Elisabetta Capello

Demyelination, Inflammation, and Neurodegeneration in Multiple Sclerosis Deep Gray Matter

Gray matter (GM) lesions are recognized as important components of the pathology of multiple sclerosis (MS), and involvement of the deep gray matter (DGM) is suggested by magnetic resonance imaging. The aims of this study were to determine the frequency and distribution of lesions and characterize the inflammatory and neurodegenerative changes in DGM of MS patients. Histochemistry, immunohistochemistry, and morphometry were performed on whole coronal sections of 14 MS and 12 control (6 normal, 6 from amyotrophic lateral sclerosis patients) brains. Demyelinating lesions were frequent in MS DGM; most often in the thalamus and caudate, but they were also seen in the putamen, pallidum, claustrum, amygdala, hypothalamus, and substantia nigra. Most DGM lesions involved both GM and white matter. Inflammation in active DGM lesions was similar to that in lesions only in white matter but was less intense, and there was a preponderance of activated microglia, scarce myelin-laden macrophages, and a lesser extent of axonal damage. Neuronal loss was observed both in DGM lesions and nondemyelinated DGM with neuron atrophy in nondemyelinated DGM. In conclusion, demyelination and neurodegenerative changes are common in MS DGM and may contribute to clinical impairment. Inflammation in DGM lesions is intermediate between the destructive inflammation of white matter lesions and the minimal inflammation of cortical lesions. We hypothesize that alterations of glutamate reuptake mechanisms may contribute to these differences.

Phenotypic and functional analysis of T cells homing into the CSF of subjects with inflammatory diseases of the CNS.

The recruitment of lymphocytes across the blood brain barrier (BBB) is mediated by adhesion molecules and chemokines. The expression of activation markers and of chemokine receptors on T cells homing to the nervous system (NS) may help define their functional state. In the cerebrospinal fluid (CSF) of subjects with inflammatory neurological diseases (IND), including multiple sclerosis, we observed an increased number of T cells coexpressing CXCR3 and CCR5 as well as T cells with a CD45RO+ CCR7+ CD27+ memory phenotype. A subset of CCR7+ T cells coexpressed CXCR3 and CCR5. We also detected an increased number of interferon‐γ‐producing T cells in the CSF compared with peripheral blood, mostly but not exclusively in the CD45RO+ CCR7− CD27− compartment. T helper 1 (Th1) clones, established from the CSF of individuals with IND and from a healthy subject, similarly migrated to CXCL10, CXCL12, and CCL5. CXCL10, CXCL12, and CCL19 were increased in the CSF of individuals with neuroinflammation. These findings suggest that CSF is enriched in Th1‐polarized memory T cells capable of differentiating into effector cells upon antigen encounter. These cells are recruited into the CSF by inducible chemokines. Thus, CSF represents a transitional station for T cells trafficking to and from the NS.

Myelin/Oligodendrocyte Glycoprotein-Induced Autoimmune Encephalomyelitis in Common Marmosets: The Encephalitogenic T Cell Epitope pMOG24–36 Is Presented by a Monomorphic MHC Class II Molecule

Immunization of common marmosets (Callithrix jacchus) with a single dose of human myelin in CFA, without administration of Bordetella pertussis, induces a form of autoimmune encephalomyelitis (EAE) resembling in its clinical and pathological expression multiple sclerosis in humans. The EAE incidence in our outbred marmoset colony is 100%. This study was undertaken to assess the genetic and immunological basis of the high EAE susceptibility. To this end, we determined the separate contributions of immune reactions to myelin/oligodendrocyte glycoprotein (MOG) and myelin basic protein to the EAE induction. Essentially all pathological features of myelin-induced EAE were also found in animals immunized with MOG in CFA, whereas in animals immunized with myelin basic protein in CFA clinical and pathological signs of EAE were lacking. The epitope recognition by anti-MOG Abs and T cells were assessed. Evidence is provided that the initiation of EAE is based on T and B cell activation by the encephalitogenic phMOG14–36 peptide in the context of monomorphic Caja-DRB*W1201 molecules.

Altered glutamate reuptake in relapsing-remitting and secondary progressive multiple sclerosis cortex: correlation with microglia infiltration, demyelination, and neuronal and synaptic damage.

Cortical involvement in multiple sclerosis (MS) is emerging as an important determinant of disease progression. The mechanisms responsible for MS cortical pathology are not fully characterized. The objective of this study was to assess the role of excitotoxicity in MS cortex, evaluating excitatory amino acid transporter (EAAT) expression and its relationship with demyelination, inflammation, gliosis, and neuronal and synaptic pathology. EAATs are essential in maintaining low extracellular glutamate concentrations and preventing excitotoxicity. Ten MS brains (3 relapsing-remitting MS cases and 7 secondary progressive MS cases) were evaluated by immunohistochemistry for myelin basic protein, CD68, HLA-DR, EAAT1, EAAT2, glial fibrillary acidic protein, phosphorylated c-Jun N-terminal kinase (pJNK), synaptophysin, and neurofilaments. Cortical lesions were frequently observed in MS brains in variable numbers and extensions. In cortical lesions, activated microglia infiltration correlated with focal loss of EAAT1, EAAT2, and synaptophysin immunostaining, and with neuronal immunostaining for pJNK, a protein involved in response to excitotoxic injury. No reduction of EAATs or synaptophysin immunostaining was observed in demyelinated cortex in the absence of activated microglia. Alterations of the mechanisms of glutamate reuptake are found in cortical MS lesions in the presence of activated microglia and are associated with signs of neuronal and synaptic damage suggestive of excitotoxicity. Excitotoxicity may be involved in the pathogenesis of demyelination and of neuronal and synaptic damage in MS cortex.

Autologous stem cell transplantation as rescue therapy in malignant forms of multiple sclerosis

Malignant forms of multiple sclerosis (MS) represent a limited group of very aggressive demyelinating diseases, which rapidly progress to severe disability leading often to life-threatening conditions. On these clinical entities, currently available therapies for MS are not very effective. Recently, it has been demonstrated that intense immunosuppression followed by autologous stem cell transplantation (ASCT) can affect the clinical course of individuals with severe MS and completely abrogate the inflammatory activity detected by magnetic resonance imaging (MRI). We report on the treatment with intense immune ablation followed by ASCT of three patients with malignant MS whose clinical course indicated a dramatically poor prognosis. This procedure succeeded in halting the rapidly worsening course of disease. The effect was long lasting, as demonstrated by a sustained efficacy over a two-year period in two subjects and 12 months in the third case. In addition, a striking effect on inflammation-related MRI findings was obtained. These results support a role for intense immunosuppression followed by ASCT as treatment in rapidly evolving malignant MS cases unresponsive to conventional therapies.

Demyelination and axonal damage in a non-human primate model of multiple sclerosis

The demyelinating plaque is the paradigmatic lesion of multiple sclerosis (MS), but only recently attention has been given to axonal damage and to its role in the pathophysiology of disease. Albeit the possible relevance of axonal loss in MS and its experimental models, the amount and timing of axonal sufferance has been addressed only in experimental autoimmune encephalomyelitis (EAE) of rodents. In this report we observed that, in the marmoset model of EAE, axonal damage occurs early during the demyelinating process as assessed by immunoreactivity for amyloid precursor protein (APP) and non-phosphorylated neurofilaments (SMI-32 positive) detected mostly in early active lesions compared to late active and normal appearing white matter. The rare occurrence of morphological features of axonal transection, such as APP or SMI-32 positive spheroids and swellings, as well as an increase of neurofilament density in the demyelinated axons without accumulation of electron dense organelles or osmiophilic bodies, at electron microscopy, suggests that early axonal damage may be, at least in part, a reversible process. These findings are of relevance for the development of therapies, which can protect axons and enhance their function and survival.

Autologous haematopoietic stem cell transplantation with an intermediate intensity conditioning regimen in multiple sclerosis: the Italian multi-centre experience

Background: Over recent years numerous patients with severe forms of multiple sclerosis (MS) refractory to conventional therapies have been treated with intense immunosuppression followed by autologous haematopoietic stem cell transplantation (AHSCT). The clinical outcome and the toxicity of AHSCT can be diverse, depending on the various types of conditioning protocols and on the disease phase. Objectives: To report the Italian experience on all the consecutive patients with MS treated with AHSCT with an intermediate intensity conditioning regimen, named BEAM/ATG, in the period from 1996 to 2008. Methods: Clinical and magnetic resonance imaging outcomes of 74 patients were collected after a median follow-up period of 48.3 (range = 0.8–126) months. Results: Two patients (2.7%) died from transplant-related causes. After 5 years, 66% of patients remained stable or improved. Among patients with a follow-up longer than 1 year, eight out of 25 subjects with a relapsing–remitting course (31%) had a 6–12 months confirmed Expanded Disability Status Scale improvement > 1 point after AHSCT as compared with one out of 36 (3%) patients with a secondary progressive disease course (p = 0.009). Among the 18 cases with a follow-up longer than 7 years, eight (44%) remained stable or had a sustained improvement while 10 (56%), after an initial period of stabilization or improvement with median duration of 3.5 years, showed a slow disability progression. Conclusions: This study shows that AHSCT with a BEAM/ATG conditioning regimen has a sustained effect in suppressing disease progression in aggressive MS cases unresponsive to conventional therapies. It can also cause a sustained clinical improvement, especially if treated subjects are still in the relapsing–remitting phase of the disease.

Marburg type and Balò's concentric sclerosis: rare and acute variants of multiple sclerosis

Abstract.Marburg type and Balò’s concentric sclerosis are considered particular variants of multiple sclerosis, clinically characterized by a severe, rapidly evolving clinical course, and by unusual neuropathological changes, often large tumor-like demyelinating plaques in Marburg type and concentric layers of partial demyelination alternating with demyelinating bands in Balò’s concentric sclerosis. These forms now can be diagnosed in vivo, with complete clinical and laboratory examination and MRI follow up. Although in some patients a favourable clinical course has been described, the poor prognosis of the majority of these cases justifies a treatment with intense immunosuppression.

The 14-3-3 protein in multiple sclerosis: a marker of disease severity

Context: In multiple sclerosis (MS) axonal damage is an early event and is probably to be considered the most relevant cause of permanent and progressive disability. Objectives: To investigate the value of the increase of 14-3-3 and tau proteins in the cerebrospinal fluid (CSF) as peripheral markers of axonal pathology and predictors of disease evolution. Patients and methods: In the CSF samples obtained from 63 patients with demyelinating diseases (DD), including 20 clinically isolated syndromes (CIS) and 43 clinically defined MS, as well as from 56 controls, we analysed the presence of 14-3-3 reactivity by immunoblot analysis along with the concentration of tau protein by sandwich ELISA. Results: The percentage of DD subjects showing a positive 14-3-3 protein CSF reactivity (38%) was significantly higher than the one previously detected, and was correlated in the MS patients with a more severe clinical phenotype in terms of degree of disability and rate of disease progression, during a 10-month mean clinical follow-up. On the contrary, the levels of the CSF-tau protein were highly variable in DD and control subjects, and the mean CSF-tau concentration was similar in both groups. Conclusions: The immunoblot analysis of 14-3-3 protein in the CSF could be a useful marker to identify a subgroup of DD patients with high risk of developing severe disability.

Involvement of the choroid plexus in multiple sclerosis autoimmune inflammation: A neuropathological study

An immunological function has been proposed for the choroid plexus (CP). In multiple sclerosis (MS) brains, CPs show (immunohistochemistry to HLA-DR, CD3, CD20, CD68, VCAM-1, CD138) T lymphocytes in vessels and stroma, VCAM-1 expression on endothelia, intense HLA-DR immunostaining on cells in CP stroma, among CP epithelium and on epiplexus cells. CPs in control or amyotrophic lateral sclerosis brains do not show such inflammatory changes. Intense CP inflammation is observed in viral encephalitis. Changes in MS CPs suggest persisting immune activation, with intensity similar to acute encephalitis, even in MS phases in which neurodegeneration prevails. In MS, CPs could represent a site for lymphocyte entry in the CSF and for CSF antigens presentation.