Acquired Neuropathies Figure 1. Transverse semi-thin sections of the musculo-cutane- ous nerve of a control and patients with chronic acquired neuropathies. A. Normal density of myelinated fibers in a nerve without lesion. B. In a CIDP nerve (Patient 12), several hypomyelinated fibers (arrowheads) and an onion bulb formation (arrow) are visualized. C. In CIAP nerve (Patient 13), loss in myelinated fibers and numerous regenerating clusters (arrowheads) are present. Scale bar 30 mm. doi:10.1371/journal.pone.0014533.g001 impossible to differentiate from CIAP on the basis of clinical and electrophysiological examinations, a situation that requires a nerve biopsy [10]. This procedure must be performed by trained physicians and examined in a laboratory with expertise in nerve pathology, as it requires sophisticated approaches, such as electron microscopy of transverse sections of peripheral nerve [10,11]. Myelinated fibers are characterized by highly differentiated domains along the axon, centered by nodes of Ranvier [12,13]. The voltage-gated Na+ channels (Nav), essential for the rapid saltatory conduction of action potentials, are concentrated at nodes of Ranvier in association with ankyrin G, a cytoplasmic protein interacting with cortical cytoskeleton and cell adhesion molecules [14], and KCNQ2 potassium channels [15]. The paranodal septate- like junctions separate the nodal and juxtaparanodal regions, attach the glial loops to the axonal membrane, and restrict the lateral diffusion of axonal membrane proteins between the node and the internodal space. Several proteins involved in axoglial contacts have been identified [12,16], including paranodin [17], also known as Caspr [18]. Nodal and paranodal proteins are altered in mouse models of CMT [19,20], and have been investigated in skin biopsies from CMT patients [21]. However, the possible alterations of nodal and paranodal regions in CIDP and CIAP are not known. The purpose of this study was to investigate the nodal and paranodal regions in these polyneuropathies using both electron microscopy and immunofluorescence in order to better understand their pathophysiology and refine their diagnosis. Results Nodal and paranodal ultrastructure is differentially altered in Schwann cells and axons in CIDP and CIAP Superficial peroneal nerve biopsies were studied in 12 patients diagnosed with CIDP (Table S1) and 10 with CIAP (Table S2). We first examined toluidine blue staining of transverse semi-thin sections (Fig. 1). In contrast with a sample with no sign of peripheral neuropathy which was considered as a control (Fig. 1A), hypomyelination (Fig. 1B, arrowheads) and onion bulbs (Fig. 1B, arrow) were visible in a typical CIDP sample. The CIAP sample displayed a decreased number of fibers and regeneration clusters (Fig. 1C, arrowheads). Since, to our knowledge, the nodal regions have not been explored in CIDP or CIAP, we used electron microscopy to determine how these regions were altered in chronic acquired neuropathies. Ten nodes of Ranvier were examined for either pathology, in longitudinal sections of nerves from two patients with CIDP and two with CIAP. In CIDP myelinated fibers, the abaxonal cytoplasm and the perinodal extensions of Schwann cells were abnormal as well as the paranodal glial loops (Fig. 2). The outer Schwann cell cytoplasm contained multiple large vacuoles some- times enclosing granular structures (Fig. 2A–D, arrows). The axons were not entirely normal and the nodal axoplasm included some vacuolar structures (Fig. 2B). At the level of paranodes, adherens junctions between adjacent glial loops (Fig. 2C) and axoglial septate junctions (Fig. 2C, arrowheads) were relatively preserved. However, some paranodal loops were of irregular size and some contained lamellar and multivesicular bodies (Fig. 2D, arrowheads). PLoS ONE | www.plosone.org 2 January 2011 | Volume 6 | Issue 1 | e14533