Transcription factors as targets of cytokine signals.
Explore the source record for details and available documents.
Acute-Phase Reaction↗
Biomedical subjects
J Huang
Publications and source records attributed to J Huang.
Distribution of glutathione and glutathione-related enzyme systems in mitochondria and cytosol of cultured cerebellar astrocytes and granule cells.
The cellular and regional distribution of glutathione (GSH) and GSH-related enzyme systems involved in cellular defense against reactive oxygen species and electrophilic xenobiotics in the nervous system has been extensively studied. However, little is known about the subcellular distribution of GSH systems in brain tissue and cultured neural cells. The present study investigates the distribution of mitochondrial and cytosolic GSH and GSH-related enzymes in cultured cerebellar astrocytes and granule cells, and compares them with levels in the adult rat cerebellum. Cytosolic GSH levels and cytosolic activities of glutathione reductase (GR), glutathione peroxidase (GPX) and glutathione-S-transferase (GST) in astrocytes were 57, 153, 245, and 92% higher than those found in granule cells, respectively. In contrast, granule cells contained significantly higher mitochondrial GSH levels than astrocytes. Granule cells also demonstrated comparable mitochondria/cytosolic concentrations of GSH and GR, GPX and GST activities to those observed in the cerebellar tissue, whereas ratios in astrocytes were markedly lower. Although in vitro treatments with 100 microM ethacrynic acid depleted both cytosolic and mitochondrial GSH in cultured astrocytes and granule cells in a time-dependent fashion, cellular GSH in granule cells was more resistant to the GSH-depleting agent than astrocytes. These results suggest that although GSH and GSH-related enzymes are abundant in cytosolic compartments of astrocytes, mitochondrial pools are relatively small. Since brain mitochondria are sites of significant hydrogen peroxide generation, the mitochondrial localization of GSH and its associated enzymes in neural cells provide important defenses against toxic oxygen species in the nervous system. Differences in subcellular distribution of GSH systems in individual neural cell types may provide a basis for selective cellular and/or subcellular expression of neurotoxicity.
Animals↗
Induction of interleukin 6 (IL-6) by hypoxia in vascular cells. Central role of the binding site for nuclear factor-IL-6.
The pathologic picture in ischemic tissue injury shares features with the inflammatory response, including production of proinflammatory cytokines. Hypoxia-mediated induction of interleukin-6 (IL-6), a cytokine with anti-inflammatory properties, could set in motion mechanisms limiting inflammation in ischemia. Exposure of cultured endothelial cells (ECs) to H (pO2 approximately 12-16 torr) increased transcription of IL-6, elevated levels of IL-6 mRNA, and induced elaboration of IL-6 antigen in a time-dependent manner. Exposure of mice to hypoxia increased IL-6 transcripts in the lung, and immunostaining revealed a striking increase in IL-6 antigen in pulmonary vasculature. Transfection of ECs with deletion chimeric IL-6 promoter-chloramphenicol acetyl-transferase (CAT) constructs showed hypoxia-mediated 9-11-fold induction with -1200/+13, -596/+13, and -225/+13 but no induction with -111/+13. Electrophoretic mobility shift assays (EMSAs) using -225/-111 as the labeled probe demonstrated enhanced binding activity in nuclear extracts of hypoxic ECs and lung; the appearance of the gel shift band was prevented by excess unlabeled probe (-225/-111), and hypoxia-mediated enhancement of the band was blocked by a probe corresponding to the nuclear factor (NF)-IL-6 site (-158/-145). The hypoxia-enhanced band on EMSA displayed a supershift with antibody to CCAAT-enhancer-binding protein beta (C/EBP-beta), but antibody to C/EBP-alpha or -delta was without effect. Transfection of ECs with a construct comprising thymidine kinase promoter, -225/-111 in either the 5' to 3' to 5' orientation, and the reporter CAT showed this region to be an enhancer (approximately 8-fold) under hypoxia. EMSA with the NF-IL-6 probe revealed a prominent induction of binding activity with nuclear extracts from hypoxic ECs and whole lung. Constructs with -158/-145 and the CAT reporter gene showed induction when transfected into hypoxic ECs, whereas a similar construct with the NF-IL-6 motif mutationally inactivated failed to display hypoxia-induced expression. Furthermore, the tumor necrosis factor (TNF) gene, whose product contributes to ischemic pathology and contains a putative regulatory NF-IL-6 site, demonstrated enhanced binding activity for its NF-IL-6 motif and induction of TNF mRNA based on analysis of hypoxic lung. These data indicate that hypoxia induces expression of IL-6, most likely a result of hypoxic activation at the NF-IL-6 site, and suggest that other genes with regulatory NF-IL-6 sites may also be induced by a similar mechanism.
Animals↗
Angiographic stenosis progression and coronary events in patients with 'stabilized' unstable angina.
BACKGROUND: Recent studies suggest that angiographically complex coronary stenoses are associated with an adverse short-term outcome. It is not known, however, if this applies to unstable angina patients who stabilize on medical therapy. METHODS AND RESULTS: We prospectively studied 85 consecutive patients with unstable angina who stabilized on medical therapy but were found to require angioplasty for treatment of obstructive coronary disease. Angiography was carried out at admission, and patients were restudied 8 +/- 4 months (mean +/- SD) after the first angiogram. Ischemia-related stenoses were identified and classified as "complex" (irregular borders, overhanging edges, or thrombus) or "smooth" (absence of complex features). Stenosis progression (> or = 20% diameter reduction or new total occlusion) was assessed by automated edge detection. At initial angiography, there were 198 stenoses (> or = 50%, 102), of which 85 (54 complex and 31 smooth) were ischemia related. At restudy, 21 ischemia-related stenoses and 8 non-ischemia-related stenoses progressed (25% versus 7%, P = .001). Seventeen of the 21 ischemia-related stenoses that progressed developed into total occlusion compared with 3 of the 8 non-ischemia-related stenoses (P = .02). Changes in average stenosis severity and in absolute stenosis diameter were significantly larger in ischemia-related stenoses than in non-ischemia-related stenoses (P = .03). Eighteen (34%) complex stenoses progressed, compared with 3 (10%) smooth lesions (P = .02). During follow-up, 1 patient died (myocardial infarction) and 25 patients had nonfatal coronary events that were associated with progression of ischemia-related stenoses in 14 (56%). CONCLUSIONS: In unstable angina patients who stabilize medically, subsequent short-term stenosis progression and coronary events are common. The unstable coronary lesion (particularly complex stenoses) is often not stabilized and will continue to progress over the ensuing months.
Angina, Unstable↗
Identification and characterization of variants of tick anticoagulant peptide with increased inhibitory potency toward human factor Xa.
Tick anticoagulant peptide (TAP) is a specific and potent inhibitor of factor Xa (fXa), a central enzyme in the blood clotting cascade. As such, TAP is a potential antithrombotic agent. Site-directed mutagenesis studies were undertaken to determine the feasibility of increasing the inhibitory potency of TAP toward fXa. The amino acid substitutions Tyr-1 to Trp (Y1W) and Asp-10 to Arg (D10R) increased inhibitory potency toward human fXa by 2.5- and 4-fold, respectively. The increased inhibitory potency reflected a decrease in the rate constant for dissociation of the final fXa-TAP inhibitory complex. The double mutant, Y1W/D10R, exhibited an inhibition constant of 10 pM, a 37-fold enhancement of inhibitory potency toward human fXa. The improvement in inhibitory potency was less pronounced (12-fold) with dog fXa wherein Kis of 220 and 18 pM were observed for wild-type TAP and the double mutant, respectively. Mutation of Tyr-1 to Glu (Y1E) generated a weaker inhibitor (Ki = 2 nM) that bound human fXa more slowly. However, no change in inhibitory potency toward human fXa was observed when Tyr-1 was replaced by Phe. Taken together, these observations are consistent with the view that a hydrophobic amino acid at the N-terminus of TAP may be a determinant of inhibitory potency. Decreases by 3-4 orders of magnitude in inhibitory potency were noted upon mutation of Asn-2 and Leu-4 of TAP, further implicating the N-terminal domain as an important determinant of inhibitory potency.(ABSTRACT TRUNCATED AT 250 WORDS)
Amino Acid Sequence↗
WT1, the Wilms' tumor suppressor gene product, represses transcription through an interactive nuclear protein.
The Wilms' tumor suppressor gene, wt1, encodes a transcription factor of the zinc finger family. Mutations in WT1 have been detected in subsets of Wilms' tumor and in patients with the Denys-Drash Syndrome. In order to determine how WT1 regulates transcription and perhaps the consequences that mutations in WT1 may have, we established that residues 85-124 and 181-250 of WT1 constitute domains that function independently with a DNA binding domain to repress or activate transcription, respectively, and function equally effectively with heterologous promoters, suggesting the activator and repressor domains interact with nuclear components of general importance. To seek evidence for such components, increasing concentrations of WT1 repressor domain without a zinc finger DNA binding domain were co-transfected with fixed concentrations of wild-type (wt) WT1 and PDGF A-chain promoter/reporter gene constructs. As levels of the repressor domain were increased, a progressive loss of wt WT1 repressor activity and a progressive increase in its activation were observed, suggesting that the repressor domain of WT1 competes with wt WT1 for an interactive protein that is an essential component of the repressor activity of wt WT1. Because the most common mutation associated with Denys-Drash Syndrome disrupts the zinc finger domains of WT1, the results also suggest that the mutant WT1 may have aberrant DNA binding activity and perhaps function as a dominant negative effector of wt WT1.
3T3 Cells↗
Reduced activation of RAF-1 and MAP kinase by a fibroblast growth factor receptor mutant deficient in stimulation of phosphatidylinositol hydrolysis.
Signaling via the fibroblast growth factor receptor 1 (FGFR1, flg) was analyzed in Ba/F3 hematopoietic cells expressing either wild-type or a mutant FGF receptor (Y766F) unable to activate phospholipase C-gamma (PLC-gamma) and stimulate phosphatidylinositol (PI) hydrolysis. Stimulation of cells expressing wild-type or mutant FGFR with acidic FGF (aFGF) caused similar activation of Ras. However, an approximately 3-fold reduced activation of Raf-1 and MAP kinase was observed in aFGF-stimulated cells expressing mutant receptors as compared to cells expressing wild-type FGF receptors. Comparison of phosphopeptide maps of Raf-1 immunoprecipitated from the two cell types activated by either aFGF or the phorbol ester (12-O-tetradecanoylphorbol-13-acetate) suggests that Raf-1 is phosphorylated by both Ras-dependent and PLC-gamma-dependent mechanisms. In spite of the differential effect on Raf-1 and MAP kinase activation, aFGF stimulated similar proliferation of cells expressing wild-type or mutant receptors indicating that Ras-dependent activation of Raf-1 and MAP kinase is sufficient for transduction of FGFR mitogenic signals. Ras may also activate signal transduction pathways that are complementary or parallel to the MAP kinase pathway to stimulate cell proliferation.
Amino Acid Sequence↗
Products of alternatively spliced transcripts of the Wilms' tumor suppressor gene, wt1, have altered DNA binding specificity and regulate transcription in different ways.
The Wilms' tumor susceptibility gene, wt1, encodes a transcription factor of the zinc finger protein family. Mutations in the WT1 gene product have been detected in both sporadic and familial Wilms' tumors, suggesting that alterations in WT1 may disrupt its normal function as a transcriptional regulator. The transcripts of wt1 are alternatively spliced; however, roles of the alternatively spliced forms have not been defined. The major transcript of wt1 encodes a WT1 protein [WT1(+KTS)+17AA] that contains three amino acids (+KTS) between the third and fourth zinc fingers and a serine-rich, 17 amino acid (+17AA) domain N-terminal to the zinc finger region. We now show that the WT1 (+KTS) forms functionally bind to a unique G+C-rich sequence within the PDGF A-chain promoter. We also show that WT1 (+KTS)+17AA functions as a strong transcriptional repressor and that +17AA alone fused to the zinc-finger domain of WT1 or to the heterologous DNA binding domain of GAL4 functions independently as a repressor. Deletion of four serine residues within +17AA abolishes the repressor activity of +17AA. These results indicate that wt1 products with +17AA contain an additional dominant repressor domain and that the presence or absence of +KTS determines alternative DNA binding specificity.
3T3 Cells↗
Giant magnetostriction of amorphous TbxFe1-x (0.10<x<0.45) thin films and its correlation with perpendicular anisotropy.
Explore the source record for details and available documents.
Journal Article↗
Cellular localization and hormonal regulation of inducible nitric oxide synthase in cycling mouse uterus.
Nitric oxide (NO), a potent and versatile free radical, is synthesized in macrophages and mast cells as well as in other types of cells by the inducible form of nitric oxide synthase (iNOS). In this study, cells containing iNOS were identified in the uteri of cycling mice by using a rabbit antibody generated to an iNOS-specific peptide. Macrophages were identified in semiserial sections of the same tissues with the monoclonal antibody, F4/80, and mast cells were identified by toluidine blue staining. In tissue sections of uteri obtained from mice in the four stages of the estrous cycle (8 to 11 mice per stage), iNOS immunoreactivity was strongest in diestrus-I uteri and weakest in diestrus-II uteri. Myometrial mast cells and endometrial epithelial cells were prominent locations of iNOS, and specific protein was also present in myometrial smooth muscle and macrophage-like cells in the endometrial stroma. Because cyclic variations suggested regulation of iNOS expression by ovarian steroid hormones, studies were done using ovariectomized mice. Seven days after ovariectomy, immunoreactive iNOS was low but detectable in mast cells and luminal epithelial cells. In the uteri of ovariectomized, estradiol-17 beta (E2)-treated mice, mast cells were iNOS+ after 24 h whereas epithelial cells were negative; the reverse was observed in progesterone (P4)-treated mice. Both mast cells and epithelial cells were iNOS+ in the uteri of mice that had received a combination of E2 + P4. These results indicate that several types of uterine cells produce iNOS and that expression of this enzyme in specific cell lineages is governed by ovarian steroid hormones. The data are consistent with the postulate that NO derived from uterine leukocytes and other types of cells plays a role in uterine cyclicity and preparation for pregnancy.
Amino Acid Oxidoreductases↗
Functional analysis of a recently originating, atypical presequence: mitochondrial import and processing of GUS fusion proteins in transgenic tobacco and yeast.
A gene family of at least five members encodes the tobacco mitochondrial Rieske Fe-S protein (RISP). To determine whether all five RISPs are translocated to mitochondria, fusion proteins containing the putative presequences of tobacco RISPs and Escherichia coli beta-glucuronidase (GUS) were expressed in transgenic tobacco, and the resultant GUS proteins were localized by cell fractionation. The amino-terminal 75 and 71 residues of RISP2 and RISP3, respectively, directed GUS import into mitochondria, where fusion protein processing occurred. The amino-terminal sequence of RISP4, which contains an atypical mitochondrial presequence, can translocate the GUS protein specifically into tobacco mitochondria with apparently low efficiency. Consistent with the proposal of a conserved mechanism for protein import in plants and fungi, the tobacco RISP3 and RISP4 presequences can direct import and processing of a GUS fusion protein in yeast mitochondria. Plant presequences, however, direct mitochondrial import in yeast less efficiently than the yeast presequence, indicating subtle differences between the plant and yeast mitochondrial import machineries. Our studies show that import of RISP4 may not require positively charged amino acid residues and an amphipathic secondary structure; however, these structural properties may improve the efficiency of mitochondrial import.
Amino Acid Sequence↗
Respiratory effects and skin allergy in workers exposed to tetrachloroisophthalonitrile.
Explore the source record for details and available documents.
Adult↗
Heart rate variability depression in patients with unstable angina.
The degree of reduction in heart rate variability (HRV) after myocardial infarction has been shown to have prognostic significance, but HRV has not been studied extensively in patients with unstable angina. We assessed spectral and nonspectral measurements of HRV in 52 patients with unstable angina, 52 patients with acute myocardial infarction, and 41 normal subjects. The spectral bands of 0.04 to 0.15 Hz (low frequency), 0.15 to 0.4 (high frequency), and nonspectral parameters SDNN, SDANN, SDNN index, rMSSD, and pNN50 were calculated from continuous 24-hour ECGs. All measures of HRV were reduced in patients with acute coronary syndromes compared to normal controls (p < 0.001), and there was no significant difference in measure of HRV between unstable angina and myocardial infarction patients. In patients with unstable angina who stabilized after admission, HRV had increased by the second 24 hours of monitoring. In contrast, HRV was further depressed in patients who had episodes of chest pain or transient ST-segment depression during the second 24 hours. rMSSD, pNN50, and SDNN index were lower in patients with unstable angina who had transient silent ischemia compared to those without silent ischemia. Of the patients with unstable angina, 4 died and 1 had nonfatal acute myocardial infarction within 11 months. HRV was lower in these patients than in patients without further cardiac events.
Aged↗
B cell responses to acetylcholine receptor in rats orally tolerized against experimental autoimmune myasthenia gravis.
Oral administration of acetylcholine receptor (AChR) to Lewis rats prior to myasthenogenic immunization with AChR and complete Freund's adjuvant (CFA) results in the prevention of experimental autoimmune myasthenia gravis (EAMG), and decreased serum levels of anti-AChR antibodies. Using an ELISPOT assay, we have now determined numbers of cells in the popliteal, inguinal and mesenteric lymph nodes, spleen and thymus secreting anti-AChR IgG antibodies. Except for mesenteric lymph nodes, a marked diminution of such cells was detected in these lymphoid organs in rats orally tolerized with AChR compared to buffer-fed or vehicle-fed control rats with EAMG. Of note is that, after AChR feeding, the B cell response to AChR in thymus was diminished to the same low level as in CFA-injected, buffer-fed control rats. The relative affinity of serum anti-AChR IgG antibodies measured by KSCN-ELISA was lower in the orally tolerized rats compared to buffer-fed or vehicle-fed rats. The observations showed that oral administration of AChR, besides preventing clinical EAMG, also counteracts the development of AChR-specific B cells, especially those with high affinity antibody production, in most lymphoid organs.
Administration, Oral↗
Toluene induces behavioral activation without affecting striatal dopamine metabolism in the rat: behavioral and microdialysis studies.
We examined the effects of toluene on the release of dopamine (DA) and its metabolites in rat striatum using microdialysis. Intraperitoneal injection of 800 mg/kg toluene significantly increased motor activity in rats, as did methamphetamine (MAP) (1 mg/kg). However, 800 mg/kg toluene did not affect the extracellular levels of DA, 3,4-dihydroxyphenylacetic acid, homovanillic acid, or 5-hydroxyindoleacetic acid. This is in contrast to MAP, which significantly increased extracellular DA and decreased the extracellular levels of its metabolites. These results suggest that toluene-induced behavioral augmentation may not be associated with alterations in DA or serotonin neurochemistry such as are associated with MAP-induced behavioral augmentation.
Animals↗
Structure-acute toxicity relationship of aromatic hydrocarbons in mice.
The structure-acute toxicity relationship of aromatic hydrocarbons was examined in mice. In all test compounds, the acute toxicity was determined under 2 conditions: control LD50 (LD50-cont) and carbon tetrachloride (CCl4)-pretreated LD50 (LD50-CCl4). The CCl4-pretreatment was done in order to evaluate the toxic potency of compound itself without the influence of metabolism. Both log (1/LD50-cont) and log (1/LD50-CCl4) were functions of the log P, n-octanol/water partition coefficient, i.e., log (1/LD50-cont) = 0.080 log P - 1.532 and log (1/LD50-CCl4) = -0.040(logP)2 + 0.157 log P - 1.373. Both equations were statistically significant (P < 0.01). The ratio of LD50-cont/LD50-CCl4 indicated that metabolic activation is more evident in hydrophobic compounds than in hydrophilic compounds. The results suggest that hydrophobicity of the aromatic hydrocarbons plays an important role in determining their acute toxicity.
Animals↗
Inhibition of mucin synthesis by benzyl-alpha-GalNAc in KATO III gastric cancer and Caco-2 colon cancer cells.
Previous studies from our laboratory have shown that benzyl-alpha-GalNAc inhibits the glycosylation of mucin in colon cancer cells. In this study, we determined whether benzyl-alpha-GalNAc inhibits mucin glycosylation in KATO III gastric cancer cells. We also examined its effects on expression of mucin antigens, and compared the mucins made by KATO III with those of a colonic cancer cell line, Caco-2. Results of these experiments suggest that benzyl-alpha-GalNAc (2 mM) inhibited [3H]glucosamine labelling of mucins by 82% in KATO III and by 70% in Caco-2. For both cell lines, the mucin secreted in the presence of benzyl-alpha-GalNAc was less acidic. Both cell lines secreted benzyl-oligosaccharides, but those from KATO III (8-9 sugars) were larger than those from Caco-2 (6-7 sugars). In mucins purified from the medium of treated cells, peripheral carbohydrate antigens (sialyl Lex in KATO III and terminal fucose in Caco-2) were decreased (compared with control), while core carbohydrate antigens (T antigen in both cell lines and sialyl Tn in Caco-2) were increased. Western blots of cell homogenates showed differences between KATO III and Caco-2 in MUC 1 apomucin protein antigens, in sialyl Lex and in sialyl Tn antigens. We conclude that benzyl-alpha-GalNAc does inhibit the glycosylation of mucin in KATO III gastric cancer cells as in human colon cancer cells, but that alterations in mucin antigens occur in a cell line-specific manner.
Acetylgalactosamine↗
Pingyangmycin as a 99mTc carrier in tumor localization.
A simple method of preparation of 99mTc-pingyangmycin (PYM) for clinical use has been established using super-micro-amounts of SnCl2 as a reductant under dark conditions. The labeling efficiency was higher than 96%, and further purification was not necessary. The effect of ascorbic acid on the distribution of 99mTc-PYM had been investigated. Ascorbic acid increased uptake of 99mTc-PYM in the tumor. Tumor uptake increased with increasing concentration of ascorbic acid. Tumor-blood, tumor-liver, tumor-lung ratios at 1.5 h after 99mTc-PYM administration were 5.19 +/- 1.64, 2.71 +/- 0.51 and 4.15 +/- 0.57, respectively. Preliminary clinical trials in nine patients showed that 99mTc-PYM is a potentially useful tracer for tumor detection with good sensitivity and specificity (true positive 7/7, true negative 1/1, and false positive 1/1).
Aged↗