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Biomedical subjects

Y Yang

Publications and source records attributed to Y Yang.

At least 253 records · Page 14Linked to original sources
PubMed2001

Group B Streptococcus capsular polysaccharide-cholera toxin B subunit conjugate vaccines prepared by different methods for intranasal immunization.

Group B Streptococcus (GBS) type III capsular polysaccharide (CPS III) was conjugated to recombinant cholera toxin B subunit (rCTB) using three different methods which employed (i) cystamine and N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP), (ii) carbodiimide with adipic acid dihydrazide (ADH) as a spacer, or (iii) reductive amination (RA). The CPS III-rCTB conjugates were divided into large- and small-molecular-weight (M(r)) fractions, and the immunogenicities of the different preparations after intranasal (i.n.) immunization were studied in mice. Both large- and small-M(r) conjugates of CPS III-rCTB(RA) or CPS III-rCTB(ADH) induced high, almost comparable levels of CPS-specific immunoglobulin G (IgG) in serum, lungs, and vagina that were generally superior to those obtained with CPS III-rCTB(SPDP) conjugates or a CPS III and rCTB mixture. However, the smaller-M(r) conjugates of CPS III-rCTB(RA) or CPS III-rCTB(ADH) in most cases elicited a lower anti-CPS IgA immune response than the large-M(r) conjugates, and the highest anti-CPS IgA titers in both tissues and serum were obtained with the large-M(r) CPS III-rCTB(RA) conjugate. Serum IgG anti-CPS titers induced by the CPS III-rCTB(RA) conjugate had high levels of specific IgG1, IgG2a, IgG2b, and IgG3 antibodies. Based on the effectiveness of RA for coupling CPS III to rCTB, RA was also tested for conjugating GBS CPS Ia with rCTB. As for the CPS III-rCTB conjugates, the immunogenicity of CPS Ia was greatly increased by conjugation to rCTB. Intranasal immunization with a combination of CPS Ia-rCTB and CPS III-rCTB conjugates was shown to induce anti-CPS Ia and III immune responses in serum and lungs that were fully comparable with the responses to immunization with the monovalent CPS Ia-rCTB or CPS III-rCTB conjugates. These results suggest that the GBS CPS III-rCTB and CPS Ia-rCTB conjugates prepared by the RA method may be used in bivalent and possibly also in multivalent mucosal GBS conjugate vaccines.

Administration, Intranasal↗
PubMed2001

Genome of the bacterium Streptococcus pneumoniae strain R6.

Streptococcus pneumoniae is among the most significant causes of bacterial disease in humans. Here we report the 2,038,615-bp genomic sequence of the gram-positive bacterium S. pneumoniae R6. Because the R6 strain is avirulent and, more importantly, because it is readily transformed with DNA from homologous species and many heterologous species, it is the principal platform for investigation of the biology of this important pathogen. It is also used as a primary vehicle for genomics-based development of antibiotics for gram-positive bacteria. In our analysis of the genome, we identified a large number of new uncharacterized genes predicted to encode proteins that either reside on the surface of the cell or are secreted. Among those proteins there may be new targets for vaccine and antibiotic development.

Bacterial Proteins↗
PubMed2001

Positively charged termini of the L2 minor capsid protein are necessary for papillomavirus infection.

Coexpression of bovine papillomavirus L1 with L2 mutants lacking either eight N-terminal or nine C-terminal amino acids that encode positively charged domains resulted in wild-type levels of viral genome encapsidation. Despite wild-type binding to the cell surface, the resulting virions were noninfectious. An L2 mutant encoding a scrambled version of the nine C-terminal residues restored infectivity, in contrast to an L2 mutant encoding a scrambled version of the N-terminal residues.

Animals↗
PubMed2001

Non-specific interstitial pneumonia as pulmonary involvement of systemic sclerosis.

The pathological features of lung disease in nine patients with systemic sclerosis (SSc) were evaluated. The patients comprised one man and eight women, with a median age of 58 years. SSc was diagnosed according to the criteria of the American Rheumatism Association. In all patients, high resolution computed radiographic scanning of the lungs (HRCT) was performed, and apparent honeycomb formation was seen in four patients. Pathologically, four patients were diagnosed with usual interstitial pneumonia (UIP), three with non-specific interstitial pneumonia (NSIP) group II, one NSIP group II-III, and one NSIP group II with diffuse alveolar damage. HRCT showed no apparent honeycomb formations in patients diagnosed with NSIP. This is the first report describing NSIP as a pulmonary complication of SSc.

Adult↗
PubMed2001

Corneal endothelial NKCC: molecular identification, location, and contribution to fluid transport.

Although Na(+)-K(+)-2Cl(-) cotransport has been demonstrated in cultured bovine corneal endothelial cells, its presence and role in the native tissue have been disputed. Using RT-PCR we have now identified a partial clone of the cotransporter protein in freshly dissected as well as in cultured corneal endothelial and epithelial cells. The deduced amino acid sequence of this protein segment is 99% identical to that of the bovine isoform (bNKCC1). [(3)H]bumetanide binding shows that the cotransporter sites are located in the basolateral membrane region at a density of 1.6 pmol/mg of protein, close to that in lung epithelium. Immunocytochemistry confirms the basolateral location of the cotransporter. We calculate the turnover rate of the cotransporter to be 83 s(-1). Transendothelial fluid transport, determined from deepithelialized rabbit corneal thickness measurements, is partially inhibited (30%) by bumetanide in a dose-dependent manner. Our results demonstrate that Na(+)-K(+)-2Cl(-) cotransporters are present in the basolateral domain of freshly dissected bovine corneal endothelial cells and contribute to fluid transport across corneal endothelial preparations.

Amino Acid Sequence↗
PubMed2001

Gut expression and regulation of FAT/CD36: possible role in fatty acid transport in rat enterocytes.

Fatty acid translocase (FAT)/CD36 is one of several putative plasma membrane long-chain fatty acid (LCFA) transport proteins; however, its role in intestinal absorption of LCFA is unknown. We hypothesized that FAT/CD36 would be differentially expressed along the longitudinal axis of the gut and during intestinal development, suggesting specificity of function. We found that intestinal mucosal FAT/CD36 mRNA levels varied by anatomic location along the longitudinal gut axis: stomach 45 +/- 7, duodenum 173 +/- 29, jejunum 238 +/- 17, ileum 117 +/- 14, and colon 9 +/- 1% (means +/- SE with 18S mRNA as control). FAT/CD36 protein levels were also higher in proximal compared with distal intestinal mucosa. Mucosal FAT/CD36 mRNA was also regulated during intestinal maturation, with a fourfold increase from neonatal to adult animals. In addition, FAT/CD36 mRNA levels and enterocyte LCFA uptake were rapidly downregulated by intraduodenal oleate infusion. These findings suggest that FAT/CD36 plays a role in the uptake of LCFA by small intestinal enterocytes. This may have important implications in understanding fatty acid absorption in human physiological and pathophysiological conditions.

Animals↗
PubMed2001

IL-1beta enhances beta2-adrenergic receptor expression in human airway epithelial cells by activating PKC.

Protein kinase C (PKC)-activated signal transduction pathways regulate cell growth and differentiation in many cell types. We have observed that interleukin (IL)-1beta upregulates beta2-adrenergic receptor (beta2-AR) density and beta2-AR mRNA in human airway epithelial cells (e.g., BEAS-2B). We therefore tested the hypothesis that PKC-activated pathways mediate IL-1beta-induced beta-AR upregulation. The role of PKC was assessed from the effects of 1) the PKC activator phorbol 12-myristate 13-acetate (PMA) on beta-AR density, 2) selective PKC inhibitors (calphostin C and Ro-31-8220) on beta-AR density, and 3) IL-1beta treatment on the cellular distribution of PKC isozymes. Recombinant human IL-1beta (0.2 nM for 18 h) increased beta-AR density to 213% of control values (P < 0.001). PMA (1 microM for 18 h) increased beta-AR density to 225% of control values (P < 0.005), whereas Ro-31-8220 and calphostin C inhibited the IL-1beta-induced upregulation of beta-AR in dose-dependent fashion. PKC isozymes detected by Western blotting included alpha, betaII, epsilon, mu, zeta, and lambda/iota. IL-1beta increased PKC-mu immunoreactivity in the membrane fraction and had no effect on the distribution of the other PKC isozymes identified. These data indicate that IL-1beta-induced beta-AR upregulation is mimicked by PKC activators and blocked by PKC inhibitors and appears to involve selective activation of the PKC-mu isozyme. We conclude that signal transduction pathways activated by PKC-mu upregulate beta2-AR expression in human airway epithelial cells.

Cell Line, Transformed↗
PubMed2001

Nuclear factor-kappa B augments beta(2)-adrenergic receptor expression in human airway epithelial cells.

Interleukin (IL)-1 beta increases beta(2)-adrenergic receptor (beta(2)-AR) mRNA and density by protein kinase C (PKC)-dependent mechanisms in human airway epithelial cells. The present study examined the role of several nuclear transcription factors in the PKC-activated upregulation of beta(2)-AR expression. BEAS-2B cells were exposed to the PKC activator phorbol 12-myristate 13-acetate (PMA; 0.1 microM for 2-18 h). PMA had no effect on activator protein (AP)-2 or cAMP response element binding protein DNA binding activity but markedly increased nuclear factor (NF)-kappa B and AP-1 binding as assessed by electrophoretic gel mobility shift assay. PMA also increased the activity of a beta(2)-AR promoter-luciferase reporter construct in transiently transfected cells. These effects were inhibited by the PKC inhibitors Ro-31-8220 and calphostin C. Furthermore, with increasing Ro-31-8220, beta(2)-AR promoter-reporter activity correlated closely with both NF-kappa B and AP-1 activities (r > 0.89 for both). Finally, the selective NF-kappa B inhibitor MG-132 dose dependently reduced NF-kappa B binding and beta(2)-AR promoter activity but increased AP-1 binding. We conclude that PKC-induced upregulation of beta(2)-AR expression in human airway epithelial cells appears to be mediated, at least in part, by increases in NF-kappa B activity.

Cell Line↗
PubMed2001

Bleomycin upregulates gene expression of angiotensin-converting enzyme via mitogen-activated protein kinase and early growth response 1 transcription factor.

Pulmonary fibrosis is a progressive disorder characterized by the loss of alveolar architecture through epithelial and endothelial cell apoptosis and fibroblast proliferation. Recent studies showed that angiotensin-converting enzyme (ACE) activity is increased in fibrotic tissues, and ACE inhibitors administered in vivo ameliorate fibrosis, suggesting that ACE may play a critical role. However, the regulation of ACE expression is not well understood. In the present study, we demonstrate that bleomycin, a chemotherapeutic agent which induces pulmonary fibrosis in animals and humans, increases gene expression of ACE. Treatment of primary bovine pulmonary artery endothelial cells with 0.1 to 1.0 microg/ml bleomycin increased ACE enzymatic activity and ACE mRNA, as monitored by hippuryl-L-histidyl-L-leucine assay and competitive quantitative reverse transcriptase polymerase chain reaction (RT-PCR), respectively. Luciferase reporter constructs showed that upregulation of ACE transcription by bleomycin is mediated through element(s) in the 97-bp ACE promoter. Bleomycin activated p42/p44 mitogen-activated protein kinase (MAPK) and induced nuclear translocation and activation of the early growth response (Egr)-1 transcription factor, a factor previously shown to positively regulate ACE expression. The MAPK kinase1/2 (MEK1/2) inhibitor U0126 blocked MAPK and Egr-1 activation by bleomycin, suggesting that Egr-1 activation is MAPK dependent. These data provide the first evidence that bleomycin activates ACE gene expression through the MAPK pathway and Egr-1.

Animals↗
PubMed2001

Cloning and characterization of the mouse and human enamelin genes.

Enamelin is likely to be essential for proper dental enamel formation. It is secreted by ameloblasts throughout the secretory stage and can readily be isolated from the enamel matrix of developing teeth. The gene encoding human enamelin is located on the long arm of chromosome 4, in a region previously linked to an autosomal-dominant form of amelogenesis imperfecta (AI). To gain information on the structure of the enamelin gene and to facilitate the future assessment of the role of enamelin in normal and diseased enamel formation, we have cloned and characterized the mouse and human enamelin genes. Both genes are about 25 kilobases long. The enamelin gene has 10 exons interrupted by 9 introns. Translation initiates in exon 3 and terminates in exon 10. All of the intron/exon junctions within the mouse and human enamelin coding regions are between codons, so there are no partial codons in any exon, and deletion of one or more coding exons by alternative RNA splicing would not shift the downstream reading frame.

Ameloblasts↗
PubMed2001

Genetic manipulation of hedgehog signaling in the endochondral skeleton reveals a direct role in the regulation of chondrocyte proliferation.

Indian hedgehog (Ihh), one of the three mammalian hedgehog (Hh) proteins, coordinates proliferation and differentiation of chondrocytes during endochondral bone development. Smoothened (Smo) is a transmembrane protein that transduces all Hh signals. In order to discern the direct versus indirect roles of Ihh in cartilage development, we have used the Cre-loxP approach to remove Smo activity specifically in chondrocytes. Animals generated by this means develop shorter long bones when compared to wild-type littermates. In contrast to Ihh mutants (Ihh(n)/Ihh(n)), chondrocyte differentiation proceeds normally. However, like Ihh(n)/Ihh(n) mice, proliferation of chondrocytes is reduced by about 50%, supporting a direct role for Ihh in the regulation of chondrocyte proliferation. Moreover, by overexpressing either Ihh or a constitutively active Smo allele (Smo*) specifically in the cartilage using the bigenic UAS-Gal4 system, we demonstrate that activation of the Ihh signaling pathway is sufficient to promote chondrocyte proliferation. Finally, expression of cyclin D1 is markedly downregulated when either Ihh or Smo activity is removed from chondrocytes, indicating that Ihh regulates chondrocyte proliferation at least in part by modulating the transcription of cyclin D1. Taken together, the present study establishes Ihh as a key mitogen in the endochondral skeleton.

Animals↗
PubMed2001

Characterization and regulation of the major histocompatibility complex-encoded proteins Hsp70-Hom and Hsp70-1/2.

Vertebrate cells contain at least 12 different genes for Hsp70 proteins, 3 of which are encoded in the major histocompatibility complex (MHC) class III region. In the human MHC, these are named Hsp70-1, -2, and -Hom. To characterize these proteins, we have determined their substrate binding specificity, their cellular and tissue distribution, and the regulation of their expression. We show for the first time (1) peptide binding specificity of Hsp70-Hom; (2) endogenous expression of Hsp70-Hom in human cell lines; (3) cytoplasmic location of Hsp70-Hom protein under basal conditions and concentration in the nucleus after heat shock; (4) unique RNA expression profiles in human tissues for each of the MHC-encoded Hsp70s, significantly different from that for the constitutive Hsc70; (5) a relative increase in levels of Hsp70-Hom protein, compared with other Hsp70s, in response to interferon gamma; and (6) a specific increase on lipopolysaccharide (LPS) treatment of in vivo messenger RNA levels for the MHC-encoded Hsp70s and the DnaJ homologue, hdj2, relative to other chaperones. The unique tissue distributions and specific up-regulation by LPS of the MHC-encoded Hsp70s suggest some specialization of functions for these members of the Hsp70 family, possibly in the inflammatory response.

Antibodies↗
PubMed2001

Functional regulation of immunoproteasomes and transporter associated with antigen processing.

The central event in the cellular immune response to invading pathogens is the presentation of non-self antigenic peptides by major histocompatibility complex (MHC) class I molecules to cytotoxic T lymphocytes (CTLs). As peptide binding and transport proteins, MHC class I molecules have evolved distinct biochemical and cellular strategies for acquiring antigenic peptides, providing CTLs an extracellular representation of the intracellular antigen content. Whereas efficient generation of MHC class I binding peptides depends on the intracellular, immunoproteasome-mediated proteolysis machinery, translocation of peptides into the lumen of the endoplasmic reticulum requires the endoplasmic reticulum-resident, adenosine 5'-triphosphate (ATP) binding cassette transporter associated with antigen processing (TAP). Here we show, for the first time, that immunoproteasomes, TAP complexes, and MHC class I molecules are physically associated, providing an effective means of transporting MHC class I binding peptides from their sites of generation into the lumen of the endoplasmic reticulum for loading onto MHC class I molecules. In this review, we assess the current understanding of the functional regulation of immunoproteasomes and transporter associated with antigen processing.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Changes in regional energy metabolism after closed head injury in the rat.

We examined in the present investigation regional ATP, glucose, and lactate content in the cortical and subcortical structures, in a rat model of closed head injury (CHI). In serial tissue sections bioluminescence imaging of ATP, glucose, and lactate was performed at 4 h, 12 h and 24 h (n = 4/5 per time point with) after the induction of CHI or sham surgery. Bioluminescence images were analyzed by computer-assisted densitometry, at the lesion site, in remote cortical areas, and in the subcortical structures (thalamus and caudate nucleus). ATP content was significantly decreased at the lesion site after 4 h and in the remote cortex at 12 h post-injury. At 12 h, the ATP content reached baseline levels on the ipsilateral side and at 24 h also at remote lateral parietal sites. In the contralateral cortex, ATP increased transiently above the baseline at 12 h. No significant changes in ATP were found in the thalamus and caudate nucleus. Cortical glucose and lactate contents could not be discerned over time. Following CHI there is an acute and progressive, yet transient, ischemic cortical profile, which is not reflected in subcortical areas.

Adenosine Triphosphate↗
PubMed2001

Intrinsic defects in the T-cell lineage results in natural killer T-cell deficiency and the development of diabetes in the nonobese diabetic mouse.

T-cell-mediated autoimmune diabetes in nonobese diabetic (NOD) mice is closely associated with natural killer T (NKT)-cell deficiency. To determine whether intrinsic defects of the T-cell lineage contribute to the pathogenesis of the disease and NKT cell deficiency, we reconstituted the T-cell compartment in NOD.scid or BALB.scid mice with T-cells from NOD, nonobese diabetes-resistant (NOR), or AKR thymic precursor cells and examined the development of the NKT cell population. NKT cells developed well from AKR thymic precursor cells but not from other precursor cells in both recipient strains. Insulitis and diabetes developed only in the NOD.scid recipients of NOD or NOR precursor cells. When thymic precursor cells of beta2-microglobulin gene-deficient AKR mice, which have a deficient NKT population, were introduced into NOD.scid recipients, both CD4(+) and CD8(+) T-cell populations developed and the recipient mice developed insulitis and diabetes. We conclude that NKT cells originate from a T-cell-committed thymic precursor population and that the deficiency in the NKT cell population in NOD mice results from intrinsic defects within the T-cell lineage and plays a major role in the development of autoimmune diabetes in the presence of both the NOD thymus and antigen-presenting cells.

Animals↗
PubMed2001

Angiographic evaluation of middle cerebral artery reperfusion caused by platelet glycoprotein IIb/IIIa receptor complex antagonist murine 7E3 F(ab')2 in a model of focal cerebral ischemia in rats.

OBJECT: Antagonists of the glycoprotein IIb/IIIa (GPIIb/IIIa) receptor complex are currently used for the treatment of acute coronary syndromes. The platelet GPIIb/IIIa mediates platelet aggregation, and blocking this receptor complex can reduce or prevent arterial thrombosis. To study the recanalization efficacy of a GPIIb/IIIa antagonist in treating cerebral ischemia, we investigated the therapeutic effects of murine 7E3 F(ab'), in a focal embolic cerebral ischemia model in rats. METHODS: Focal cerebral ischemia was produced by introducing an autologous thrombus into the right side of the middle cerebral artery (MCA). Thirty male Wistar rats were randomly divided into three groups of 10 rats each: control, 7E3 F(ab')2 administered 1 hour postischemia, and 7E3 F(ab')2 administered 3 hours postischemia. Animals in the therapeutic groups received intravenous infusion of 6 mg/kg 7E3 F(ab')2 at 1 or 3 hours following cerebral embolization. Brain infarct volume, neurobehavioral scores, duration of bleeding, and findings on angiograms of the MCA (before and after infusion) were assessed in all animals. Angiographic evaluation revealed full MCA recanalization in three of 10 animals in each 7E3 F(ab')2 treatment group. Animals in these groups exhibited a significant reduction in infarct volume when compared with animals in the control group: 1) infarct volume 1 hour postischemia, 22 +/- 13.9% (p = 0.005); 2) infarct volume 3 hours postischemia, 22.1 +/- 14.8% (p = 0.008); and 3) infarct volume in control animals, 42.4 +/- 16%. Postischemia treatment with 7E3 F(ab')2 also improved the animal's neurobehavioral performance. The duration of bleeding significantly increased by more than two times, but there was no associated increase in intracerebral hemorrhage in any group. CONCLUSIONS: On the basis of their findings, the authors conclude that murine 7E3 F(ab'), is a potent and safe antiplatelet agent in this experimental focal embolic cerebral ischemia model. Neuronal lesions were significantly reduced when the treatment was delayed up to 3 hours.

Animals↗