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Biomedical subjects

B Liu

Publications and source records attributed to B Liu.

At least 361 records · Page 20Linked to original sources
PubMed1998

Structure of human G protein G gamma 5 gene GNG5.

Heterotrimeric G-proteins consist of alpha, beta and gamma subunits that together function in numerous signal transduction pathways. Ggamma subunits form dimers with Gbeta-subunits which together can significantly contribute to signal transduction. So far 11 Ggamma subunit isoforms have been found. Ggamma5 appears to be the most prominently occurring Ggamma-subunit and we now report the genomic structure of human GNG5. The gene spans 6 kb and consists of four exons that range from 92 bp to 290 bp and three introns that range from 134 bp to 3.3 kb. Intron-exon boundaries conform to GT-AG consensus splice junctions. Alternative transcriptional start sites that result from two separate, closely linked promoters in the 5'-flanking region of the gene were confirmed by luciferase reporter-gene assay.

5' Untranslated Regions↗
PubMed1998

Efficiency of signalling through cytokine receptors depends critically on receptor orientation.

Human erythropoietin is a haematopoietic cytokine required for the differentiation and proliferation of precursor cells into red blood cells. It activates cells by binding and orientating two cell-surface erythropoietin receptors (EPORs) which trigger an intracellular phosphorylation cascade. The half-maximal response in a cellular proliferation assay is evoked at an erythropoietin concentration of 10 pM, 10(-2) of its Kd value for erythropoietin-EPOR binding site 1 (Kd approximately equal to nM), and 10(-5) of the Kd for erythropoietin-EPOR binding site 2 (Kd approximately equal to 1 microM). Overall half-maximal binding (IC50) of cell-surface receptors is produced with approximately 0.18 nM erythropoietin, indicating that only approximately 6% of the receptors would be bound in the presence of 10 pM erythropoietin. Other effective erythropoietin-mimetic ligands that dimerize receptors can evoke the same cellular responses but much less efficiently, requiring concentrations close to their Kd values (approximately 0.1 microM). The crystal structure of erythropoietin complexed to the extracellular ligand-binding domains of the erythropoietin receptor, determined at 1.9 A from two crystal forms, shows that erythropoietin imposes a unique 120 degrees angular relationship and orientation that is responsible for optimal signalling through intracellular kinase pathways.

Crystallography, X-Ray↗
PubMed1998

[Joint effects of arsenic and lead on lipid peroxidation of human and rat erythrocytes].

The authors observed the joint effects of arsenic and lead on lipid peroxidation(LPO) in vitro and in vivo. In vitro, the authors used the extracted erythrocyte membrane of health adult men and divided them into three exposed groups. The first group was exposed to As2O3 (0, 0.4, 2.0 and 10.0 mmol/L). The second group was exposed to PbAc2 (0, 0.08, 0.4 and 2.0 mmol/L). The third group was exposed to As(10 mmol/L) + Pb (0.08 mmol/L). The results show that there is interaction between As and Pb on LPO of erythrocyte membrane. In vivo, the authors measured LPO levels of rat's plasma, blood glutathione(GSH) and superoxide dismutase(SOD) activity. By using 2 x 2 factorial design, Wistar rats were gavaged with As2O3 2.4 mg/(kg.d) and PbAc2 30 mg/(kg.d) as well as both arsenic and lead in water for 10 days. The results show that in the As-Pb treated rats, the levels of LPO did not change significantly, the GSH levels and SOD activity decreased significantly. The results of factorial analysis revealed a synergic effect of As-Pb on SOD activity.

Adult↗
PubMed1998

MUC4 is a major component of salivary mucin MG1 secreted by the human submandibular gland.

High molecular weight salivary mucin (MG1) is an important component of saliva, contributing to the lubricative and tissue-protective functions of this biological fluid. We have shown previously that the human mucin gene MUC5B is expressed at high levels in sublingual gland and is a significant constituent of MG1. Since many epithelia express multiple mucin genes, it seemed likely that MG1 in salivary secretions is also a heterogeneous mixture of mucin gene products. The aim of this study was to determine whether MUC4, a mucin shown in Northern blotting experiments to be expressed in salivary glands, was a significant protein component of MG1 in salivary secretions. Two cDNA clones containing MUC4 tandem repeats were isolated from a human submandibular gland cDNA library. In addition, recombinant MUC4 produced in a bacterial expression system cross-reacted with an antibody directed against deglycosylated MG1. This shows conclusively that human salivary mucin MG1 contains both MUC5B and MUC4 gene products suggesting that each mucin may perform distinct functions in the oral cavity.

Amino Acid Sequence↗
PubMed1998

Phosphorylation and coupling of zeta-chains to activated T-cell receptor (TCR)/CD3 complexes from peripheral blood T-cells of elderly humans.

Aging is often accompanied by altered T-cell signaling and functions. Signals mediated through the T-cell receptor (TCR)/CD3 complex are associated with tyrosine phosphorylations of zeta-chains by the regulated activities of protein tyrosine kinases p56(lck) and p59(fyn) as well as protein tyrosine phosphatases. In the present investigation, the coupling and phosphorylation of zeta-chains to TCR/CD3 immunocomplexes were examined in peripheral blood T-cells from 13 elderly and young humans stimulated by ligation of the TCR/CD3 with cross-linked anti-CD3epsilon monoclonal antibody OKT3. Western blots analyzing the non-covalent coupling of zeta-chains to TCR/CD3 immunocomplexes from Brij-96 detergent lysates of anti-CD3 ligated T-cells showed that the levels of zeta-chains within TCR/CD3 immunocomplexes from T-cells of elderly and young subjects did not significantly differ. By contrast, the levels of phosphorylated zeta-chains generated during in vitro phosphorylations of TCR/CD3 immunocomplexes from elderly subjects were significantly reduced and averaged 44% of those observed for anti-CD3epsilon ligated T-cells from young subjects. Analyses of the levels of zeta-chain coupling and phosphorylations in T-cells from each of the 13 elderly individuals also showed that the reductions in zeta-chain phosphorylations were heterogeneous and unrelated to modest reductions in coupling. Furthermore, the age-related decreases in zeta-chain phosphorylations were not due to diminished frequencies of CD3epsilon+ cells or densities of CD3epsilon surface receptors and could be observed without reductions in epsilon-chain phosphorylations. These results suggest that aberrancies of zeta-chain phosphorylations can occur in T-cells of elderly humans independent from any uncoupling of zeta-chains to activated TCR/CD3 complexes.

Aged↗
PubMed1998

Inhibition of Stat1-mediated gene activation by PIAS1.

STAT (signal transducer and activator of transcription) proteins are latent cytoplasmic transcription factors that become activated by tyrosine phosphorylation in response to cytokine stimulation. Tyrosine phosphorylated STATs dimerize and translocate into the nucleus to activate specific genes. Different members of the STAT protein family have distinct functions in cytokine signaling. Biochemical and genetic analysis has demonstrated that Stat1 is essential for gene activation in response to interferon stimulation. Although progress has been made toward understanding STAT activation, little is known about how STAT signals are down-regulated. We report here the isolation of a family of PIAS (protein inhibitor of activated STAT) proteins. PIAS1, but not other PIAS proteins, blocked the DNA binding activity of Stat1 and inhibited Stat1-mediated gene activation in response to interferon. Coimmunoprecipitation analysis showed that PIAS1 was associated with Stat1 but not Stat2 or Stat3 after ligand stimulation. The in vivo PIAS1-Stat1 interaction requires phosphorylation of Stat1 on Tyr-701. These results identify PIAS1 as a specific inhibitor of Stat1-mediated gene activation and suggest that there may exist a specific PIAS inhibitor in every STAT signaling pathway.

Amino Acid Sequence↗
PubMed1998

Use of selective serotonin-reuptake inhibitors or tricyclic antidepressants and risk of hip fractures in elderly people.

BACKGROUND: Tricyclic antidepressants (TCAs) are associated with an increased risk of falls and hip fractures in elderly people. Selective serotonin-reuptake inhibitors (SSRIs) are reported to be better tolerated than TCAs. We investigated the risk of hip fractures associated with SSRIs and TCAs. METHODS: This case-control study used administrative healthcare data from the province of Ontario, Canada. 8239 cases-patients aged 66 years or older, treated in hospital between April, 1994, and March, 1995, for hip fracture-were each matched for age and sex to five controls. Logistic regression was used to calculate the odds ratio for hip fracture with adjustment for potential confounding effects produced by concomitant drug use and comorbidity. FINDINGS: With participants who had no exposure to antidepressants as the reference category, the adjusted odds ratio for hip fracture was 2.4 (95% CI 2.0-2.7) for exposure to SSRIs, 2.2 (1.8-2.8) for exposure to secondary-amine TCAs, and 1.5 (1.3-1.7) for exposure to tertiary-amine TCAs. For all types of antidepressants, current use was associated with a higher risk of hip fracture than former use. The odds ratios for hip fracture were higher for new current users than for continuous current users in all three drug classes. The proportion of current use in the low-dose range was 22% for SSRIs, 50% for secondary-amine TCAs, and 58% for tertiary-amine TCAs. INTERPRETATION: Exposure to any of the three classes of antidepressants is associated with a significant increase in the risk of hip fracture. Despite differences in dose distribution, this analysis suggests that SSRIs do not offer an advantage over TCAs in terms of risk of hip fracture.

Accidental Falls↗
PubMed1998

Glutathione regulation of neutral sphingomyelinase in tumor necrosis factor-alpha-induced cell death.

Tumor necrosis factor-alpha (TNFalpha)-induced cell death involves a diverse array of mediators and regulators including proteases, reactive oxygen species, the sphingolipid ceramide, and Bcl-2. It is not known, however, if and how these components are connected. We have previously reported that GSH inhibits, in vitro, the neutral magnesium-dependent sphingomyelinase (N-SMase) from Molt-4 leukemia cells. In this study, GSH was found to reversibly inhibit the N-SMase from human mammary carcinoma MCF7 cells. Treatment of MCF7 cells with TNFalpha induced a marked decrease in the level of cellular GSH, which was accompanied by hydrolysis of sphingomyelin and generation of ceramide. Pretreatment of cells with GSH, GSH-methylester, or N-acetylcysteine, a precursor of GSH biosynthesis, inhibited the TNFalpha-induced sphingomyelin hydrolysis and ceramide generation as well as cell death. Furthermore, no significant changes in GSH levels were observed in MCF7 cells treated with either bacterial SMase or ceramide, and GSH did not protect cells from death induced by ceramide. Taken together, these results show that GSH depletion occurs upstream of activation of N-SMase in the TNFalpha signaling pathway. TNFalpha has been shown to activate at least two groups of caspases involved in the initiation and "execution" phases of apoptosis. Therefore, additional studies were conducted to determine the relationship of GSH and the death proteases. Evidence is provided to demonstrate that depletion of GSH is dependent on activity of interleukin-1beta-converting enzyme-like proteases but is upstream of the site of action of Bcl-2 and of the execution phase caspases. Taken together, these studies demonstrate a critical role for GSH in TNFalpha action and in connecting major components in the pathways leading to cell death.

Cell Death↗
PubMed1998

Overexpression of both p185c-erbB2 and p170mdr-1 renders breast cancer cells highly resistant to taxol.

We recently found that overexpression of p185c-erbB2 in c-erbB2 transfected MDA-MB-435 breast cancer cells (435.eB transfectants) confers a 5-9-fold increase in Taxol resistance. To examine whether Taxol resistance is a common phenomenon in other c-erbB2 overexpressing breast cancer cell lines, we tested a panel of human breast cancer cell lines established from different patients and expressing pl85c-erbB2 at different levels for their sensitivity to Taxol and Taxotere, a synthetic taxoid. Higher expression of p185c-erbB2 in these breast cancer cell lines indeed correlated well with resistance to Taxol and Taxotere, and the degree of resistance was about 100-fold that in c-erbB2-overexpressing 435.eB transfectants, demonstrating that these breast cancer cells are highly resistant to Taxol. Since mdr-1-encoded p-glycoprotein (p170mdr-1) has been implicated in Taxol resistance, we next examined the p170mdr-1 levels in these breast cancer cell lines that are highly resistant to Taxol. Higher levels of p170mdr-1 expression were found in several breast cancer cell lines that are highly resistant to Taxol. Since these same breast cancer cell lines also expressed higher levels of p185c-erbB2, we sought to determine the relative contribution of p185c-erbB2 and p170mdr-1 overexpression to Taxol resistance. We first specifically down-regulated cell surface p185c-erbB2 using anti-p185c-erbB2 monoclonal antibodies and assayed sensitivity of these cells to Taxol. We next specifically inactivated p170mdr-1 function using p170mdr-1 blockers (thioridazine or verapamil) and again assayed Taxol sensitivity. Both p185c-erbB2 down-regulation and p170mdr-1 blockade significantly sensitized the breast cancer cell lines to Taxol. The results indicate that overexpression of either p185c-erbB2 or p170mdr-1 renders human breast cancer cells resistant to Taxol. Furthermore, p185c-erbB2 synergizes with p170mdr-1 conferring higher degrees of Taxol resistance. Finally, combination therapy (down-regulation of p185c-erbB2 plus blocking p170mdr-1 plus administration of Taxol) may be beneficial to breast cancer patients whose tumors express high levels of both p185c-erbB2 and p170mdr-1.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Oxidized LDL damages endothelial cell monolayer and promotes thrombocyte adhesion.

The influence of oxidized low density lipoprotein (LDL) on a human endothelial cell monolayer was examined. The resulting contraction of the oxidized LDL-damaged endothelial cells lets intercellular spaces become enlarged and therefore visible via light microscopy. Electron microscopy reveals that the structural damage facilitates thrombocyte adhesion and formation of microthrombi. Oxidized LDL appears to play a pivotal role in initiating and deteriorating thromboembolic complications.

Blood Platelets↗
PubMed1998

Torsional control of double-stranded DNA branch migration.

DNA branched junctions are analogues of Holliday junction recombination intermediates. Partially mobile junctions contain a limited amount of homology flanking the branch point. A partially mobile DNA branched junction has been incorporated into a synthetic double-stranded circular DNA molecule. The junction is flanked by four homologous nucleotide pairs, so that there are five possible locations for the branch point. Two opposite arms of the branched junction are joined to form the circular molecule, which contains 262 nucleotides to the base of the junction. This molecule represents a system whereby torque applied to the circular molecule can have an impact on the junction, by relocating its branch point. Ligation of the molecule produces two topoisomers; about 87% of the product is a relaxed molecule, and the rest is a molecule with one positive supercoil. The position of the branch point is assayed by cleaving the molecule with endonuclease VII. We find that the major site of the branch point in the relaxed topoisomer is at the maximally extruded position in the relaxed molecule. Upon the addition of ethidium, the major site of the branch point migrates to the minimally extruded position.

Base Sequence↗
PubMed1998

Crystal structure of recombinant human tissue kallikrein at 2.0 A resolution.

Human tissue kallikrein, a trypsin-like serine protease involved in blood pressure regulation and inflammation processes, was expressed in a deglycosylated form at high levels in Pichia pastoris, purified, and crystallized. The crystal structure at 2.0 A resolution is described and compared with that of porcine kallikrein and of other trypsin-like proteases. The active and S1 sites (nomenclature of Schechter I, Berger A, 1967, Biochem Biophys Res Commun 27:157-162) are similar to those of porcine kallikrein. Compared to trypsin, the S1 site is enlarged owing to the insertion of an additional residue, cis-Pro 219. The replacement Tyr 228 --> Ala further enlarges the S1 pocket. However, the replacement of Gly 226 in trypsin with Ser in human tissue kallikrein restricts accessibility of substrates and inhibitors to Asp 189 at the base of the S1 pocket; there is a hydrogen bond between O delta1Asp189 and O gammaSer226. These changes in the architecture of the S1 site perturb the binding of inhibitors or substrates from the modes determined or inferred for trypsin. The crystal structure gives insight into the structural differences responsible for changes in specificity in human tissue kallikrein compared with other trypsin-like proteases, and into the structural basis for the unusual specificity of human tissue kallikrein in cleaving both an Arg-Ser and a Met-Lys peptide bond in its natural protein substrate, kininogen. A Zn+2-dependent, small-molecule competitive inhibitor of kallikrein (Ki = 3.3 microM) has been identified and the bound structure modeled to guide drug design.

Amino Acid Sequence↗
PubMed1998

Myosin-reactive autoantibodies in rheumatic carditis and normal fetus.

EBV-transformed B cells from a 20-week human fetal spleen and from blood of patients with poststreptococcal rheumatic carditis were studied. Most antibodies from nine fetal and six patient myosin-reactive B cell clones were multireactive (reacting with cardiac myosin, Streptococcus pyogenes, and rat cardiac myocytes) which supports a role for molecular mimicry in stimulation of these autoantibodies. Sequence analysis revealed that fetal and patient anti-myosin repertoires were composed of unrelated clones with diverse V gene usages. Fetal and patient antibodies had reduced VH CDR3 length on average and reduced light chain N region addition with a low rate of somatic mutation in the variable region genes, characteristics generally associated with fetal B cells but also with some adult B cells. Five of six myosin-reactive patient clones used VH3, whereas only two of nine fetal clones used VH3, suggesting skewing from the average 50-60% VH3 gene usage found in randomly selected adult and fetal antibodies.

Adult↗
PubMed1998

I-2190A is a potent immunosuppressive drug for vascularized heart transplantation in rats.

The effect of a new immunosuppressant-I-2190A was tested in a rodent heart allograft model. Grafts were transplanted to recipients heterotopically. There were 5 groups: group 1 received no immunosuppressive agents; group 2 was given CsA (2.0 mg/kg, i.p.); group 3 was administered I-2190A (0.1 mg/kg, i.p.) in carboxymethyl cellulose (CMC); group 4 received injection of I-2190A (0.5 mg/kg, i.p. in CMC); group 5 received the combination treatment of I-2190A (0.1 mg/kg) and CsA (2.0 mg/kg). Immunosuppressants were discontinued 14 days after operation. No statistically significant difference in grafts median survival time (MST) was found between group 2 (9.5 days) and group 1 (9 days). The MSTs of grafts in group 3 (22 days, P < 0.05), group 4 (> 100 days, P < 0.01) and group 5 (> 100 days, P < 0.01) were significantly prolonged compared with control group 1 (9 days). Our results suggest that I-2190A is a potent immunosuppressant able to significantly prolong heart allograft survival in rats after a short time treatment, Low-dose I-2190A could potentiate the effect of sub-therapeutic dose of CsA as well.

Animals↗
PubMed1998

Molecular screening of Batten disease: identification of a missense mutation (E295K) in the CLN3 gene.

Batten disease, the juvenile form of neuronal ceroid lipofuscinosis, is a prevalent neuron degenerative disorder of childhood. A 1.02-kb genomic deletion in the Batten disease gene CLN3 has been determined to be a common mutation. We developed a PCR method to screen for this deletion and tested 43 Batten disease probands. We found 36% (31/86) of Batten disease chromosomes did not carry the 1.02-kb deletion. Of the three heterozygotes for the 1.02-kb deletion, a novel G-to-A missense mutation at nucleotide 1020 of the CLN3 cDNA sequence was found on two of the non-1.02-kb deletion chromosomes. The missense mutation resulted in a substitution of glutamic acid (E) by lysine (K) at position 295 (E295 K). The E295 K mutation causes a change in predicted local protein conformation. This glutamic acid is a highly conserved acidic amino acid, being present in human, mouse, dog and yeast, which suggests it may play an important role in the function of the Batten disease protein.

Alternative Splicing↗
PubMed1998

Alphitol, a phenolic substance from Alphitonia zizyphoides which inhibits prostaglandin biosynthesis in vitro.

The new phenolic compound, 3,5-dihydroxy-4-methoxy phenethyl alcohol, named alphitol, and betulinic acid were from the bark of Alphitonia zizyphoides. The chemical structure of alphitol was determined by mass spectrometry in combination with one and two dimensional NMR, including HMBC. Both compounds inhibited prostaglandin biosynthesis in vitro, alphitol with an IC50 value of 0.66mM, which is of the same magnitude as acetyl salicylic acid.

Magnetic Resonance Spectroscopy↗
PubMed1998

The pathogenesis of cutaneous fibrosis.

Cutaneous fibrosis is an integral component of a variety of human disorders including keloids, hypertrophic scar, and most notably, scleroderma. Each has its own etiology and unique clinical characteristics, but all involve the dysregulation of connective tissue metabolism, in particular, the activation of dermal fibroblasts. In this review, we examine various molecular events in scleroderma that may lead to fibroblast activation, and propose a new model to explain the persistence of such activation by scleroderma fibroblasts in the apparent absence of exogenous stimuli.

Animals↗