A Thiol Proteinase Highly Elevated in and Around the Plaques of Multiple Sclerosis

Abstract

Abstract— A thiol dependent proteolytic enzyme (tentatively identified as carboxypeptidase B) which liberates phenylalanine from CBZ‐glutamyl‐phenylalanine at pH 5.3 was shown, by a sensitive micromethod, to be greatly increased in activity in and around MS plaques. These increases exceeded those of other hydrolases previously measured. Plaque tissue, on the basis of lipid‐free dry weight, is up to 3‐fold richer in this enzyme than control white matter, and most samples of apparently uninvolved MS white matter already show elevated activities. The enzyme is highly dependent on the presence of dithiothreitol. It is unaffected by diisopropyl fluorophosphate and pepstatin, but inhibited by iodoacetate and leupeptin. Macrophages or lymphocytic infiltrations in the tissue do not appear to be the main source of the enzyme. In conjunction with measurements of DNA, reflecting gliosis, invasion by hematogenom cells and proliferation of phagocytes as well as oligodendrocyte loss, and acid lipase‐esterase, indicative of the survival or degeneration of oligodendrocytes, these results are interpreted as probably reflecting predominantly the activity of astrocytes. The incidental finding that most samples of unaffected white matter from MS patients contain more DNA per unit lipid free dry weight than average control white matter is considered significant in pointing to more widespread tissue changes independent of or preceding plaque formation.

Glycerol Phosphate Dehydrogenase: Reduced Activity in Multiple Sclerosis Plaques Confirms Localization in Oligodendrocytes

“…6, where the plaques were highly cellular and evidently of recent origin by histological criteria, as they contained mononuclear infiltrates, macrophages, gitter cells, and reactive astrocytes. A similar reduction in the plaques of patients 1, 3, and 5 has previously been documented for acid lipaseesterase (Hirsch and Parks, 1979); in patient 6 this enzyme also parallels GPDH, showing only an 11% reduction in plaque no. 2 (Hirsch and Parks, unpublished experiments).…”

“…The low activities of these two enzymes and that of the m y e l i n -s p e c i f i c 2 ' -3 ' -c y c l i c n u c l e o t i d e 3 'phosphodiesterase (CNP-ase) (Roytta et al, 1976) in older plaques contrast with those of other enzymes so far measured in these lesions, e.g., lactic dehydrogenase, glucose-6-phosphate dehydrogenase, and various acid hydrolases, all of which are either unchanged or elevated (Hirsch et al, 1976;Hirsch and Parks, 1979). Since oligodendrocytes as well as their myelin disappear from MS plaques and older plaques consist largely of fibrous astrocytes and axons (Adams, 1977), a localization in oligodendrocytes or myelin can be inferred for these en- Values are given as mol per kg lipid-free dry weight per hour.…”

“…All human tissues were obtained at autopsy within 6 h of death and were quick-frozen in liquid nitrogen. The sources of the tissues, both MS patients and nonneurological controls, as well as the histology of the plaques, have been described in previous publications (Hirsch et al, 1976;Hirsch and Parks, 1979). In MS patients nos.…”

The Immunological Identification of Brain Proteins on Cellulose Nitrate in Human Demyelinating Disease

“…It has been reported recently that a decrease in the myelin-associated glycoprotein (MAG) precedes that of BP (Itoyama et al, 1980). In a study of macroscopically normal white matter in MS, a high proportion of samples was found to be histologically abnormal (Allen and McKeown, 1979), and raised activities of several lysosomal hydrolases have been demonstrated in histologically and histochemically normal white matter (Einstein et al, 1972; Arstila et al, 1973;Hirsch and Parks, 1979; Allen and McKeown, 1979). The separation of sodium dodecylsulphate (SDS)-solubilized brain particulate fractions by polyacrylamide gel electrophoresis (PAGE) revealed changes in a number of protein bands in normal-appearing MS white matter, as well as in plaques (Newcombe et al,1 9 8 0~) .…”

Myelin

“…Bowen and Davison (1974) argued that the elevated cathepsin A, which is 30-fold more active in macrophages than in brain tissue, was an indicator of the infiltrating phagocytes. Other investigators have concluded that many of the enzymes, especially acid phosphatase, acid proteinase, carboxypeptidase B, and N-acetyl-ß-glucosaminidase, are derived from reactive astrocytes (Cuzner el al., 1976;Hirsch el al., 1976;Hirsch and Parks, 1979;McKeown and Allen, 1978;Allen el al., 1979;Hirsch, 1980Hirsch, , 1981.…”