Increased expression of distinct galectins in multiple sclerosis lesions

M. Stancic, J. van Horssen, V. L. Thijssen, H.‐J. Gabius, P. van der Valk, D. Hoekstra and W. Baron (2011) Neuropathology and Applied Neurobiology 37, 654–671 Increased expression of distinct galectins in multiple sclerosis lesions Aims: Multiple sclerosis (MS) is a chronic progressive degenerative disorder of the central nervous system, characterized by inflammation, demyelination, ultimate failure of remyelination and axonal loss. Current research identifies galectins, adhesion/growth‐regulatory effectors binding β‐galactosides, peptide motifs and lipids, as important immunomodulators in diverse inflammatory diseases. However, little is known about their expression, cellular localization and role in human central nervous system tissue. To identify a potential role of galectins in MS, their expression and localization in control white matter (CWM) and demyelinated MS lesions were examined. Methods: qPCR, Western blot and immunohistochemical analyses were performed on human post mortem CWM and MS lesions at different stages. Cultured astrocytes, derived from healthy subjects and MS patients, were analysed similarly. Results: Among 11 different galectins tested, galectins‐1, ‐3, ‐8 and ‐9 were present at detectable levels in CWM, and, interestingly, significantly enhanced in active MS lesions. On the cellular level, galectins localized to microglia/macrophages, astrocytes and endothelial cells. Intriguingly, galectin‐9 displayed a distinctly different intracellular localization in microglia/macrophages when comparing active and inactive MS lesions, being restricted to the nuclei in active lesions, and primarily localizing in the cytoplasm in inactive lesions. Furthermore, enhanced levels of galectin‐1, detected as dimers in Western blot analysis, were released by cultured astrocytes from MS patients. Conclusions: This study provides a detailed analysis of galectins in MS lesions and assigns distinct galectins to different aspects of the disease. Thus, besides being known as modulators of inflammatory processes, our findings suggest additional association of distinct galectins with MS pathology.     

Matrix metalloproteinase-19 is highly expressed in active multiple sclerosis lesions

Matrix metalloproteinases (MMPs) are proteases known for their capacity to degrade extracellular matrix (ECM) components. MMPs have been implicated in several central nervous system (CNS) diseases, including multiple sclerosis (MS). Microarray analysis has demonstrated significant increased mRNA levels of MMP-19 in chronic MS lesions, suggesting a role of MMP-19 in MS pathogenesis. Therefore, in this study, we investigated the expression pattern and cellular localization of MMP-19 protein in various well-characterized MS lesion stages. In normal control patient white matter, MMP-19 was constitutively expressed by microglia throughout the brain parenchyma, suggesting a physiological role for this MMP family member. Likewise, MMP-19 was expressed by microglia in (p)reactive MS lesions, albeit more intense. In highly active demyelinating MS lesions, parenchymal and perivascular myelin-laden macrophages were strongly immunoreactive for MMP-19, whereas reactive astrocytes were occasionally immunopositive. Astrocytes in chronic inactive lesions were weakly stained for MMP-19. In vitro, MMP-19 was expressed in cultures of primary human microglia, not in astrocyte cultures. As MMP-19 is able to degrade basement membrane constituents and other ECM proteins, it is conceivable that this relatively novel MMP family member contributes to MS pathology by remodelling the ECM of the CNS, thereby influencing leucocyte infiltration, axonal regeneration and astrogliosis.     

TROY and LINGO-1 expression in astrocytes and macrophages/microglia in multiple sclerosis lesions

Nogo constitutes a family of neurite outgrowth inhibitors contributing to a failure of axonal regeneration in the adult central nervous system (CNS). Nogo-A is expressed exclusively on oligodendrocytes where Nogo-66 segment binds to Nogo receptor (NgR) expressed on neuronal axons. NgR signalling requires a coreceptor p75(NTR) or TROY in combination with an adaptor LINGO-1. To characterize the cell types expressing the NgR complex in the human CNS, we studied demyelinating lesions of multiple sclerosis (MS) brains by immunohistochemistry. TROY and LINGO-1 were identified in subpopulations of reactive astrocytes, macrophages/microglia and neurones but not in oligodendrocytes. TROY was up-regulated, whereas LINGO-1 was reduced in MS brains by Western blot. These results suggest that the ternary complex of NgR/TROY/LINGO-1 expressed on astrocytes, macrophages/microglia and neurones, by interacting with Nogo-A on oligodendrocytes, might modulate glial-neuronal interactions in demyelinating lesions of MS.     

Cytokines decrease expression of interleukin‐6 signal transducer and leptin receptor in central nervous system glia

Multiple sclerosis (MS) lesion formation is modulated by cytokines secreted within the central nervous system (CNS). Th1 lymphocytes and monocyte/macrophages (MM) likely induce lesion formation, whereas Th2 lymphocytes may inhibit formation. To explore the role of cytokines in MS lesions, we used gene arrays to investigate effects of cytokines representative of Th1 and Th2 cells and M/M on gene expression in cultured CNS glia; at 6 hr, all three increased expression of the interleukin‐6 (IL‐6) gene and decreased expression of the leptin receptor gene (obr), which mediates IL‐6 production and other inflammatory responses. However, expression of a closely related gene, the interleukin‐6 signal transducer or gp130 (IL‐6st), showed no changes at 6 hr. IL‐6st is an essential component of receptor complexes for IL‐6 and other cytokines and growth factors that play critical roles in CNS inflammation, protection, and/or regeneration. To analyze expression of IL‐6st and leptin receptor over time, we incubated rat CNS glial cultures for 6 hr to 5 days with the cytokines. All three cytokine mixtures down‐regulated both IL‐6st and leptin receptor mRNA and protein for up to 5 days. Immunocytochemical staining showed expression of both IL‐6st and leptin receptor in all three types of glia, with lower IL‐6st expression by 3 days. Down‐regulation of IL‐6st and leptin receptor in glia by cytokines could lead to decreased signaling by the proinflammatory IL‐6 and reduced responses to regenerative/protective growth factors such as leukemia inhibitory factor and ciliary neurotrophic factor, potentially affecting the disease course in MS. © 2009 Wiley‐Liss, Inc.     

Enhanced expression of MMP-7 and MMP-9 in demyelinating multiple sclerosis lesions

The pathology of multiple sclerosis (MS) is characterised by breakdown of the blood-brain barrier accompanied by infiltration of macrophages and T cells into the central nervous system (CNS). Myelin is degraded and engulfed by the macrophages, producing lesions of demyelination. Some or all of these mechanisms might involve proteinases, and here we have studied the cellular localisation and distribution of two matrix metalloproteinases (MMPs), MMP-7 (matrilysin) and MMP-9 (92-kDa gelatinase), in the normal human CNS and active demyelinating MS lesions. Cryostat sections of CNS samples were immunostained with antisera to MMP-7 and MMP-9. In addition, non-radioactive in situ hybridisation (ISH) was performed using a digoxygenin-labelled riboprobe to detect the expression of MMP-7. MMP-7 immunoreactivity was weakly detected in microglial-like cells in normal brain tissue sections, and was very strong in parenchymal macrophages in active demyelinating MS lesions. This pattern of expression was confirmed using ISH. MMP-7 immunoreactivity was not detected in macrophages in spleen or tonsil, indicating that it is specifically induced in infiltrating macrophages in active demyelinating MS lesions. MMP-9 immunoreactivity was detected in a few small blood vessels in normal brain tissue sections, whereas many blood vessels stained positive in CNS tissue sections of active demyelinating MS lesions. The up-regulation of MMPs in MS may contribute to the pathology of the disease. Received: 19 December 1996 / Revised: 22 May 1997 / Accepted: 4 June 1997     

Heterogeneity of cortical lesions in multiple sclerosis: an MRI perfusion study

In this study, dynamic susceptibility contrast-magnetic resonance imaging (DSC-MRI) was used to quantify the cerebral blood flow (CBF), the cerebral blood volume (CBV), and the mean transit time (MTT) and to analyze the changes in cerebral perfusion associated with the cortical lesions in 44 patients with relapsing-remitting multiple sclerosis. The cortical lesions showed a statistically significant reduction in CBF and CBV compared with the normal-appearing gray matter, whereas there were no significant changes in the MTT. The reduced perfusion suggests a reduction of metabolism because of the loss of cortical neurons. A small population of outliers showing an increased CBF and/or CBV has also been detected. The presence of hyperperfused outliers may imply that perfusion could evolve during inflammation. These findings show that perfusion is altered in cortical lesions and that DSC-MRI can be a useful tool to investigate more deeply the evolution of cortical lesions in multiple sclerosis.     

The relative number of macrophages/microglia expressing macrophage colony-stimulating factor and its receptor decreases in multiple sclerosis lesions

The activation of macrophages/microglia in multiple sclerosis (MS) lesions plays a central role in the effector phase of myelin breakdown. The precise patterns of macrophage/microglia activation during demyelination have not yet been defined. The growth and activating factor macrophage-colony stimulating factor (M-CSF) and its specific receptor (M-CSFR) may be involved in this process. The present study investigated the expression of M-CSF and M-CSFR mRNA by in situ hybridization in 60 lesions from 32 MS patients. In the control and periplaque white matter, microglia was almost completely M-CSFR positive. Irrespective of the demyelinating activity, an increased number of cells expressed M-CSF or M-CSFR mRNA within the lesions. However, despite the tremendous increase in macrophages/microglia within the lesions, the relative number of these cells expressing M-CSF or M-CSFR decreased. There was no correlation of M-CSF or M-CSFR expression with active myelin breakdown. The correlation between the clinical course and the expression of M-CSF or M-CSFR mRNA revealed significant differences with the lowest expression in primary progressive MS. These results suggest a downregulation of M-CSF and M-CSFR inside the MS plaque probably due to the high amount of macrophage-derived cytokines or mediators. Nevertheless, the differences in the relative number of cells expressing the M-CSF/M-CSFR pathway implicate that this pathway may be an important contributory factor in different forms of MS pathology.     

Acute and chronic pain syndromes in multiple sclerosis

A representative sample of 117 patients with definite multiple sclerosis (MS) was interviewed on pain syndromes. Chronic syndromes lasting more than one month included dysaestesthesia, low back pain, spasms, tonic seizures, tightening and painful sensations in the extremities. Acute syndromes included neuralgia, L'Hermitte's sign and pain associated with optic neuritis. Thirty-five per cent were pain-free. Of the remaining patients had 45% pain at the time of the examination, 32% indicated pain among the most severe symptoms of MS and 23% had pain at the onset of MS. The number of patients with pain at the time of the examination increased with age and duration of disease. Patients with pain were significantly more often spastic and significantly more often sought alternative treatment forms. No difference was found for mean age, sex, physical impairment, duration of disease from onset of MS, depressive score and score of delayed verbal memory.     

Insulin-like growth factors and binding proteins in multiple sclerosis plaques

Insulin-like growth factors (IGFs) play an important role in development and myelination in the central nervous system (CNS) as well as in the proliferation and differentiation of cells of the immune system. To assess the influence of this growth factor family on demyelination and repair in multiple sclerosis (MS), the expression of IGF-I, IGF-II, insulin, IGF binding proteins (IGFBP) 1-3 and IGF-I receptor (IGF-IR) in CNS tissue from MS and normal control cases was studied by immunocytochemistry. In active MS lesions, the expression of IGF-I, insulin and IGFBP1 was detected in hypertrophic astrocytes while that of IGF-II and IGFBP2 and 3 was confined to foamy macrophages within lesions and activated microglia in adjacent white matter. IGF-IR, the major IGF receptor, was immunolocalized in macrophages and an astrocyte subpopulation in plaques. Oligodendrocytes in normal-appearing white matter expressed only IGFBP1, not IGFs or IGF-IR. As the remyelinating capacity of oligodendrocytes could be impaired owing to the absence of IGF-IR, the prevailing role of IGFs in inflammatory demyelination may be to promote phagocytosis of myelin and astrogliosis.     

MR peri‐CSF lesions and CSF oligoclonal bands in Italian multiple sclerosis patients

Objectives – We investigated the association between brain lesion distribution and the presence of oligoclonal IgG bands (OCBs) in Italian multiple sclerosis (MS) patients. Materials and methods – We retrospectively selected brain magnetic resonance imaging (MRI) uniformly performed in 56 relapsing patients (41 patients OCB positive). Results – Brain lesions in periventricular areas occurred in 92.86% of the patients (100% OCB+ and 73.33% OCB?) (P = 0.004), but we did not find a significant difference for their median volume (P = 0.553) and median number (P = 0.606) between the two groups. Parenchymal lesions occurred in 76.8% of the patients with a similar distribution (P = 1.00) and no significant difference in the median volume (P = 0.818) and number (P = 0.643) between the two groups. Conclusions – The present study on cohort of Italian MS patients demonstrated a lack of correlation between lesion distribution and OCBs, suggesting that B cells producing them could be localized both in meningeal niches and cerebral parenchyma.     

PECAM-1 and gelatinase B coexist in vascular cuffs of multiple sclerosis lesions

In multiple sclerosis (MS), the matrix metalloprotease (MMP) gelatinase B/MMP-9 and platelet endothelial cell adhesion molecule (PECAM)-1 have both been implicated in trans-endothelial infiltration of leucocytes into the brain, but their functional connection has not yet been investigated. We investigated the expression of gelatinase B and PECAM-1 in post mortem brains of MS patients by immunohistochemistry. Because increased soluble PECAM-1 serum levels have been observed in MS patients, we also tested in vitro whether this could be due to cleavage of PECAM-1 by gelatinase B or matrilysin-1/MMP-7. Constitutive expression of PECAM-1 was found on brain endothelial cells, whilst in active MS lesions cell-bound PECAM-1 was highly up-regulated on foamy macrophages in perivascular infiltrates and co-localized with gelatinase B. However, human THP-1 monocyte-bound or soluble recombinant PECAM-1 were both resistant to proteolytic cleavage by gelatinase B or matrilysin-1 in vitro, as demonstrated by Western blot analysis and flow cytometry. These results suggest that PECAM-1 and gelatinase B may complement each other during the transmigration of the blood-brain barrier by mononuclear cells.     

Pseudo‐quadriceps sparing in multiple sclerosis

Background: Sparing of the quadriceps muscle has been reported in various myopathies. In multiple sclerosis (MS) and pyramidal syndromes, in general, such a differential involvement of distinct muscle groups has not been described. Methods: Muscle power was evaluated in 127 patients with chronic pyramidal syndrome caused by MS and 37 patients with acute or chronic paraparesis from other etiologies (mainly cerebro‐vascular events). Results: A striking difference in muscle power of the quadriceps (spared) and the iliopsoas (significantly weakened) was found in the patients suffering from chronic pyramidal syndrome caused by MS. The mean muscle power of the iliopsoas was 1.68 ± 1.1 and that of the quadriceps 4.06 ± 1.4 (P < 0.00005). In the control group, the mean muscle power was 2 ± 1.2 and 2.4 ± 1.4 (difference not significant), for the iliopsoas and the quadriceps, respectively. Conclusions: Quadriceps muscle remains relatively spared in patients with MS, even with severe and long‐standing paraparesis. Various neuroanatomical, neurophysiological, and rehabilitational mechanisms may be involved and explain this phenomenon. This observation may contribute to the building of more reliable and linear scales for the assessment of motor disability and disease progression in MS.     

Comparison of patient‐reported outcome measures in multiple sclerosis

Background

Patient‐reported outcome measurements (PROMS) have been proposed sensitive outcome parameters in multiple sclerosis (MS). In this study, we assessed a German version of the Multiple Sclerosis Impact Scale (MSIS‐29) and a revised version of the Hamburg Quality of Life Questionnaire in Multiple Sclerosis (HAQUAMS) in comparison with rater‐ and physician‐based tools.

Methods

Consecutive MS patients (n = 117) of the MS outpatient unit were included. In addition to MSIS‐29 and HAQUAMS, the following parameters were obtained: Expanded Disability Status Scale (EDSS) and modified Multiple Sclerosis Functional Composite (MSFC) [9‐hole peg test (9HPT), 25‐foot walk test and symbol digit modalities test]. We investigated validity, internal consistency and test–retest reliability as well as correlation between these measures.

Results

Internal consistency (Cronbach's α ≤ 0.96) and test–retest coefficients (ICC ≤ 0.87) of both scales were high and satisfied psychometric standards. Convergent and discriminant validity was supported by direction, magnitude and pattern of correlation with other rater‐based measures depending on the functional subdomain. Both MSIS‐29 and HAQUAMS correlated with EDSS (ρ = 0.55 vs 0.62), but stronger correlation was found between MSIS‐29 and HAQUAMS total score (ρ = 0.90). Both scales distinguished between patient groups of varied disease severity and cognitive impairment.

Conclusion

Patient‐reported outcome measurements as MSIS‐29 and HAQUAMS seem to be valid instruments to detect different impairment levels in comparison with traditional rater‐based instruments like EDSS or MSFC.     

Sphingosine 1‐phosphate receptor 1 and 3 are upregulated in multiple sclerosis lesions

Sphingolipids are a class of biologically active lipids that have a role in multiple biological processes including inflammation. Sphingolipids exert their functions by direct signaling or through signaling by their specific receptors. Phosphorylated FTY720 (FTY720P) is a sphingosine 1‐phosphate (S1P) analogue that is currently in trial for treatment of multiple sclerosis (MS), which targets all S1P receptors but S1P₂. To date, however, it remains unknown whether FTY720P may exert direct anti‐inflammatory effects within the central nervous system (CNS), because data concerning S1P receptor expression and regulation under pathological conditions in the human brain are lacking. To investigate potential regulation of S1P receptors in the human brain during MS, we performed immunohistochemical analysis of S1P receptor 1 and 3 expression in well‐characterized MS lesions. A strong increase in S1P receptor 1 and 3 expression on reactive astrocytes was detected in active and chronic inactive MS lesions. In addition, we treated primary cultures of human astrocytes with the proinflammatory cytokine tumor necrosis factor‐alpha to identify the regulation of S1P_1/3 on astrocytes under pathological conditions. Importantly, we demonstrate that FTY720P exerts an anti‐inflammatory action on human astrocytes by limiting secretion of proinflammatory cytokines. Our data demonstrate that reactive astrocytes in MS lesions and cultured under proinflammatory conditions strongly enhance expression of S1P receptors 1 and 3. Results from this study indicate that astrocytes may act as a yet‐unknown target within the CNS for the anti‐inflammatory effects observed after FTY720P administration in the treatment of MS. © 2010 Wiley‐Liss, Inc.     

Non‐adherence to interferon‐beta therapy in Swedish patients with multiple sclerosis

Cunningham A, Gottberg K, von Koch L, Hillert J. Non‐adherence to interferon‐beta therapy in Swedish patients with multiple sclerosis.
Acta Neurol Scand: 2010: 121: 154–160.
© 2009 The Authors Journal compilation © 2009 Blackwell Munksgaard. Objectives – To explore the occurrence and reasons for stopping, switching or continuing first prescribed interferon‐beta therapy in patients with multiple sclerosis in Sweden, with respect to demographic, clinical and/or therapy‐related factors. Materials and methods – A retrospective study reviewing the medical charts of 259 patients with multiple sclerosis, comparing patients continuing therapy for at least 3 years with those switching or stopping therapy. Results – Sixty 9% stopped (15%), or switched (54%), interferon‐beta therapy within 3 years. Stoppers had longer disease duration before starting therapy (P = 0.002), less frequently relapsing‐remitting multiple sclerosis (P = 0.046), and more often Expanded Disability Status Scale scores 6–9.5 (P = 0.045) compared to Switchers. The most common reasons for switching/stopping therapy were perceived lack of effect and side‐effects. Conclusions – Adherence to initial immune‐modulating therapy is low; identification of patients at higher risk of stopping therapy and provision of adequate support are essential.