A rapid gas chromatographic assay for determining oxyradical scavenging capacity of antioxidants and biological fluids.

Herein, we report a new, rapid,and reliable method for measuring the protective antioxidant potential of pure antioxidant solutions or biological tissues. Peroxyl radicals generated by thermal homolysis of 2,2'-azobis-amidinopropane (ABAP) cause the oxidation of alpha-keto-gamma-methiolbutyric acid (KMBA) to ethylene; ethylene formation is monitored by gas chromatographic analysis of head space from the reaction vessel. The partial inhibition of ethylene formation in the presence of antioxidants that compete with KMBA for oxyradicals is the basis of the Total Oxyradical Scavenging Capacity Assay (TOSCA).

Leuprolide, a gonadotropin-releasing hormone agonist, reestablishes spermatogenesis after 2,5-hexanedione-induced irreversible testicular injury in the rat, resulting in normalized stem cell factor expression.

2,5-Hexanedione (2,5-HD) exposure in the rat produces irreversible testicular atrophy, a model of human male infertility that can be used for mechanistic and therapeutic studies. Following testicular injury by 2,5-HD, stem cell factor (SCF), a Sertoli cell-derived growth factor that binds the c-kit receptor on spermatogonia, is altered in its expression, changing from predominantly membrane SCF to predominantly soluble SCF. The goals of this study were 2-fold: first, evaluate leuprolide, a GnRH agonist, as a therapy for 2,5-HD-induced testicular atrophy, and second, examine changes in SCF expression during testicular injury and following recovery from injury.

Adenovirus-mediated gene transfer to rat testis in vivo.

To study transgene expression in the adult rat testis in vivo, an adenovirus vector carrying a lacZ transgene with a nuclear localization signal was used as a marker. The adenovirus vector was first tested on rat Sertoli cell-germ cell cocultures in vitro; it efficiently mediated transgene expression in Sertoli cells but not germ cells. This vector was then delivered to the interstitial compartment of adult rat testes by intratesticular injection, resulting in Leydig cells expressing the transgene.

In vivo analysis of DNA-protein interactions on the human erythropoietin enhancer.

The erythropoietin (EPO) gene is one of the best examples of a mammalian gene controlled by oxygen tension. The DNA elements responsible for hypoxia-induced transcription consist of a short region of the proximal promoter and a <50-bp 3' enhancer. The elements act cooperatively to increase the transcriptional initiation rate approximately 100-fold in response to low oxygen tension in Hep3B cells.

Exogenous stem cell factor (SCF) compensates for altered endogenous SCF expression in 2,5-hexanedione-induced testicular atrophy in rats.

2,5-Hexanedione (2,5-HD) is a Sertoli cell toxicant that causes irreversible testicular atrophy in rats. After toxicant exposure, only Sertoli cells, stem cells, and a few committed type A spermatogonia remain in the seminiferous epithelium. A majority of the stem cell progeny differentiate into type A spermatogonia, but then, rather than continuing to differentiate, undergo apoptosis.

Fate of germ cells in 2,5-hexanedione-induced testicular injury. I. Apoptosis is the mechanism of germ cell death.

2,5-hexanedione (2,5-HD) is a Sertoli cell toxicant which causes germ cell loss and testicular atrophy in the rat. The mechanism of germ cell death over the course of 2,5-HD treatment is not known nor is the reason why residual germ cells do not repopulate the seminiferous epithelium following toxicant withdrawal. In the current study, the role of apoptosis in germ cell loss was studied.

Pulmonary activation and toxicity of cyclopentadienyl manganese tricarbonyl.

Cyclopentadienyl manganese tricarbonyl (CMT) produces acute pulmonary injury following cytochrome P450 mixed-function oxidase (CYP450) activation. The current studies were designed to characterize the role of hepatic and/or pulmonary CMT activation and the subsequent pneumotoxicity of this compound following subcutaneous injection in the male Sprague-Dawley rat. Both pulmonary and hepatic tissues were capable of CYP450-dependent CMT metabolism in vitro.

Molecular cloning, chromosomal mapping, and characterization of the human cardiac-specific homeobox gene hCsx.

Background: Csx/Nkx2.5, a murine nonclustered homeobox gene expressed primarily in the heart, has significant sequence similarity to the Drosophila tinman gene. Tinman is essential for heart and gut formation in Drosophila.

Development and implementation of a perinatal education consortium.

The North Puget Sound Perinatal Education Consortium provides a collaborative framework for planning, implementing, and evaluating a basic orientation program for newly hired and cross-trained perinatal nurses. The program currently provides 9 days of prenatal, labor and delivery, mother-infant couplet, and neonatal didactic orientation at no cost to registrants from participating hospitals. Consortium hospitals supply instruction time and additional services to support this high-quality, regional, perinatal orientation program.

Effect of lymphokines on 35sulfate uptake by the glomerular basement membrane.

We have previously shown that lymphocytes from idiopathic minimal-lesion nephrotic patients produce a lymphokine (supernatant factor) that increases the 35sulfate uptake in the glomerular basement membrane (GBM). The purpose of this report was to further characterize the supernatant factor by studying the effects of interleukins (IL) 2-4, 6, and 8, granulocyte-macrophage colony stimulating factor, and tumor necrosis factor on the 35sulfate incorporation by rat glomeruli in vitro. A significant increase in GBM 35sulfate uptake was only seen when the glomeruli were cultured with the addition of IL-8 as compared with control cultures: 10.

A 57-year-old man with bruising and joint pain.

The differential diagnosis for a 57-year-old man with joint pain, bruising, and diarrhea is discussed in the setting of a Clinical Pathological Conference at Louisiana State University Medical Center in Shreveport, La. The pathophysiology and etiology of malabsorptive states are presented.

C9-mediated killing of bacterial cells by transferred C5b-8 complexes: transferred C5b-9 complexes are nonbactericidal.

The formation of the C5b-9 complex on the outer membrane of complement-sensitive cells of Escherichia coli results in inhibition of inner membrane function and the death of the cell. Cells bearing a precursor of the C5b-9 site, the C5b-8 complex, suffer no loss in viability. Antibiotic-sensitive, complement-sensitive donor cells bearing precursor C5b-8 complexes were incubated with equal numbers of antibiotic-resistant, complement-sensitive acceptor cells that had not been exposed to a complement source.

Effects of m-xylene on rat nasal cytochrome P450 mixed function oxidase activities.

The effect of m-xylene on the rat nasal cytochrome P450 (P450) mixed function oxidase system was analyzed in vitro utilizing microsomes isolated 2, 12, and 24 h following intraperitoneal administration of this solvent in vivo. For comparative purposes, pulmonary and hepatic activities were also measured. Benzyloxyresorufin O-deethylation (BROD) and ethoxyresorufin O-deethylation (EROD), catalytic activities linked with P450 isozymes IIB1 and IA1, respectively, were inhibited in nasal tissue at all times following m-xylene administration.

A physiologically based pharmacokinetic model for nasal uptake and metabolism of nonreactive vapors.

A PB-PK model has been developed for nasal nonreactive vapor uptake in the F344 rat which incorporates nasal enzyme distribution as well as nasal airflow patterns. Nasal tissue is separated into respiratory and olfactory mucosal areas with each area containing mucus, epithelial, and submucosal compartments. Metabolic activities are distributed among the compartments in accordance with published histochemical data, and intercompartmental transfer rate constants are based on molecular diffusivity.

Regulation of the erythropoietin gene.

Erythropoietin (Epo), the hormone that stimulates red blood cell production, is induced by hypoxia. We have utilized the human hepatoma cell line, Hep3B, to investigate the regulation of the Epo gene. We present evidence that the oxygen sensor in Hep3B cells is a heme protein.

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Hypoxic induction of the human erythropoietin gene: cooperation between the promoter and enhancer, each of which contains steroid receptor response elements.

Transcription of the human erythropoietin (Epo) gene is stimulated by exposure to hypoxia and/or cobalt in whole animals and in Hep3B cells. We have systematically investigated the promoter and 3' enhancer elements necessary for this induction by transient transfection of Hep3B cells. We define a promoter region of 53 bp and an enhancer region of 43 bp that confer hypoxia and cobalt inducibility.

Clonality in myeloproliferative disorders.

The myeloproliferative disorders are a group of hematologic diseases that are believed to arise from somatic mutations in an early hematopoietic stem cell. This statement is based on the demonstration of monoclonal involvement of terminally differentiated myeloid and lymphoid elements. The techniques for establishing clonal derivation of cells are discussed and the application of these techniques to myeloproliferative diseases is reviewed.

Upper respiratory tract deposition of inspired acetaldehyde.

Inhalation exposure of rodents to high concentrations of acetaldehyde produces lesions in the upper respiratory tract (URT, all regions of the respiratory tract anterior to and including the larynx). Information on the inhalation dosimetric relationships for this vapor are needed for a comprehensive understanding of its inhalation toxicity. Toward this end, uptake of acetaldehyde was measured in the surgically isolated URT of the urethane-anesthetized male F344 rat under unidirectional (50, 100, 200, or 300 ml/min) and cyclic (100 ml/min) flow conditions at inspired concentrations of 1, 10, 100, or 1000 ppm.

Development of Lhermitte's sign after bone marrow transplantation.

The authors observed Lhermitte's sign in four patients after bone marrow transplantation (BMT) for hematologic malignancies. Three patients had acute myelogenous leukemia (AML), and one had chronic myelogenous leukemia. Before BMT, the patients with AML received daunorubicin, cytosine arabinoside and etoposide, whereas the patient with chronic myelogenous leukemia received hydroxyurea.

Clonality in myeloproliferative disorders: analysis by means of the polymerase chain reaction.

The myeloproliferative syndromes are acquired disorders of hematopoiesis that provide insights into the transition from somatic cell mutation to neoplasia. The clonal origin of specific blood cells can be assessed in patients with X chromosome-linked polymorphisms, taking advantage of random inactivation of the X chromosome. We have adapted the PCR for determination of clonality on as few as 100 cells, including individual colonies grown in culture.

Clonality in acquired hematologic disorders.

Clonal populations of cells can be identified by a number of different independent approaches, including analyses of karyotype, gene rearrangements, deletions or point mutations, X-linked polymorphisms, and integration of virus into the genome. Assessment of clonality has yielded valuable and surprising clues to the pathogenesis of acquired hematologic disorders. In the myeloproliferative states, clonal expansion of a mutated pluripotent stem cell can induce production of blood cells having a normal phenotype.

Analysis of cytokine mRNA and DNA: detection and quantitation by competitive polymerase chain reaction.

The expression of two cytokines, granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 3 (IL-3), has been investigated in MLA-144 cells before and after induction with phorbol 12-myristate 13-acetate. We describe an adaptation of the polymerase chain reaction (PCR) for highly accurate quantitation of mRNA or DNA from a small number of cells. Aliquots of the PCR mixture containing cDNA copies of the RNA to be assayed were added to serial dilutions of a competitor DNA fragment that differed from the cDNA of interest by having either a small intron or a mutated internal restriction enzyme site.