Biomedical subjects

J Shen

Publications and source records attributed to J Shen.

At least 145 records · Page 8Linked to original sources

PubMed2000

A bis-cyclodextrin diselenide with glutathione peroxidase-like activity.

A diselenide, 2,2'-diseleno-bis-beta-cyclodextrin (2-SeCD), was synthesized to imitate the antioxidant enzyme glutathione peroxidase (GPX). The GPX mimic accepts a variety of hydroperoxides as substrates. The GPX activities, reduction of H(2)O(2), tert-butyl hydroperoxide and cumenyl hydroperoxide by glutathione, are 7.4, 4.5 and 10.2 U/micromol, respectively. In contrast to ebselen (PZ51), the diselenide displays high GPX-like activity. The reduction of hydroperoxide by glutathione in the presence of a radical trap shows that the mimic catalyzes the reaction via a non-radical mechanism. A ping-pong mechanism was observed in the steady-state kinetic studies of the 2-SeCD-catalyzed reaction.

Animals↗

PubMed2000

Spectrum of mutations in long-QT syndrome genes. KVLQT1, HERG, SCN5A, KCNE1, and KCNE2.

BACKGROUND: Long-QT Syndrome (LQTS) is a cardiovascular disorder characterized by prolongation of the QT interval on ECG and presence of syncope, seizures, and sudden death. Five genes have been implicated in Romano-Ward syndrome, the autosomal dominant form of LQTS: KVLQT1, HERG, SCN5A, KCNE1, and KCNE2. Mutations in KVLQT1 and KCNE1 also cause the Jervell and Lange-Nielsen syndrome, a form of LQTS associated with deafness, a phenotypic abnormality inherited in an autosomal recessive fashion. METHODS AND RESULTS: We used mutational analyses to screen a pool of 262 unrelated individuals with LQTS for mutations in the 5 defined genes. We identified 134 mutations in addition to the 43 that we previously reported. Eighty of the mutations were novel. The total number of mutations in this population is now 177 (68% of individuals). CONCLUSIONS: KVLQT1 (42%) and HERG (45%) accounted for 87% of identified mutations, and SCN5A (8%), KCNE1 (3%), and KCNE2 (2%) accounted for the other 13%. Missense mutations were most common (72%), followed by frameshift mutations (10%), in-frame deletions, and nonsense and splice-site mutations (5% to 7% each). Most mutations resided in intracellular (52%) and transmembrane (30%) domains; 12% were found in pore and 6% in extracellular segments. In most cases (78%), a mutation was found in a single family or an individual.

Adolescent↗

PubMed2000

New long-wavelength perylenequinones: synthesis and phototoxicity of hypocrellin B derivatives.

Five new derivatives of hypocrellin B were obtained from the reactions of hypocrellin B with ammonia and ethanolamine. Their photophysical and photochemical properties were investigated. The phototoxicity of one compound on AH cells irradiated with red light (lambda = 600-700 nm) was also studied. Their significantly enhanced red absorptivities at wavelengths longer than 600 nm and singlet oxygen-generating function qualify them as promising photodynamic therapy agents.

Ammonia↗

PubMed2000

Activation of ATF6 and an ATF6 DNA binding site by the endoplasmic reticulum stress response.

ATF6 is a member of the basic-leucine zipper family of transcription factors. It contains a transmembrane domain and is located in membranes of the endoplasmic reticulum. ATF6 has been implicated in the endoplasmic reticulum (ER) stress response pathway since it can activate expression of GRP78 and other genes induced by the ER stress response. ER stress appears to activate ATF6 by cleavage from the ER membrane and translocation to the nucleus. However, direct DNA binding by ATF6 had not been demonstrated. In this report, we have identified a consensus DNA binding sequence for ATF6. This site is related to but distinct from ATF1/CREB binding sites. The site was placed in a reporter gene and was specifically activated by ATF6 overexpression and was strongly induced by the ER stress response. A dominant negative form of ATF6 blocked ER stress induction of both ATF6 site and GRP78 reporter genes. We further found that GAL4-ATF6 could be activated by ER stress. These results demonstrate that ATF6 is a direct target of the ER stress response. A proximal sensor of the ER stress response, human IRE1 (hIRE1), was sufficient to activate the ATF6 reporter gene, while a dominant negative form of hIRE1 blocked ER stress activation, suggesting that hIRE1 is upstream of ATF6 in the ER stress signaling pathway.

Activating Transcription Factor 6↗

PubMed2000

Interleukin-6 expression in gut of parasite challenged sheep.

Following challenge with Trichosirongylus colubrifonizis, increased numbers of T-cells and immunoglobulin responses are seen in the intestine of sheep immunised by repeated infection with live worms. IL-6 mRNA expression in the small intestine from T. colubriformis-immunised and naive sheep was determined by in situ hybridisation, whereas CD4(+), IgA(+), IgG(+) cells in the gut were evaluated by immunohistochemistry. There was constitutive expression of IL-6 mRNA by cells in the naive gut, and the number of these cells was increased by parasite challenge. There were corresponding increases in numbers of CD4(+) and TCR gamma/delta(+) T-cells and IgG(+) B-cells. Our data are consistent with a role for IL-6, perhaps produced by CD4(+) and/or TCR gamma/delta(+) T-cells or B-cells, in B-cell terminal differentiation. Infiltration of B-cells, particularly IgG(+) B-cells, may reflect parasite immunity in the host.

Animals↗

PubMed2000

Free entropy and property T factors.

We show that a large class of finite factors has free entropy dimension less than or equal to one. This class includes all prime factors and many property T factors.

Journal Article↗

PubMed2000

Molecular cloning and analysis of a group of genes differentially expressed in cells which overexpress the Hoxa-1 homeobox gene.

The homeobox gene Hoxa-1 is transcriptionally regulated by retinoic acid (RA) and encodes a transcription factor which has been shown to play important roles in cell differentiation and embryogenesis. In order to clone and characterize target genes of Hoxa-1, we utilized differential hybridization screening and cDNA subtractive hybridization methods to identify genes which are differentially expressed in F9-10, a murine F9 teratocarcinoma stem cell line which expresses high levels of exogenous Hoxa-1, compared to F9 wild-type stem cells, which do not express endogenous Hoxa-1 mRNA in the absence of RA. Twenty-eight candidate genes were identified; these genes encode very diverse proteins, including signaling molecules such as BMP-4, the enzyme superoxide dismutase, the cell adhesion molecule cadherin-6, proteins involved in gene transcription such as HMG-1 and SAP18, homeodomain-containing proteins Gbx-2 and Evx-2, and cell cycle regulatory proteins such as the retinoblastoma binding protein-2. Clone 104 encodes a novel protein; the expression of the clone 104 mRNA is also regulated in a fashion very similar to that of the exogenous Hoxa-1 gene in another F9 cell line, called F9-tet-Hoxa1-8, in which the exogenous Hoxa-1 mRNA expression is tightly regulated by a Tet-off gene expression system. These data strongly suggest that clone 104 is a direct downstream target of the transcription factor Hoxa-1. The cDNA sequence of clone 104 is related to that of human ubiquitin carboxyl-terminal hydrolase T. Further characterization of these putative Hoxa-1 target genes will aid in delineating the functions of the Hoxa-1 protein in the differentiation processes which occur during embryogenesis.

Animals↗

PubMed2000

Homodimeric peptides displayed by the major coat protein of filamentous phage.

Peptide libraries displayed by filamentous bacteriophage have proven a powerful tool for the discovery of novel peptide agonists, antagonists and epitope mimics. Most phage-displayed peptides are fused to the N terminus of either the minor coat protein, pIII, or the major coat protein, pVIII. We report here that peptides containing cysteine residues, displayed as N-terminal fusions to pVIII, can form disulfide-bridged homodimers on the phage coat. Phage clones were randomly selected from libraries containing one or two fixed Cys residues, and surveyed for the presence of peptide-pVIII homodimers by SDS-PAGE analysis that involved pretreatment of the phage with reducing or thiol-modifying agents. For all phage whose recombinant peptide contained a single Cys residue, a significant fraction of the peptide-pVIII molecules were displayed as dimers on the phage coat. The dimeric form was in greater abundance than the monomer in almost all cases in which both forms could be reliably observed. Occasionally, peptides containing two Cys residues also formed dimers. These results indicate that, for a given pVIII-displayed peptide bearing a single Cys residue, a significant fraction of the peptide (>40 %) will dimerize regardless of its sequence; however, sequence constraints probably determine whether all of the peptide will dimerize. Similarly, only occasionally do peptides bearing two Cys residues form intermolecular disulfide bridges instead of intramolecular ones; this indicates that sequence constraints may also determine dimerization versus cyclization. Sucrose-gradient analysis of membranes from cells expressing pVIII fused to a peptide containing a single Cys residue showed that dimeric pVIII is present in the cell prior to its assembly onto phage. A model of the peptide-pVIII homodimer is discussed in light of existing models of the structure and assembly of the phage coat. The unique secondary structures created by the covalent association of peptides on the phage surface suggest a role for homo- and heterodimeric peptide libraries as novel sources of bioactive peptides.

Amino Acid Sequence↗

PubMed2000

Detection of tau phosphorylated at threonine 231 in cerebrospinal fluid of Alzheimer's disease patients.

A new sandwich ELISA is described which shows specificity for tau phosphorylated at threonine 231 and preferentially reacts with Alzheimer's disease (AD) brain extracts relative to other dementias. This assay was used to analyze 58 antemortem cerebrospinal fluid samples. Twenty-three of 27 AD samples (85% sensitivity) yielded signals greater than the cutoff, while only one of 31 non-AD samples (97% specificity) were greater. This indicates that detection of phosphotau in cerebrospinal fluid with this sandwich ELISA could prove useful in the diagnosis of AD.

Aged↗

PubMed2000

Identification and characterization of a potent, selective, and orally active antagonist of the CC chemokine receptor-1.

The CC chemokine receptor-1 (CCR1) is a prime therapeutic target for treating autoimmune diseases. Through high capacity screening followed by chemical optimization, we identified a novel non-peptide CCR1 antagonist, R-N-[5-chloro-2-[2-[4-[(4-fluorophenyl)methyl]-2-methyl-1-piperazinyl ]-2-oxoethoxy]phenyl]urea hydrochloric acid salt (BX 471). Competition binding studies revealed that BX 471 was able to displace the CCR1 ligands macrophage inflammatory protein-1alpha (MIP-1alpha), RANTES, and monocyte chemotactic protein-3 (MCP-3) with high affinity (K(i) ranged from 1 nm to 5.5 nm). BX 471 was a potent functional antagonist based on its ability to inhibit a number of CCR1-mediated effects including Ca(2+) mobilization, increase in extracellular acidification rate, CD11b expression, and leukocyte migration. BX 471 demonstrated a greater than 10,000-fold selectivity for CCR1 compared with 28 G-protein-coupled receptors. Pharmacokinetic studies demonstrated that BX 471 was orally active with a bioavailability of 60% in dogs. Furthermore, BX 471 effectively reduces disease in a rat experimental allergic encephalomyelitis model of multiple sclerosis. This study is the first to demonstrate that a non-peptide chemokine receptor antagonist is efficacious in an animal model of an autoimmune disease. In summary, we have identified a potent, selective, and orally available CCR1 antagonist that may be useful in the treatment of chronic inflammatory diseases.

Administration, Oral↗

PubMed2000

Roles for Fgf signaling during zebrafish fin regeneration.

Following amputation of a urodele limb or teleost fin, the formation of a blastema is a crucial step in facilitating subsequent regeneration. Using the zebrafish caudal fin regeneration model, we have examined the hypothesis that fibroblast growth factors (Fgfs) initiate blastema formation from fin mesenchyme. We find that fibroblast growth factor receptor 1 (fgfr1) is expressed in mesenchymal cells underlying the wound epidermis during blastema formation and in distal blastemal tissue during regenerative outgrowth. fgfr1 transcripts colocalize with those of msxb and msxc, putative markers for undifferentiated, proliferating cells. A zebrafish Fgf member, designated wfgf, is expressed in the regeneration epidermis during outgrowth. Furthermore, we show that a specific inhibitor of Fgfr1 applied immediately following fin amputation blocks blastema formation, without obvious effects on wound healing. This inhibitor blocks the proliferation of blastemal cells and the onset of msx gene transcription. Inhibition of Fgf signaling during ongoing fin regeneration prevents further outgrowth while downregulating the established expression of blastemal msx genes and epidermal sonic hedgehog. Our findings indicate that zebrafish fin blastema formation and regenerative outgrowth require Fgf signaling.

Amputation, Surgical↗

PubMed2000

Analysis of domains in the IKKalpha and IKKbeta proteins that regulate their kinase activity.

The IkappaB kinases IKKalpha and IKKbeta are critical in activating the NF-kappaB pathway. Although these proteins have a similar structure that includes kinase, leucine zipper, and helix-loop-helix domains, they exhibit marked differences in their kinase activity and functional properties. For example, IKKbeta has a 10-20-fold higher level of kinase activity for IkappaBalpha than does IKKalpha. Furthermore, disruption of the murine IKKbeta gene, but not the IKKalpha gene, results in severe defects in activating the NF-kappaB pathway. Mice lacking IKKbeta succumb to severe hepatic apoptosis because of failure to activate the NF-kappaB pathway, whereas mice deficient in IKKalpha exhibit skin and skeletal abnormalities and an embryonic lethal phenotype. To better characterize differences in the functional properties of these kinases, hybrid IKK proteins were constructed by domain swapping, and their kinase activity was assayed. These studies demonstrated that differences in the IKKalpha and IKKbeta helix-loop-helix domains are primarily responsible for differences in their kinase activity. In contrast, their kinase and leucine zipper domains exhibited relatively conserved function. These studies further define the properties of IKKalpha and IKKbeta, which are involved in their unique regulatory roles.

Animals↗

PubMed2000

Leptin inhibits bone formation through a hypothalamic relay: a central control of bone mass.

Gonadal failure induces bone loss while obesity prevents it. This raises the possibility that bone mass, body weight, and gonadal function are regulated by common pathways. To test this hypothesis, we studied leptin-deficient and leptin receptor-deficient mice that are obese and hypogonadic. Both mutant mice have an increased bone formation leading to high bone mass despite hypogonadism and hypercortisolism. This phenotype is dominant, independent of the presence of fat, and specific for the absence of leptin signaling. There is no leptin signaling in osteoblasts but intracerebroventricular infusion of leptin causes bone loss in leptin-deficient and wild-type mice. This study identifies leptin as a potent inhibitor of bone formation acting through the central nervous system and therefore describes the central nature of bone mass control and its disorders.

Animals↗

PubMed2000

Induction of lef1 during zebrafish fin regeneration.

Because the transcription factor Lef1 is important for development of several vertebrate organs but has not been investigated for involvement in epimorphic regeneration, we examined its expression during regeneration of amputated adult zebrafish caudal fins. We found that lef1 is markedly up-regulated in the newly formed wound epidermis of the fin regenerate and is maintained in the basal epidermal layer during formation of the regeneration blastema. During regenerative outgrowth, lef1 expression is strongest in epidermal cells adjacent to newly aligned scleroblasts that secrete bone matrix, while it is low or undetectable in epidermis adjacent to mesenchymal areas with either mature bone or proliferative distal blastema cells. This localization is similar to that of the putative fin ray patterning signal Shh. In addition, brief treatments of fin regenerates with retinoic acid or the synthetic Fgfr1 inhibitor SU5402 down-regulate epidermal lef1, similar to their effects on shh. These results suggest a role for Lef1 in scleroblast alignment analogous to that proposed for Shh. Other Wnt signaling pathway members wnt3a, wnt5, and beta-catenin are also expressed in the fin regenerate. Our data suggest that Lef1 has specific roles in inducing and patterning vertebrate regenerating tissue.

Animals↗

PubMed2000

Spectroscopic imaging of glutamate C4 turnover in human brain.

One-dimensional spectroscopic imaging of (13)C-4-glutamate turnover is performed in the human brain with a 6 cc nominal voxel resolution at 4T. Data were acquired with an indirect detection approach using a short spin echo single quantum (1)H-(13)C heteronuclear editing method and a 7 cm surface coil with quadrature (13)C decoupling coils. To analyze the data as a function of tissue type, T(1)-based image segmentation through the surface coil was performed to determine the gray and white matter contributions to each voxel. The tricarboxylic acid (TCA) cycle rate in gray and white matter was then determined using a two-compartment model with the tissue fractionation derived from the image segmentation. The mean values for the TCA cycle rate for occipital gray and white matter from three volunteers was 0.88 +/- 0.12 and 0.28 +/- 0.13 respectively, in agreement with literature data.

Brain↗

PubMed2000

Genotype-phenotype correlation in two frequent mutations and mutation update in type III glycogen storage disease.

Deficiency of glycogen debranching enzyme (AGL) activity causes glycogen storage disease type III (GSD-III). Generalized loss of AGL activity results in GSD-IIIa, and muscle-specific retention of AGL activity results in GSD-IIIb. To date, no common mutation has been described among GSD-III patients, except for three alleles; two linked specifically with GSD-IIIb, and the third found only in North African Jews with GSD-IIIa. Here we report two frequent mutations, each of which was found in the homozygous state in multiple patients, and each of which was associated with a subset of clinical phenotype in those patients with that mutation. A novel point mutation of a single T deletion at cDNA position 3964 (3964delT) was first detected in an African American patient, who has a severe phenotype and early onset of clinical symptoms. The second mutation was an A to G transition at position -12 upstream of the 3' splice site of intron 32 (IVS32-12A > G). This lesion, previously implicated as a IIIb mutation in a Japanese patient, was identified in a confirmed GSD-IIIa Caucasian patient presenting with mild clinical symptoms. These two mutations together account for more than 12% of the molecular defects in the GSD-III patients tested. Our molecular and clinical data suggest a genotype-phenotype correlation for each of these mutations. Furthermore, this current study, coupled with our previous reports, describes the molecular tools necessary for the development of a DNA-based diagnostic test for GSD-III.

Adult↗

PubMed2000

Development of a surface acoustic wave LB membrane immunity sensor.

A new kind of biosensor for immunology was developed by ultrasonic technique and LB membrane. A double delay-line resonator was made by using ST-cut quartz crystal with working frequency of 149.7 MHz. Then a layer of LB membrane was covered on it. When anti-IgM antibody of various concentrations was added to it, the sensor can be used to detect IgM antigen. The biosensor was highly sensitive, small and light. The experimental results showed that the working frequency change of the sensor was proportional to the concentration of antibody with its dilution ratio between 1:10,000 and 1:100. It was also first observed that the frequency curve of the sensor resulting from the reaction of IgM antigen and antibody undulated in the experiment.

Antigen-Antibody Reactions↗

PubMed2000

Study of immortalization and malignant transformation of human embryonic esophageal epithelial cells induced by HPV18 E6E7.

In order to study the effect of viruses and tumor promoters on the tumorigenicity of the esophagus, human embryonic esophageal epithelial cells were infected with human papilloma virus HPV18 E6E7-AAV in synergy with 12-O-tetradecanoylphorbol 13-acetate (TPA) to observe their malignant transformation. The cultured esophageal epithelial cells incubated with HPV18 E6E7-AAV were divided into two groups: the SHEEC1 group was exposed to TPA (5 ng/ml) for 4 weeks at the 5th passage of the cells; the SHEE group served as the control and was cultured in the same medium without TPA. The morphological phenotype, the DNA content during the cell cycle and the chromosomes were analyzed. The tumorigenicity was assessed by colony formation after cultivation in soft agar and transplanting the cells into nude mice. HPV18 E6E7 DNA was assayed by fluorescent in situ hybridization (FISH) and the polymerase chain reaction (PCR). The SHEE group, at its 20th passage, grew as a monolayer with the cells showing anchorage dependence and contact inhibition. The chromosome analysis showed diploidy, and soft-agar cultivation and injection into nude mice showed the cells to be non-tumorigenic. They were therefore immortalized cells. In contrast, the SHEEC1 group (TPA group) showed increased DNA synthesis and a proliferative index that was higher (45%) than that of the SHEE group (34%). The number of large colonies of dense multilayer cells (positively transformed foci) in soft agar was high in SHEEC1 group (4.0%) but low in the SHEE group (0.1%). Tumors resulting from transplantation were observed in all six nude mice injected subcutaneously with cells of the SHEEC1 group but no tumor developed in mice receiving cells of the SHEE group. In both groups of cells, HPV18 E6E7 DNA was positively detected by FISH and PCR. The malignant transformation of human embryonic epithelial cells was induced in vitro by HPV18 E6E7 in synergy with TPA. This is a good evidence for the close relationship between HPV and the etiology and pathogenicity of esophageal carcinoma. It is also a reliable model for studying the cellular and molecular mechanisms of carcinogenesis of esophageal carcinoma.

Cell Transformation, Neoplastic↗