Biomedical subjects

Q Chen

Publications and source records attributed to Q Chen.

At least 541 records · Page 30Linked to original sources

PubMed1996

The quaking gene product necessary in embryogenesis and myelination combines features of RNA binding and signal transduction proteins.

The mouse quaking gene, essential for nervous system myelination and survival of the early embryo has been positionally cloned. Its sequence implies that the locus encodes a multifunctional gene used in a specific set of developing tissues to unite signal transduction with some aspect of RNA metabolism. The quaking(viable) (qkv) mutation has one class of messages truncated by a deletion. An independent ENU-induced mutation has a nonconservative amino acid change in one of two newly identified domains that are conserved from the C. elegans gld-1 tumour suppressor gene to the human Src-associated protein Sam68. The size and conservation of the quaking gene family implies that the pathway defined by this mutation may have broad relevance for rapid conveyance of extracellular information directly to primary gene transcripts.

Adaptor Proteins, Signal Transducing↗

PubMed1996

The complete BRCA2 gene and mutations in chromosome 13q-linked kindreds.

Breast carcinoma is the most common malignancy among women in developed countries. Because family history remains the strongest single predictor of breast cancer risk, attention has focused on the role of highly penetrant, dominantly inherited genes in cancer-prone kindreds (1). BRCA1 was localized to chromosome 17 through analysis of a set of high-risk kindreds (2), and then identified four years later by a positional cloning strategy (3). BRCA2 was mapped to chromosomal 13q at about the same time (4). Just fifteen months later, Wooster et al. (5) reported a partial BRCA2 sequence and six mutations predicted to cause truncation of the BRCA2 protein. While these findings provide strong evidence that the identified gene corresponds to BRCA2, only two thirds of the coding sequence and 8 out of 27 exons were isolated and screened; consequently, several questions remained unanswered regarding the nature of BRCA2 and the frequency of mutations in 13q-linked families. We have now determined the complete coding sequence and exonic structure of BRCA2 (GenBank accession #U43746), and examined its pattern of expression. Here, we provide sequences for a set of PCR primers sufficient to screen the entire coding sequence of BRCA2 using genomic DNA. We also report a mutational analysis of BRCA2 in families selected on the basis of linkage analysis and/or the presence of one or more cases of male breast cancer. Together with the specific mutations described previously, our data provide preliminary insight into the BRCA2 mutation profile.

BRCA2 Protein↗

PubMed1996

BRCA2 germline mutations in male breast cancer cases and breast cancer families.

The breast cancer susceptibility gene, BRCA2 on chromosome 13q12-13, was recently isolated. Mutations in BRCA2 are thought to account for as much as 35% of all inherited breast cancer as wall as a proportion of inherited ovarian cancer. Many BRCA2-linked families also contain cases of male breast cancer. We have analysed germline DNA from 50 males with breast cancer (unselected for family history) and 26 individuals from site-specific female breast and breast-ovarian cancer families for mutations in BRCA2. All 17 breast-ovarian cancer families have been screened for BRCA1 coding region mutations and none were detected. Conformation-sensitive gel electrophoresis (CSGE) analysis of PCR-amplified DNA followed by direct sequencing was used to detect sequence variants. Three of eleven individuals carry the same mutation, all are of Ashkenazi Jewish descent, supporting the observation by Neuhausen et al. in this issue that there is a common mutation in this population. Eleven truncating mutations and nine polymorphisms were identified -- all were coding region variants. No loss-of-transcript mutations were identified in the sixteen samples for which this analysis was possible. Seven of the nine disease-associated mutations were detected in the 50 men with breast cancers; for thus in our series, BRCA2 mutations account for 14% of male breast cancer, all but one of which had a family history of male and/or female breast cancer.

BRCA2 Protein↗

PubMed1996

Low incidence of BRCA2 mutations in breast carcinoma and other cancers.

Inherited mutant alleles of familial tumour suppressor genes predispose individuals to particular types of cancer. In addition to an involvement in inherited susceptibility to cancer, these tumour suppressor genes are targets for somatic mutations in sporadic cancers of the same type found in the familial forms. An exception is BRCA1, which contributes to a significant fraction of familial breast and ovarian cancer, but undergoes mutation at very low rates in sporadic breast and ovarian cancers. This finding suggests that other genes may be the principal targets for somatic mutation in breast carcinoma. A second, recently identified familial breast cancer gene, BRCA2 (refs 5-8), accounts for a proportion of breast cancer roughly equal to BRCA1. Like BRCA1, BRCA2 behaves as a dominantly inherited tumour suppressor gene. Individuals who inherit one mutant allele are at increased risk for breast cancer, and the tumours they develop lose the wild-type allele by heterozygous deletion. The BRCA2 coding sequence is huge, composed of 26 exons that span 10,443 bp. Here we investigate the rate of BRCA2 mutation in sporadic breast cancers and in a set of cell lines that represent twelve other tumour types. Surprisingly, mutations in BRCA2 are infrequent in cancers including breast carcinoma. However, a probable germline mutation in a pancreatic tumour cell line suggests a role for BRCA2 in susceptibility to pancreatic cancer.

BRCA2 Protein↗

PubMed1996

Generation and application of monoclonal antibody to human pancreatic carcinoma.

An IgG1 mouse monoclonal antibody (MAb) YPC3 against human pancreatic carcinoma was generated by a hybridoma technique. When human tissue sections were examined with immunohistochemical staining, YPC3 MAb reacted with 28 of 32 (87.5%) cases of pancreatic carcinoma tissue; it failed to react with all 11 cases of normal pancreas examined. When 99mTC-radiolabeled YPC3 MAb was administrated intravenously to the nude mice bearing xenograft of pancreatic carcinoma, clear scintigraphic visualization of tumor xenograft was found with SPECT 24 h after injection. Administration of either YPC3 MAb or LAK cells combined with YPC3 MAb could inhibit the growth of the xenografts in nude mice. These results suggest that YPC3 MAb might be an ideal carrier for radioimmunolocalization and immunotherapy of human pancreatic carcinoma.

Animals↗

PubMed1996

Detection of 8-oxo-2'-deoxyguanosine, a marker of oxidative DNA damage, in culture medium from human mesothelial cells exposed to crocidolite asbestos.

Crocidolite asbestos is associated with the development of mesothelioma. Although chromosomal changes have been documented in mesothelial cells, the mechanisms of interaction of crocidolite with DNA remain obscure. Since human mesothelial cells are exquisitely sensitive to asbestos, oxidative DNA damage was measured in an asbestos-exposed human mesothelial cell line (MET5A) by assaying oxidized guanine bases [8-oxo-2'-deoxyguanosine (oxo8dG), 8-oxoguanine (oxo8G), and 8-oxoguanosine (oxo8Gua)] excreted into the spent culture medium after DNA repair or turnover. At growth inhibitory, but not cytolytic concentrations, asbestos caused significant elevation of all bases in the spent medium over a 48-h period. In contrast, riebeckite, a chemically similar, nonfibrous analog of crocidolite did not cause increased adduct release. Results show that oxidative RNA and DNA bases are produced in response to asbestos in target cells of asbestos-induced cancers.

8-Hydroxy-2'-Deoxyguanosine↗

PubMed1996

Construction, properties and specific fluorescent labeling of a bovine prothrombin mutant engineered with a free C-terminal cysteine.

To define the role of phosphatidylserine-induced conformational changes in prothrombin activation during blood coagulation, a recombinant bovine prothrombin was constructed, characterized and shown to have a globally native-like conformation. We introduced a cysteine to replace the penultimate residue (Gly581) of a previously constructed active site mutant, and expressed the double mutant in Chinese hamster ovary cells at the level of 0.6 microgram/ml of cell culture medium. Specific labeling with fluorescein maleimide was accomplished by limited reduction with dithiothreitol to free the engineered cysteine while maintaining the native-like functional properties of the molecule. The average stoichiometry of labeling was 0.84 probe/protein. The location of the probe at the C-terminus was confirmed by proteolysis by native thrombin, by Taipan venom, and by carboxypeptidase Y. Both the double mutant and labeled prothrombin could be activated by snake venoms and the prothrombinase but, as expected, the double mutant meizothrombin did not autolyze as does native meizothrombin. Thus, for the first time, a native-like but specifically labeled prothrombin has been constructed. This molecule will be an essential tool for elucidating the structural role of membranes during prothrombin activation. In addition, the methods described might be usefully applied to labeling of an odd, engineered cysteine in other disulfide bond-containing proteins.

Amino Acid Sequence↗

PubMed1996

Isolation and characterization of glutamine synthetase genes in Chlamydomonas reinhardtii.

To elucidate the role of glutamine synthetase (GS) in nitrogen assimilation in the green alga Chlamydomonas reinhardtii we used maize GS1 (the cytosolic form) and GS2 (the chloroplastic form) cDNAs as hybridization probes to isolate C. reinhardtii cDNA clones. The amino acid sequences derived from the C. reinhardtii clones have extensive homology with GS enzymes from higher plants. A putative amino-terminal transit peptide encoded by the GS2 cDNA suggests that the protein localizes to the chloroplast. Genomic DNA blot analysis indicated that GS1 is encoded by a single gene, whereas two genomic fragments hybridized to the GS2 cDNA probe. All GS2 cDNA clones corresponded to only one of the two GS2 genomic sequences. We provide evidence that ammonium, nitrate, and light regulate GS transcript accumulation in green algae. Our results indicate that the level of GS1 transcripts is repressed by ammonium but induced by nitrate. The level of GS2 transcripts is not affected by ammonium or nitrate. Expression of both GS1 and GS2 genes is regulated by light, but perhaps through different mechanisms. Unlike in higher plants, no decreased level of GS2 transcripts was detected when cells were grown under conditions that repress photorespiration. Analysis of GS transcript levels in mutants with defects in the nitrate assimilation pathway show that nitrate assimilation and ammonium assimilation are regulated independently.

Amino Acid Sequence↗

PubMed1996

A PC-based ultrasonic data acquisition system for computer-aided prosthetic socket design.

A PC-based ultrasound data acquisition system has been developed which uses compound scanning techniques to image a residual limb in a water tank. From the received ultrasonic eco data, the system produces cross-sectional images and reconstructs a three-dimensional (3-D) model of the limb. A commercial software for computer-aided prosthetic socket design was modified so that it can display both the external shape and cross-sectional image of the limb and allow the prosthetist to perform socket design with the help of a visualization of the limb's internal structure. The image resolution and measurement accuracy of the system were tested using a wire phantom and a contrast tissue mimicking phantom, respectively. Preliminary results from amputee patients are presented and the sources of measurement error are discussed.

Computer-Aided Design↗

PubMed1996

The AngR protein and the siderophore anguibactin positively regulate the expression of iron-transport genes in Vibrio anguillarum.

Vibrio anguillarum virulence is associated with the presence of a plasmid-mediated iron-uptake system expressed under iron-limiting conditions, which consists of the siderophore anguibactin and specific iron-transport proteins. This system is maximally expressed under iron-limiting conditions and requires the AngR protein that acts as a positive regulator of anguibactin biosynthesis and also possess an EntE-like enzymatic function that may play a role in anguibactin biosynthesis. In this work, we demonstrate that in addition to possessing these functions related to anguibactin production, AngR also positively regulated transcription of the iron-transport genes fatA and fatB. We also show that transcription of angR is repressed by Fur under iron-rich conditions. In addition, we present evidence that anguibactin itself enhanced transcription of the iron-transport genes fatA and fatB, independently of AngR and the trans-acting factor (TAF) product(s). The presence of either AngR (together with the TAF product(s)) or anguibactin alone led to a partial level of expression of the iron-transport genes fatA and fatB, while full expression is achieved when AngR, the TAF products and anguibactin are all present.

Bacterial Outer Membrane Proteins↗

PubMed1996

Use of post-hospital care by Medicare patients.

BACKGROUND: Medicare's introduction of the Prospective Payment System for hospitals has shortened hospital stays and, as a consequence, has increased the use of post-hospital care. Medicare coverage provides for various types of post-hospital care. This paper examines the characteristics of patients, cities, and hospitals associated with discharge to these different types of post-hospital care. METHODS: A Total of 2248 consecutive Medicare patients having one of five diagnosis related groups (DRGs), who were about to be discharged from 52 hospitals in three cities in 1988-1989, were enrolled in the study. These DRGs comprised approximately one-eighth of all Medicare hospital discharges and 40% of all Medicare-paid post-hospital care. Patients were interviewed in person before discharge and again 6 weeks after discharge. Clinical severity measures were developed from information abstracted from each patient's medical record. For each DRG, multinomial logit regression equations were developed to identify factors associated with the choice of one of four possible discharge locations: home with no formal care, home health care, nursing home care, or rehabilitation. RESULTS: Discharge location could be predicted correctly in 52 to 71% of cases, depending on the DRG. This level of predictive accuracy was significantly greater than relying on the modal discharge location, which accounted for 33 to 62% of cases. Most of the predictive power came from information gathered at the discharge interview. The variables associated significantly with the discharge location varied with the DRG and location examined. Living alone and functional dependency at discharge were the significant predictors found most often. CONCLUSIONS: Rather than assuming that everyone is discharged to the modal location, patient discharge location can be predicted. Much of the explanation can be traced to a few variables such as functional status and living situation. The lack of greater accuracy suggests that factors other than those identified as important by clinical panels are involved in discharge planning for Medicare patients.

Activities of Daily Living↗

PubMed1996

Do rehabilitative nursing homes improve the outcomes of care?

OBJECTIVES: To compare the differences in outcomes of Medicare patients discharged from hospital to two types of nursing homes, rehabilitative and regular, and to rehabilitative facilities. DESIGN: Criteria for distinguishing rehabilitative nursing homes (RNHs) from ordinary nursing homes (NH), based on staffing criteria, were developed by an expert panel and validated on a national sample of nursing homes. Those criteria that significantly discriminated the two types of NHs were then applied to a sample of nursing homes from a study of the outcomes of care for more than 2500 Medicare patients to classify the nursing homes in which patients were discharged. Actual discharge outcomes were compared with optimal outcomes based of predictive equations for different types of treatment (ordinary NH care, RNH care, and formal rehabilitative care). PARTICIPANTS: Medicare patients with strokes an hip fractures discharged from 52 hospitals in three cities. MEASUREMENTS: A disability scale that weights components of ADL measures was used as the primary outcome indicator. Nursing homes were classified as rehabilitative on the basis of the extent of staffing in rehabilitative areas. RESULTS: Patients discharged to various types of care varied on several parameters. After adjusting for these differences, stroke patients fared better when treated in rehabilitative facilities; there was no substantial benefit for RNH care over NH care. Healthier hip fracture patients who received RNH care fared better, but functional change for sicker hip fracture patients was not different among the three groups. CONCLUSIONS: The study suggests that at least a preliminary distinction among NHs can be made on the basis of staffing patterns and that the benefits of the additional staffing may vary with the problem under consideration. More work is needed to establish just what sorts of patients are most likely to benefit from the higher level of NH care.

APACHE↗

PubMed1996

Damage threshold of normal rat brain in photodynamic therapy.

Normal brain tissue response to photodynamic therapy (PDT) must be quantified in order to implement PDT as a treatment of brain neoplasm. We therefore calculated the threshold for PDT-induced tissue necrosis in normal brain using Photofrin (porfimer sodium, Quadralogic Technologies Inc., Vancouver, BC) as the photosensitizer. The absolute light fluence-rate distribution for superficial irradiation and effective attenuation depth were measured in vivo using an invasive optical probe. Photosensitizer uptake in cerebral cortex was measured with chemical extraction and fluorometric analysis. Photodynamic therapy-induced lesion depths at various drug dose levels were measured as a biological end point. The PDT threshold for normal brain necrosis was calculated as in the magnitude of 10(16) photons/cm3. Thus normal rat brain is extremely vulnerable to PDT damage. This suggests that extra precautions must be exercised when PDT is used in brain.

Animals↗

PubMed1996

Tumor oxygenation changes post-photodynamic therapy.

Tumor oxygenation after a photodynamic therapy (PDT) treatment is a critical factor for understanding the posttreatment metabolic pathway of the tumor. It also provides important information for designing combination therapy of PDT and other oxygen-dependent anticancer modalities. In this study, mammary carcinoma in flank and hind leg of C3H mice were subjected to PDT at either subcurative or curative level (12.5 mg/kg Photofrin; 200 or 600 J/cm2, respectively). The before and post-PDT tumor oxygenation was measured with an oxygen-sensitive microelectrode. The data revealed that tumor oxygenation at the time of PDT has a profound effect on posttreatment tumor oxygenation, which may largely be due to an interplay between direct PDT cytotoxicity and PDT damage to the tumor microvasculature. Transient reoxygenation occurred after PDT, which may provide a window for improved combination therapy for other oxygen-dependent modalities.

Adenocarcinoma↗

PubMed1996

Physiological changes and alk gene instability in Pseudomonas oleovorans during induction and expression of alk genes.

The alk genes of Pseudomonas oleovorans, which is able to metabolize alkanes and alkenes, are organized in alkST and alkBFGHJKL clusters, in which the expression of alkBFGHJKL is positively regulated by AlkS. Growth of the wild-type strain GPo1 and P. oleovorans GPo12 alk recombinants on octane resulted in changes of cellular physiology and morphology. These changes, which included lower growth rates and a reduction of the number of CFU due to filamentation, were also seen when the cells were grown on aqueous medium, and the alk genes were induced with dicyclopropylketone, a gratuitous inducer of the alk genes. These effects were seen only for recombinants carrying both alkST and alkBFGHJKL operons. Deletion of parts of either alkB or alkJ, which encode two major Alk proteins located in the cytoplasmic membrane, modified but did not eliminate the effects described above, suggesting that they were due to induction and expression of several alk genes. Continuous growth of the cells in the presence of dicyclopropylketone for about 10 generations led to inactivation, but not elimination, of the alk genes. This resulted in a return of the recombinants to normal physiology and growth.

Alkanes↗

PubMed1996

Regulation of the G-protein-coupled alpha-factor pheromone receptor by phosphorylation.

The alpha-factor pheromone receptor activates a G protein signaling cascade that stimulates MATa yeast cells to undergo conjugation. The cytoplasmic C terminus of the receptor is not necessary for G protein activation but instead acts as a regulatory domain that promotes adaptation to alpha-factor. The role of phosphorylation in regulating the alpha-factor receptor was examined by mutating potential phosphorylation sites. Mutation of the four most distal serine and threonine residues in the receptor C terminus to alanine caused increased sensitivity to alpha-factor and a delay in recovering from a pulse of alpha-factor. 32PO4 labeling experiments demonstrated that the alanine substitution mutations decreased the in vivo phosphorylation of the receptor. Phosphorylation apparently alters the regulation of G protein activation, since neither receptor number nor affinity for ligand was significantly altered by mutation of the distal phosphorylation sites. Furthermore, mutation of the distal phosphorylation sites in a receptor mutant that fails to undergo ligand-stimulated endocytosis caused increased sensitivity to alpha-factor, which suggests that regulation by phosphorylation can occur at the cell surface and is independent of endocytosis. Mutation of the distal serine and threonine residues of the receptor also caused a slight defect in alpha-factor-induced morphogenesis, but the defect was not as severe as the morphogenesis defect caused by truncation of the cytoplasmic C terminus of the receptor. These distal residues in the C terminus play a special role in receptor regulation, since mutation of the next five adjacent serine and threonine residues to alanine did not affect the sensitivity to alpha-factor. Altogether, these results indicate that phosphorylation plays an important role in regulating alpha-factor receptor function.

Binding Sites↗

PubMed1996

Membrane cholesterol alters gallbladder muscle contractility in prairie dogs.

The cause and effect relationship between membrane cholesterol and gallbladder muscle contractility was examined by altering membrane cholesterol to phospholipid mole ratio using cholesterol-rich or cholesterol-free liposomes. Gallbladder single muscle cells, from prairie dogs that were fed either a regular or high-cholesterol (1.2%) diet, were isolated enzymatically with collagenase. Plasma membranes of gallbladder muscle were purified in sucrose gradient. Cholesterol was measured using the cholesterol oxidase method. Phospholipids were measured with the method of G.R. Bartlett (J. Biol. Chem. 234: 466-468, 1959). The results of this experiment are 1) after high-cholesterol feeding, cholesterol contents and cholesterol/ phospholipid mole ratio in plasma membranes of gallbladder muscle increased 90%, and muscle cell contraction in response to cholecystokinin octapeptide decreased 58%; 2) similar changes were observed when normal gallbladder muscle cells were incubated with cholesterol-rich liposomes for 2 h; and 3) the changes induced either in vivo or in vitro were reversed when muscle cells were subsequently incubated with cholesterol-free liposomes for 2-6 h. We conclude that gallbladder muscle may incorporate excess cholesterol into its plasma membrane when exposed to a cholesterol-rich environment, that excess membrane cholesterol impairs muscle contractility, and that these changes appear to be reversible.

Animals↗

PubMed1996

Automated DNA sequencing requiring no DNA template purification.

A critical component in automated fluorescent DNA sequencing is good quality of DNA template. In an effort to reduce the dependence of sequencing success on DNA template quality, the effect of heat-soaked PCR on automated sequencing reactions has been examined. We have found that the heat-soaked PCR protocol considerably improves the overall quality of sequence data and significantly reduces the dependence on the quality of DNA templates. The improvement is corroborated by our ability to obtain over 500 bp of readable sequence per reaction using DNA from E. coli lysates as template obviating DNA purification.

Automation↗