C. J. A. De Groot

Axonal loss in multiple sclerosis lesions: Magnetic resonance imaging insights into substrates of disability

Magnetic resonance imaging (MRI) monitoring of disease progression in multiple sclerosis is limited by the lack of correlation of abnormalities seen on T2‐weighted imaging, and disability. We studied the histopathology of multiple sclerosis lesions, as depicted by MRI, in a large postmortem sample, focusing on axonal loss. Tissue samples from 17 patients were selected immediately postmortem for histopathological analysis on the basis of T2‐weighted imaging, including normal appearing white matter and T1 hypointense lesions. In each region, we measured magnetization transfer ratios (MTR), T1 contrast ratio, myelin, and axonal density. T2 lesions (109 samples) were heterogeneous with regard to MRI appearance on T1 and MTR, whereas axonal density ranged from 0% (no residual axons) to 100% (normal axonal density). Of 64 T2 lesions, 17 were reactive (mild perivascular inflammation only), 21 active, 15 chronically active, and 11 chronically inactive. MTR and T1 contrast ratio correlated strongly with axonal density. Also in normal appearing white matter (24 samples), MTR correlated with axonal density. In conclusion, postmortem tissue sampling by using MRI revealed a range of pathology, illustrating the high sensitivity and low specificity of T2‐weighted imaging. T1 hypointensity and MTR were strongly associated with axonal density, emphasizing their role in monitoring progression in multiple sclerosis.

Staging of multiple sclerosis (MS) lesions: pathology of the time frame of MS

Several processes take place during an attack of demyelination in multiple sclerosis (MS). The timing of these various processes, and thus of the attack in its entirety, is important if therapeutic stratagies are to be planned. Attempts have been made to introduce and investigate variables relevant to timing the disease processes, leading to staging systems for MS. Here, the terminology and the various parameters used are reviewed, including inflammatory cells, glial cells, axonal loss and myelin staining; then the different systems are compared, including the system put forward by Bö and Trapp, our own modification of that, the Brück and Lassmann system and the recent consensus reached at a Vienna meeting. It is concluded that an ideal staging system does not yet exist, and that, more than anything else, the material dictates the choice for a staging system. The terminology of the Vienna consensus could be used as a reference to facilitate international comparison.

Cyclooxygenase expression in microglia and neurons in Alzheimer's disease and control brain

Abstract. Epidemiological studies suggest that non-steroidal anti-inflammatory drugs (NSAIDs) lower the risk of developing Alzheimers disease (AD). Most NSAIDs act upon local inflammatory events by inhibiting the expression or activation of cylooxygenase (COX). In the present study the expression of COX-1 and COX-2 in AD and non-demented control temporal and frontal cortex was investigated using immunohistochemistry. COX-1 expression was detected in microglial cells, while COX-2 expression was found in neuronal cells. In AD brains, COX-1-positive microglial cells were primarily associated with amyloid β plaques, while the number of COX-2-positive neurons was increased compared to that in control brains. No COX expression was detected in astrocytes. In vitro, primary human microglial and astrocyte cultures, and human neuroblastoma cells (SK-N-SH) were found to secrete prostaglandin E2 (PGE2), especially when stimulated. PGE2 synthesis by astrocytes and SK-N-SH cells was stimulated by interleukin-1β. Microglial cell PGE2 synthesis was stimulated by lipopolysaccharide only. Although astrocytes are used in studies in vitro to investigate the role of COX in AD, there are no indications that these cells express COX-1 or COX-2 in vivo. The different distribution patterns of COX-1 and COX-2 in AD could implicate that these enzymes are involved in different cellular processes in the pathogenesis of AD.

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.

Monocyte infiltration is highly associated with loss of the tight junction protein zonula occludens in HIV-1-associated dementia

In human immunodeficiency virus (HIV)‐1‐associated dementia (HAD), consequences of interactions between infiltrating monocytes and brain endothelial cells are not yet fully understood. This study investigated whether the blood–brain barrier is affected in brain tissue of patients suffering from HAD and whether it was possible to find a correlation with the presence or absence of monocytic cells, which have been suggested to play a major role in HAD. Immunohistochemical analysis for zonula occludens 1, a tight junction protein, and CD68, a macrophage marker, revealed that loss of tight junction immunoreactivity was highly correlated with monocyte infiltration and with HAD. This suggests that the presence of perivascular macrophages cells is associated with breakdown of the blood–brain barrier thereby facilitating infiltration of more monocytic cells hence enhancing disease progression.

Macrophage inflammatory protein‐1α (MIP‐1α), MIP‐1β, and RANTES mRNA semiquantification and protein expression in active demyelinating multiple sclerosis (MS) lesions

MS is a demyelinating disease characterized by infiltration of monocytes and lymphocytes into the brain parenchyma, destruction of oligodendrocytes and loss of myelin. Since chemokines play a major role in the migration of monocytes and T cells, we here investigated the expression of the CC chemokines MIP‐1α, MIP‐1β, and RANTES in brain tissue from MS patients using reverse transcriptase‐polymerase chain reaction techniques. Both MIP‐1β as well as RANTES were found to be significantly elevated in brain tissue of MS patients. In addition, MIP‐1α was also increased, although not significantly. Immunohistochemistry revealed that, whereas RANTES was mainly localized in reactive astrocytes, MIP‐1α and MIP‐1β immunoreactivity was predominantly found in perivascular and parenchymal macrophages, containing myelin degradation products. Thus, chemokines appear to be associated with MS and an increased chemokine expression may further enhance disease progression by attracting more leucocytes into the brain parenchyma and by activation of effector functions of astrocytes and microglial cells.

Isolation and characterization of adult microglial cells and oligodendrocytes derived from postmortem human brain tissue

The present study provides a detailed description of the simultaneous establishment and immunocytochemical characterization of highly enriched human adult microglial cell cultures as well as of oligodendrocyte cultures. For this study, brain tissue specimens were collected at autopsy with relatively short postmortem times (3-9 h) from various regions of the CNS of Alzheimers disease, Picks disease and non-demented control cases. Although methods to isolate viable glial cells from human adult brain tissue have been described, these human brain specimens were often derived from surgical resections, i.e., in order to treat intractable epilepsy, brain tumors or cardiovascular diseases involving the brain. However, for the study of many neurological disorders, surgical material is not available. Furthermore, for obvious reasons, there is a limit to the number of central nervous system (CNS) regions from which (enough) tissue can be obtained at surgery. The adherent primary microglial cells, isolated according to the here described procedures consisted of proliferating, phagocytotic cells that expressed various microglia/macrophage-specific markers as judged by immunocytochemical analysis. Non-adherent cells isolated from the same brain tissue samples expressed oligodendrocyte-specific markers. The current described culture system may provide a valuable tool in studying human CNS biology and disease.

Therapeutic effect of the D2-dopamine agonist bromocriptine on acute and relapsing experimental allergic encephalomyelitis.

We examined the effect of bromocriptine (BCR) treatment on the duration and severity of neurological symptoms of acute experimental allergic encephalomyelitis (EAE), an animal model for demyelinating diseases, particularly multiple sclerosis. To mimic the clinical situation, BCR treatment was started after the onset of clinical signs. Furthermore, the effect of BCR treatment on the course of a chronic relapsing form of EAE was studied. BCR was injected at daily intervals in a dose that resulted in sustained suppression of plasma concentrations of prolactin, a pituitary hormone that plays a role in immunoregulation. In acute EAE, BCR therapy reduced both severity and duration of the clinical signs. In chronic relapsing EAE, BCR treatment did not affect the severity and duration of the first attack, but reduced the duration of the subsequent, second attack. Thus, BCR treatment improves the clinical course in animals with ongoing disease. These findings may have implications for the search for new therapeutic approaches in multiple sclerosis.

Complement C1-inhibitor expression in Alzheimer's disease

Abstract In situ and in vitro studies suggest that activation of locally produced complement factors may act as a mediator between amyloid deposits and neurodegenerative changes seen in Alzheimer’s disease (AD). C1-esterase inhibitor (C1-Inh), which regulates activation of C1 of the complement classical pathway, can be detected immunohistochemically in its inactivated form in activated astrocytes and dystrophic neurites in AD plaque areas. In this study, designed to investigate the cellular source of C1-Inh, C1-Inh was found to be secreted in a functionally active form by astrocytes cultured from postmortem human brain specimens as well as by neuroblastoma cell lines. Recombinant human interferon-γ (IFN-γ), which stimulates C1-Inh synthesis in various cell types, several-fold stimulated C1-Inh protein secretion by cultured human astrocytes derived from different regions of the central nervous system and by one (SK-N-SH) of two neuroblastoma cell lines (SK-N-SH and IMR-32) included in this study. In contrast to IFN-γ, other cytokines [interleukin (IL)-1β, IL-6 and tumor necrosis factor (TNF)-α] that can be found in brain areas affected by AD, did not stimulate C1-Inh secretion by astrocytes or neuroblastomas in vitro. This inability to secrete C1-Inh is probably due to unresponsiveness at the transcriptional level, since C1-Inh secretion paralleled the expression of the 2.1-kb C1-Inh mRNA. In situ hybridization with a C1-Inh RNA antisense probe labeled neurons rather than astrocytes, suggesting a role for neurons as producers of complement regulatory proteins in vivo. Since IFN-γ is apparently lacking in the brain parenchyma, and amyloid plaque-associated cytokines (IL-1β, IL-6, TNF-α) do not stimulate C1-Inh expression in vitro, the nature of the stimulus responsible for neuronal C1-Inh expression in AD brains remains to be investigated.

The Expression of Ia Antigen in the Course of Acute and Relapsing Experimental Allergic Encephalomyelitis in the Lewis Rat

Acute experimental allergic encephalomyelitis (AEAE) can be induced in Lewis rats by immunization with nervous tissue or myelin basic protein (MBP) in Freud’s complete adjuvant.1 The course of the disease is usually monophasic and animals recover without remnant clinical signs. After treatment with low doses of cyclosporin A in the initial phase of the disease, the course of the disease becomes relapsing-remitting (CREAE).2