An In Vitro Micromethod for the Quantitative Assessment of Central Demyelination

Abstract

We report the development of a simple and reliable method for the study of demyelination in vitro based on the measurement of 2':3'-cyclic nucleotide 3'-phosphodiesterase in isolated myelin. Using only small quantities of myelin (equivalent to 100 micrograms of myelin protein) the system was tested under conditions that are believed to approximate those found at the site of an inflammatory demyelinating lesion. Treatment with a combination of trypsin, phospholipase A2, and lysophosphatidylcholine was used to evaluate the method. This microsystem has the potential not only for testing the myelinotoxicity of soluble factors but also for investigating the involvement of inflammatory cells in the demyelinating process. Myelin degradation by elicited peritoneal macrophages could be demonstrated at relatively high densities of these cells. Nylon wool purified lymph node T cells from myelin basic protein-primed SJL/J mice, after selective expansion with antigen and interleukin 2, failed to induce any significant myelin breakdown unless a limited number of syngeneic activated macrophages were also present. T cells from mice that had been inoculated with keyhole limpet haemocyanin failed to show any effect. The advantages of this technique over other in vitro systems are that it enables the study of demyelination using syngeneic sources of myelin and defined cell populations.

Theiler's virus replication in brain macrophages cultured in vitro

“…Specific T cells can release lymphokines which lead to the recruitment, accumulation, and activation of additional brain macrophages in the CNS tissue. Brain macrophages can also produce tumor necrosis factor alpha, which induces oligodendrocyte necrosis and demyelination (6,17,36), prostaglandin E (16), and other mediators such as leukotrienes and phospholipases, proteinases that have been shown to have high myelinolytic activity (7,42). The amount of tumor necrosis factor alpha (37) and of IL-1 (40) in the cerebrospinal fluid and sera of patients with MS has been correlated with the intensity of the disease.…”

Degradation of human myelin in vitro by leucocytes from patients with multiple sclerosis

“…However, there is some evidence that a relapse may be initiated peripherally, for recent studies on lymphocyte migration have found CD2-positive cells crossing the blood-brain barrier from the blood into the CSF in progressive MS (Hafler & Weiner, 1987). It has also been suggested that cells may circulate from the CNS to the blood (Oehmichen et al, 1979;Wekerle et al, 1986). Therefore, the Correspondence: Samantha J. Owen, Department of Neurochemistry, Institute of Neurology, London WCIN 3BG, UK.…”