Search PubMed⌕ Search

Biomedical subjects

Y Wan

Publications and source records attributed to Y Wan.

At least 55 records · Page 3Linked to original sources
PubMed2001

Separable features of the ligand-binding domain determine the differential subcellular localization and ligand-binding specificity of glucocorticoid receptor and progesterone receptor.

Glucocorticoid receptor (GR) and progesterone receptor (PR) are closely related members of the steroid receptor family of transcription factors. The two receptors share a similar domain structure, substantial sequence identity, DNA binding specificity, and the ability to induce many of the same genes. Despite these similarities, the unliganded GR is localized predominantly in the cytoplasm, while unliganded PR is found predominantly in the nucleus. By expressing green fluorescent protein (GFP)-tagged receptors and assessing subcellular localization in living cells by confocal microscopy, we have investigated the structural basis for the differential localization of GR and PR. By constructing a series of GFP-tagged receptor chimeras between GR and PR, we have shown that multiple features in the N-terminal half of the ligand-binding domain (LBD) are the critical determinants that mandate the differential localization of GR and PR. Replacement of residues encompassing helices 1-5 of GR with those of PR yields a receptor that is nuclear. However, this domain is unable to mediate nuclear import by itself when removed from the context of the receptor. The chimeric receptors also indicate that regions encompassing helices 6 and 7 are key determinants of the ligand binding potential and the transactivation potential of receptors. Thus, the determinants specifying localization of hormone-free receptors are separable from those governing ligand binding character.

Animals↗
PubMed2001

Role of dendritic cell-derived cytokines in immune regulation.

Over the last several years, it has become increasingly clear that dendritic cells (DC) are not only critical for the initiation of T cell immunity, but these cells also determine the course of the subsequent immune response (i.e. tolerance vs. immunity, Th1 vs. Th2). However, the mechanisms by which DC can influence the final outcome of a given immune response remain to be understood. Currently, the ability of DC to direct immunity has been linked to: (1) hematopoeitic lineage, (2) maturation stage, and (3) environmental stimuli. While the literature supports each possibility, one common feature of all three hypotheses is that immunological outcome is directly correlated to the profile of DC-derived cytokines. A review of the existing scientific literature strongly suggests that the capacity of DC to orchestrate the immune responses is not an intrinsic quality of the cell, but rather it is the result of environmental stimulation which is reflected in their cytokine and chemokine production.

Cell Communication↗
PubMed2001

Association of H. pylori infection with gastric carcinoma: a Meta analysis.

AIM: To follow the principles of evidence based medicine to reach the integrated results of these studies. METHODS: Twenty-one papers of case-control studies were selected, including 11 on gastric cancer,7 on precancerous lesion of stomach and 3 on lymphoma of stomach. Meta analysis was used to sum up the odds ratios (OR) of these studies. RESULTS: H. pylori vs gastric cancer (intestinal and diffuse type): the odds ratio from the fixed effect model is 3.0016 (95% CI: 2.4197-3.7234, P<0.001). H. pylori vs precancerous lesion of stomach: a random effect model was used to calculate the summary odds ratio and its value is 2.5635 (95% CI: 1.8477-3.5566, P<0.01). H. pylori vs lymphoma of stomach: though the quantity of literature is too small to make Meta analysis, the data of these 3 studies show that lymphoma of stomach is highly associated with H. pylori infections. CONCLUSION: Since it had been revealed that H. pylori infection pre-exists in gastric carcinoma and precancerous lesions, the results of Meta analysis present a strong evidence to support the conclusion that H. pylori infection is a risk factor for gastric carcinoma.

Case-Control Studies↗
PubMed2001

Transmodulation of epidermal growth factor receptor mediates IL-1 beta-induced MMP-1 expression in cultured human keratinocytes.

Ultraviolet (UV) irradiation causes human skin aging and skin cancer through the activation of matrix metalloproteinases (MMPs) which are responsible for the degradation of collagen and tumor progression in human skin. The molecular mechanisms of UV-induced MMPs are yet to be defined. Our previous studies and others suggest that i) the transient activation of cell surface receptors and subsequent activation of MAP kinase cascade contributes to the transcriptional up-regulation of MMPs; and ii) UV-induced expression of pro-inflammatory cytokines such as IL-1 beta and TNF-alpha may also account for the expression of MMPs. However, signaling pathway through which cytokines induce MMP expression remains to be unraveled. In this study, we investigated the pathway that leads to the IL-1 beta-induced up-regulation of MMP-1 in human keratinocytes. IL-1 beta activated epidermal growth factor (EGF) receptor in cultured human keratinocytes in a time- and dose-dependent manner. IL-1 beta-induced EGF receptor tyrosine phosphorylation started at 5 min and peaked at 10 min and remained elevated up to 40 min post IL-1 beta treatment. EGF receptor kinase inhibitor PD153035 and AG1478 inhibited IL-1 beta-induced EGF receptor tyrosine phosphorylation. To test the effect of EGF receptor transactivation on downstream components, we examined the ERK activation by IL-1 beta. We found that IL-1 beta-induced ERK phosphorylation, PD153035 and MEK inhibitor PD98059 blocked IL-1 beta-induced ERK activity. Furthermore, both inhibitors also dramatically reduced IL-1 beta-induced expression of c-jun and c-fos mRNA which are required for up-regulation of MMPs. EGF receptor kinase inhibitor PD153035 and AG1478 and MEK inhibitor PD98059 also blocked IL-1 beta induction of MMP-1 in cultured human keratinocytes. Collectively, our data indicate that IL-1 beta-induced expression of MMP-1 is mediated by transactivation of EGF receptor and through ERK pathway in human keratinocytes.

Blotting, Western↗
PubMed2001

Cloning and characterization of a novel human histamine receptor.

Histamine exerts its numerous physiological functions through interaction with G protein-coupled receptors. Three such receptors have been defined at both the pharmacological and molecular level, while pharmacological evidence hints at the existence of further subtypes. We report here the cloning and characterization of a fourth histamine receptor subtype. Initially discovered in an expressed-sequence tag database, the full coding sequence (SP9144) was subsequently identified in chromosome 18 genomic sequence. This virtual coding sequence exhibited highest homology to the H(3) histamine receptor and was used to generate a full-length clone by polymerase chain reaction (PCR). The distribution of mRNA encoding SP9144 was restricted to cells of the immune system as determined by quantitative PCR. HEK-293 cells transiently transfected with SP9144 and a chimeric G protein alpha-subunit (Galpha(q/i1,2)) exhibited increases in intracellular [Ca(2+)] in response to histamine but not other biogenic amines. SP9144-transfected cells exhibited saturable, specific, high-affinity binding of [(3)H]histamine, which was potently inhibited by H(3) receptor-selective compounds. The rank order and potency of these compounds at SP9144 differed from the rank order at the H(3) receptor. Although SP9144 apparently coupled to Galpha(i), HEK-293 cells stably transfected with SP9144 did not exhibit histamine-mediated inhibition of forskolin-stimulated cAMP levels. However, both [(35)S]GTPgammaS binding and phosphorylation of mitogen-activated protein kinase were stimulated by histamine via SP9144 activation. In both of these assays, SP9144 exhibited evidence of constitutive activation. Taken together, these data demonstrate that SP9144 is a unique, fourth histamine receptor subtype.

Amino Acid Sequence↗
PubMed2001

TGF-beta-induced p38 activation is mediated by Rac1-regulated generation of reactive oxygen species in cultured human keratinocytes.

p38 has been shown to be involved in TGF-beta-induced gene expression, but the upstream of the signaling pathway leading to the activation of p38 is left undefined. We investigated the pathway in cultured human keratinocytes (HaCat cells). Western blot analysis revealed that TGF-beta induced the activation of p38 within 1 h post TGF-beta treatment. H2O2 also strongly induced p38 activation in a time dependent manner. We also observed that TGF-beta-induced p38 activation was inhibited by PDTC, pyrrolidinedithiocarbamate, a known antioxidant, and DPI, diphenylene iodonium chloride, one of the known NADPH oxidase inhibitors. In contrast, TGF-beta-induced Smad2 phosphorylation was not affected. To test whether reactive oxygen species (ROS) is involved in TGF-beta-induced p38 activation, we examined the generation of ROS and activation of NADPH oxidase. FACS analysis showed that TGF-beta induced generation of ROS in time-dependent manner. DPI, an inhibitor of NADPH oxidase, inhibited TGF-beta-induced ROS production. Lucigenin-based NADPH oxidase assay indicated that TGF-beta-induced NADPH oxidase activity started as early as 5 min following treatment and peaked at about 15 min with induction of about 2-folds. The activity remained elevated up to 1 h. Immunofluorescence microscopy study showed that Rac1, one of the subunits of NADPH oxidase, translocated from cytoplasm to the membrane within 5 min. Pretreatment with DPI dramatically reduced TGF-beta-induced NADPH oxidase activity. Collectively, our data suggest that TGF-beta-induced p38 activation is mediated by Rac1-regulated generation of reactive oxygen species in cultured human keratinocytes.

Biological Transport↗
PubMed2001

Arsenite activation of P13K/AKT cell survival pathway is mediated by p38 in cultured human keratinocytes.

BACKGROUND: Arsenic has been considered as a carcinogen. Recently the issue of arsenic in drinking water raised an unprecedented social concern on human health, and yet the molecular mechanisms through which arsenic induces cancer remain unknown. Activation of cell survival pathway leading to the activation of eNOS has been associated with various types of cancer. The objective of this study was to investigate the pathway leading to the activation of eNOS in response to arsenite using human keratinocytes. MATERIALS AND METHODS: Cultured keratinocytes (HaCat cells) were exposed to arsenite with or without pretreatment of various inhibitors. Western blot analysis was performed to determine the activation of p38, AKT, eNOS. EGFR tyrosine phosphorylation was detected by immunoprecipitation and Western blot analysis. pNPP assay was used to measure phosphatase activity in cell lysate. FACS analysis was performed for the determination of generation of reactive oxygen species. RESULTS: Arsenite induced the activation of AKT at both Ser473 and Thr308, and its downstream effector eNOS in cultured human keratinocytes. Arsenite also induced phosphorylation of p38. PI-3-kinase inhibitors, Wortmannin and LY294002 inhibited arsenite-induced phosphorylation of AKT and eNOS but had no effect on phosphorylation of p38. Interestingly, however, SB203580, a known p38 inhibitor, completely inhibited arsenite-induced phosphorylation of AKT and eNOS. Arsenite induced generation of reactive oxygen species and inactivated phosphatase activity, but did not activate EGF receptor tyrosine phosphorylation. CONCLUSIONS: Collectively, our data indicate that arsenite induces activation of AKT and eNOS, via PI-3-kinase and p38 pathway, likely bypassing the activation of EGF receptor in cultured human keratinocytes.

Arsenites↗
PubMed2001

Potentiation of docetaxel antitumor activity by batimastat against mouse forestomach carcinoma.

OBJECTIVE: To improve the treatment of tumors, we studied the combined effects of docetaxel and batimastat (BB-94) on mouse forestomach carcinoma (MFC), and compared them with doxorubicin. METHODS AND RESULTS: In vitro, growth curve analysis, MTT assay and clonogenic assay used to determine the cytotoxic effect of docetaxel or/and BB-94 on MFC cell showed that docetaxel but not BB-94 had a significant cytotoxicity, and the effect of docetaxel wasn't enhanced by BB-94. In early stage MFC tumor model, obvious antitumor effect of docetaxel or doxorubicin given i.v. at maximum tolerated dose (MTD, docetaxel: 20 mg/kg; doxorubicin: 6 mg/kg) every 4 days for 3 injections (q4d x 3), even that of BB-94 (30 mg/kg i. p. qd x 20) was observed. Tumor growth inhibition was greater for docetaxel-batimastat (96.0%) than for doxorubicin-batimastat (88.0%), docetaxel (89.0%), doxorubicin (68.0%) and BB-94 (33.0%), and the effect of docetaxel could be potentiated by BB-94. Docetaxel also showed activity against advanced stage MFC tumor in dose-dependent manner, and was more effective at MTD than doxorubicin with 4/5 regressions, 46.5 days tumor growth delay and 2.8log(10) tumor cell kill. CONCLUSION: Our results suggest that in the MFC model with dose and schedule used, docetaxel is an effective cytotoxic new drug against MFC tumor and BB-94 enchances the antitumor activity of docetaxel.

Animals↗
PubMed2000

Molecular assembly and disassembly: novel photolabile molecular hosts

A new approach to the assembly and photochemical disassembly of molecular hosts is developed. It is based on photoinduced fragmentation in hydroxyalkyl dithianes and utilizes a novel spiro-bis-dithiane as a photolabile molecular tether to link two formylated macromolecular blocks, e.g., formyl calixarenes or formyl dibenzocrown ethers. A key feature of this molecular system is that after an assembly-disassembly cycle the starting macromolecular blocks are recovered intact and can be used again.

Journal Article↗
PubMed2000

Genetically modified dendritic cells prime autoreactive T cells through a pathway independent of CD40L and interleukin 12: implications for cancer vaccines.

Genetic immunization through ex vivo transduction of dendritic cells has been suggested as an effective approach to enhance antitumor immunity by activating both CD4+ and CD8+ T cells. Immunizing mice with dendritic cells transduced with an adenovirus expressing the human melanoma antigen glycoprotein 100 (DCAdhgp100) as a cancer vaccine, we demonstrated complete protective immunity and a potent CTL response against melanomas expressing murine glycoprotein 100 in a CD4+ cell-dependent manner. Surprisingly, however, effective tumor rejection was not the result of cooperation between CD4+ and CD8+ T cells. Protective immunity was completely lost when CD4+ cells were depleted immediately before tumor challenge, whereas it was unaffected by removal of CD8+ cells, establishing a principal role for CD4+ cells in the effector phase of tumor rejection. Neither protective immunity nor CTL generation in this model required interleukin 12, in spite of high levels of IFN-gamma secretion by tumor-reactive T cells. Most notably, the DCAdhgp100 vaccine could elicit protective antitumor CD4+ cells in the absence of CD40 ligand, although it does not bypass the need for CD40-mediated signals to generate melanoma-reactive CTLs. Thus, in contrast to the current thinking that the optimal cancer vaccine should include determinants for both CD4+ and CD8+ cells, the potency of the DCAdhgp100 vaccine appears to be a result of its ability to directly prime autoreactive CD4+ cells through a process that does not require interleukin 12 and CD40 signals.

Adenoviridae↗
PubMed2000

Characterization and modulation of [125I]iberiotoxin-D19Y/Y36F binding in the guinea-pig urinary bladder.

The radioligand binding characteristics of the Ca(2+)-activated K(+) channel ligand [125I]iberiotoxin-D19Y/Y36F were examined in guinea-pig urinary bladder membranes. Saturation analysis revealed a single class of high affinity binding sites in the bladder with a K(D) value of 45.6 pM and a B(max) value of 112 fmol/mg protein. Specific binding was displaced by unlabeled iberiotoxin and penitrem A, but not by blockers of other classes of K(+) channels including alpha-dendrotoxin, margatoxin and apamin. The indole alkaloids, paxilline and verruculogen, significantly increased binding by 4.5- and 4.3-fold, respectively. Tetraacetic acid derivatives such as ethylenediamine tetraacetic acid and ethyleneglycoltetraacetic acid enhanced specific [125I]iberiotoxin-D19Y/Y36F binding about 2.5-fold, which was not attributable to calcium chelation. This increase was due to a significant change in ligand binding affinity (K(D)=6.3 pM), but not due to a change in the B(max), indicating that these compounds may enhance toxin binding via allosteric interactions. Collectively, these results demonstrate that the binding sites for [125I]iberiotoxin-D19Y/Y36F present in the urinary bladder shows a pharmacological profile typical of maxi-K(+) channels and can be modulated, not only by previously known indole alkaloids, but also by tetraacetic acid analogs.

Animals↗
PubMed2000

Prostatic levels of tocopherols, carotenoids, and retinol in relation to plasma levels and self-reported usual dietary intake.

This study evaluated how prostatic levels of antioxidants relate to plasma levels and self-reported usual dietary intake. Definition of these relations may aid in interpreting studies of antioxidant exposure and prostate cancer risk. Between July 1996 and April 1997, plasma and prostatic tissue levels of tocopherols, carotenoids, and retinol were measured in 47 men undergoing radical prostatectomy or transurethral prostatectomy at Loyola University Medical Center, Maywood, Illinois, and an affiliate hospital. Dietary intake was measured by using a 122-item version of the Block Health Habits and History Questionnaire, and correlations were assessed with Pearson's coefficients. Prostatic levels of tocopherols and carotenoids (but not retinol) were significantly correlated with plasma levels (r= 0.31-0.56, p < 0.05-0.0001); the strongest correlations were associated with lycopene, beta-carotene, and gamma-tocopherol (0.56, 0.54, and 0.52, respectively; p < 0.0001). Relative concentrations of tocopherols and carotenoids in prostate tissue were proportionate to those in plasma. No correlation between prostatic levels and reported dietary intake was observed (r = -0.09 to 0.16, p < not significant). Adjustment for energy intake, body mass index, and serum lipids did not impact these relations. These results suggest that plasma levels of tocopherols and carotenoids better reflect prostatic exposure than self-reported usual dietary intake.

Adenocarcinoma↗
PubMed2000

Receptor reserve analysis of the human CCR3 receptor in eosinophils and CCR3-transfected cells.

A novel pharmacological study of CCR3 receptor reserve in a CCR3-transfected cell (CREM3) and human eosinophils was done; functional responses measured were increases in intracellular calcium and chemotaxis. Eotaxin, eotaxin-2, monocyte chemoattractant protein-4 (MCP-4), RANTES, and MCP-3 induced similar maximal eosinophil chemotaxis, whereas MCP-3 and RANTES induced submaximal calcium responses in eosinophils compared to eotaxin, MCP-4, and eotaxin-2. This suggested a receptor reserve in the chemotaxis response. Receptor reserve was quantitated for eotaxin. Occupancy of all CCR3 receptors was required for a maximal calcium response in both CREM3 and eosinophils (reserve = 1.0 or 0.17, respectively); the stimulus-calcium response relationship was linear, indicating no receptor reserve. In contrast, in eosinophils a large receptor reserve (6.5) was found for chemotaxis, where occupancy of 15% receptors drove half-maximal responses. These studies indicate that CCR3 interacts with G-proteins that are poorly coupled to the calcium response, whereas coupling efficiency and/or amplification to the chemotaxis apparatus in human eosinophils is significantly greater.

Animals↗
PubMed2000

Synergy in induction of increased renal allograft survival after portal vein infusion of dendritic cells transduced to express TGFbeta and IL-10, along with administration of CHO cells expressing the regulatory molecule OX-2.

Dendritic cells (DC), generated from C57BL/6 mouse bone marrow cells cultured with GM-CSF and IL-4 for 9 days, were engineered to express constitutively the cytokines TGFbeta, IL-10, and IL-12, using adenovirus vectors constructed using an E1-deleted replication-deficient recombinant adenovirus carrying the appropriate cDNA for the relevant cytokines (Ad-TGFbeta, Ad-IL-12, or Ad-IL-10). C3H mice receiving nontransduced DC or pretransplant infusion of DC-Ad-LacZ showed increased survival of C57BL/6 renal grafts relative to that of control nonimmunized mice. Transfusion of Ad-IL-12-transduced DC abolished this increased survival, leading to a graft survival equivalent to that of controls with no DC. Optimal graft survival was seen in the group receiving a mixture of DC transduced with constructs for both IL-10 and TGFbeta. There was a correlation between increased graft survival and both inhibition of the induction of CTL and enhancement of a polarization to produce type-2 cytokines (IL-4, IL-10, and TGFbeta) on antigen-specific restimulation in vitro. These effects were more pronounced following concomitant infusion of CHO cells transfected with a full-length cDNA for murine OX-2.

Adenoviridae↗
PubMed2000

Macrophage cell-conditioned medium promotes cholinergic differentiation of undifferentiated progenitors and synergizes with nerve growth factor action in the developing basal forebrain.

Conditioned medium from stimulated microglia and from the monocyte/macrophage cell line (RAW 264.7; MC-CM) promotes the differentiation of cholinergic neurons from undifferentiated progenitors in the septal nuclei and adjacent basal forebrain (BF). We have studied the regulation of this process by measuring the activity of choline acetyltransferase (ChAT) in cultured BF taken from embryonic day 16 rat brain. Inhibition of either xanthine oxidase with allopurinol or nitric oxide synthase with N(G)-monomethyl-l-arginine produces a small but significant improvement in the efficacy of MC-CM while inclusion of pyrrolidine dithiocarbamate, a hydroxyl radical scavenger widely used as an antioxidant, lowers MC-CM-induced ChAT activity. Addition of nerve growth factor (NGF) but not brain-derived neurotrophic factor or glial-derived neurotrophic factor together with MC-CM has a synergistic effect on both ChAT activity and ChAT mRNA, raising ChAT activity as much as 29-fold and ChAT mRNA almost 15-fold. While MC-CM raised mRNA for trkA, the effect was not synergistic with NGF. mRNA for the common neurotrophin receptor (p75NTR) showed a modest synergistic increase. Blockade of the Ras/Raf/ERK [extracellular signal-regulated kinase, also known as mitogen-activated protein [(MAP) kinase] signal transduction pathway with either PD28059 (an inhibitor of MAP kinase/ERK kinase kinase or MEK) or N-acetyl-S-farnesyl-l-cysteine (an inhibitor of Ras farnesylation and, hence, activation) inhibited the action of MC-CM. Moreover, a subpopulation of cells responded rapidly to MC-CM with an increased appearance of phosphorylated ERK. Because NGF also utilizes this pathway, synergy may occur along this signal transduction pathway.

Acetylcysteine↗