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Biomedical subjects

M B Lees

Publications and source records attributed to M B Lees.

At least 73 records · Page 4Linked to original sources
PubMed1983

Chronic experimental allergic encephalomyelitis produced by bovine proteolipid apoprotein: immunological studies in rabbits.

A chronic experimental allergic encephalomyelitis (EAE) has been produced in rabbits sensitized with bovine white matter proteolipid apoprotein. Eleven of 12 animals developed clinical disease one to six months after immunization with a single dose of the apoprotein. The clinical course was characterized by posterior ataxia, flaccid paralysis progressing to spastic paralysis, and incontinence. Spontaneous relapses and remissions were observed in 3 rabbits. Histologically, acute and chronic encephalomyelitis accompanied by primary demyelination were observed. Serum antibody production, assayed by both an enzyme-linked immunosorbent assay and an electroblot procedure, did not correlate with either the clinical course or the histopathological findings. Delayed hypersensitivity to proteolipid apoprotein was observed in all rabbits prior to the onset of clinical signs. The data suggest that lymphocytes specifically sensitized to the proteolipid may be involved in the pathogenesis of the demyelination in chronic EAE.

Animals↗
PubMed1982

A hydrophobic tryptic peptide from bovine white matter proteolipid.

A hydrophobic, chloroform-soluble tryptic peptide with a molecular weight of approximately 4000 has been purified from the bovine white matter proteolipid protein. Its primary structure was obtained by a combination of solid-phase Edman degradation and mass spectrometry. A major part of the tryptic peptide appears to be inaccessible to the action of proteolytic enzymes. The peptide spans the three cyanogen bromide peptides located by Jollès et al. (Biochem. Biophys. Res. Commun. (1979) 87, 619--626) at the COOH-terminal region of the intact protein. Secondary structure calculations for this region indicate a segregation into discrete domains, with most of the tryptic peptide corresponding to a highly ordered, hydrophobic domain; an equal probability for alpha-helical or beta-structure is predicted for this region.

Amino Acid Sequence↗
PubMed1982

Electroblot analysis of the myelin proteolipid protein.

The myelin proteolipid has been studied by the electroblot method of Towbin et al [1979]. Samples were separated by SDS-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and incubated with proteolipid antibody. The proteolipid band could be identified immunologically in CNS myelin and in whole brain homogenates. No proteolipid was detected in PNS myelin. Proteolipid from human, bovine, rat, and mouse myelin all cross-react when analyzed by this method. No cross-reactivity was indicated between proteolipid and myelin basic protein.

Animals↗
PubMed1982

Interactions of dicyclohexylcarbodiimide with myelin proteolipid.

Dicyclohexylcarbodiimide (DCCD) is known to bind preferentially to a proteolipid subunit of proton-translocating systems and thereby to inhibit proton transport. In the present study we show that, in an aqueous medium, DCCD binds to the bovine white matter proteolipid apoprotein, the major protein of central nervous system myelin. The binding is dependent on time, temperature, and concentration and is not inhibited by the hydrophilic carbodiimide 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide. By contrast, when the incubation is carried out in chloroform/methanol no labeling by DCCD is demonstrable. In isolated rat myelin, DCCD binds specifically to the proteolipid and not to the myelin basic proteins. Liposomes reconstituted with the myelin proteolipid apoprotein transport protons, as assayed by quenching of the fluorescence of 9-aminoacridine. Preincubation of proteolipid-containing liposomes with DCCD results in an inhibition of transport. These studies have important implications for a possible ionophoric function of the myelin proteolipid and for the occurrence of transport processes within myelin.

Animals↗
PubMed1982

Chronic experimental allergic encephalomyelitis induced in rabbits with bovine white matter proteolipid apoprotein.

A chronic, progressive form of experimental allergic encephalomyelitis was produced by immunization of rabbits with bovine brain white matter proteolipid apoprotein. Clinical signs appeared 4 to 13 months after sensitization, and were characterized by ataxia and limb paresis which progressed to flaccid paralysis and incontinence. Light and electron microscopic observations showed both acute and chronic nonsuppurative myelitis or encephalomyelitis accompanied by primary demyelination. Myelin damage was most evident in the spinal cord but was also present in the optic nerve and brain. The neuropathology was consistent with lesions of chronic experimental allergic encephalomyelitis produced by central nervous system tissue, and resembled lesions of multiple sclerosis as well. These observations suggest that protein may be involved in the pathophysiology of demyelinating diseases. A mechanism for the chronic course of the disease is discussed.

Animals↗
PubMed1982

Proteolipids.

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Amino Acids↗
PubMed1982

Solid-phase immunoassays for quantitation of antibody to bovine white matter proteolipid apoprotein.

Two solid-phase immunoassays have been developed for quantitation of antibodies to bovine white matter proteolipid apoprotein. Conditions were established for optimal specific antibody binding. Water-soluble proteolipid apoprotein was bound to microtiter plates and plates were incubated with test serum. Goat anti-rabbit IgG conjugated with horseradish peroxidase was used as the second antibody for an enzyme-linked immunospecific assay and 125I-labeled protein A for a radioimmunoassay. Both procedures have been used to follow the time course of anti-proteolipid antibody production in rabbits and to compare different immunization protocols.

Animals↗
PubMed1981

A study of elastase peptides from bovine white matter proteolipid.

Bovine white matter proteolipid has been digested with elastase in the presence of deoxycholate. After acidification, the digest was separated into an acid-soluble and an acid-insoluble fraction. The acid-insoluble fraction was enriched in nonpolar amino acids and, by a combination of solvent fractionation and chromatography, a fraction was obtained which consisted of a mixture of two peptides with a molecular weight of approximately 4000 daltons. The acid-soluble peptides were separated by molecular sieve, ion exchange and high performance liquid chromatography (HPLC) in the reverse phase mode. The purified peptides were smaller than expected on the basis of their elution position from a molecular sieve column, suggesting they were in an aggregated state during the initial chromatography. Reverse phase HPLC was shown to be useful for fingerprinting these peptide mixtures. The data demonstrate the difficulties associated with the study of this proteolipid and emphasize the tendency of both the protein and the peptides derived from it to aggregate.

Amino Acids↗
PubMed1981

Is Na + ATPase a myelin-associated enzyme?

The Na + K ATPase activity associated with purified myelin has been investigated. On the basis of marker enzyme studies, the Na + K ATPase activity of myelin was higher than could be accounted for by microsomal contamination. Fractions prepared from white matter-enriched areas of rat brain showed a threefold enrichment in Na + K ATPase activity in myelin as compared with the white matter homogenate. The ATPase activity in myelin was stimulated fourfold by treatment with sodium deoxycholate, but the activity in the whole brain homogenate and the microsomal fraction was only doubled. This discontinuity temperature for Na + K ATPase activity was significantly higher for the myelin fraction (29 degrees C) than for the microsomal fraction (21 degrees C), but the energies of activation, both above and below the discontinuity temperature, were the same for both fractions, Myelin Na + K ATPase had a lower affinity for strophanthidin than the microsomal enzyme, but both fractions were inhibited to the same extent by 10-3 M-strophanthidin. The evidence thus indicated that much of the ATPase activity of myelin is not the result of microsomal contamination. Although the possibility of axolemmal contamination cannot be ruled out conclusively, indirect evidence suggest that this is not a significant factor and that Na + K ATPase may be a myelin-associated enzyme.

Animals↗
PubMed1981

Production and purification of antibody to bovine white matter proteolipid apoprotein.

Circulating antibody to the bovine white matter proteolipid apoprotein was detected in rabbits 1 month ater a single injection of the water-soluble form of the apoprotein. By double immunodiffusion, the antiserum reacted specifically with the delipidated proteolipid apoprotein and the crude proteolipid fraction containing complex lipids; after exposure of the proteolipid apoprotein to sodium dodecyl sulfate (SDS), no reactivity was observed. The antiserum did not react with other myelin components, i.e., basic protein, cerebroside or GM1 ganglioside, nor was there reactivity with non-neural proteolipids. The anti-apoprotein antibody was purified by affinity chromatography. The antibody-antigen interaction is apparently very hydrophobic, since elution of the antibody from the affinity column requires buffer containing 0.5% Triton X-100-4 M-urea.

Animals↗
PubMed1981

Hydrophobic compounds interfere in radioimmunoassay for basic protein in myelin.

Hydrophobic compounds influenced the accuracy of the radioimmunoassay for myelin basic protein when lipids (stearic acid, phosphatidylcholine, cholesterol, cerebroside, sulfatide, or GM1 ganglioside) or proteolipids (white-matter proteolipid apoprotein, kidney proteolipid apoproteins, or heart proteolipid apoproteins) were added to a known amount of basic protein and the samples assayed. All of these interfere with the assay, but the direction of the error depends on the quantity added: low concentrations of lipid decrease apparent basic protein, high concentrations enhance it. Obviously, results of basic-protein assays must be interpreted carefully.

Animals↗
PubMed1981

Jordi Folch-Pi.

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History, 20th Century↗
PubMed1980

Developmental changes in carbonic anhydrase and adenylate cyclase in quaking mice.

The present investigation focuses on the developmental changes in the activity and levels of carbonic anhydrase, adenylate cyclase and 3',5'-cyclic adenosine monophosphate (cAMP) in the Quaking mouse mutant in different regions of the brain. Carbonic anhydrase activity was found to be lower than normal in the forebrain of the C57/B Quaking mouse. The deficit in forebrain carbonic anhydrase was restricted to subcortical structures and was reflected equally in the membrane and soluble factions, indicating that neither pool was affected selectively. However, no difference in carbonic anhydrase activity was observed in purified myelin from Quaking and control mice. Investigation of the changes in carbonic anhydrase activity as a function of age showed a cessation in enzyme accumulation in Quaking mice at around 20 days postnatally, suggesting an abnormality in cellular development. A tritiated acetazolamide binding assay was used to quantitate the amount of enzyme present. The amount of carbonic anhydrase parallelled enzyme activity, suggesting that the defect in the Quaking animals was at the level of the control of enzyme synthesis. Similar studies on cyclic AMP metabolism showed a higher than normal adenylate cyclase activity in the upper brain stem region of Quaking mice of ages between 19 and 40 days. Adenylate cyclase activity was stimulated by norepinephrine in both control and Quaking animals. The increased adenylate cyclase activity in the Quaking mice was in contrast to a lower cyclic AMP level and could not be accounted for by an alteration in phosphodiesterase activity.

Adenylyl Cyclases↗
PubMed1980

Carbonic anhydrase and 2',3' cyclic nucleotide 3'-phosphohydrolase activity in normal human brain and in demyelinating diseases.

The activities of carbonic anhydrase and 2',3' cyclic nucleotide 3'-phosphohydrolase (CNPase) were measured in gray and white matter and in myelin from human brains obtained at autopsy. Carbonic anhydrase activity increased with age, and at all ages a major part of the activity was associated with membrane fractions. The percentage of membrane-bound carbonic anhydrase was lower than normal in white matter from Krabbe disease and adrenoleukodystrophy; isolated myelin had a low specific activity in these diseases. CNPase activity was decreased in both white matter and myelin, but was somewhat higher than normal in gray matter. Although the yield of myelin from a case of metachromatic leukodystrophy was markedly reduced, changes in the enzymes activities were minimal.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1979

Interactions of Folch-Lees proteolipid apoprotein with planar lipid bilayers.

Water-soluble Folch-Lees proteolipid apoprotein from bovine CNS white matter induces a voltage-dependent conductance in black lipid membranes. Na+ is required for the induced conductance change but the established conductance has very low ionic selectivity. The induced conductance fluctuates with a minimum amplitude of 10(-11)--10(-10) mho. The magnitude of the conductivity change is dependent on protein concentration and on the composition of lipid bilayers. At a fixed voltage the induced conductance of a phosphatidylcholine-cholesterol membrane is proportional to the sixth power of the protein concentration and the first power of Na+ concentration. The interactions between the apoprotein and the lipids are both electrostatic and hydrophobic, but the interaction leading to the conductance increase appears to be mainly hydrophobic. Both the increase in conductance and the current fluctuations remain after extensive washing of the chambers to remove the protein. Furthermore, pronase or glutaraldehyde added to either the cis or trans side of the membrane does not affect the apoprotein-established conductance. However, if the bilayer is formed in the presence of both the apoprotein and pronase or if the apoprotein is treated with pronase prior to its addition to the chamber, no conductance change is observed. The association of the apoprotein with the membrane thus appears to render the protein inaccessible to proteolytic digestion, suggesting that the apoprotein is at least partially imbedded in the membrane interior.

Animals↗
PubMed1978

Regional developmental and fractional studies on myelin and other carbonic anhydrases in rat CNS.

Myelin carbonic anhydrase (CA) was studied with respect to its development in various brain regions and light and heavy myelin (LM and HM). The data indicate that the specific activity of myelin CA has a clear neuraxial distribution, increasing rostrally. The absolute activities and relative distribution are invariant with age; this suggests the CA activity in myelin is independent of stage and degree of myelination. The studies on HM and LM illustrate that HM, like total myelin, has a constant CA activity during development. In contrast, LM although equal to HM at 14 days, progressively decays to an adult level which is one-fourth that of HM. The distribution of CA in myelin was further investigated by comparing the activity in myelin with that present in the SN4 fraction. The activity in this latter fraction, which is derived from heavy myelin, was found to be 2.2 times higher than that in the myelin fraction. Thus, in the adult there exists an almost ten-fold range of activities among the various myelin fractions, SN4 greater than HM greater than LM. This may indicate a segregation of activity towards the outer lamellae. This segregation may have physiological importance in that it is this region of the sheath which should be integrally involved in control of myelin edema. Evidence indicates that there is an interaction of chloride with the enzyme, and maybe the primary ion moved by CA in order to initiate an osmotic flux out of the sheath. The interaction of chloride with the enzyme is dependent on the CA complex with the membrane in that solubilization and partial (60-fold) purification results in a preparation which is refractory to anions.

Aging↗