Biomedical subjects

Y J Liu

Publications and source records attributed to Y J Liu.

At least 55 records · Page 3Linked to original sources

PubMed2000

[Expression of FAS and FASL genes in brain and effect of flunarizine on these expressions in rats after global ischemia-reperfusion].

AIM: To explore the expression of Fas and FasL genes after ischemia-reperfusion in rats and the effect of flunarizine. METHODS: Ischemia was induced by four-vessel occlusion for 30 min following reperfusion in rats. The biopsy tissues from brain were immunohistochemically assayed with Fas and FasL genes polyclonal antibody. RESULTS: The expression of Fas was increased as early as 6 h after the onset of reperfusion. The peak of the expression of Fas occurred 24-48 h after ischemia-reperfusion. The expression of FasL was observed 12 h after ischemia-reperfusion and peaked at 48-72 h. The expression of Fas and FasL gene was quite obvious in the cortex and hippocampus CA1, the more sensitive areas to ischemic injury. Flunarizine i.p. 10 mg.kg-1 and 20 mg.kg-1 obviously inhibited the expression of Fas and FasL in dose-dependent manner. CONCLUSION: Expression of Fas and FasL in cerebral cortex and hippocampus can be induced by global ischemia-reperfusion. Flunarizine significantly inhibited the expression of Fas and FasL genes following ischemia-reperfusion.

Animals↗

PubMed1999

Lead generation using pharmacophore mapping and three-dimensional database searching: application to muscarinic M(3) receptor antagonists.

By using a pharmacophore model, a geometrical representation of the features necessary for molecules to show a particular biological activity, it is possible to search databases containing the 3D structures of molecules and identify novel compounds which may possess this activity. We describe our experiences of establishing a working 3D database system and its use in rational drug design. By using muscarinic M(3) receptor antagonists as an example, we show that it is possible to identify potent novel lead compounds using this approach. Pharmacophore generation based on the structures of known M(3) receptor antagonists, 3D database searching, and medium-throughput screening were used to identify candidate compounds. Three compounds were chosen to define the pharmacophore: a lung-selective M(3) antagonist patented by Pfizer and two Astra compounds which show affinity at the M(3) receptor. From these, a pharmacophore model was generated, using the program DISCO, and this was used subsequently to search a UNITY 3D database of proprietary compounds; 172 compounds were found to fit the pharmacophore. These compounds were then screened, and 1-[2-(2-(diethylamino)ethoxy)phenyl]-2-phenylethanone (pA(2) 6.67) was identified as the best hit, with N-[2-(piperidin-1-ylmethyl)cycohexyl]-2-propoxybenz amide (pA(2) 4. 83) and phenylcarbamic acid 2-(morpholin-4-ylmethyl)cyclohexyl ester (pA(2) 5.54) demonstrating lower activity. As well as its potency, 1-[2-(2-(diethylamino)ethoxy)phenyl]-2-phenylethanone is a simple structure with limited similarity to existing M(3) receptor antagonists.

Animals↗

PubMed1999

The nature of the principal type 1 interferon-producing cells in human blood.

Interferons (IFNs) are the most important cytokines in antiviral immune responses. "Natural IFN-producing cells" (IPCs) in human blood express CD4 and major histocompatibility complex class II proteins, but have not been isolated and further characterized because of their rarity, rapid apoptosis, and lack of lineage markers. Purified IPCs are here shown to be the CD4(+)CD11c- type 2 dendritic cell precursors (pDC2s), which produce 200 to 1000 times more IFN than other blood cells after microbial challenge. pDC2s are thus an effector cell type of the immune system, critical for antiviral and antitumor immune responses.

CD40 Ligand↗

PubMed1999

The CC chemokine receptor-7 ligands 6Ckine and macrophage inflammatory protein-3 beta are potent chemoattractants for in vitro- and in vivo-derived dendritic cells.

Dendritic cell migration to secondary lymphoid tissues is critical for Ag presentation to T cells necessary to elicit an immune response. Despite the importance of dendritic cell trafficking in immunity, at present little is understood about the mechanisms that underlie this phenomenon. Using a novel transwell chemotaxis assay system, we demonstrate that the CC chemokine receptor-7 (CCR7) ligands 6Ckine and macrophage inflammatory protein (MIP)-3 beta are selective chemoattractants for MHC class IIhigh B7-2high bone marrow-derived dendritic cells at a potency 1000-fold higher than their known activity on naive T cells. Furthermore, these chemokines stimulate the chemotaxis of freshly isolated lymph node dendritic cells, as well as the egress of skin dendritic cells ex vivo. Because these chemokines are expressed in lymphoid organs and 6Ckine has been localized to high endothelial venules and lymphatic endothelium, we propose that they may play an important role in the homing of dendritic cells to lymphoid tissues.

Animals↗

PubMed1999

Toward a role of dendritic cells in the germinal center reaction: triggering of B cell proliferation and isotype switching.

We have reported previously that in vitro generated dendritic cells (DC) can directly regulate B cell responses. Recently, germinal center DC (GCDC) were identified within B cell follicles. Due to their particular localization, we have tested in the present study whether GCDC could contribute to key events characteristic of the GC reaction. Our present results demonstrate that 1) ex vivo GCDC induce a dramatic GC B cell expansion upon CD40 and IL-2 activation and drive plasma cell differentiation, 2) this property is shared by GCDC and blood DC, but not by Langerhans cells, 3) IL-12 production by GCDC is critical in GC B cell expansion and differentiation, and 4) importantly, GCDC also induce IL-10-independent isotype switching toward IgG1. These observations support the novel concept that GCDC directly contribute to the germinal center reaction.

Animals↗

PubMed1999

Reciprocal control of T helper cell and dendritic cell differentiation.

It is not known whether subsets of dendritic cells provide different cytokine microenvironments that determine the differentiation of either type-1 T helper (TH1) or TH2 cells. Human monocyte (pDC1)-derived dendritic cells (DC1) were found to induce TH1 differentiation, whereas dendritic cells (DC2) derived from CD4+CD3-CD11c- plasmacytoid cells (pDC2) induced TH2 differentiation by use of a mechanism unaffected by interleukin-4 (IL-4) or IL-12. The TH2 cytokine IL-4 enhanced DC1 maturation and killed pDC2, an effect potentiated by IL-10 but blocked by CD40 ligand and interferon-gamma. Thus, a negative feedback loop from the mature T helper cells may selectively inhibit prolonged TH1 or TH2 responses by regulating survival of the appropriate dendritic cell subset.

Apoptosis↗

PubMed1999

Nucleotide sequence and thermostability of pND324, a 3.6-kb plasmid from Lactococcus lactis.

A 3.6-kb plasmid, designated pND324, was isolated from Lactococcus lactis subsp. lactis LL57-1. Sequence analysis revealed the presence of three open reading frames, rep324, orfX1 and orfX2, which are flanked by two non-coding regions, ori324 and cisE. The minimal replication region of pND324 consists of ori324 and rep324, which is closely related to the lactococcal theta-type replicons of the pWV02/pCI305 family. pND324 was stable at both 30 degrees C and 37 degrees C, whereas derivatives that lack cisE were highly unstable at 37 degrees C, indicating that cisE is essential for thermostability. Sequences that are similar to orfX1 are commonly present in the lactococcal theta-type plasmids. The orfX2 product is homologous to TrfA, a 43-kDa protein of the E. coli theta-type plasmid RK2 required for replication and maintenance. Plasmid deletion and stability analyses showed that orfX2 is involved in the thermostability of pND324. Based on the minimal replication region of pND324, an integrative cloning vector, designated pND421, was constructed. In L. lactis LM0230, cells that carried pND421 integrated into its host chromosomal DNA could be recovered readily following incubation at 37 degrees C for 40 generations. The integrated plasmid was totally stable for at least 100 generations without selection at 30 degrees C.

Base Sequence↗

PubMed1999

Influence of placental 11beta-hydroxysteroid dehydrogenase (11beta-HSD) inhibition on glucose metabolism and 11beta-HSD regulation in adult offspring of rats.

Placental 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2) converts glucocorticoids to 11-keto-products and is believed to play an important role in protecting fetuses from higher maternal glucocorticoid levels. Recent reports have speculated that prenatal glucocorticoid exposure leads to fetal growth retardation and adult offspring hypertension and hyperglycemia. To investigate the effects of placental 11beta-HSD2 inhibition on glucose metabolism and the 11beta-HSD system in adult offspring, pregnant rats were treated with daily injections of carbenoxolone (CBX), an inhibitor of 11beta-HSD. The offspring of the maternal CBX treatment group showed reduced birth weight (treated v control, 5.6 +/- 0.5 v 6.4 +/- 0.4 g, P < .0001). In adult offspring of the maternal CBX treatment group, plasma hemoglobin A1c was significantly increased (7.3% +/- 1.8% v 4.8% +/- 0.3%, P < .01) and glucose intolerance was shown on the oral glucose tolerance test. The gene expression of hepatic 11beta-HSD1 and renal 11beta-HSD2 was decreased 87.6% (P < .05) and 52.3% (P < .01) in adult offspring of the maternal CBX treatment group, whereas renal 11beta-HSD1 was not significantly altered. The change in 11beta-HSD activity corresponded to the change in the gene expression. These results suggest that inhibition of placental 11beta-HSD2 causes growth retardation, glucose intolerance, and partial suppression of the 11beta-HSD system in the offspring.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1999

Iontophoretic current and intradermal microdialysis recovery in humans.

When microdialysis (MD) is used to study dermal delivery by iontophoresis, the effects of current may alter MD recovery through an increase in temperature, a change of pH, hyperemia, and dermal hydration. The objective of this work is to assess whether these effects of current may cause a measurable change in the retrodialysis of a model compound (sodium fluorescein, Fl). Two linear MD-probes were inserted in the forearm dermis of healthy human volunteers and perfused with Ringer's solution containing Fl. Two identical iontophoresis chambers (IC, filled with NaCl in propylene glycol) were placed over the MD-probes. Each IC included a laser Doppler flowmetry probe to monitor skin blood flow. At one IC, current was applied for two periods of 30 min each, separated by 30 min of no current. No current was applied to the control site. Dialysate samples were collected every 5 min and analyzed for Fl by HPLC. Skin blood flow increased in response to iontophoresis, on average, 570% compared to the control site. However, there was no difference in the recovery of Fl between the current-active site versus the control site, and between the period with applied current versus the period with no current. In conclusion, iontophoretic current did not affect intradermal MD recovery.

Administration, Cutaneous↗

PubMed1999

Phylogenetic relationships among ascomycetes: evidence from an RNA polymerse II subunit.

In an effort to establish a suitable alternative to the widely used 18S rRNA system for molecular systematics of fungi, we examined the nuclear gene RPB2, encoding the second largest subunit of RNA polymerase II. Because RPB2 is a single-copy gene of large size with a modest rate of evolutionary change, it provides good phylogenetic resolution of Ascomycota. While the RPB2 and 18S rDNA phylogenies were highly congruent, the RPB2 phylogeny did result in much higher bootstrap support for all the deeper branches within the orders and for several branches between orders of the Ascomycota. There are several strongly supported phylogenetic conclusions. The Ascomycota is composed of three major lineages: Archiascomycetes, Saccharomycetales, and Euascomycetes. Within the Euascomycetes, plectomycetes, and pyrenomycetes are monophyletic groups, and the Pleosporales and Dothideales are distinct sister groups within the Loculoascomycetes. We confirm the placement of Neolecta within the Archiascomycetes, suggesting that fruiting body formation and forcible discharge of ascospores were characters gained early in the evolution of the Ascomycota. These findings show that a slowly evolving protein-coding gene such as RPB2 is useful for diagnosing phylogenetic relationships among fungi.

Ascomycota↗

PubMed1999

Ca2+ signaling in mouse pancreatic polypeptide cells.

Ca2+ signaling was studied in pancreatic polypeptide (PP)-secreting cells isolated from mouse islets of Langerhans. After measuring the cytoplasmic Ca2+ concentration ([Ca2+]i), the cells were identified by immunocytochemistry. Most PP-cells reacted to carbachol and epinephrine with prompt and reversible elevation of [Ca2+]i, often manifested as slow oscillations. The carbachol effect was muscarinic, because it was inhibited by atropine. Beta-adrenergic elevation of cAMP explains the epinephrine stimulation, which was mimicked by an activator of adenylate cyclase and blocked by an inhibitor of protein kinase A. The responses to carbachol and epinephrine apparently involve depolarization with opening of voltage-dependent Ca2+ channels, because the effects were prevented by the Ca2+ channel antagonist methoxyverapamil and by diazoxide, which activates ATP-dependent K+ (K(ATP)) channels. Being equipped with K(ATP) channels, the PP-cells often responded to tolbutamide or high concentrations of glucose with elevation of [Ca2+]i. Somatostatin reversed the [Ca2+]i elevation obtained by carbachol, epinephrine, tolbutamide, and glucose. These preliminary studies support the idea that glucose has a direct stimulatory effect on the PP-cells, which can be masked by locally released somatostatin. Expressing both K(ATP) channels and voltage-dependent Ca2+ channels, the PP-cells share fundamental regulatory mechanisms with other types of islet cells.

Animals↗

PubMed1998

Identification of functional human splenic memory B cells by expression of CD148 and CD27.

Memory B cells isolated from human tonsils are characterized by an activated cell surface phenotype, localization to mucosal epithelium, expression of somatically mutated immunoglobulin (Ig) variable (V) region genes, and a preferential differentiation into plasma cells in vitro. In spleens of both humans and rodents, a subset of memory B cells is believed to reside in the marginal zone of the white pulp. Similar to tonsil-derived memory B cells, splenic marginal zone B cells can be distinguished from naive follicular B cells by a distinct cell surface phenotype and by the presence of somatic mutations in their Ig V region genes. Although differences exist between human naive and memory B cells, no cell surface molecules have been identified that positively identify all memory B cells. In this study, we have examined the expression of the receptor-type protein tyrosine phosphatase CD148 on human B cells. CD148(+) B cells present in human spleen exhibited characteristics typical of memory B cells. These included an activated phenotype, localization to the marginal zone, the expression of somatically mutated Ig V region genes, and the preferential differentiation into plasma cells. In contrast, CD148(-) B cells appeared to be naive B cells due to localization to the mantle zone, the expression of surface antigens typical of unstimulated B cells, and the expression of unmutated Ig V region genes. Interestingly, CD148(+) B cells also coexpressed CD27, whereas CD148(-) B cells were CD27(-). These results identify CD148 and CD27 as markers which positively identify memory B cells present in human spleen. Thus, assessing expression of these molecules may be a convenient way to monitor the development of memory B cell responses in immunocompromised individuals or in vaccine trials.

B-Lymphocyte Subsets↗

PubMed1998

Antigen receptor engagement turns off the V(D)J recombination machinery in human tonsil B cells.

The germinal center (GC) is an anatomic compartment found in peripheral lymphoid organs, wherein B cells undergo clonal expansion, somatic mutation, switch recombination, and reactivate immunoglobulin gene V(D)J recombination. As a result of somatic mutation, some GC B cells develop higher affinity antibodies, whereas others suffer mutations that decrease affinity, and still others may become self-reactive. It has been proposed that secondary V(D)J rearrangements in GCs might rescue B cells whose receptors are damaged by somatic mutations. Here we present evidence that mature human tonsil B cells coexpress conventional light chains and recombination associated genes, and that they extinguish recombination activating gene and terminal deoxynucleotidyl transferase expression when their receptors are cross-linked. Thus, the response of the recombinase to receptor engagement in peripheral B cells is the opposite of the response in developing B cells to the same stimulus. These observations suggest that receptor revision is a mechanism for receptor diversification that is turned off when antigen receptors are cross-linked by the cognate antigen.

Animals↗

PubMed1998

The normal counterpart of IgD myeloma cells in germinal center displays extensively mutated IgVH gene, Cmu-Cdelta switch, and lambda light chain expression.

Human myeloma are incurable hematologic cancers of immunoglobulin-secreting plasma cells in bone marrow. Although malignant plasma cells can be almost eradicated from the patient's bone marrow by chemotherapy, drug-resistant myeloma precursor cells persist in an apparently cryptic compartment. Controversy exists as to whether myeloma precursor cells are hematopoietic stem cells, pre-B cells, germinal center (GC) B cells, circulating memory cells, or plasma blasts. This situation reflects what has been a general problem in cancer research for years: how to compare a tumor with its normal counterpart. Although several studies have demonstrated somatically mutated immunoglobulin variable region genes in multiple myeloma, it is unclear if myeloma cells are derived from GCs or post-GC memory B cells. Immunoglobulin (Ig)D-secreting myeloma have two unique immunoglobulin features, including a biased lambda light chain expression and a Cmu-Cdelta isotype switch. Using surface markers, we have previously isolated a population of surface IgM-IgD+CD38+ GC B cells that carry the most impressive somatic mutation in their IgV genes. Here we show that this population of GC B cells displays the two molecular features of IgD-secreting myeloma cells: a biased lambda light chain expression and a C&mu-Cdelta isotype switch. The demonstration of these peculiar GC B cells to differentiate into IgD-secreting plasma cells but not memory B cells both in vivo and in vitro suggests that IgD-secreting plasma and myeloma cells are derived from GCs.

B-Lymphocytes↗

PubMed1998

Origin of slow and fast oscillations of Ca2+ in mouse pancreatic islets.

1. Pancreatic islets exposed to 11 mM glucose exhibited complex variations of cytoplasmic Ca2+ concentration ([Ca2+]i) with slow (0.3-0.9 min-1) or fast (2-7 min-1) oscillations or with a mixed pattern. 2. Using digital imaging and confocal microscopy we demonstrated that the mixed pattern with slow and superimposed fast oscillations was due to separate cell populations with the respective responses. 3. In islets with mixed [Ca2+]i oscillations, exposure to the sarcoplasmic-endoplasmic reticulum Ca2+-ATPase inhibitors thapsigargin or 2,5-di-tert-butylhydroquinone (DTBHQ) resulted in a selective disappearance of the fast pattern and amplification of the slow pattern. 4. In addition, the protein kinase A inhibitor RP-cyclic adenosine 3',5'-monophosphorothioate sodium salt transformed the mixed [Ca2+]i oscillations into slow oscillations with larger amplitude. 5. Islets exhibiting only slow oscillations reacted to low concentrations of glucagon with induction of the fast or the mixed pattern. In this case the fast oscillations were also counteracted by DTBHQ. 6. The spontaneously occurring fast oscillations seemed to require the presence of cAMP-elevating glucagon, since they were more common in large islets and suppressed during culture. 7. Image analysis revealed [Ca2+]i spikes occurring irregularly in time and space within an islet. These spikes were preferentially observed together with fast [Ca2+]i oscillations, and they became more common after exposure to glucagon. 8. Both the slow and fast oscillations of [Ca2+]i in pancreatic islets rely on periodic entry of Ca2+. However, the fast oscillations also depend in some way on paracrine factors promoting mobilization of Ca2+ from intracellular stores. It is proposed that such a mobilization in different cells within a tightly coupled islet syncytium generates spikes which co-ordinate the regular bursts of action potentials underlying the fast oscillations.

Animals↗

PubMed1998

The cytokine profile expressed by human dendritic cells is dependent on cell subtype and mode of activation.

In the present study, we have analyzed the pattern of cytokines expressed by two independent dendritic cell (DC) subpopulations generated in vitro from human cord blood CD34+ progenitors cultured with granulocyte-macrophage CSF and TNF-alpha. Molecularly, we confirmed the phenotypic differences discriminating the two subsets: E-cadherin mRNA was only detected in CD1a+-derived DC, whereas CD68 and factor XIIIa mRNAs were observed exclusively in CD14+-derived DC. Semiquantitative reverse-transcriptase PCR analysis revealed that both DC subpopulations spontaneously expressed IL-1alpha, IL-1beta, IL-6, IL-7, IL-12 (p35 and p40), IL-15, IL-18, TNF-alpha, TGF-beta, macrophage CSF, and granulocyte-macrophage CSF, but not IL-2, IL-3, IL-4, IL-5, IL-9, and IFN-gamma transcripts. Both subpopulations were shown to secrete IL-12 after CD40 triggering. Interestingly, only the CD14+-derived DC secreted IL-10 after CD40 activation, strengthening the notion that the two DC subpopulations indeed represent two independent pathways of DC development. Furthermore, both DC subpopulations expressed IL-13 mRNA and protein following activation with PMA-ionomycin, but not with CD40 ligand, in contrast to IL-12 and IL-10, revealing the existence of different pathways for DC activation. Finally, we confirmed the expression of IL-7, IL-10, and IL-13 mRNA by CD4+ CD11c+ CD3- DC isolated ex vivo from tonsillar germinal centers. Thus, CD14+-derived DC expressing IL-10 and factor XIIIa seemed more closely related to germinal center dendritic cellsGCDC than to Langerhans cells.

Antigens, CD1↗

PubMed1998

Somatic hypermutation introduces insertions and deletions into immunoglobulin V genes.

During a germinal center reaction, random mutations are introduced into immunoglobulin V genes to increase the affinity of antibody molecules and to further diversify the B cell repertoire. Antigen-directed selection of B cell clones that generate high affinity surface Ig results in the affinity maturation of the antibody response. The mutations of Ig genes are typically basepair substitutions, although DNA insertions and deletions have been reported to occur at a low frequency. In this study, we describe five insertion and four deletion events in otherwise somatically mutated VH gene cDNA molecules. Two of these insertions and all four deletions were obtained through the sequencing of 395 cDNA clones (approximately 110,000 nucleotides) from CD38+IgD- germinal center, and CD38-IgD- memory B cell populations from a single human tonsil. No germline genes that could have encoded these six cDNA clones were found after an extensive characterization of the genomic VH4 repertoire of the tonsil donor. These six insertions or deletions and three additional insertion events isolated from other sources occurred as triplets or multiples thereof, leaving the transcripts in frame. Additionally, 8 of 9 of these events occurred in the CDR1 or CDR2, following a pattern consistent with selection, and making it unlikely that these events were artifacts of the experimental system. The lack of similar instances in unmutated IgD+CD38- follicular mantle cDNA clones statistically associates these events to the somatic hypermutation process (P = 0.014). Close scrutiny of the 9 insertion/deletion events reported here, and of 25 additional insertions or deletions collected from the literature, suggest that secondary structural elements in the DNA sequences capable of producing loop intermediates may be a prerequisite in most instances. Furthermore, these events most frequently involve sequence motifs resembling known intrinsic hotspots of somatic hypermutation. These insertion/deletion events are consistent with models of somatic hypermutation involving an unstable polymerase enzyme complex lacking proofreading capabilities, and suggest a downregulation or alteration of DNA repair at the V locus during the hypermutation process.

Amino Acid Sequence↗

PubMed1998

Effects of thyroid hormone (thyroxine) and testosterone on hepatic 11beta-hydroxysteroid dehydrogenase mRNA and activity in pubertal hypothyroid male rats.

To investigate the effects of thyroid hormone and testosterone on 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1), we measured changes in hepatic 11beta-dehydrogenase activity and its mRNA levels in pubertal methimazole (MMI)-induced hypothyroid male rats following treatment with thyroxine ([T4] 50 microg/kg/d) or testosterone (250 microg/d) for 14 days. Hypothyroidism in male rats markedly reduced hepatic 11beta-HSD1 mRNA levels and serum testosterone concentrations (P < .01). Subcutaneous injection of T4 in the hypothyroid rats significantly (P < .01) increased hepatic 11beta-HSD1 mRNA to approximately normal levels and simultaneously increased serum testosterone levels. However, the same daily dose of T4 administered to castrated male hypothyroid rats for 14 days did not elevate hepatic 11beta-HSD1 activity. Treatment with testosterone for 14 days in castrated hypothyroid male rats and rats without gonadectomy significantly (P < .01) increased the enzyme activity without administration of T4. Variations in hepatic 11beta-HSD1 activity were demonstrated to be accompanied by changes in serum testosterone levels in the rats following alteration of the thyroid hormone state. These results suggest that the effect of T4 in increasing the subnormal 11beta-HSD1 gene expression in hypothyroid male rats is mediated by its ability to increase testosterone production in these rats, because in castrated hypothyroid rats, T4 does not elevate 11beta-HSD1 gene expression.

11-beta-Hydroxysteroid Dehydrogenases↗