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Biomedical subjects

J S Nye

Publications and source records attributed to J S Nye.

12 recordsLinked to original sources
PubMed1994

Human cerebral cortical cell lines from patients with unilateral megalencephaly and Rasmussen's encephalitis.

Continuous cerebral cortical cell lines have been developed from two patients, an 11-month-old with unilateral megalencephaly and a seven-year-old with Rasmussen's encephalitis, designated HCN-1 and HCN-2, respectively. The two cell lines stain for neuronal markers such as neurofilament and neuron-specific enolase but not for non-neuronal markers such as glial fibrillary acidic protein and S-100 protein. In the presence of appropriate growth factors, the cells extend long, branched processes resembling neurons. Differentiation of HCN-1 cells can be induced with nerve growth factor, dibutyryl cyclic AMP and isobutylmethylxanthine, while for HCN-2 cells nerve growth factor, isobutylmethylxanthine and the phorbol ester 12-O-tetradecaoylphorbol-13-acetate are most effective. Immunohistochemical staining of both differentiated cell lines reveals intense staining for GABA, glutamate, somatostatin, cholecystokinin-8 and methionine enkephalin. Two human cortical neuronal cell lines have been developed which represent neuronal precursors. These cell lines propagate in culture and are capable of differentiating upon the addition of a variety of growth factors and chemical agents. These cell lines should prove to be useful models for the study of in vitro neuronal processes.

Blotting, Western↗
PubMed1990

Human cortical neuronal cell line: establishment from a patient with unilateral megalencephaly.

A cell line has been established in continuous culture of human cerebral cortical neurons obtained from a patient with unilateral megalencephaly, a disorder associated with continued proliferation of immature neuronal cells. When differentiated in the presence of nerve growth factor, 1-isobutyl-3-methylxanthine, and dibutyryl adenosine 3',5'-monophosphate (cAMP), the cells display mature neuronal morphology with numerous long, extensively branched processes with spines and varicosities. The cells stain positively for neurofilament protein and neuron-specific enolase (selective neuronal markers) but are negative for glial markers, such as glial fibrillary acidic protein, S-100, and myelin basic protein. The cells also stain positively for the neurotransmitters gamma-aminobutyric acid (GABA), glutamate, somatostatin, cholecystokinin-8, and vasoactive intestinal polypeptide. These cells may facilitate characterization of neurons in the human central nervous system.

1-Methyl-3-isobutylxanthine↗
PubMed1989

High-affinity cannabinoid binding site: regulation by ions, ascorbic acid, and nucleotides.

The high-affinity cannabinoid site in rat brain is an integral component of brain membranes that recognizes cannabinoids with inhibitory constants (Ki) in the nanomolar range. To clarify its physiological role, we studied the regulation of [3H]5'-trimethylammonium delta 8-tetrahydrocannabinol ([3H]TMA) binding. The site is inhibited by heavy metal ions, such as La3+, at low micromolar concentrations; divalent cations, such as Ca2+ and Mg2+, inhibit [3H]TMA binding, though at somewhat higher concentrations. In contrast, [3H]TMA binding is stimulated by Fe2+, Cu2+, and Hg2+ ions. Ascorbic acid and its analogs are also stimulators of cannabinoid binding at low micromolar concentrations. Stimulation of [3H]TMA binding by ascorbate or ions is dependent upon molecular oxygen, but is not inhibited by metabolic poisons. Metabolically stable nucleoside triphosphate analogs enhance [3H]TMA binding by different mechanisms, with hydrolysis of a high-energy phosphate bond apparently requisite for these influences. These results suggest that the cannabinoid binding site is associated with a nucleotide-utilizing protein possessing multiple regulatory subsites.

Adenosine Triphosphate↗
PubMed1988

Drug solubilizers to aid pharmacologists: amorphous cyclodextrin derivatives.

Conversion of crystalline alpha-, beta-, or gamma-cyclodextrins into amorphous mixtures of water soluble derivatives yields non-toxic solubilizers which dissolve drugs through the formation of inclusion complexes. From these types of compounds 2-hydroxypropyl ethers of cyclodextrins have presently been investigated and the ranges for the safe use in working with (a) receptor binding assays on membrane preparations, (b) cells in vitro, and (c) parenteral use in mice were established for these compounds. The drugs which were investigated were dissolved in amounts linearly proportionate to the concentration of the solubilizers used and did not precipitate upon dilution by aqueous media. These solubilizers may considerably facilitate pharmacological evaluation of new, water insoluble potential drugs.

Adenylyl Cyclases↗
PubMed1988

Myelin basic protein is an endogenous inhibitor of the high-affinity cannabinoid binding site in brain.

Radioligand binding studies with the water-soluble cannabinoid [3H]5'-trimethylammonium delta 8-tetrahydrocannabinol ([3H]TMA) have revealed a saturable high-affinity site in brain that is specific for cannabinoids. To determine whether endogenous compounds of brain might act upon the site physiologically, we sought inhibitors in extracts of brain. An endogenous inhibitor has been purified to homogeneity by acid extraction of rat brain followed by adsorption to a reverse-phase matrix and gel filtration chromatography. The purified inhibitor has a subunit molecular mass of 14,500 daltons by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Inhibition of [3H]TMA binding by the purified inhibitor occurs with a Ki of about 4 nM in a noncompetitive manner. The molecular weight, abundance, and extraction properties are the same as a species of myelin basic protein (MBP). The MBPs of rat, rabbit, pig, and cow also inhibit [3H]TMA binding noncompetitively with similar potencies. The purified inhibitor comigrates with rat MBP-small form on SDS-PAGE, has a similar amino acid composition, and is recognized by antibody directed against MBP. Studies of fragments of rabbit MBP suggest that the determinants of affinity for the [3H]TMA site are contained primarily within the C-terminal half of the rabbit MBP. Synthetic polycationic peptides such as polylysine and polyarginine mimic the effects of MBP, suggesting that the high-affinity cannabinoid binding site recognizes large polycations. The identification of the endogenous inhibitor of [3H]TMA binding as MBP suggests that MBP interacts physiologically with the high-affinity cannabinoid site.

Amino Acids↗
PubMed1987

Porphyrins are endogenous ligands for the mitochondrial (peripheral-type) benzodiazepine receptor.

"Peripheral-type" benzodiazepine receptors are localized to the outer mitochondrial membrane. We have identified potent competitive inhibitors of these receptors and purified them from human blood and from several rat organs. TLC analysis of the purified inhibitor from erythrocytes displays a single peak of inhibitory activity with an absorbance spectrum identical to hemin. All of the inhibitory activity in extracts of several tissues can be accounted for by their porphyrin and metalloporphyrin content. Pure hemin and protoporphyrin IX competitively inhibit mitochondrial benzodiazepine binding with Ki values of 41 and 15 nM, respectively, and are less active by a factor of 1000 at central-type benzodiazepine receptors. Thus, porphyrins appear to be endogenous ligands for mitochondrial benzodiazepine receptors.

Animals↗
PubMed1985

High-affinity cannabinoid binding sites in brain membranes labeled with [3H]-5'-trimethylammonium delta 8-tetrahydrocannabinol.

The binding of [3H]-5'-trimethylammonium delta 8-tetrahydrocannabinol (THC) [( 3H]TMA) to rat neuronal membranes was studied. TMA is a positively charged analog of delta 8THC modified on the 5' carbon, a portion of the molecule not important for its psychoactivity. Unlabeled TMA inhibits field-stimulated contractions of the guinea-pig ileum (IC50 = 1 microM) in the same presynaptic manner as delta 9THC. [3H]TMA binds saturably and reversibly to brain membranes with high affinity (KD = 89 nM) to apparently one class of site (Hill coefficient, 1.1). Highest binding site density occurs in the brain, but several peripheral organs also display specific binding. Detergent solubilizes the sites without affecting their pharmacological properties. Molecular sieve chromatography reveals a bimodal peak of [3H]TMA binding activity of approximately 60,000 daltons apparent molecular weight. delta 9THC competitively inhibits [3H]TMA binding potently (Ki = 27 nM) and stereoselectively. For some cannabinoids potency in behavioral and physiological tests parallels their affinity for the [3H]TMA binding site. However, several nonpsychotropic cannabinoids are active at the binding site.

Animals↗
PubMed1982

Mutant lambda phage repressor with a specific defect in its positive control function.

The lambda phage repressor is both a positive and a negative regulator of gene transcription. We describe a mutant lambda phage repressor that has specifically lost its activator function. The mutant binds to the lambda phage operator sites and represses the lambda phage promoters PR and PL. However, it fails to stimulate transcription from the promoter PRM. The mutation lies in that portion of repressor--namely, the amino-terminal domain--that has been shown [Sauer, R. T., Pabo, C. O., Meyer, B. J., Ptashne, M. & Backman, K. C. (1979) Nature (London) 279, 396-400] to mediate stimulation of PRM. We suggest that the mutation has altered that region of repressor which, in the wild-type, contacts RNA polymerase to activate transcription from PRM.

Bacteriophage lambda↗
PubMed1979

Induction of sarcomas in nude mice by implantation of Syrian hamster fetal cells exposed in vitro to nickel subsulfide.

In vitro exposure of Syrian hamster fetal cells to nickel subsulfide (alpha Ni3S2) yielded positive colony assays for morphological transformation. A dose-response relationship was found between the concentration of alpha Ni3S2 and the incidence of morphological transformation. Exposures of alpha Ni3S2 induced morphological transformation at concentrations (0.1 or 1.0 microgram/ml of culture medium) which did not impair cell plating efficiency. Nickel monosulfide (NiS) did not induce morphological transformation of Syrian hamster fetal cells under the same conditions. Clones of alpha Ni3S2-transformed cells were able to grow in soft agar medium and demonstrated increased basal and induced activities of ornithine decarboxylase. Undifferentiated sarcomas developed in 26 of 27 nude mice at the site of s.c. injection of clones of alpha Ni3S2-transformed cells. No tumors developed in 19 control nude mice which were given s.c. injections of nontransformed Syrian hamster fetal cells which had not been exposed to alpha Ni3S2. This study demonstrates that fetal cells which undergo transformation following exposure to alpha Ni3S2 are capable of producing malignant tumors in nude mice.

Animals↗