Studies on the influence of the internal environment on autoantibody production by B cells

Purified splenic B cells from autoimmune NZB and nonautoimmune DBA/2 mice were transferred to unmanipulated H-2 compatible xid recipients. The number of autoantibody-secreting clones present in recipient mice was quantitated at varying times after transfer using a splenic fragment assay. We found that NZB and DBA/2 B cells expanded equally well in equivalent xid environments. Cells from either donor expanded significantly better in autoimmune-prone NZB.xid as compared with DBA/2.xid recipients. Moreover, clones producing antibodies reactive with T cell surface antigens, bromelain-treated mouse red cells, or DNA expanded more rapidly than did cells producing antibodies to the nonautoantigen TNP-KLH. Serum autoantibody levels rose in concert with the increased numbers of autoantibody-producing lymphocytes. We conclude that factors present in the internal milieu of autoimmune-prone NZB.xid mice, rather than an intrinsic B cell defect, facilitate the expansion of (auto)antibody-secreting B cells.     

Herbal medicine among the miskito of Eastern Nicaragua

Medicinal plants identified by Miskito informants in Awastara, Nicaragua, were collected in the field. They are listed and botanically identified in this paper. Particularly interesting among the collection of 23 plant species are those used to cure snakebite and athlete’s foot, as observed in the field.     

The species composition,occurrence and temporal stability of submerged aquatic macrophyte patches along the main channel border of pool 5A,Upper Mississippi River

Species composition, relative abundance, distribution and physical habitat associations of submerged aquatic macrophytes in the main channel border (MCB) habitat of Pool 5A, Upper Mississippi River (UMR) were investigated during the summers of 1980 and 1983. The submerged aquatic macrophytes in Pool .5A MCB were a small and stable component of the river ecosystem. Submerged plants occurred primarily in small, monospecific clumps. Clumps in close proximity to each other formed plant patches. Plant patches were stable in location and number between 1980 and 1983; 82.5% of the patches first observed in 1980 were present in 1983. Submerged macrophytes covered about 10–12 ha of the 201 ha MCB in Pool 5A. Submerged plants were most common in the lower two-thirds of the pool. Ten species of aquatic macrophytes occurred on rock channel-training structures and eleven occurred on non-rock substrates in the MCB. The most common submerged plants, in order of abundance, were Vallisneria americana Michx., Heteranthra dubia Jacq., Potamogeton pectinatus L., Ceratophyllum demersum L. and Potamogeton americanus C. & S.     

Magnesium and Manganese Content of Halophilic Bacteria

Magnesium and manganese contents were measured by atomic absorption spectrophotometry in bacteria of several halophilic levels, in Vibrio costicola, a moderately halophilic eubacterium growing in 1 M NaCl, Halobacterium volcanii, a halophilic archaebacterium growing in 2.5 M NaCl, Halobacterium cutirubrum, an extremely halophilic archaebacterium growing in 4 M NaCl, and Escherichia coli, a nonhalophilic eubacterium growing in 0.17 M NaCl. Magnesium and manganese contents varied with the growth phase, being maximal at the early log phase. Magnesium and manganese molalities in cell water were shown to increase with the halophilic character of the logarithmically growing bacteria, from 30 mmol of Mg per kg of cell water and 0.37 mmol of Mn per kg of cell water for E. coli to 102 mmol of Mg per kg of cell water and 1.6 mmol of Mn per kg of cell water for H. cutirubrum. The intracellular concentrations of manganese were determined independently by a radioactive tracer technique in V. costicola and H. volcanii. The values obtained by ⁵⁴Mn loading represented about 70% of the values obtained by atomic absorption. The increase of magnesium and manganese contents associated with the halophilic character of the bacteria suggests that manganese and magnesium play a role in haloadaptation.     

Stereochemical probes of bovine plasma amine oxidase: evidence for mirror image processing and a syn abstraction of hydrogens from C-1 and C-2 of dopamine

Bovine plasma amine oxidase (PAO) has previously been shown to catalyze a nonstereospecific loss of tritium from [2(R)-3H]- and [2(S)-3H]dopamines, attributed to multiple, catalytically active binding sites for substrate [Summers, M. C., Markovic, R., & Klinman, J. P. (1979) Biochemistry 18, 1969-1979]. Analysis of products formed from incubation of dopamine with PAO in tritiated water indicates a stereospecific, pro-R, incorporation of label at C-2. Thus, tritium washout (random) and washin (pro-R) are not the microscopic reverse of one another. We conclude that the (enamine) intermediates leading to tritium washin are nonequivalently bound. The observation of pro-R incorporation has provided a straightforward synthetic route to [1(R)-2H,2(R)-3H]- and [1(S)-2H,2(R)-3H]dopamines, which upon oxidation with PAO are expected to be processed preferentially by 1S and 1R cleavage, respectively. From previously measured isotope effects, we predict the loss of tritium from the 1(R)-2H and 1(S)-2H samples to be 74:8 for a syn relationship between cleavage at C-1 and C-2 vs. 21:90 for an anti relationship. The observation of a 68:18 ratio at 100% conversion provides strong evidence for a syn cleavage. The data support a mechanism in which a single base catalyzes a 1,3-prototrophic shift of hydrogen from C-1 of the substrate to cofactor, followed by exchange from C-2. Additionally, the results confirm the presence of alternate binding modes for dopamine at the active site of bovine plasma amine oxidase. This interaction of dopamine with plasma amine oxidase is a rare example of mirror-image catalysis in which a single substrate has two functional binding orientations on an enzyme surface.     

Measurement of Endogenous Noradrenaline Release in the Rat Cerebral Cortex In Vivo by Transcortical Dialysis: Effects of Drugs Affecting Noradrenergic Transmission

The release of endogenous noradrenaline was measured in the cerebral cortex of the halothane-anesthetized rat by using the technique of brain dialysis coupled to a radioenzymatic assay. A thin dialysis tube was inserted transversally in the cerebral cortex (transcortical dialysis) and perfused with Ringer medium (2 microliter min-1). Under basal conditions, the cortical output of noradrenaline was stable over a period of at least 6 h and amounted to 8.7 pg/20 min (not corrected for recovery). Histological control of the perfused area revealed very little damage and normal morphology in the vicinity of the dialysis tube. Omission of calcium from the perfusion medium caused a marked drop in cortical noradrenaline output. Bilateral electrical stimulation (for 10 min) of the ascending noradrenergic pathways in the medial forebrain bundle caused a frequency-dependent increase in cortical noradrenaline output over the range 5-20 Hz. Stimulation at a higher frequency (50 Hz) resulted in a levelling off of the increase in cortical noradrenaline release. Systemic administration of the dopamine-beta-hydroxylase inhibitor bis-(4-methyl-1-homopiperazinylthiocarbonyl) disulfide (FLA 63) (25 mg/kg i.p.) markedly reduced, whereas injection of the monoamine oxidase inhibitor pargyline (75 mg/kg i.p.) resulted in a progressive increase in, cortical noradrenaline output. d-Amphetamine (2 mg/kg i.p.) provoked a sharp increase in cortical noradrenaline release (+450% over basal values within 40 min). Desmethylimipramine (10 mg/kg i.p.) produced a twofold increase of cortical noradrenaline release. Finally, idazoxan (20 mg/kg i.p.) and clonidine (0.3 mg/kg i.p.), respectively, increased and decreased the release of noradrenaline from the cerebral cortex.(ABSTRACT TRUNCATED AT 250 WORDS)     

Mechanism-based inhibition of dopamine beta-monooxygenase by aldehydes and amides

A mechanism for beta-chlorophenethylamine inhibition of dopamine beta-monooxygenase has been postulated in which enzyme-bound alpha-aminoacetophenone is generated, followed by an intramolecular redox reaction to yield a ketone-derived radical cation as the enzyme inhibitory species (Mangold, J. B., and Klinman, J. P. (1984) J. Biol. Chem. 259, 7772-7779). If correct, additional compounds capable of producing enzyme-bound (formula; see text) reductant should inhibit dopamine beta-monooxygenase. Phenylacetaldehyde was chosen to test this model, since beta-hydroxyphenylacetaldehyde is expected to function as a reductant in a manner analogous to alpha-aminoacetophenone. Phenylacetaldehyde exhibits the properties of a mechanism-based inhibitor. Kinetic parameters are comparable to beta-chlorophenethylamine under both initial velocity and inactivation conditions. Since phenylacetaldehyde bears little resemblance to beta-chlorophenethylamine, its analogous inhibitory action provides support for an intramolecular redox reaction (via beta-hydroxyphenylacetaldehyde oxidation to a radical cation) in dopamine beta-monooxygenase inactivation. beta-Hydroxyphenylacetaldehyde was identified as the enzymatic product of phenylacetaldehyde turnover. As predicted, this product behaves both as a time-dependent inhibitor of dopamine beta-monooxygenase and as an electron donor in enzyme-catalyzed hydroxylation of tyramine to octopamine. Phenylacetamide and p-hydroxyphenylacetamide are also found to be mechanism-based inhibitors of dopamine beta-monooxygenase. In this case the product of hydroxylation (beta-hydroxyphenylacetamide) is redox inactive and, therefore, is unable to function as either a reductant or an inhibitor. Thus, mechanism-based inhibitors are divided into two types: type I, which undergoes hydroxylation prior to inactivation, and type II, which only requires hydrogen atom abstraction. A general mechanism for dopamine beta-monooxygenase inactivation is described, in which a common mechanistic radical intermediate is formed from both pathways.