy of the drug, but rituximab seems to be a very promising new therapeutic option in patients with refractory CIDP.

Balance disturbances represent a disabling symptom in CMT disease, which may determine postural instability with falls. Different factors contribute to CMT imbalance. Previous studies have suggested that both distal weakness of lower limbs and proprioceptive sensory deficits may influence the balance in these patients. However, the basis of CMT imbalance remains controversial. The aim of this study was to investigate in a group of CMT1A patients the balance during upright standing by using a baropodometric platform. We evaluated static and stabilometric parameters in 21 CMT1A patients (mean age = 38.8 ± 16.9; M/F = 10/11) and 24 mean age-matched healthy controls (mean age = 43 ± 20.8; M/F = 13/11). For static parameters, during upright standing, we considered load and plantar surface of forefoot, midfoot and barefoot. For stabilometric parameters we evaluated, during upright standing, sway area, velocity of center of pressure (CoP) and length of the sway path at both open and closed eyes. Clinical disability was evaluated by using CMT examination score (CMTES). Muscle strength at lower limbs was assessed by MRC scale and maximal isometric strength of dorsi- and plantar-flexion was tested using a dynamometer. We also recorded Achilles’ tendon retraction, presence of foot deformity and scoliosis. Stabilometric data were correlated with all clinical features and with data from static analysis. Static baropodometric analysis showed that CMT patients had a higher load on the forefoot (right foot: 26.1 ± 4.9% vs. 19.3 ± 6.8%, p = 0.001; left foot: 26.4 ± 6% vs. 19.6 ± 5.3%, p < 0.001) and a smaller

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Research Center of Cardiovascular Biology, University of Genova, Italy.

MORPHOMETRICAL ANALYSIS IN DORSAL ROOT GANGLIA (DRG) CULTURES: AN “ALL-AROUND” TOOL FOR DRUG SCREENING IN MYELIN DISORDERS

Demyelination and remyelination are major issues for physicians working on myelin disorders, but fast, reliable and low-cost methods to monitor myelin rearrangements still lack both in humans and experimental models. Myelin is mainly composed by lipids and particularly enriched in glycosphingolipids. Indeed, these types of lipids are essential for the proper biochemical and biophysical organization of myelin membranes. Moreover, in a recent attempt to overview inborn errors of metabolism involving lipid biosynthesis and remodeling, more than 20 genes have been shown to cause several disorders in many of which peripheral neuropathy has been described. Lipids may therefore be used as marker for myelin membranes integrity and associated nerve fiber function. Thus, we established a high-throughput fluorescence-based assay to readily quantify sphingomyelin in tissue homogenates and biological fluids to use sphingomyelin dosage as a marker of myelin loss, lack and eventually recovery (Italian Patent: TO2014A001001). In this assay, sphingomyelin is hydrolyzed by enzymatic reactions in a single 100-microl mixture for 20 min. The use of 96-well plates allows automated and sensitive quantification by a fluorescence detector. To validate this method, we first demonstrated in different experimental settings, i.e., rat and mouse tissues, central and peripheral nervous system, genetic and inducible injury that sphingomyelin dosage reliably resembles myelin remodeling. Then, we moved to human patients to verify our hypothesis. We found that our assay was able to detect sphingomyelin either in serum and cerebrospinal fluid (CSF) of these patients. Then we quantified sphingomyelin in a large cohort of patients with suspected peripheral neuropathy. Interestingly, following laboratory and clinical confirmation of the diagnosis, we found the highest levels of sphingomyelin in patients affected by demyelinating neuropathies, namely chronic inflammatory polyradiculoneuropathy and Guillain-Barré syndrome which inversely correlated with their motor conduction velocities. Moreover, by correlation of sphingomyelin levels with the blood brain barrier (BBB) injury index QAlb, we were able to discriminate demyelinating neuropathies either from the controls or from other non-demyelinating neuropathies with BBB damage. Thus, sphingomyelin dosage is a reliable fast, inexpensive and laboratory-friendly method to improve the management of patients affected by demyelinating neuropathies.

Visigalli D1 , Capodivento G1 , Abbate M2 , Schenone A1 , Nobbio L1 . 1 Dept. of Neurosciences, Rehabilitation, Ophthalmology Genetics and Maternal-Infantile Sciences and CEBR, University of Genoa; 2 Immagini & Computer snc, Milan, Italy. Mixed glial cultures are more complicated than single-cell cultures to establish and use but they can serve as an intermediate culture system for examination of demyelinating and remyelinating agents between simpler single-cell preparations and the resource-consuming in vivo models. Myelinating dorsal root ganglia cultures (DRG) have been extensively used to study molecular mechanisms and therapeutic strategies for a variety of diseases affecting the peripheral nervous system, including Charcot-Marie-Tooth disease type 1A (CMT1A) neuropathy. However, most of the analyses performed on these in vitro models, especially when they were used in pre-clinical studies to test the pro-remyelinating effects of different molecules, are quite rough being confined to the mere quantification of myelin density, without taking into account the structural complexity of the DRG cultures and myelinated fibers. In this study, we developed a semi-automated morphometric analysis able to evaluate several myelin and axon parameters within these in vitro myelinating systems. DRG cultures were stained with different antibodies specific for the myelin membrane, the axon and myelinated fiber domains essential for their proper physiological function. Images from the whole DRG cultures were then acquired and scanned by an ad-hoc macro that we developed to quantify several parameters, including: i) myelinated area; ii) internode, axon and node density, length and thickness, and their relative frequency functions. Moreover, we compared these morphometric indexes, in wild type and CMT1A rat DRG cultures, either in basal conditions and following pharmacological treatments. Interestingly, we were not only able to discriminate the pathological phenotype from the wild type one but also to find a different and specific effect of each treatment on these morphometric indexes which we completely lacked with conventional analyses. As we currently lack outcome measures and defined guidelines to assess these in vitro models, the development of our morphometric analysis system can be able to select truly effective drugs targeting demyelination and neurodegeneration processes and thus, speed up the translation of these therapeutic interventions to in vivo models and eventually human patients.

NOVEL OUTCOME MEASURES FOR CHARCOT-MARIE-TOOTH DISEASE Vita G, Padua L, Pareyson D, Schenone A, Fabrizi GM, Santoro L, Gemignani F, Quattrone A, Mazzeo A, Russo M, Stancanelli C, Gentile L, Pazzaglia C, Iacovelli C, Simbolotti C, Piscosquito G, Calabrese D, Aiello A, Bolla S, Cavallaro T, Manganelli F, Pisciotta C, Vitetta F, Contini M, Valentino P. Messina, Rome, Milan, Genoa, Verona, Naples, Parma, Catanzaro – Italy.

SPHINGOMYELIN LEVELS IN BIOLOGICAL FLUIDS AS A BIOMARKER FOR DYSIMMUNE DEMYELINATING NEUROPATHIES Visigalli D1 , Capodivento G1 , Garnero M1 , Parodi G1 , Grandis M1 , Ursino G1 , Ferrara DM1 , Garibaldi S2 , Nobili F1 , Capello E1 , Schenone A1 , Nobbio L1 . 1 Dept. of Neurosciences, Rehabilitation, Ophthalmology Genetics and Maternal-Infantile Sciences and CEBR, University of Genova; 2 Cardiology, Dept. of Internal Medicine,

Despite few clinical trials and natural history studies performed in CMT disease, responsiveness of most

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Abstracts


used outcome measures (OM) is low due to slow disease progression, confirming the need to find new and more sensitive-to-change OM. Two tools that most likely reflect accurately patient motor performances in daily living, but not yet validated in CMT patients, are the 6-minute walk test (6MWT) and the StepWatchTM Activity Monitor (SAM). They have been widely employed to assess functional exercise capacity in neurological diseases and in some other neuromuscular disorders. We performed a prospective follow-up (baseline and after 6 and 12 months) of 180 CMT patients (n. 100 with CMT1A, n. 40 with CMT1B, n. 40 with X-linked CMT) to evaluate reliability and validity of 6MWT and SAM compared with already validated OM previously used in CMT patients, specifically: CMTNS, 10-meter timed walking test, distal arm and leg strength measured by MVIC and SF-36 health related quality of life scale. Age of patient population (52% women) had a normal distribution. We present here only the baseline data. 6 and 12 months results are still under analysis. 6MWT resulted highly inversely related to the 10MWT (r = -0.54; p = -7.2E-14). Moreover 6MWT showed high inverse relationship with either CMTNS total score (r = -0.55; p = 9.35E-09) or subscore (CMTES r = -0.53; p = 1.79E-12). The 6MWT showed also high significant relationship with all the hand myometer measures (minimal significance: r = 0.46; p = 1.00E-09). As expected, the 6MWT

was always highly related also to foot myometer measures, including both plantar flexion and dorsiflexion (minimal significance: r = 0.45; p = 2.69E-09). Finally, regarding quality of life, the 6MWT showed high positive relationship with the main physical domain of the SF36 (r = 0.68; p = 8.45E-23) and to the mental domain (r = 0.20; p = 1.34E-02). Concerning SAM results we found a more complex pattern, likely because the different measures provided by the SAM represent very different kind of activities. Globally it seems that those measures assessing the best activity in a short time are more related with myometer measures of dorsal and plantar foot flexion and with CMTNS. One of the main steps of validity and reliability process, the construct validity, showed that 6MWT is very well related to previously used OM for CMT. Diversely, only some SAM measures, likely related to high activity on a short time, seem to be related to results from previously validated measures. However, more detailed statistical analysis will tell us if some single parameters obtained from both 6MWT and SAM can be better indicators of disease status. In addition, the analysis of results at 6 and 12 months can offer the opportunity to reveal if the two new tools are more sensitive to changes over time relating to disease progression. Funded by a Telethon-UILDM grant (GUP10008).

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