PubMed Health⌕ Search

Biomedical subjects

J R Hoidal

Publications and source records attributed to J R Hoidal.

At least 55 records · Page 3Linked to original sources
PubMed1996

Effect of intracellular pH on ferret pulmonary arterial smooth muscle cell calcium homeostasis and pressure.

In this study, we investigated the role of Na+/H+ antiport in regulating cytosolic (intracellular) pH (pHi) in isolated and cultured ferret pulmonary arterial smooth muscle cells (PSMC). We also studied the effects of modulating pHi on the cytosolic (intracellular) calcium concentration ([Ca2+]i) in the PSMC and on the pulmonary arterial pressure (Ppa) of isolated ferret lungs. pHi was modulated by the NH4Cl washout method. To eliminate the contribution of Cl-/HCO3- exchangers, the PSMC and isolated lungs were perfused in HCO3- free buffer. Blocking the Na+/H+ antiporter decreased baseline pHi and prevented the recovery from NH4Cl washout-induced intracellular acidosis. Intracellular alkalinization caused an initial transient increase in both [Ca2+]i and Ppa that were dependent on extracellular Ca2+ entry. Maintaining cytosolic alkalinization caused another increase in Ppa that was not associated with an increase in [Ca2+]i. Intracellular acidosis also caused an increase in [Ca2+]i and Ppa. The cytosolic acidosis-induced increase in [Ca2+]i and Ppa were mediated by both extracellular Ca2+ influx and release of stored intracellular Ca2+. Cytosolic acidosis also appears to have a direct effect on the smooth muscle contractile elements. Both cytosolic alkalosis and acidosis increased vascular reactivity.

Acidosis↗
PubMed1996

Potassium currents in cultured human pulmonary arterial smooth muscle cells.

In this study, using whole cell and single-channel configurations of the patch-clamp technique, we characterized K+ currents (IK) in cultured human pulmonary arterial smooth muscle cells. The net whole cell outward membrane current (IKo) was activated at potentials positive to -60 mV. One component of IKo, IK(dr), was inhibited by 4-aminopyridine (4-AP) and high concentrations of tetraethylammonium (TEA) but was Ca2+ and charybdotoxin (CTX) insensitive. The other component of IKo, IK(Ca), was voltage and Ca2+ dependent and was inhibited by CTX and low concentrations of TEA. Activation of IKo in single-channel recordings was voltage dependent and demonstrated a high-conductance channel (245 +/- 2 pS) that was Ca2+ and CTX sensitive [IK(Ca)] and a low-conductance channel (109 +/- 2 pS) that was inhibited by 4-AP [IK(dr)] but was insensitive to low concentrations of TEA or to an increase in intracellular [Ca2+]. In isolated pulmonary arterial rings, TEA and 4-AP caused an additive increase in arterial tension. To our knowledge these data provide the first characterization of the IK in human pulmonary arterial smooth muscle cells and indicate that IK(Ca) and IK(dr) play an important role in maintaining pulmonary vascular tone. The data confirm previous observations in pulmonary smooth muscle cells of animal models.

Calcium↗
PubMed1996

Regulation of Ca(2+)-activated K+ channels in pulmonary vascular smooth muscle cells: role of nitric oxide.

Nitric oxide (NO.) is believed to mediate nitrovasodilators and acetylcholine-induced vasodilatation via increasing intracellular guanosine 3',5'-cyclic monophosphate (cGMP) levels. The cellular mechanisms involved in No.-mediated pulmonary vasodilatation are complex and include membrane hyperpolarization. Using the patch-clamp technique in cell-attached and inside-out configurations, we examined the effect of NO. gas, 3-morpholinosydnomimine hydrochloride (SIN-1), and perfusate from ACh-stimulated human pulmonary arterial endothelial cells, or endothelium-derived relaxing factors (EDRF), on the Ca(2+)-dependent K+ (KCa) channels in isolated cultured human pulmonary arterial smooth muscle cells (HPSMC). NO., SIN-1, and EDRF caused similar increases in KCa channel activity. Inhibiting cGMP generation with methylene blue or inhibiting the effect(s) of cGMP with the cGMP antagonist 8-bromoguanosine 3',5'-cyclic monophosphorothioate Rp isomer Rp-cGMPS prevented the NO.- and SIN-1-mediated activation of KCa channels, respectively. Treating the human pulmonary arterial endothelial cells with methylene blue blocked the EDRF-mediated activation of KCa channels in HPSMC. The cGMP analogue 8-bromo-cGMP increased KCa channel activity in intact cells and in excised inside-out HPSMC membrane patches. In the presence of cGMP and ATP, the alpha-isozyme of the cGMP-dependent protein kinase (I alpha-cGMP-PK) significantly increased KCa channel activity, and the channel activation was further increased on addition of the protein phosphatase inhibitors okadaic acid and calyculin A. Furthermore, the cGMP-mediated KCa channel activation was reduced by the cyclic nucleotide-dependent protein kinase inhibitor N-[2-methylamino)ethyl]-5-isoquinlinesulfonamide (H-8). Thus, in HPSMC, the mechanism of NO.- and native EDRF-induced KCa channel activation appears to be mediated via cGMP-I alpha-cGMP-PK phosphorylation of KCa channels.

Calcium↗
PubMed1995

The Gabriel-Colman rearrangement in biological systems: design, synthesis and biological evaluation of phthalimide and saccharin derivatives as potential mechanism-based inhibitors of human leukocyte elastase, cathepsin G and proteinase 3.

The results of a structure-activity relationship study focusing on the interaction of a series of phthalimide and saccharin derivatives with leukocyte elastase, cathepsin G and proteinase 3 are described. The phthalimide derivatives were found to be inactive while some of the saccharin derivatives were found to be fair inhibitors of these enzymes.

Cathepsin G↗
PubMed1995

Inhaled nitric oxide prevents the increase in pulmonary vascular permeability caused by hydrogen peroxide.

Given the interest in using inhaled nitric oxide (NO.) to treat acute lung injury and the importance of oxygen radicals in its pathogenesis, we studied the effects, in buffer-perfused isolated rabbit lungs, of inhaled NO. (24 ppm) on the injury caused by generating hydrogen peroxide with glucose and glucose oxidase (GOX). Experiments were performed at a constant pulmonary arterial pressure. GOX substantially augmented vascular permeability, as demonstrated by an increase in the lung-to-perfusate 125I-labeled albumin ratio, lavage-to-perfusate 125I-albumin ratio, wet-to-dry lung weight ratio, and pulmonary vascular filtration coefficient. Lungs treated with inhaled NO. before perfusion with GOX had lung-to-perfusate and lavage-to-perfusate 125I-albumin ratios that were not significantly different from control values and intermediate between the control and GOX groups. Inhaled NO. also prevented the increase in wet-to-dry lung weight ratio and pulmonary vascular filtration coefficient caused by GOX.. Thus inhaled NO. substantially reduced in the isolated lung the increase in pulmonary vascular permeability produced by the intravascular generation of hydrogen peroxide.

Administration, Inhalation↗
PubMed1995

Inhibitory properties of recombinant human monocyte/neutrophil elastase inhibitor.

Human monocyte/neutrophil elastase inhibitor (HEI) is a serpin superfamily protein that rapidly and irreversibly inhibits neutrophil and pancreatic elastase. We generated a recombinant baculovirus that supports production of HEI in insect cells at approximately 400 times the level in monocytes. Recombinant HEI was found to be indistinguishable from monocyte HEI in its physicochemical properties and ability to inhibit neutrophil elastase and pancreatic elastase. The recombinant protein was used to test for additional functions. HEI was shown to inhibit proteinase 3, an important neutrophil inflammatory protease, by the classical serpin mechanism of forming a covalent protease-protease inhibitor complex. Preliminary evidence suggests that HEI also inhibits neutrophil cathepsin G. On the other hand, HEI does not inhibit u-plasminogen activator (urokinase). These findings suggest that HEI functions as a class-specific regulator of the neutrophil serine proteases characteristically found at inflammatory sites.

Animals↗
PubMed1994

Assignment of human xanthine dehydrogenase gene to chromosome 2p22.

Xanthine dehydrogenase (XDH, EC 1.1.1.204) oxidizes a variety of purines, pterins, and other heterogenic nitrogen compounds, serving as a rate-limiting enzyme in nucleic acid degradation. The genetic defect of XDH results in hereditary xanthinuria and other disorders in purine metabolism. Based on the cloning and sequencing results of human XDH cDNA in our laboratory, we studied the localization and sublocalization of the XDH gene. A Version 3.0 human-hamster somatic cell hybrid PCRable DNA panel and specific PCR primers derived from human XDH cDNA for amplification were used to assign the XDH gene to human chromosome 2. The fidelity of the PCR product was confirmed by nucleotide sequencing the PCR product. The assignment of the XDH gene to chromosome 2 at band p22 was established by fluorescence in situ hybridization on human metaphase chromosomes using a clone from a pWE 15 cosmid library containing the XDH gene. The results should be useful for further studies of the molecular basis for hereditary xanthinuria and other genetic disorders related to abnormal XDH activity.

Animals↗
PubMed1994

Xanthine dehydrogenase and xanthine oxidase activity and gene expression in renal epithelial cells. Cytokine and steroid regulation.

Reactive oxygen species have been implicated in the tissue injury and loss of epithelial barrier function associated with a number of clinical disorders in which disregulated inflammation seems to be a dominant event, such as endotoxemia and viral syndromes. In these disorders, xanthine oxidase (XO) contained within the epithelial cell has been proposed as a major source of injurious reactive oxygen species. This study was undertaken in an effort to understand the regulation of xanthine dehydrogenase (XDH)/XO expression at both the activity and gene expression levels in the epithelial cell under conditions associated with the inflammatory response. The results indicate that TNF, IFN-gamma, IL-6, IL-1, and dexamethasone induce XDH/XO activity in bovine renal epithelial cells (MDBK). This pattern of XDH/XO regulation by cytokines and steroids is analogous to the profile of response seen by acute phase reactants. Metabolic labeling and immunoprecipitation revealed the increase in XDH/XO activity requires new protein synthesis. By Northern analysis, all cytokines and dexamethasone increased the level of the 5-kb XDH/XO mRNA. This increase was not detectable in the presence of actinomycin D but was further induced in the presence of cycloheximide, consistent with the major site of XDH/XO up-regulation occurring at the transcriptional level. XDH/XO mRNA was very stable, with no indication that the rates of transcript degradation contributed to differences in mRNA accumulation or ultimate activity levels. In addition to providing information on the regulation of XDH/XO, the data presented furthers the understanding of the epithelial cell's potential to actively respond to immunomodulators associated with injury/inflammation.

Animals↗
PubMed1994

DNA strand breaks following in vitro exposure to asbestos increase with surface-complexed [Fe3+].

Surface functional groups on silicate dusts complex iron cations which can cycle through reduction and oxidation states to generate free radicals. These oxidants have a capacity to produce DNA strand breaks and mutations which are primary events in cancer induction. A differential in the capacity of fibrous silicates to produce carcinoma is recognized with the amphiboles demonstrating a greater biologic effect than the serpentine fiber chrysotile. We tested the hypothesis that the differences in genotoxicity of these fibrous silicates correspond to varying concentrations of iron complexed to the surface. Relative to chrysotile, the amphibole fibers complexed greater amounts of iron cations from both inorganic and in vivo sources. Increased concentrations of surface-complexed iron were associated with greater oxidant generation, measured as thiobarbituric acid-reactive products of deoxyribose, and more covalently closed, circular DNA strand scission. These results indicate that genotoxic effects of these fibers may correspond to their capacity to complex iron at the surface.

Asbestos↗
PubMed1994

Molecular cloning, tissue expression of human xanthine dehydrogenase.

Xanthine dehydrogenase (XDH, EC 1.1.1.204) is a molybdenum iron-sulphur flavin hydroxylase which oxidizes a variety of purines, pterins and other heterogenic nitrogen compounds, serving as a rate-limiting enzyme in nucleic acid degradation. In this work, we have isolated and sequenced cDNA clones of human liver XDH. The obtained cDNA covers 4577 bases of human liver XDH mRNA with a 63 bp 5'-end untranslated region and a 515 bp 3'-end untranslated region. A termination codon TGA and a polyadenylation signal AATAAA were identified. An open reading frame encodes 1333 amino acid residues. The assignment of the N-terminal was confirmed by directly sequencing that region of purified human milk XDH. Northern blot analysis shows that the human XDH gene is widely expressed in human tissues.

Amino Acid Sequence↗
PubMed1994

Complexation of iron cation by sodium urate crystals and gouty inflammation.

Gouty inflammation can be suppressed by an iron chelator. We therefore hypothesized that arthritis associated with sodium urate crystal deposition could follow the incomplete complexation of iron cation with subsequent oxidant generation as the metal cycles through reduced and oxidized states. Urate crystals adsorbed Fe3+ in vitro and crystals collected from a human tophus had significant concentrations of ionizable iron. Urate crystals oxidized deoxyribose to a thiobarbituric acid (TBA)-reactive product, augmented luminol chemiluminescence by neutrophils, released leukotriene B4 from neutrophils, activated complement, and promoted neutrophil chemotaxis. All of these events increased with the concentration of complexed iron and were suppressed by the iron chelator deferoxamine or the .OH scavenger dimethylthiourea. These results suggest that some portion of gouty inflammation after urate crystal deposition could result from the incomplete complexation of iron with subsequent catalytic generation of reactive oxygen species.

Chemotaxis, Leukocyte↗
PubMed1994

Pathogenesis of chronic bronchitis.

Inflammation of the central airways is a prominent feature in subjects with chronic bronchitis. The pathology of chronic bronchitis includes an inflammatory mononuclear cell infiltrate in the airway wall and a neutrophil influx into the airway lumen. The molecular events that produce the inflammation and its pathogenetic role in causing mucus hypersecretion are beginning to be elucidated. The inflammatory cell recruitment to the airways likely involves chemotactic agents derived not only from tissue fluid and invading microbes but also generated by the diseased bronchial epithelium. For example, bronchial epithelial cells synthesize interleukin (IL-8), a potent chemoattractant and activator of neutrophils and lymphocytes. Adhesion of infiltrating leukocytes to resident parenchymal cells in the bronchi and to extracellular matrix also is crucial for the development of airway inflammation. The resultant inflammation likely plays a direct role in the clinical features of the disorder. There is growing evidence incriminating neutrophil and lymphocytes constituents in the initiation and maintenance of cough and mucus expectoration that occurs in subjects with chronic bronchitis.

Bronchitis↗
PubMed1993

Localization of the gene encoding proteinase-3 (the Wegener's granulomatosis autoantigen) to human chromosome band 19p13.3.

Proteinase-3 (PR3) is a polymorphonuclear leukocyte serine proteinase that is a potent inducer of emphysema in experimental models. PR3 is also the target antigen associated with Wegener's granulomatosis. By analysis of a panel of somatic cell hybrids, the gene encoding PR3 was localized previously to chromosome 19; we have used fluorescence in situ hybridization to sublocalize this gene to 19p13.3.

Autoantigens↗
PubMed1993

Interaction of secretory leukocyte protease inhibitor with proteinase-3.

Secretory leukocyte protease inhibitor (SLPI) is a 12 kD nonglycosylated serine antiproteinase secreted by cells of mucosal surfaces. In human lung, SLPI is present in the respiratory epithelium. It is the major barrier to tissue destruction mediated by the polymorphonuclear leukocyte (PMN) serine proteinases, elastase and cathepsin G, within the upper respiratory tract. We have recently described a third PMN serine proteinase, proteinase-3, that like elastase causes lung matrix destruction and experimental emphysema. The current studies examine interactions between SLPI and proteinase-3. The results show that: (1) SLPI and its reactive-site variants have no or minimal inhibitory activity against proteinase-3; (2) native SLPI does not complex with proteinase-3; (3) proteinase-3 selectively degrades both native and oxidized SLPI; (4) the cleavage of SLPI by proteinase-3 occurs at the peptide bond COOH-terminal to Ala-16 in the NH2-terminal domain of SLPI.

Amino Acid Sequence↗
PubMed1993

Expression and regulation of tumor necrosis factor in macrophages from cystic fibrosis patients.

Cachexia and anorexia commonly occur in patients with cystic fibrosis (CF), particularly those with severe pulmonary compromise and heavy tracheobronchial colonization with Pseudomonas aeruginosa. Current understanding of the pathophysiology of cachexia attributes much of the anorexia and weight loss to the effects of the cytokine tumor necrosis factor (TNF), which is secreted by endotoxin-stimulated macrophages. It has further been suggested that TNF may play a role in the pathobiochemistry of CF cachexia, secondary to the localized inflammatory response in the lung or wider systemic activation of cells of the monocyte-macrophage series in response to endotoxin. This study investigates TNF production and gene expression by peripheral blood monocyte-derived macrophages from CF patients, compared with normals (NL). The results indicate that although both cell populations responded dose-dependently to lipopolysaccharide (LPS); CF macrophages, upon stimulation with LPS at concentrations of 1 to 1,000 ng/ml, consistently produced substantially higher amounts of TNF than NL macrophages. At the molecular level, Northern blot analysis also revealed that both macrophage populations expressed TNF mRNA in response to LPS in a dose-dependent manner. However, at the same LPS concentrations, CF macrophage TNF mRNA expression was 2- to 4-fold greater than that of NL macrophages. LPS had no effect in either macrophage population on mRNA for CHO-B, a constitutive probe. To investigate differences between NL and CF macrophage TNF regulation, nuclear run-on/half-life studies as well as studies addressing potential differences in LPS membrane interactions and signal transduction were performed.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗