Brain Advance Access originally published online on April 22, 2003
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Brain, Vol. 126, No. 7, 1638-1649,
July 2003
© 2003 Guarantors of Brain
doi: 10.1093/brain/awg151
Changes in the expression and localization of the paranodal protein Caspr on axons in chronic multiple sclerosis
Netherlands Institute for Brain Research, Amsterdam, The Netherlands
Correspondence to: Guus Wolswijk, Netherlands Institute for Brain Research, Meibergdreef 33, 1105AZ Amsterdam, The Netherlands E-mail: g.wolswijk{at}hetnet.nl
Received October 22, 2002. Revised January 15, 2003. Accepted February 15, 2003.
 |
Summary |
|---|
 Top Summary IntroductionMaterial and methodsResultsDiscussionReferences |
|---|
The presence of intact paranodal junctions on myelinated axons in the CNS and PNS is crucial for both myelin sheath attachment and saltatory impulse conduction. The axonal glycoprotein contactin-associated protein (Caspr) is expressed in the paranodal region and plays an important role in the creation and maintenance of these adhesive junctions. In the present study, antibodies to Caspr were used to assess the integrity of paranodal junctions on myelinated axons in brain and spinal cord tissue from subjects with longstanding multiple sclerosis, a neurological disorder that affects both myelin and axons. Triple immunofluorescence combined with confocal laser scanning microscopy showed that axons in the demyelinated centre of the 36 brain and 16 spinal cord multiple sclerosis lesions studied were devoid of Caspr immunoreactivity, suggesting that axons down regulate the expression of Caspr following demyelination. Additional data indicated that Caspr reappears in the paranodal region with the formation of new myelin sheaths. Immuno labelling further revealed that Caspr on myelinated axons in border regions was often no longer concentrated in the paranodal region, but was also present in the internodal region—a phenomenon particularly common in the borders of the more chronic lesions in the collection. Myelinated axons with long Caspr-positive stretches were often present at a considerable distance from the lesion edges. These findings raise the possibility that the aberrant location of Caspr is an early sign of impending myelin loss. This would imply that demyelination continues at a slow rate in established lesions. The diameters of Caspr-positive structures on some myelinated axons near the lesion edges were also increased. Moreover, the gap between individual myelin sheaths on these apparently swollen axons was widened occasionally and a very small myelin sheath plus additional Caspr-positive structures had sometimes formed in the enlarged space. This finding thus suggests that the formation of new myelin in multiple sclerosis is not only induced following the loss of complete internodes but also in response to broadening of the nodal region. Interestingly, alterations in the expression and localization of Caspr were observed in tissue from both subjects with the primary and secondary progressive form of multiple sclerosis. In summary, the present study provides immunohistochemical evidence that paranodal junctions on some myelinated axons in the borders of lesions of patients with chronic progressive multiple sclerosis are no longer intact. This may impair saltatory impulse conduction and lead to further myelin loss, thereby contributing to disease progression in multiple sclerosis.
Keywords: axon pathology; Caspr; demyelination; paranodal junction; saltatory impulse conduction
Abbreviations: Caspr= contactin-associated protein; HLA = human leukocyte antigen; MOG = myelin oligodendrocyte glycoprotein
|
Introduction |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Multiple sclerosis is an acquired, immune-mediated and often progressive disorder of the CNS characterized by the presence of multiple, chronically demyelinated lesions, axonal loss and astrocytic scarring (Lassmann, 1998
; Prineas et al., 2002). The damage to myelin, oligodendrocytes (the myelin-forming cells of the CNS) and axons occurs predominantly during the active phase of lesion formation, but continuously at a slow rate in more chronic lesions (Ferguson et al., 1997; Trapp et al., 1998; Bitsch et al., 2000; Kornek et al., 2000; Lucchinetti et al., 2000; Prineas et al., 2001; Kuhlmann et al., 2002). Several pathogenic mechanisms underlying the demise of oligodendrocytes and myelin in multiple sclerosis have been identified (Lassmann, 1998; Lucchinetti et al., 2000), but much less is known about the mechanisms that cause axons to degenerate in this disease (Coleman and Perry, 2002), a process that results in further loss of myelin. Early symptoms experienced by multiple sclerosis patients are thought to result directly from the disruption or removal of the insulating sheath of myelin around axons, leading to the slowing or blockade of saltatory impulse conduction, while more permanent neurological deficits have been attributed to cumulative axonal loss (Davie et al., 1995; De Stefano et al., 1998; Trapp et al., 1998; Wingerchuk et al., 2001; Compston and Coles, 2002).
The cytoplasm-filled myelin loops of oligodendrocytes in the CNS and Schwann cells in the PNS adhere tightly to the axolemma in domains flanking nodes of Ranvier—termed paranodal junctions (Salzer, 1997; Peles and Salzer, 2000; Scherer and Arroyo, 2002). These septate-like, adhesive junctions are also critical for the propagation of action potentials via saltatory conduction because they form a barrier to the flow of ionic currents between the voltage-gated Na+ channels at the node and the delayed-rectifier K+ channels in the juxtaparanodal region (Chiu and Ritchie, 1980; Wang et al., 1993; Waxman and Ritchie, 1993; Mi et al., 1995). Moreover, the paranodal junctions prevent molecules from entering the peri-axonal space and they are thought to represent a site for communication between myelinating glia and axons (Hirano and Llena, 1995; Salzer, 1997; Brophy, 2001). The absence of proper paranodal junctions leads to a profound reorganization of the axonal membrane, including the displacement of K+ channels to the paranodal region and, as a result, a severe reduction in nerve conduction velocity and neurological deficits (Dupree et al., 1999; Bhat et al., 2001; Boyle et al., 2001). Thus, the presence of both myelin and intact paranodal junctions is crucial for the rapid propagation of action potentials from node to node.
A neuronal molecule that is highly enriched in the paranodal region of mature myelinated axons in the CNS and PNS and that has been implicated in the formation and maintenance of the paranodal junctions is the Drosophila Neurexin IV-related molecule contactin-associated protein (Caspr), also termed paranodin (Einheber et al., 1997; Menegoz et al., 1997; Peles et al., 1997a,b). The present study has examined in detail the expression of this paranodal glycoprotein on nerve fibres in multiple sclerosis lesions at various stages of evolution using indirect immunofluorescence techniques and confocal laser scanning microscopy. It was hypothesized that disease-associated disturbances in neuronal and/or axonal functioning might affect the expression of Caspr on the surface of axons and thus the attachment of the myelin loops to the axolemma and the integrity of the barrier between the nodal Na+ channels and the juxtaparanodal K+ channels.
|
Material and methods |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Tissue fixation and treatment
Spinal cord and brain tissue with a short post-mortem delay (3 h 45 min to 16 h 45 min) were obtained from the Netherlands Brain Bank (co-ordinator, R. Ravid); the Netherlands Brain Bank received permission for performing autopsies, for the use of tissue and for access to medical records for research purposes from the Ethical Committee of the Free University Medical Centre, Amsterdam, The Netherlands. At autopsy, blocks of spinal cord and brain tissue (mostly periventricular tissue) from subjects who died of non-neurological conditions (Case 1, 38-year-old male; Case 2, 46-year-old female; Case 3, 69-year-old male; Case 4, 77-year-old female; Case 5, 78-year-old female; Case 6, 78-year old-male; Case 7, 89-year-old female) and from subjects with long-standing multiple sclerosis (Table 1) were placed in a solution of 4% paraformaldehyde in phosphate-buffered saline (pH 7.4), stored for 1–7 days at 4°C and incubated in a solution of 30% sucrose for 1–3 days at 4°C under constant rotation (Wolswijk, 1998
, 2000, 2002). The tissue was then placed in a boat prepared from aluminium foil containing Tissue-Tek O.C.T embedding compound (Sakura Finetek Europe B.V., Zoeterwoude, The Netherlands), frozen on dry ice and stored at –80°C. Dr W. Kamphorst, Department of Pathology, The Free University Medical Centre, Amsterdam, The Netherlands, carried out the routine neuropathological analysis on the control and multiple sclerosis tissue.
|
Immunohistochemistry
Ten micrometer thick sections were cut from each tissue block using a Reichert-Jung 2800 cryostat (Reichert-Jung, Vienna, Austria) (cutting temperature –25°C), mounted onto SuperFrost*/Plus microscope slides (Menzel-Gläser, Braunsweig, Germany), stored at –20°C and then immunolabelled using indirect immunofluorescence or immunoperoxidase techniques as described previously (Wolswijk, 1998, 2000, 2002). Sections were incubated for 1–3 days at 4°C in the primary antibody solutions, rinsed several times in Tris-buffered saline (pH 7.6) and then incubated for 1–3 days at 4°C with the fluorochrome (FITC, TRITC, Cy3 or Cy5)-conjugated or biotinylated anti-rabbit or anti-mouse immunoglobulin G (H+L) or anti-mouse immunoglobulin subclass-specific antibodies (purchased from either Southern Biotechnology Associates, Inc., Birmingham, AL, USA or Jackson ImmunoResearch, West Grove, PA, USA). The binding of the biotinylated antibodies was visualized by incubating sections in the presence of the Vectastain ABC kit reagents A and B (Vector Laboratories, Inc., Burlingham, CA, USA) followed by substrate [a filtered solution of 0.42 mg/ml 3-amino-9-ethylcarbazole dissolved in dimethyl formamide (Merck, Darmstadt, Germany) and 0.01% hydrogen peroxide (Merck) in 0.05M sodium acetate (Sigma Chemical Company, St. Louis, MO, USA) buffer, pH 5.0] or by incubating sections in the presence of fluorochrome-coupled streptavidin (Vector Laboratories). The main primary antibodies used in the present study were: (i) a mouse anti-myelin oligodendrocyte glycoprotein (MOG) monoclonal antibody (a generous gift of Dr S. Piddlesden; Piddlesden et al., 1993
); (ii) a mouse anti-neurofilament monoclonal antibody [the RT97 antibody (Wood and Anderton, 1981); Chemicon International, Inc., Temecula, California, USA]; (iii) a mouse anti-non-phosphorylated neurofilament monoclonal antibody (the SMI32 antibody; Sternberger Monoclonals, Inc., Maryland, USA); (iv) a mouse anti-human leukocyte antigen (HLA) HLA-DP, DQ, DR monoclonal antibody (DAKO A/S, Glostrup, Denmark); and (v) a rabbit antiserum to Caspr (a generous gift of Dr Elior Peles, The Weizmann Institute of Science, Rehovot, Israel; Peles et al., 1997b; Poliak et al., 1999). Antibodies were diluted in Tris-buffered saline containing 0.125% Triton X-100 (Sigma) and 2.5% heat-inactivated calf serum (Sigma). At the end of the staining procedure, a drop of glycerol containing 22mM 1,4-diazobicyclo [2,2,2] octane (Sigma) was placed on the section (to reduce fading of the fluorochromes), followed by a glass cover slip. The excess glycerol was removed and the cover slip was then sealed using clear nail varnish. Sections were viewed on a Zeiss Axiophot microscope equipped with phase-contrast, Nomarski, bright-field and dark-field optics, epi-UV illumination and selective filters optimized for distinguishing between FITC, TRITC/Cy3 and Cy5 emission or on a Zeiss 410 inverted confocal laser scanning microscope equipped with lasers emitting at 488, 543 and 633 nm to excite FITC, TRITC/Cy3 and Cy5, respectively, and with bright-field optics (Carl Zeiss B.V., Sliedrecht, The Netherlands).
Measurements of the dimensions of Caspr-positive structures and data analysis
The length and diameter of individual Caspr-positive structures in control white matter, normal-appearing multiple sclerosis white matter and the borders of multiple sclerosis lesions were measured with the help of printed confocal laser scanning microscope-generated images (stacks of seven optical sections, 1 µm apart; image size: 68 x 68 µm); 1 mm in the printed images represented 0.267 µm. Microcal Origin 4.1 software (Microcal Software, Inc, Northampton, MA, USA) was used to analyse the data.
|
Results |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Caspr expression on axons in control brain and spinal cord tissue
Caspr-positive, paranodal-like structures on axons in periventricular brain white matter and spinal cord tissue from subjects without neurological disease were elongated, sharply demarcated and cylindrical in shape (Figs 1 and 2); tissue from seven subjects were analysed in these studies. The structures were small and numerous in brain white matter and in grey matter areas of the spinal cord, but were larger and less numerous in white matter regions of the spinal cord (Figs 1 and 2). Immunolabelling involving antibodies to Caspr, myelin (antibodies to MOG) and nerve fibres (antibodies to neurofilament, a class of intermediate filaments found in axons) established that Caspr on myelinated axons in control tissue is expressed in regions flanking the nodes of Ranvier and that bundles of neurofilaments run through the centre of the Caspr-positive structures (Fig. 2). The studies further revealed that Caspr-positive, paranodal-like structures were also present on axons in the spinal nerve tissue attached to the control spinal cord samples (data not shown).
|
|
5 µm, i.e. over eight times wider than the average diameter of a control Caspr-positive structure. The nodal region is indicated with an arrow. No labelling with antibodies to HLA-DP, DQ, DR (blue colour) was present in this area of the section. (H) Myelinated axon with a cluster of three Caspr-positive structures in the border of a lesion from multiple sclerosis Case 19. A short MOG-positive myelin sheath (Caspr expression in brain tissue with demyelination
Virtually all demyelinated axons in the centre of the four active, 17 chronic active and 15 chronic inactive brain lesions studied lacked Caspr immunoreactivity (Fig. 2); these lesions were derived from 26 subjects with longstanding multiple sclerosis [8–52 years of clinical disease duration (Table 1)]. Demyelinated axons expressing Caspr were, however, observed in and close to the borders of many of the lesions (Fig. 2). The Caspr immunoreactivity on these demyelinated axons was present often in the form of long stretches and, sometimes, a small gap was visible between two stretches—possible representing the location of what previous had been the nodal domain. Demyelinated axons expressing Caspr were most common in the borders of lesions obtained from multiple sclerosis Cases 2, 4 (lesion c), 17 and 24. Heminodes at the lesion edges occasionally had Caspr immunoreactivity on their demyelinated side (Fig. 2).
The borders of the multiple sclerosis lesions harboured immunoreactive structures that were considerably larger than those present in control and normal-appearing multiple sclerosis white matter. Measurements on Caspr-positive structures in confocal laser scanning microscope-generated images of control brain white matter showed that the control structures were on average 2.0 ± 0.7 µm in length and 0.6 ± 0.2 µm in diameter (n = 90) (Fig. 3). The dimensions of Caspr-positive structures in normal-appearing multiple sclerosis brain white matter did not differ significant from those of control structures [length: 1.6 ± 0.6 µm; diameter: 0.6 ± 0.2 µm (n = 90) (Fig. 3)]. In contrast, many immunoreactive structures in the borders of multiple sclerosis lesions were > 4 µm in length and/or >1.1 µm in diameter (Fig. 3) and they were generally also more strongly labelled with antibodies to Caspr than their counterparts in control and normal-appearing multiple sclerosis white matter (Figs 1 and 2). In total, the sizes of 327 enlarged structures in the borders of two active, seven chronic active and four chronic inactive lesions were measured (Fig. 3). Forty percent of these structures had significantly increased lengths (average length: 6.8 ± 2.9 µm, 22% had significantly increased diameters (average diameter: 1.8 ± 0.5 µm), while 38% had both increased lengths and diameters (average length: 7.7 ± 3.9 µm; average diameter: 2.0 ± 0.7 µm) (Figs 3 and 4). Myelinated axons with long stretches of Caspr immunoreactivity were most common in the borders of lesions derived from multiple sclerosis Cases 2, 4, 17 and 24 (Table 2). Immunoreactive structures with clearly increased diameters were relatively common in the borders of lesions from multiple sclerosis Cases 4, 16 and 17 (Table 2). The enlarged structures were intermingled with Caspr-positive structures that had normal dimensions and were found predominantly near the lesion edges. The width of the zone around the lesions containing Caspr-positive structures with increased lengths and/or widths varied in size from <0.5 mm to >1.5 mm (Table 2 and Fig. 4). There were no striking differences between the dimensions and frequencies of atypical Caspr-positive structures in the borders of brain lesions from subjects with primary progressive or secondary progressive multiple sclerosis (Table 2).
|
|
|
Caspr-positive structures with increased dimensions appeared to be more common in the borders of chronic active and chronic inactive lesions than in the borders of active lesions (Table 2 and Fig. 4). For example, <10% of the immunoreactive structures were enlarged in microscope fields close to the borders of active lesions from multiple sclerosis Case 7 (lesion a) and Case 9 (lesion a), while >80% of the Caspr-positive structures had increased dimensions in microscope fields close to the borders of chronic inactive lesions from multiple sclerosis Cases 17 and 24 (see Fig. 1). In addition, the average diameter of the enlarged structures, but not their length, tended to increase gradually with maturation of the lesions [1.1 ± 0.5 µm for structures in the borders of active lesions (n = 24), 1.3 ± 0.6 µm for structures in the borders of chronic active lesions (n = 194) and 1.8 ± 0.9 µm for structures in the borders of chronic inactive lesions (n = 109); lengths were on average, 5.7 ± 2.5 µm, 6.3 ± 3.5 µm and 6.4 ± 3.7 µm, respectively].
Confocal laser scanning microscopical analyses suggested that the Caspr-positive structures with increased lengths were generally not demarcated sharply as on control axons. The Caspr labelling appeared instead to be present in a gradient with higher concentrations found nearest to the nodal region (Fig. 2). The stretches of Caspr labelling were sometimes >20 µm in length, i.e. on average >10 times longer than those on control axons. The redistribution of Caspr appeared to occur on axons with normal diameters as well as on those with increased diameters (Figs. 2 and 3).
A number of myelinated axons with apparently increased diameters had rows of three or four (and sometimes more) sharply demarcated immunoreactive structures in their (para)nodal region (Fig. 2). Immunolabelling involving antibodies to Caspr, MOG and neurofilament revealed that extremely short myelin segments were frequently present in the centre of these clusters [average length internodes: 8.3 ± 5.4 µm (n = 16); range 2.1–23.4 µm]. Myelinated axons with such clusters were observed relatively frequently in the borders of lesions derived from multiple sclerosis Cases 4 (lesion c), 14, 16, 17 and 26 (Table 2 and Fig. 4). Due to the varying orientations of axons in the brain tissue, the presence of large numbers of myelinated axons in border areas and the thinness of the sections (10 µm), it proved difficult to determine whether lesion borders harboured myelin segments that were larger than those associated with clusters of Caspr-positive structures, but were still unusually short; the presence of short myelin segments is indicative of remyelination (Prineas and Connell, 1979; Lassmann, 1983; Prineas et al., 1993, 2002). However, segments with a length >25 µm were observed occasionally (Fig. 2).
To explore the possibility that enlarged Caspr-positive structures were associated specifically with injured axons, some sections were immunolabelled with antibodies to Caspr, myelin and non-phosphorylated neurofilaments (the SMI32 monoclonal antibody), which are thought to be enriched in axons following damage (Trapp et al., 1998). These studies provided evidence that Caspr-positive structures with increased lengths and/or widths were present on the surface of myelinated axons that contained non-phosphorylated neurofilaments, as well as on those that lacked such neurofilaments (Fig. 2). However, demyelinated axons that still expressed Caspr were generally devoid of non-phosphorylated neurofilaments (Fig. 2).
Caspr expression in spinal cord tissue with demyelination
The expression of Caspr was also analysed in sections cut from 16 blocks of spinal cord tissue obtained from 13 subjects with chronic multiple sclerosis (Table 1). The extent of demyelination in these samples varied from <5% to >95% of an entire cross-section, and nine of the samples contained significant numbers of macrophages laden with myelin debris (Wolswijk, 2002).
As was the case with the brain lesions, demyelinated axons expressing Caspr were virtually absent from the completely demyelinated areas of the spinal cord samples studied (Fig. 2). However, stretches of Caspr immunoreactivity were present on some demyelinated axons in white and grey matter areas with reduced numbers of myelin segments. Two of the six spinal cord samples that still had many myelinated axons in grey matter areas (spinal cord samples from multiple sclerosis Cases 4 and 9) contained many Caspr-positive structures with increased lengths and/or widths, clusters of labelled structures and short internodes (Fig. 2). Due to the large variation in diameters of myelinated axons in white matter areas of the spinal cord, changes in the diameter and/or length of the Caspr-positive structures were less obvious than in brain tissue with demyelination. However, labelled structures on large calibre myelinated axons in affected white matter areas were often more irregular in shape in cross-section than those on control axons and those on axons in unaffected regions; some appeared to be composed of several layers. Caspr-positive structures with long stretches of Caspr immunoreactivity as well as clusters of immunoreactive structures with short myelin segments were also observed in longitudinal sections cut from samples obtained from multiple sclerosis Cases 15 and 25. Additional immunolabellings revealed that atypical Caspr-positive structures were associated with both non-phosphorylated neurofilament-positive and -negative axons (Fig. 2).
|
Discussion |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
The present study has examined in detail the expression of the paranodal glycoprotein Caspr on axons in the healthy mature human CNS and in the brain and spinal cord of subjects who died during the chronic progressive stage of multiple sclerosis. Caspr on myelinated axons in control and normal-appearing tissue surrounding the multiple sclerosis lesions was present in the form of distinct, cylindrical structures flanking the nodes of Ranvier with bundles of neurofilament passing through their centre; peripheral axons were also found to harbour Caspr-positive structures. These findings are consistent with previous studies in rodents indicating that Caspr is highly enriched in the paranodal region of myelinated axons in both the CNS and PNS (Einheber et al., 1997
; Menegoz et al., 1997; Peles et al., 1997b; Rasband and Trimmer, 2001)—a site at which the paranodal myelin loops adhere tightly to the axonal plasma membrane and which forms a physical barrier between the nodal Na+ channels and the juxtaparanodal K+ channels.
The analysis of 36 brain and 16 spinal cord lesions from 29 subjects with chronic progressive multiple sclerosis indicated that the expression of Caspr is down regulated on the surface of chronically demyelinated parts of axons in multiple sclerosis tissue. Because naked nerve fibres with stretches of Caspr were often present in the borders of the multiple sclerosis lesions studied, it is possible that Caspr persists for some time following the loss of myelin. Caspr may be re-expressed or re-directed to the paranodal region during the process of remyelination because short internodes, which are thought to represent newly formed myelin sheaths (Prineas and Connell, 1979; Lassmann, 1983; Prineas et al., 1993), had bands of Caspr immunoreactivity on either ends. This may occur in a manner similar to that taking place in the developing nervous system, i.e. Caspr is expressed initially diffusely along the length of the axon and then becomes restricted to the paranodal region (Einheber et al., 1997; Menegoz et al., 1997; Rasband et al., 1999; Rios et al., 2000). The observation that the new Caspr-positive paranodes were often sharply demarcated suggests that they were proper junctions—an essential requirement for the restoration of saltatory impulse conduction on the affected axons.
The redistribution of Caspr may precede the loss of myelin because myelinated axons with long stretches of Caspr–and thus apparently internodal expression of Caspr–were common in border areas that also contained numerous demyelinated Caspr-positive nerve fibres. If this is indeed the case, then the displacement of Caspr may be an early sign of impending myelin loss. Moreover, it would suggest that even established lesions expand slowly over time (Prineas et al., 2001). Axons with Caspr-positive structures with increased lengths were more widespread in the borders of more chronic lesions than in the borders of active lesions, suggesting that the redistribution of Caspr on axons is not an acute phenomenon. The data further suggest that paranodal junctions on some myelinated axons at the edges of multiple sclerosis lesions may no longer be intact. If this were indeed the case, it would have consequences for the segregation of the Na+ channels at the node and the K+ channels at the juxtaparanode (and thus impulse conduction) and for the attachment of the myelin sheaths to the axon. The increased expression and aberrant localization of Caspr may occur as a consequence of disease-associated changes in axons, or may be a reflection of pathological changes in oligodendrocytes affecting the synthesis and localization of the molecule(s) in the myelin loops that together with Caspr form the septate-like, paranodal junctions (Collinson et al., 1998; Tait et al., 2000; Brophy, 2001; Charles et al., 2002).
Myelinated axons at the lesion edges often had increased diameters, including those with short myelin sheaths, and some of these apparently swollen axons contained non-phosphorylated neurofilaments, suggesting that they were damaged (Trapp et al., 1998). The swelling of some axons may have resulted in an increase in the width of the nodal gap and the subsequent formation of very short internodes and additional paranodes. It is also possible that this occurred because of the detachment of the myelin loops in the paranodal region and/or because of the destruction of specifically the paranodal myelin loops (Blakemore, 1978). Thus, it appears that new myelin segments are not only formed following the loss of complete internodes, but also following widening of the nodal region. Prineas and Connell (1979) found that some of the myelin segments they measured were as short as 11 µm. This raises the possibility that some of the short internodes segments identified by Prineas and Connell had also been formed in response to widening of the nodal region.
The generation of maintenance of the paranodal septate-like junctions is a complex process involving axonal molecules as wells as molecules expressed by myelinating glia. This notion has come from observations that paranodal junctions fail to form properly not only in Caspr null mice (Bhat et al., 2001), but also in mice that lack contactin (Boyle et al., 2001), with which Caspr forms a cis complex (Rios et al., 2000) and which is necessary for the expression of Caspr on the axon surface (Faivre-Sarrailh et al., 2000); unlike Caspr, contactin is also expressed in oligodendrocytes (Einheber et al., 1997; Koch et al., 1997). In addition, paranodal junctions are also frequently disrupted in rodents with genetic defects in oligodendrocytes (Dupree et al., 1999; Rasband et al., 1999; Mathis et al., 2001; Arroyo et al., 2002; Jenkins and Bennett, 2002). The identification of additional molecules (especially those that are involved specifically in the creation and maintenance of the paranodal junctions on CNS axons) and the analysis of their expression in multiple sclerosis tissue and the manipulation of their expression levels in experimental animals, including those with experimental allergic encephalomyelitis [a model for inflammatory CNS demyelination (Lassmann, 1983)], may provide novel insights into the pathogenesis and pathophysiology of multiple sclerosis.
|
Acknowledgements |
|---|
We wish to thank Elior Peles and Sara Piddlesden for their generous gift of antibodies and the team of the Netherlands Brain Bank for collecting the control and multiple sclerosis tissue, for analysing the patients’ medical records and for advice. We also wish to thank Elior Peles and Robert Weissert for their comments on the manuscript. Financial support for this study came from the Netherlands Foundation ‘stichting Vrienden MS Research’.
|
References |
|---|
|
TopSummaryIntroductionMaterial and methodsResultsDiscussionReferences |
|---|
Arroyo EJ, Xu T, Grinspan J, et al. Genetic dysmyelination alters the molecular architecture of the nodal region. J Neurosci 2002; 22: 1726–37.
Bhat MA, Rios JC, Lu Y, et al. Axon-glia interactions and the domain organization of myelinated axons requires neurexin IV/Caspr/Paranodin. Neuron 2001; 30: 369–83.[CrossRef][ISI][Medline]
Bitsch A, Schuchardt J, Bunkowski S, et al. Acute axonal injury in multiple sclerosis. Correlation with demyelination and inflamma tion. Brain 2000; 123: 1174–83.
Blakemore WF. Partial demyelination of cat spinal cord after X-irradiation and surgical interference. Neuropathol Appl Neurobiol 1978; 4: 381–92.[ISI][Medline]
Boyle ME, Berglund EO, Murai KK, et al. Contactin orchestrates assembly of the septate-like junctions at the paranode in myelinated peripheral nerve. Neuron 2001; 30: 385–97.[CrossRef][ISI][Medline]
Brophy PJ. Axoglial junctions: separate the channels or scramble the message. [Review]. Curr Biol 2001; 11: R555–7.
Brück W, Porada P, Poser S, et al. Monocyte/macrophage differentiation in early multiple sclerosis lesions. Ann Neurol 1995; 38: 788–96.[CrossRef][ISI][Medline]
Charles P, Tait S, Faivre-Sarrailh C, et al. Neurofascin is a glial receptor for the paranodin/Caspr-contactin axonal complex at the axoglial junction. Curr Biol 2002; 12: 217–20.[CrossRef][ISI][Medline]
Chiu SY, Ritchie JM. Potassium channels in nodal and internodal axonal membrane of mammalian myelinated fibres. Nature 1980; 284: 170–1.[CrossRef][Medline]
Coleman MP, Perry VH. Axon pathology in neurological disease: a neglected therapeutic target. Trends Neurosci 2002; 25: 532–7.[CrossRef][ISI][Medline]
Collinson JM, Marshall D, Gillespie S, et al. Transient expression of neurofascin by oligodendrocytes at the onset of myelinogenesis: implications for mechanisms of axon-glial interaction. Glia 1998; 23: 11–23.[CrossRef][ISI][Medline]
Compston A, Coles A. Multiple sclerosis. [Review]. Lancet 2002; 359: 1221–31. [CrossRef][ISI][Medline]
Davie CA, Barker GJ, Webb S, et al. Persistent functional deficit in multiple sclerosis and autosomal dominant cerebellar ataxia is associated with axon loss. Brain 1995; 118: 1583–92.
De Stefano N, Matthews PM, Fu L, et al. Axonal damage correlates with disability in patients with relapsing-remitting multiple sclerosis. Results of a longitudinal magnetic resonance spectroscopy study. Brain 1998; 121: 1469–77.
Dupree JL, Girault JA, Popko B. Axo-glial interactions regulate the localization of axonal paranodal proteins. J Cell Biol 1999; 147: 1145–52.
Einheber S, Zanazzi G, Ching W, et al. The axonal membrane protein Caspr, a homologue of neurexin IV, is a component of the septate-like paranodal junctions that assemble during myelination. J Cell Biol 1997; 139: 1495–506.
Faivre-Sarrailh C, Gauthier F, Denisenko-Nehrbass N, et al. The glycosylphosphatidyl inositol–anchored adhesion molecule F3/contactin is required for surface transport of paranodin/contactin-associated protein (caspr). J Cell Biol 2000; 149: 491–502.
Ferguson B, Matyszak MK, Esiri MM, et al. Axonal damage in acute multiple sclerosis. Brain 1997; 120: 393–9.
Hirano A, Llena JF. Morphology of central nervous system axons. In: Waxman S, Kocsis J, Stys P, editors. The axon. New York: Oxford University Press; 1995. p. 49–67.
Jenkins SM, Bennett V. Developing nodes of Ranvier are defined by ankyrin-G clustering and are independent of paranodal axoglial adhesion. Proc Natl Acad Sci USA 2002; 99: 2303–8.
Koch T, Brugger T, Bach A, Gennarini G, et al. Expression of the immunoglobulin superfamily cell adhesion molecule F3 by oligodendrocyte-lineage cells. Glia 1997;19: 199–212.
Kornek B, Storch MK, Weisert R, et al. Multiple sclerosis and chronic autoimmune encephalomyelitis: a comparative quantitative study of axonal injury in active, inactive, and remyelinated lesions. Am J Pathol 2000; 157: 267–76.
Kuhlmann T, Lingfeld G, Bitsch A, et al. Acute axonal damage in multiple sclerosis is most extensive in early disease stages and decreases over time. Brain 2002; 125: 2202–12.
Lassmann H. Comparative neuropathology of chronic experimental allergic encephalomyelitis and multiple sclerosis. Schriftner Neurol 1983; 25: 1–135.
Lassmann H. Pathology of multiple sclerosis. In: Compston A, Ebers G, Lassmann H, McDonald WI, Matthews B, Wekerle H, editors. McAlpine’s multiple sclerosis. 3rd ed. London: Churchill Livingstone; 1998. p. 323–58.
Lucchinetti C, Brück W, Parisi J, et al. Heterogeneity of multiple sclerosis lesions: implications for the pathogenesis of demyelination. Ann Neurol 2000; 47: 707–17.[CrossRef][ISI][Medline]
Mathis C, Denisenko-Nehrbass N, Girault J-A, et al. Essential role of oligodendrocytes in the formation and maintenance of central nervous system nodal regions. Development 2001; 128: 4881–90.
Menegoz M, Gaspar P, Le Bert M, et al. Paranodin, a glycoprotein of neuronal paranodal membranes. Neuron 1997; 19: 319–31.[CrossRef][ISI][Medline]
Mi H, Deerinck TJ, Ellisman MH, et al. Differential distribution of closely related potassium channels in rat Schwann cells. J Neurosci 1995; 15: 3761–74.[Abstract]
Peles E, Salzer JL. Molecular domains of myelinated axons. [Review]. Curr Opin Neurobiol 2000; 10: 558–65.[CrossRef][ISI][Medline]
Peles E, Joho K, Plowman GD, et al. Close similarity between Drosophila neurexin IV and mammalian Caspr protein suggests a conserved mechanism for cellular interactions. Cell 1997a; 88: 745–6.[CrossRef][ISI][Medline]
Peles E, Nativ M, Lustig M, et al. Identification of a novel contactin- associated transmembrane receptor with multiple domains implicated in protein-protein interactions. EMBO J 1997b; 16: 978–88.[CrossRef][ISI][Medline]
Piddlesden SJ, Lassmann H, Zimprich F, et al. The demyelinating potential of antibodies to myelin oligodendrocyte glycoprotein is related to their ability to fix complement. Am J Pathol 1993; 143: 555–64.[Abstract]
Poliak S, Gollan L, Martinez M, et al. Caspr2, a new member of the neurexin superfamily, is localized at the juxtaparanodes of myelinated axons and associates with K+ channels. Neuron 1999; 24: 1037–47.[CrossRef][ISI][Medline]
Prineas JW, Connell F. Remyelination in multiple sclerosis. Ann Neurol 1979; 5: 22–31.[CrossRef][ISI][Medline]
Prineas JW, Barnard RO, Kwon EE, et al. Multiple sclerosis: remyelination of nascent lesions. Ann Neurol 1993; 33: 137–51.[CrossRef][ISI][Medline]
Prineas JW, Kwon EE, Cho E-S, et al. Immunopathology of secondary-progressive multiple sclerosis. Ann Neurol 2001; 50: 646–57.[CrossRef][ISI][Medline]
Prineas JW, McDonald WI, Franklin RJM. Demyelinating diseases. In: Graham DI, Lantos PL, editors. Greenfield’s neuropathology. 7th ed. London: Arnold; 2002. p. 471–550.
Rasband MN, Trimmer JS. Subunit composition and novel localization of K+ channels in spinal cord. J Comp Neurol 2001; 429: 166–76.[CrossRef][ISI][Medline]
Rasband MN, Peles E, Trimmer JS, et al. Dependence of nodal sodium channel clustering on paranodal axoglial contact in the developing CNS. J Neurosci 1999; 19: 7516–28.
Rios JC, Melendez-Vasquez CV, Einheber S, et al. Contactin-associated protein (Caspr) and contactin form a complex that is targeted to the paranodal junctions during myelination. J Neurosci 2000; 20: 8354–64.
Salzer JL. Clustering sodium channels at the node of Ranvier: close encounters of the axon-glia kind. [Review]. Neuron 1997; 18: 843–6. [CrossRef][ISI][Medline]
Scherer SS, Arroyo EJ. Recent progress on the molecular organization of myelinated axons. [Review]. J Peripher Nerv Syst 2002; 7: 1–12. [Medline]
Tait S, Gunn-Moore F, Collinson JM, et al. An oligodendrocyte cell adhesion molecule at the site of assembly of the paranodal axo-glial junction. J Cell Biol 2000; 150: 657–66.
Trapp BD, Peterson J, Ransohoff RM, et al. Axonal transection in the lesions of multiple sclerosis. New Engl J Med 1998; 338: 278–85.
Wang H, Kunkel DD, Martin TM, et al. Heteromultimeric K+ channels in terminal and juxtaparanodal regions of neurons. Nature 1993; 365: 75–9.[CrossRef][Medline]
Waxman SG, Ritchie JM. Molecular dissection of the myelinated axon. Ann Neurol 1993; 33: 121–36.[CrossRef][ISI][Medline]
Wingerchuk DM, Lucchinetti CF, Noseworthy JH. Multiple sclerosis: current pathophysiological concepts. [Review]. Lab Invest 2001; 81: 263–81.[ISI][Medline]
Wolswijk G. Chronic stage multiple sclerosis lesions contain a relatively quiescent population of oligodendrocyte precursor cells. J Neurosci 1998; 18: 601–9.
Wolswijk G. Oligodendrocyte survival, loss and birth in lesions of chronic-stage multiple sclerosis. Brain 2000; 123: 105–15.
Wolswijk G. Oligodendrocyte precursor cells in the demyelinated multiple sclerosis spinal cord. Brain 2002; 125: 338–49.
Wood JN, Anderton BH. Monoclonal antibodies to mammalian neurofilaments. Biosci Rep 1981; 1: 263–8.[CrossRef][ISI][Medline]
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  K. Susuki and M. N. Rasband Spectrin and Ankyrin-Based Cytoskeletons at Polarized Domains in Myelinated Axons Experimental Biology and Medicine, April 1, 2008; 233(4): 394 - 400. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. K. Mathey, T. Derfuss, M. K. Storch, K. R. Williams, K. Hales, D. R. Woolley, A. Al-Hayani, S. N. Davies, M. N. Rasband, T. Olsson, et al. Neurofascin as a novel target for autoantibody-mediated axonal injury J. Exp. Med., October 1, 2007; 204(10): 2363 - 2372. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 I. Coman, M. S. Aigrot, D. Seilhean, R. Reynolds, J. A. Girault, B. Zalc, and C. Lubetzki Nodal, paranodal and juxtaparanodal axonal proteins during demyelination and remyelination in multiple sclerosis Brain, December 1, 2006; 129(12): 3186 - 3195. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. W. Howell, A. Palser, A. Polito, S. Melrose, B. Zonta, C. Scheiermann, A. J. Vora, P. J. Brophy, and R. Reynolds Disruption of neurofascin localization reveals early changes preceding demyelination and remyelination in multiple sclerosis Brain, December 1, 2006; 129(12): 3173 - 3185. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Simons and K. Trajkovic Neuron-glia communication in the control of oligodendrocyte function and myelin biogenesis J. Cell Sci., November 1, 2006; 119(21): 4381 - 4389. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. P. Garcia-Fresco, A. D. Sousa, A. M. Pillai, S. S. Moy, J. N. Crawley, L. Tessarollo, J. L. Dupree, and M. A. Bhat Disruption of axo-glial junctions causes cytoskeletal disorganization and degeneration of Purkinje neuron axons PNAS, March 28, 2006; 103(13): 5137 - 5142. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||