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Biomedical subjects

A Peterson

Publications and source records attributed to A Peterson.

At least 91 records · Page 5Linked to original sources
PubMed1988

Pituitary-specific expression and glucocorticoid regulation of a proopiomelanocortin fusion gene in transgenic mice.

The product of a single gene encoding proopiomelanocortin (POMC) is differentially processed to produce corticotropin and alpha-melanotropin in anterior and intermediate pituitary cells, respectively. Hormonal control of POMC gene transcription and of corticotropin or alpha-melanotropin release is also tissue-specific; for example, glucocorticoids specifically inhibit anterior but not intermediate pituitary POMC transcription. Outside the pituitary gland, very low levels of POMC mRNAs are present in brain, testes, ovaries, and placenta. We have used transgenic mice to identify POMC 5' flanking sequences that are sufficient for tissue-specific expression and glucocorticoid regulation in anterior and intermediate pituitary cells. Three lines of transgenic mice were established, each carrying 50-75 copies (per cell) of a chimeric rPOMCneo gene constituted of rat POMC promoter sequences and of bacterial neomycin-resistance coding sequence. High levels of rPOMCneo transcripts were detected in pituitaries of mice from all three lineages. In situ hybridization revealed that the ratio of intermediate to anterior pituitary transcripts was similar for the transgene and endogenous POMC mRNA. rPOMCneo transcripts were not detected in any other tissue except at very low levels in the testes in two transgenic lines. Endogenous mouse POMC mRNA increased in response to depletion of plasma glucocorticoids (adrenalectomy) and decreased after glucocorticoid treatment; rPOMCneo transcripts were altered to the same extent by these treatments in all three lines. Intermediate pituitary and testicular rPOMCneo transgene expression was not altered by these treatments. Thus, no more than 769 base pairs of the rat POMC promoter are required for pituitary-specific expression and for specific glucocorticoid inhibition of the POMC gene in the anterior pituitary.

Animals↗
PubMed1988

Expression of the T-cell surface molecule CD2 and an epitope-loss CD2 mutant to define the role of lymphocyte function-associated antigen 3 (LFA-3) in T-cell activation.

To define the role of the CD2-lymphocyte function-associated antigen 3 (LFA-3) interaction in T-cell activation, we have expressed a cDNA encoding the human CD2 molecule in a murine antigen-specific T-cell hybridoma. Expression of the CD2 molecule greatly enhances T-cell responsiveness to antigen; this enhancement is inhibited by anti-CD2 and anti-LFA-3 monoclonal antibodies (mAbs). CD2+ hybridomas produce interleukin 2 in response to combinations of anti-CD2 mAbs 9.6 and 9-1 and, in the presence of mAb 9-1, to sheep erythrocytes or to the LFA-3 antigen. Furthermore, hybridomas expressing a mutant CD2 molecule that has lost mAb 9.6 binding do not exhibit the enhanced response to antigen or the ability to respond to LFA-3 plus mAb 9-1, but these hybridomas retain the ability to respond to combinations of anti-CD2 mAbs. The role of the CD2-LFA-3 interaction in T-cell activation and the potential for other physiologic ligands for CD2 are discussed.

Animals↗
PubMed1987

Expression and function of CD8 in a murine T cell hybridoma.

In general, the human CD8 molecule is expressed on T cells specific for HLA class I molecules. Studies designed to delineate the function and to define the ligand of the CD8 molecule have been complicated by the fact that the presumptive ligand for CD8 is on the HLA class I molecule, the same molecule encoding the ligand for the antigen-specific T cell receptor. The ability to express genes in cells other than their natural host has produced a new technology with which to approach CD8 functional studies. The insertion of a cDNA clone for CD8 in a defective retroviral vector has allowed the transfer of CD8 by infection with the resulting defective retrovirus. CD8 was then expressed in an HLA class II-specific T cell, thus separating the ligand requirements of the TCR and CD8. By this approach, the human CD8 molecule was expressed in a murine T cell hybridoma specific for human class II antigens. The resulting CD8+ hybridomas demonstrated a 10-fold increase in IL-2 production over the parent cell line when stimulated with JY, a human B lymphoblastoid cell line expressing both class I and II HLA antigens, demonstrating that expression of CD8 increases T cell activation. mAbs directed against the CD8 molecule inhibited the response of CD8+ hybridomas to JY, supporting the conclusion that the CD8 molecule was fractional. The role of CD8 as a receptor for class I MHC antigens was addressed by stimulation with a cell line expressing HLA-DR antigens, but lacking the expression of HLA class I antigens (Daudi). Stimulation of the CD8+ hybridomas by Daudi did not result in increased IL-2 production. The response to Daudi was unaltered by the addition of anti-CD8 mAb, in contrast to the ability of anti-CD8 mAb to block JY stimulation. Furthermore, mAbs directed against the class I antigens present on JY cells were able to block the enhanced response of the CD8+ hybridomas to JY. These data support the hypothesis that HLA class I molecules are the ligands involved in the CD8-dependent enhancement of T cell activation.

Antibodies, Monoclonal↗
PubMed1987

Expression and assembly of a human neurofilament protein in transgenic mice provide a novel neuronal marking system.

To investigate the regulation of neurofilament gene expression, we have generated several lines of transgenic mice carrying multiple copies of a cloned human neurofilament (NF-L) gene. We show that a 21.5-kb DNA fragment including the human NF-L gene contains essential information for correct expression in nervous tissue of transgenic mice. The integrated genes are arranged in multiple tandem arrays, but the extent of transgene expression does not correlate with copy number nor does it influence the expression of the endogenous neurofilament genes. However, the levels of human NF-L protein recovered in neurofilament preparations from brains of transgenic mice correlate directly with the relative abundance of human NF-L mRNA detected in each line. There is an apparent delay in the accumulation of human NF-L protein during development, as determined by immunoblotting with a human-specific monoclonal antibody. Finally, immunohistochemical localization of the human NF-L protein results in the specific staining of neurons and their processes in transgenic mice.

Age Factors↗
PubMed1986

Oxidative metabolism in cultured astroglial cells from rat brain.

Growth, morphology, glutamine synthetase activity, cytochrome C oxidase activity and respiratory activity of rat brain cultures enriched in astrocytes were studied during four weeks in culture. Two different polarographic methods were used for measurement of respiratory activity: one newly developed perfusion method leaving the cellular monolayer morphologically intact and still attached to the culture dish, and one traditional stirring method involving the removal of cells from the culture vessel. Regardless of the method used, a stable respiratory activity was registrated throughout the four weeks of culturing. Also the cytochrome C oxidase activity remained unchanged. In perfusion all absolute values for respiration were found to be higher than those obtained with the stirring method. The use of the stirring technique resulted in a doubling of oxygen consumption upon succinate addition. No such effect was seen in perfusion. It can thus be concluded that the removal of cultured astrocytic cells from their substratum alters their respiratory activity and their response to added substrates.

Animals↗
PubMed1986

Depletion of T cells from bone marrow for allogeneic transplantation: method for treatment of bone marrow in bulk.

T lymphocytes were depleted from donor marrow for 23 patients undergoing allogeneic bone marrow transplantation using an anti-T-cell antibody, CT-2, and complement. The methodology is described in detail for in vitro depletion of large quantities of bone marrow. The extent of T-lymphocyte depletion using various T-cell markers, the percent of marrow lost in the processing and quantity of antibody, and complement needed are presented. These techniques for in vitro T-lymphocyte depletion were reproducible and did result in an average final yield of 47% of the harvested donor marrow.

Antibodies, Monoclonal↗
PubMed1985

Use of a perfusion technique for measurements of respiratory activity in cultured cells.

A method for measuring respiratory activity in anchorage-dependent cultured cells has been developed. This method is based on a technique that permits the perfusion of standard plastic culture dishes with attached cells. Basal respiratory activities were studied in two continuous cell lines of neural origin, neuroblastoma C1300 clone 41A3 and glioma 138MG. As compared to traditional measurements on detached cells, a fourfold increase in value was obtained. Investigations on membrane permeability suggested that the observed difference could be attributed to alterations in cell membrane integrity. Pretreatment with dibutyryl cyclic AMP, known to induce a morphological and biochemical differentiation in C1300 and 138MG cells, caused in both cell lines an enhanced respiration.

Animals↗
PubMed1985

Effects of dissolved carbon monoxide on the respiratory activity of perfused neuronal and muscle cell cultures.

In order to address the question of whether small amounts of dissolved CO may inhibit cellular respiration, cultured mouse neuroblastoma cells and primary cultures of chick neurons, rat astrocytes and chick skeletal muscle and heart cells were exposed to CO containing buffer solutions in a closed perfusion system. Oxygen uptake was measured simultaneously with two polarographic oxygen electrodes as the difference in partial pressure of oxygen between the inlet and outlet of the perfusion chamber. After registration of the basal respiratory activity, perfusion solutions containing 5 ul 02/ml were bubbled with CO or N2 at a rate of 200 ml/min for 120 sec. By this procedure the partial pressure of 02 was decreased to reach a value of about 50% of the initial 02 content for both gases. Perfusion was then continued for 30 min at a rate of 0.5 ml/min. The respiratory activity of all the perfused cell cultures, except chick neurons, was found to be inhibited (13-29%) by perfusion solutions bubbled for 120 sec with CO as compared to N2 controls. Of the cells from the nervous system, astrocytes were more sensitive than neurons. Apparently, small amounts of dissolved CO can inhibit cellular respiration in the presence of a physiologically adequate amount of oxygen.

Animals↗
PubMed1985

Mismatched bone marrow transplantation in children with hematologic malignancy using T lymphocyte depleted bone marrow.

Twenty children with hematologic malignancies were treated with bone marrow transplantation using histoincompatible donor marrow depleted of T lymphocytes. CT-2 and complement were used to deplete the T lymphocytes. Engraftment was not a major problem in those receiving increased pre- and posttransplant immunosuppression. Five of the 20 children are currently alive and disease-free. Complications in the other children included engraftment problems, cyclosporin toxicity, infections, and bleeding. However, for most children, durable engraftment was seen. Thus, the nonavailability of histocompatible donors is no longer a contraindication to allogeneic bone marrow transplantation for those at high risk of relapse.

Adolescent↗
PubMed1984

Disease expression in +-/+- ----mdg/mdg mouse chimeras: evidence for an extramuscular component in the pathogenesis of both dysgenic abnormal diaphragm innervation and skeletal muscle 16 S acetylcholinesterase deficiency.

Homozygous mdg/mdg mice die at birth and express a syndrome of abnormalities, the most striking of which is a gross failure of skeletal muscle development. Recently, additional abnormalities in the development of nerve-muscle relationships have been recognized; in particular, on muscle fibers within the diaphragm, motor end plates are inappropriately dispersed and, in all muscles, there is a paucity of the 16 S form of acetylcholinesterase (AChE). These abnormalities could result entirely as secondary consequences of the primary muscle defect or from expression of the mdg defect in additional cell types, e.g., motor neurons. To determine if the muscle genotype alone is responsible for these defects in dysgenic mice, chimeras composed of both dysgenic and normal cells have been investigated. Different glucosephosphate isomerase variants existed in the mdg/mdg and normal cells comprising these chimeras and the mutant, normal, or mosaic genotypes of chimera diaphragm and skeletal muscle was estimated by measuring the relative proportions of each isozyme. In two chimeras, the diaphragm innervation pattern was revealed by AChE cytochemistry and in both, discrete regions of abnormally dispersed and normally restricted motor end-plate zones were observed. No correlation between these patterns of innervation and the assessed genotype of the muscle fibers existing in each area was observed. The relative 16 S AChE content in the limbs of four chimeras was found to range from 2.5 to 42.0%. Here also, no correlation between 16 S AChE content and the muscle genotype was observed. The results of these investigations are not consistent with a model of mdg/mdg pathogenesis in which only the skeletal muscle is primarily affected; an extramuscular deficiency responsible for at least part of the full mdg/mdg syndrome is therefore suggested.

Acetylcholinesterase↗
PubMed1984

On the application of cultured neuroblastoma cells in chemical toxicity screening.

The acute toxic action of a number of common chemicals was tested by their ability to cause detachment of cultured mouse neuroblastoma C1300 cells. A TD25 value was obtained by graphic estimation of the concentration needed to cause 25% of the total cell number to detach. These TD25 values were compared with LD50 values obtained from the literature, and they were found to correlate with a coefficient of 0.86. For six of the tested substances-diuron, butylated hydroxytoluene, benzidine, cyclophosphamide, Na2SeO3, and KCN-a very poor correlation was obtained. These diverging results could be ascribed to deficiencies in the neuroblastoma cell detachment test and emphasize the necessity for combined in vitro test procedures.

Animals↗
PubMed1983

P1 deficiency in shiverer myelin is expressed by Schwann cells in shiverer dystrophic normal mouse chimaera nerves.

The myelin basic protein (P1) deficiency in shiverer myelin is expressed in shiverer reversible normal mouse chimaera nerves. Chimaera examined with immunocytochemical techniques have revealed populations of both densely labeled and unreacting myelinated Schwann cells. Single axons can innervate both Schwann cell types, demonstrating that the expression of P1 in Schwann cell myelin is unrelated to the shiverer or normal genotype of the neuron. The coexistence of both Schwann cell types in single nerves indicates that multiple progenitor Schwann cells are allocated to developing nerves and the mosaic patterns expressed further suggest that such cells tend to proliferate relatively small coherent clones of Schwann cells.

Animals↗
PubMed1983

Growth and morphology of neuronal cell lines cultured in perfusion.

To optimize culture conditions and gain a more reliable culturing system for studies of metabolic properties of neuronal cells, a simplified perfusion chamber was developed. It consists of two parts: a perfusion block and a standard plastic culture dish. To confirm the suitability of this chamber for continuous culturing of anchorage-dependent cells, the growth and morphology of the four neuronal cell lines, glioma C6 and glioma 138MG, neuroblastoma C1300, clones N1E115 and N18 were followed for 4 d using both traditional and perfusion techniques. A marked increase in growth and a decrease in the degree of morphological differentiation were obtained with the latter technique compared to the former.

Animals↗
PubMed1983

Acute toxicity testing in cultures of mouse neuroblastoma cells.

Cultured mouse neuroblastoma cells (C1300) may be used as models for nerve cells since they have a number of properties in common with their normal counterparts in vivo. In order to test the possibility of using C1300 cells as alternative to experimental animals when testing for acute toxicity, cells (clone 41A3) were exposed to a number of common chemicals (CH3HgCl, CdCl2,HgCl2 ppDDT, n-butanol, benzene, dioxan, n-propanol, aceton and t-butanol). The toxic effect was quantified by measuring the degree of cell detachment in the cultures. The concentrations of chemicals that caused 25% of the total cell number to detach (TD25) were used for comparison with LD50 values. In spite of the very simplified situation in culture, where the toxicity of a substance is little or not at all influenced by factors like penetration, storage, metabolism and excretion a good correlation (corr. coeff. 0,98) was obtained between TD25 values and LD50 values. Good correlations between in vitro and in vivo tests have also been reported by others. One possible explanation to these findings could be simplified in vivo toxicokinetics of these substances when tested in high doses for general effects like animal death. If so, simple in vitro tests may be used for predicting acute toxicity of certain groups of substances.

Animals↗