Shortened internodal length of dermal myelinated nerve fibres in Charcot–Marie-Tooth disease type 1A

doi:10.1093/brain/awp274 Brain 2009: 132; 3263–3273 | 3263 BRAIN A JOURNAL OF NEUROLOGY Shortened internodal length of dermal myelinated nerve fibres in Charcot–Marie-Tooth disease type 1A Mario A. Saporta,1 Istvan Katona,1 Richard A. Lewis,1 Stacey Masse,1 Michael E. Shy1,2 and Jun Li3 1 Department of Neurology, Wayne State University, Detroit 48201, USA 2 Department of Molecular Medicine and Genetics, Wayne State University, Detroit 48201, USA 3 Department of Neurology, Vanderbilt University, Nashville 37232, USA Correspondence to: Jun Li, MD, PhD, Department of Neurology, Vanderbilt University School of Medicine, AA0204F Medical Center North, 1161 21st Avenue South, Nashville, TN 37232-2551, USA E-mail: Charcot–Marie-Tooth disease type 1A is the most common inherited neuropathy and is caused by duplication of chromosome 17p11.2 containing the peripheral myelin protein-22 gene. This disease is characterized by uniform slowing of conduction velocities and secondary axonal loss, which are in contrast with non-uniform slowing of conduction velocities in acquired demyelinating disorders, such as chronic inflammatory demyelinating polyradiculoneuropathy. Mechanisms responsible for the slowed conduction velocities and axonal loss in Charcot–Marie-Tooth disease type 1A are poorly understood, in part because of the difficulty in obtaining nerve samples from patients, due to the invasive nature of nerve biopsies. We have utilized glabrous skin biopsies, a minimally invasive procedure, to evaluate these issues systematically in patients with Charcot–MarieTooth disease type 1A (n = 32), chronic inflammatory demyelinating polyradiculoneuropathy (n = 4) and healthy controls (n = 12). Morphology and molecular architecture of dermal myelinated nerve fibres were examined using immunohistochemistry and electron microscopy. Internodal length was uniformly shortened in patients with Charcot–Marie-Tooth disease type 1A, compared with those in normal controls (P50.0001). Segmental demyelination was absent in the Charcot–Marie-Tooth disease type 1A group, but identifiable in all patients with chronic inflammatory demyelinating polyradiculoneuropathy. Axonal loss was measurable using the density of Meissner corpuscles and associated with an accumulation of intra-axonal mitochondria. Our study demonstrates that skin biopsy can reveal pathological and molecular architectural changes that distinguish inherited from acquired demyelinating neuropathies. Uniformly shortened internodal length in Charcot–Marie-Tooth disease type 1A suggests a potential developmental defect of internodal lengthening. Intra-axonal accumulation of mitochondria provides new insights into the pathogenesis of axonal degeneration in Charcot–Marie-Tooth disease type 1A. Keywords: CMT1A; internodal length; Schwann cell; skin biopsy; Charcot–Marie-Tooth disease Abbreviations: Caspr = contactin-associated protein; CIDP = chronic inflammatory demyelinating polyradiculoneuropathy; CMT1A = Charcot–Marie-Tooth disease type 1A; MBP = myelin basic protein; PGP = protein gene product; PMP22 = peripheral myelin protein 22 gene Received July 30, 2009. Revised September 6, 2009. Accepted September 11, 2009 ß The Author (2009). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: 3264 | Brain 2009: 132; 3263–3273 Introduction Charcot–Marie-Tooth disease (CMT) type 1A is the most common inherited neuropathy, representing almost 50% of all CMT patients (Skre, 1974; Nelis et al., 1996). CMT1A is associated with 1.4 Mb duplication in chromosome 17p11.2 (Lupski et al., 1991; Raeymaekers et al., 1991) which includes the peripheral myelin protein 22 (PMP22) gene. PMP22 over-expression is believed to play an important role in the pathogenesis of CMT1A since over-expression of PMP22 in rodents causes a similar neuropathy. Deletion of the PMP22 gene also causes neuropathy with a different clinical phenotype, named hereditary neuropathy with liability to pressure palsies. Typical phenotypes in patients with CMT1A consist of childhood onset, distal and symmetrical weakness, muscle atrophy, sensory loss, areflexia and foot deformities (Harding and Thomas, 1980; Thomas et al., 1997). Nerve conduction studies show uniform slowing in conduction velocities, which can be observed even in young children (Nicholson, 1991; Garcia et al., 1998; Yiu et al., 2008) and is distinct from the non-uniform slowing of conduction velocities in the acquired demyelinating neuropathies (Lewis and Sumner, 1982). However, the mechanism of the uniform slowing in CMT1A is still elusive. In sural nerve biopsies of patients with CMT1A, increased myelin thickness followed by active de-/remyelination has been observed during the first years of life (Gabreels-Festen et al., 1992, 1995), with the amount of segmental demyelination significantly decreasing after the teenage years (Fabrizi et al., 1998). At the same time, onion bulbs, formed by supernumerary Schwann cells that are not attached to axons, gradually appear in the majority of myelinated fibres (Robertson et al., 2002; Hattori et al., 2003). Variable secondary axonal degeneration has been shown to gradually occur during late childhood (Gabreels-Festen et al., 1995). Axonal degeneration, but not de-/re-myelination, correlates with neurological disability in patients with CMT1A (Dyck et al., 1989; Krajewski et al., 2000). The mechanisms responsible for the axonal loss in CMT1A are yet to be determined. Acquired demyelinating neuropathies such as chronic demyelinating inflammatory polyradiculoneuropathy (CIDP) are often asymmetric, both in their clinical presentation and in their nerve conduction studies. In particular, slowing in nerve conduction velocities is non-uniform in distinction to the uniform slowing described in CMT1 (Lewis and Sumner, 1982). Pathologically, CIDP is also non-uniform in that segmental demyelination is scattered along myelinated nerves with some internodes shorter than others as a result of the de-/remyelination (Hahn, 2005). Pathological distinctions between inherited and acquired demyelinating neuropathies often require invasive procedures such as sural nerve biopsies. Furthermore, DNA testing provides reliable diagnosis and eliminates the necessity of sural nerve biopsy on patients with CMT1A. Thus, pathophysiological studies in patients with CMT have been limited by the difficulty in obtaining nerve samples due to the invasive nature of this procedure. As an alternative approach, we and others have begun to utilize skin biopsies to obtain morphological and molecular information from dermal myelinated nerve fibres (Nolano et al., 2003; Li et al., 2005; M. A. Saporta et al. Provitera et al., 2007). This approach has taken the use of skin biopsy beyond the established application of determining unmyelinated epidermal nerve fibre density in small fibre sensory neuropathies (Holland et al., 1998; Polydefkis (...truncated)