Quantitation of the Myelin-Associated Glycoprotein in Human Nervous Tissue from Controls and Multiple Sclerosis Patients

Myelin-associated glycoprotein (MAG) was measured by radioimmunoassay in the human CNS and peripheral nervous system (PNS). The level of MAG, expressed as ng/microgram of total protein, was approximately 20-fold higher in whole homogenates of cerebral white matter (4.7 +/- 0.60) than of peripheral nerve (0.12-0.28). MAG concentrations were only slightly higher in the isolated myelin fractions from these tissues: CNS myelin, 5.6 ng/microgram; PNS myelin, 0.37 ng/microgram. The levels of MAG were measured in nine plaques, periplaque regions, and areas of macroscopically normal-appearing white matter (NAWM) from six separate multiple sclerosis brains and compared with the levels of other myelin proteins in the same samples. MAG and other myelin proteins were reduced to very low levels in plaques. The levels of MAG and basic protein (BP) and the activity of 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP) in periplaque areas were significantly lower than those in control white matter, and MAG and BP levels were also significantly reduced in NAWM. In a periplaque region and NAWM from the most rapidly progressing case of multiple sclerosis examined, the MAG content was between 30 and 35% of the control level, whereas BP and PLP levels and CNP activity were between 50 and 85% of control values. The reduction of MAG content in periplaque regions from all nine multiple sclerosis plaques examined was significantly greater than the reductions of BP level and CNP activity. In NAWM samples, the mean reduction of MAG content was also greater than the reductions of BP level and CNP activity, but the difference was only statistically significant in comparison to CNP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Influence of pertussis toxin on the effects of guanine nucleotide on adenylate cyclase in rat striatal membranes

The influence of pertussis toxin on the effects of guanine nucleotide on adenylate cyclase activity were investigated in rat striatal membranes. GTP promoted and inhibited the activity at 1 and 100 microM, respectively. The inhibitory effects of GTP were abolished by pretreatment of the membranes with pertussis toxin. GppNHp (guanyl-5'-y1-beta,gamma-imidodiphosphate) exerted only stimulatory effects and pertussis toxin did not affect the effects of GppNHp. GDP at 10 and 100 microM caused significant inhibition which was completely suppressed by pertussis toxin. It is suggested that guanine nucleotide regulates the affinity of as in stimulatory GTP-binding regulatory protein to either beta gamma or catalytic units of adenylate cyclase in a flip-flop manner. Inhibitory GTP-binding regulatory protein seems to play a regulatory role in inhibiting alpha s activity supplying the beta gamma heterodimer.     

Biochemical Characterization of α-Adrenergic Receptors in Human Brain and Changes in Alzheimer-Type Dementia

Abstract Using ligand binding techniques, we studied α-adrenergic receptors in brains obtained at autopsy from seven histologically normal controls and seven patients with histopathologically verified Alzheimer-type dementia (ATD). Binding of the α-adrenergic antagonists [³H]prazosin and [³H]yohimbine to membranes of human brains exhibited characteristics compatible with α₁- and α₂-adrenergic receptors, respectively. Binding of both ligands was saturable and reversible, with dissociation constants of 0.15 nM for [³H]prazosin and 5.5 nM for [³H]yohimbine. [³H]Prazosin binding was highest in the hippocampus and frontal cortex and lowest in the caudate and putamen in the control brains. [³H]Yohimbine binding was highest in the nucleus basalis of Meynert (NbM) and frontal cortex and lowest in the caudate and cerebellar hemisphere in the control brains. Compared with values for the controls, [³H]prazosin binding sites were significantly reduced in number in the hippocampus and cerebellar hemisphere, and [³H]yohimbine binding sites were significantly reduced in number in the NbM in the ATD brains. These results suggest that α₁ and α₂-adrenergic receptors are present in the human brain and that there are significant changes in numbers of both receptors in selected regions in patients with ATD.     

(6R)-Tetrahydrobiopterin Increases the Activity of Tryptophan Hydroxylase in Rat Raphe Slices

The effects of (6R)- and (6S)-tetrahydrobiopterin (BPH4), tetrahydroneopterin, and 6-methyltetrahydropterin on the activity of tryptophan hydroxylase were investigated in rat raphe slices. The activity of tryptophan hydroxylase was estimated by measurement of 5-hydroxytryptophan (5-HTP) formation under inhibition of aromatic L-amino acid decarboxylase with use of HPLC-fluorometric detection. (6R)-BPH4 (the naturally occurring form) at 42 microM, tetrahydroneopterin at 50 microM, and 6-methyltetrahydropterin at 100 microM increased tryptophan hydroxylase activity to 350, 145, and 146% of control values, respectively. (6S)-BPH4, however, had no significant effects on tryptophan hydroxylase activity. These results suggest that tryptophan hydroxylase is subsaturating in vivo for the naturally occurring cofactor, (6R)-BPH4, and that the concentration of (6R)-BPH4 may play an important role for the regulation of tryptophan hydroxylase activity in vivo.     

Interleukin 2 acts directly on a Tac-positive cloned B cell line established from a patient with adult T cell leukemia

A Tac-positive B cell line termed K3B was established from a patient with adult T cell leukemia (ATL). This cell line had EBNA antigen and human T cell leukemic virus (HTLV) provirus besides B1 antigen and surface immunoglobulin. A cloned Tac-positive B cell line termed K3B01 was obtained from K3B by the limiting dilution method. The K3B01 cells were shown to absorb IL 2 activity in a tonsillar IL 2 preparation. By using this cloned cell line and a purified recombinant IL 2 preparation, it was shown that the proliferation of K3B01 cells was enhanced by the addition of recombinant IL 2. Moreover, this response was inhibited by anti-Tac antibody. These results demonstrate definitively that IL 2 acts directly on B cells through IL 2 receptors on them.     

The differentiation of HL-60 cells in the synthetic medium induced by GM3 ganglioside

GM3 ganglioside, added exogenously to a promyelocytic leukemia cell line (HL-60 cells) in serum-free synthetic medium, induced differentiation into macrophage-like cells. Macrophagic morphology and function of differentiation-induced cells were determined by cell growth behavior, May-GriJnwald-Giemsa staining, activities of nonspecific esterase, phagocytosis and nitroblue tetrazolium (NBT) reduction. GM3 ganglioside may play a role in triggering differentiation of HL-60 cells into macrophage-like cells.     

In vitro establishment of human fibroblasts of lysosomal diseases,GM1-gangliosidosis and Sandhoff disease,by transformation with origin-minus SV40 DNA

The permanent human cell lines preserving defects of lysosomal enzymes, G_M1-1019-SV and SA-1077-SV, were established from the respective fibroblasts from patients with G_Ml-gangliosidosis and Sandhoff disease by transfection with replication origin-minus simian virus 40 DNA. These ceils grow rapidly without entering senescence during more than 120 population doublings. The activity of -galactosidase in G_M1-1019-SV and of B-N-acetylhexosaminidase in SA-1077-SV was respectively 40- and 180-fold lower than that of normal fibroblasts.     

Formation of giant protoplasts from protoplasts of Pleurotus cornucopiae by the cell wall lytic enzyme

Summary When purified protoplasts of Pleurotus cornucopiae IFO9614 were incubated with a mixture of cell wall lytic enzymes, they were found to increase their size. Their average diameter increased from 4.3 m to 31 m after 65 h incubation at 24° C. The presence of cellulase ONOZUKARS in the enzyme mixture had a significant effect on the formation of giant protoplasts. Regeneration frequency of giant protoplasts in a medium containing 0.5 M sucrose was 3.5%, approximately six times that of normal protoplasts.     

Isolation and physical mapping of temperature-sensitive mutants defective in heat-shock induction of proteins in Escherichia coli

Summary Mutants of Escherichia coli K12 that are partially or totally defective in induction of major heat-shock proteins and cannot grow at high temperature (42° C) were isolated by localized mutagenesis. These mutants carry a single mutation in the gene htpR (formerly hin) located at min 76 on the E. coli genetic map. Some mutants exhibit delayed (partial) induction of heat-shock proteins or require a higher temperature for induction than the wild type, whereas others are not induced under any of these conditions. The maximum temperature that allows growth varies among different mutants and is correlated with the residual induction capacity. Temperature-resistant revertants obtained from each mutant are fully or partially recovered in heat-shock induction. These results indicate that the inability of htpR mutants to grow at high temperature is due to the defect in heat-shock induction. In addition, a couple of mutants was found that produce significantly higher amounts of heat-shock proteins even at 30° C.The htpR gene has been cloned into plasmid pBR322 using the above mutants, and was localized to a DNA segment of 1.6 kilobase pairs. The mutants harboring certain palsmids that carry a part of htpR produce temperature-resistant recombinants at high frequency. This permits further localization of mutations within the htpR gene. Analysis of proteins encoded by each of the recombinant plasmids including the one carrying a previously isolated amber mutation (htpR165) led to the identification of a protein with an apparent molecular weight of about 36,000 daltons as the htpR gene product.     

Hormonal Regulation of Adipose S-100 Protein Release

The release of S-100 protein from epididymal fat pads was enhanced by epinephrine in vitro, and about 50% of S-100 protein in the tissue was released into the medium after 2-h incubation at 37 degrees C with 10 microM epinephrine. Similar results were obtained with the incubation of isolated adipocytes. The S-100 protein release was also enhanced by isoproterenol, norepinephrine, ACTH, and dibutyryl cyclic AMP, which all increase the lipolysis by increasing cyclic AMP levels in the tissue. Propranolol, a beta-adrenergic blocker, could block the increase of S-100 protein release by catecholamines, indicating that the release was mediated by the beta-adrenergic effect of catecholamines. However propranolol had no suppressive effect on the enhancement of S-100 protein release by ACTH or dibutyryl cyclic AMP. Insulin had an inhibitory effect on the epinephrine-enhanced S-100 protein release. Epinephrine or ACTH could not stimulate the S-100 protein release in the absence of Ca2+, whereas the epinephrine-enhanced glycerol release was not affected under the same conditions. The increase in S-100 protein release was induced by only a pretreatment of the tissue with epinephrine. However, the lipolysis in the tissue was not enhanced by the pretreatment alone. These results indicate that the release of S-100 protein from adipocytes is regulated by the hormones that have been known to control the lipolysis with a manner slightly different from that of lipolysis.