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Biomedical subjects

L Wu

Publications and source records attributed to L Wu.

At least 451 records · Page 25Linked to original sources
PubMed1997

Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts: method and efficacy of adenovirus-mediated transduction.

OBJECTIVE: Allograft-targeted immunosuppressive gene therapy may inhibit recipient immune activation and provide an alternative to systemic immunosuppression. We studied the optimal technique and efficacy of intracoronary gene transfer of viral interleukin-10 and human transforming growth factor-beta 1 in a rabbit model of heterotopic heart transplantation. METHODS: Replication-defective adenoviral vectors were constructed, expressing viral interleukin-10 (AdSvIL10) or transforming growth factor-beta 1 (AdCMVTGF-beta 1). Intracoronary delivery of vectors was accomplished ex vivo by either bolus injection or slow infusion. The allografts were implanted heterotopically in recipient rabbits and collected 4 days after the operation. Vector dose was 4 x 10(9) to 6 x 10(10) pfu/gm of donor heart. Transfer was confirmed by DNA amplification for both genes. Gene product expression in tissue was quantified by immunoassay and visualized by immunohistochemical staining. RESULTS: Allograft viral uptake was only 9.9% +/- 2.4% with bolus injection, but increased to 80.5% +/- 6.8% at 1 ml/min infusion rate (p = 5 x 10(-14)). Uptake ratio was not affected by vector quantity or slower infusion rates. Transforming growth factor-beta 1 was consistently detected in allografts infected with AdCMVTGF-beta 1, but not with control adenovirus or AdSvIL10. Expression was proportional to infused vector quantity and reached 10 ng/gm of allograft at infused 10(10) pfu/gm. Transforming growth factor-beta 1 was also detected in recipient's serum at less than 1 ng/ml. Viral interleukin-10 was detected in minor amounts only (< 1 ng/gm) in allografts infected with AdvIL10 up to 5 x 10(10) pfu/gm. Nevertheless, it was detected in recipient serum at concentrations up to 0.4 ng/ml. CONCLUSIONS: Intracoronary gene transfer of immunosuppressive cytokines to cardiac allografts during cold preservation is feasible. Slow infusion is superior to bolus injection. In vivo effects on allograft rejection remain to be determined.

Adenoviridae↗
PubMed1997

Stem cell antigen 2 expression in adult and developing mice.

Stem cell antigen 2 (Sca-2) expression can distinguish the most immature T-lymphocyte precursors in the thymus from the hemopoietic stem cells. Sequence analysis of the Sca-2 protein showed that Sca-2 is a glycosylphosphatidylinositol (GPI) anchored molecule that shares some characteristics with the members of the Ly-6 multigene family, and that it is the same as the thymic shared antigen-1 (TSA-1). Here we extend these studies and critically reassess the expression of the Sca-2/TSA-1 antigen in hematopoietic tissues of adult and developing mice. With more sensitive methods we show that the distribution of Sca-2/TSA-1 differs from existing reports. We find especially high expression of Sca-2/TSA1 at day 14 of fetal development.

Animals↗
PubMed1997

Miz1, a novel zinc finger transcription factor that interacts with Msx2 and enhances its affinity for DNA.

Msx2 is a homeobox gene with a regulatory role in inductive tissue interactions, including those that pattern the skull. We demonstrated previously that individuals affected with an autosomal dominant disorder of skull morphogenesis (craniosynostosis, Boston type) bear a mutated form of Msx2 in which a histidine is substituted for a highly conserved proline in position 7 of the N-terminal arm of the homeodomain (p148h). The mutation behaves as a dominant positive in transgenic mice. The location of the mutation in the N-terminal arm of the homeodomain, a region which in other homeodomain proteins plays a key part in protein-protein interactions, prompted us to undertake a yeast two hybrid screen for Msx2-interacting proteins. Here we present a functional analysis of one such protein, designated Miz1 (Msx-interacting-zinc finger). Miz1 is a zinc finger-containing protein whose amino acid sequence closely resembles that of the yeast protein, Nfi-1. Together these proteins define a new, highly conserved protein family. Analysis of Miz1 expression by Northern blot and in situ hybridization revealed a spatiotemporal pattern that overlaps that of Msx2. Further, Miz1 is a sequence specific DNA binding protein, and it can function as a positive-acting transcription factor. Miz1 interacts directly with Msx2 in vitro and enhances the DNA binding affinity of Msx2 for a functionally important element in the rat osteocalcin promoter. The p148h mutation in Msx2 augments the Miz1 effect on Msx2 DNA binding, suggesting a reason why this mutation behaves in vivo as a dominant positive, and providing a potential explanation of the craniosynostosis phenotype.

Amino Acid Sequence↗
PubMed1997

Cell-autonomous defects in dendritic cell populations of Ikaros mutant mice point to a developmental relationship with the lymphoid lineage.

The transcription factor Ikaros is a major determinant of lymphocyte differentiation. Mice homozygous for an Ikaros dominant-negative (DN-/-) mutation lack all cells of lymphoid origin, including T, B, and natural killer (NK) cells. Mice homozygous for an Ikaros null allele lack B and NK cells but display specific defects in T lymphocytes. Nonetheless, both Ikaros mutant lines make an excess of monocytes and macrophages. Here we report that the production of subsets of antigen-presenting dendritic cells (DCs) is also defective. By constructing bone marrow chimeras, we demonstrate that the Ikaros-mediated defect in lymphocytes and DCs is intrinsic to their precursors and is not environment dependent. The selective defects in DCs manifested with the Ikaros null mutation suggest a tight linkage between development of T cells and CD8alpha+ DCs. The complete lack of DCs in the lymphoid organs of Ikaros DN-/- micke points to an essential role for the Ikaros gene family in the development of all DCs.

Animals↗
PubMed1997

Carbon monoxide-induced vasorelaxation and the underlying mechanisms.

1. Carbon monoxide (CO) induced a concentration-dependent relaxation of isolated rat tail artery tissues which were precontracted with phenylephrine or U-46619. This vasorelaxing effect of CO was independent of the presence of the intact endothelium. 2. The CO-induced vasorelaxation was partially inhibited by the blockade of either the cyclicGMP pathway or big-conductance calcium-activated K (KCa) channels. When both the cyclicGMP pathway and KCa channels were blocked, the CO-induced vasorelaxation was completely abolished. 3. Incubation of vascular tissues with hemin, in order to enhance the endogenous production of CO, suppressed the phenylephrine-induced vasocontraction in a time- and concentration-dependent manner. The hemin-induced suppression of the vascular contractile response to phenylephrine was abolished after the vascular tissues were co-incubated with either oxyhaemoglobin or zinc protoporphyrin-IX, suggesting an induced endogenous generation of CO from vascular tissues. 4. The effect of hemin incubation on vascular contractility did not involve the endogenous generation of nitric oxide. 5. Our results suggest that CO may activate both a cyclicGMP signalling pathway and KCa channels in the same vascular tissues, and that the endogenously generated CO may significantly affect the vascular contractile responses.

Animals↗
PubMed1997

Gene-based neurotransmitter modulation in cerebellar granule neurons.

The human glutamic acid decarboxylase (GAD) gene was transferred into rat cerebellar granule neurons. Following adenoviral-mediated gene transfer, nearly 100% of the neurons had transgene expression that persisted for the duration of their survival in culture. GABA levels were elevated both in the growth media and in lysates of GAD-modified granule neurons. In GAD-modified neurons, extracellular GABA levels steadily increased with time, whereas intracellular GABA levels peaked 10 days after gene transfer. GAD-modified neurons released both glutamate and GABA into the surrounding media before and after potassium-induced stimulation, but only the release of glutamate was sensitive to potassium stimulation. These data suggest that glutamatergic neurons, which initially contained no detectable GABA, can be genetically modified to release GABA constitutively.

Adenoviridae↗
PubMed1997

International Classification of Diseases, 9th Revision, Clinical Modification codes in discharge abstracts are poor measures of complication occurrence in medical inpatients.

OBJECTIVES: The authors tested the ability of International Classification of Diseases, 9th Revision, Clinical Modification (ICD-9-CM) codes in discharge abstracts to identify medical inpatients who experienced an in-hospital complication, using complications identified through chart review as the gold standard. METHODS: Two sets of ICD-9-CM codes were used: an inclusive set including many medical diagnoses that may also be coexistent complicating conditions on admission rather than complications and an exclusive set consisting primarily of ICD-9-CM-specified complication and adverse drug event codes. RESULTS: Neither set performed well as a diagnostic test for complication occurrence according to receiver operating characteristic analysis (ROC areas were 0.61 for the inclusive set and 0.55 for the exclusive set). Sensitivities of the ICD-9-CM codes for complications were 0.34 for the inclusive set and 0.14 for the exclusive set. Corresponding positive predictive values were 0.32 and 0.37, respectively. Sensitivities of code definitions for individual complications were generally poor, less than 0.5 in most cases. CONCLUSIONS: The authors conclude that ICD-9-CM codes in discharge abstracts are poor measures of complication occurrence.

Abstracting and Indexing↗
PubMed1997

Differential effects of platelet-derived growth factor BB in accelerating wound healing in aged versus young animals: the impact of tissue hypoxia.

Platelet-derived growth factor BB (PDGF-BB) plays a central role in wound healing. Platelet-derived growth factor has been shown to accelerate healing in preclinical and clinical studies, but its wound-healing effects in older animals have not been examined. An ischemic incisional model in young female (5 months) and aged male (60 months) rabbits was used to determine the influence of platelet-derived growth factor on day 14 postwounding with reduced blood supply. Wounds in aged ischemic animals were severely impaired in breaking strength compared with their nonischemic control wounds and wounds in young animals (p < 0.001). Topical platelet-derived growth factor (10 microg per wound) partially reversed the reduction in breaking strength in aged ischemic animals but was ineffective in young animals. Histologic studies showed that new granulation tissue deposition and mononuclear cell infiltration was dramatically lower in ischemic wounds of aged animals compared with their control wounds as well as wounds from young animals under both nonischemic and ischemic conditions. Platelet-derived growth factor partially reversed this deficit in old ischemic wounds but not in young ischemic wounds. Epithelial growth was reduced in wounds from aged animals compared with wounds from young animals after 14 days of healing. Platelet-derived growth factor treatment increased ischemic wound epithelial growth in aged ischemic animals about threefold compared with paired controls. In conclusion, wound healing in aged rabbits was severely impaired by ischemia, and a single topical platelet-derived growth factor treatment partially reversed this deficit.

Aging↗
PubMed1997

Acute and chronic animal models for excessive dermal scarring: quantitative studies.

Excessive scarring in the form of keloids and hypertrophic scars continues to be a clinical problem for some patients. The lack of an animal model for such scarring has been an obstacle to studying the cellular and molecular biology of these entities. Previous observations made by the authors that some surgical scars in the rabbit ear remain raised for months after wounding prompted us to investigate whether the rabbit ear might provide a model by which to study excessive dermal scarring. After establishing the model in preliminary study, 40 excisional wounds, 6 mm in diameter, were created over the ventral surface of rabbit ears. Elevated scars were treated with either intralesional triamcinolone acetonide or saline at day 16 postwounding. On day 22, 25 scar wounds were used for thorough histomorphometric analysis, 15 wounds were eliminated prior to analysis because of invagination of epithelial tissue, which made analysis difficult. Total area of scar and Hypertrophic Index, a ratio comparing scar prominence with the thickness of adjacent unwounded tissue, were measured for 25 (62 percent) of the resulting scars. Both total area of scar and Hypertrophic Index were found to be significantly decreased in the steroid-treated group (p < 0.02 and < 0.03, respectively). In a chronic form of this model, in which larger excisions were taken, an excessive accumulation of both new collagen and cartilage over 9 months was observed. An animal model for excessive dermal scarring that allows quantitation of scar formation and, at an early stage, can be modulated in a predictable way with intralesional corticosteroid treatment is presented. This model may parallel hypertrophic scarring in humans and thus might provide a tool by which to study its pathophysiology and objectively evaluate therapeutic modalities.

Animals↗
PubMed1997

Discovery of cyanovirin-N, a novel human immunodeficiency virus-inactivating protein that binds viral surface envelope glycoprotein gp120: potential applications to microbicide development.

We have isolated and sequenced a novel 11-kDa virucidal protein, named cyanovirin-N (CV-N), from cultures of the cyanobacterium (blue-green alga) Nostoc ellipsosporum. We also have produced CV-N recombinantly by expression of a corresponding DNA sequence in Escherichia coli. Low nanomolar concentrations of either natural or recombinant CV-N irreversibly inactivate diverse laboratory strains and primary isolates of human immunodeficiency virus (HIV) type 1 as well as strains of HIV type 2 and simian immunodeficiency virus. In addition, CV-N aborts cell-to-cell fusion and transmission of HIV-1 infection. Continuous, 2-day exposures of uninfected CEM-SS cells or peripheral blood lymphocytes to high concentrations (e.g., 9,000 nM) of CV-N were not lethal to these representative host cell types. The antiviral activity of CV-N is due, at least in part, to unique, high-affinity interactions of CV-N with the viral surface envelope glycoprotein gp120. The biological activity of CV-N is highly resistant to physicochemical denaturation, further enhancing its potential as an anti-HIV microbicide.

Acquired Immunodeficiency Syndrome↗
PubMed1997

Utilization of C-C chemokine receptor 5 by the envelope glycoproteins of a pathogenic simian immunodeficiency virus, SIVmac239.

We examined chemokine receptors for the ability to facilitate the infection of CD4-expressing cells by viruses containing the envelope glycoproteins of a pathogenic simian immunodeficiency virus, SIVmac239. Expression of either human or simian C-C chemokine receptor CCR5 allowed the SIVmac239 envelope glycoproteins to mediate virus entry and cell-to-cell fusion. Thus, distantly related immunodeficiency viruses such as SIV and the primary human immunodeficiency virus type 1 isolates can utilize CCR5 as an entry cofactor.

Animals↗
PubMed1997

Roles of Wee1 and Nim1 protein kinases in regulating the switch from mitotic division to sexual development in Schizosaccharomyces pombe.

In self-fertile strains of the fission yeast Schizosaccharomyces pombe, nitrogen starvation initiates a program of sexual development in which cells express mating pheromones and receptors, arrest cell cycle progression in G1, and conjugate. This process is dependent on Rum1, an inhibitor of the Cdc2-Cdc13 and Cdc2-Cig2 cyclin B kinases. The M-phase induction activity of Cdc2-Cdc13 is inhibited by Wee1 tyrosine kinase, which phosphorylates Cdc2 on tyrosine-15. We report here that Wee1 activity is also important for mating. This discovery arose from studies of Nim1, a kinase which promotes mitosis by inhibiting Wee1. Nim1 was previously thought to have an important role in promoting mitosis during nitrogen starvation, but our studies revealed that Nim1 protein drops to an undetectable level within 15 min of nitrogen depletion. In contrast, Wee1 remains abundant, and tyrosine-phosphorylated Cdc2 is detected for at least 4 h after resuspension of cells in nitrogen-free medium. This suggested that maintenance of Wee1 activity may be important during the early stages of nitrogen starvation, a proposal confirmed by the observation that mating efficiency is reduced ca. fivefold in wee1- cells. Transcriptional induction of genes encoding mating factors and receptors is also delayed in wee1- cells. The wee1- mating defect is suppressed by deletion of cig2+, which encodes a B-type cyclin that promotes the onset of S and inhibits conjugation. These findings indicate that Wee1 and Rum1 act jointly to inhibit Cdc2 and promote sexual development in nitrogen-starved cells.

Ammonium Chloride↗
PubMed1997

The 36-kilodalton embryonic-type cytoplasmic polyadenylation element-binding protein in Xenopus laevis is ElrA, a member of the ELAV family of RNA-binding proteins.

The translational activation of several maternal mRNAs in Xenopus laevis is dependent on cytoplasmic poly(A) elongation. Messages harboring the UUUUUAU-type cytoplasmic polyadenylation element (CPE) in their 3' untranslated regions (UTRs) undergo polyadenylation and translation during oocyte maturation. This CPE is bound by the protein CPEB, which is essential for polyadenylation. mRNAs that have the poly(U)12-27 embryonic-type CPE (eCPE) in their 3' UTRs undergo polyadenylation and translation during the early cleavage and blastula stages. A 36-kDa eCPE-binding protein in oocytes and embryos has been identified by UV cross-linking. We now report that this 36-kDa protein is ElrA, a member of the ELAV family of RNA-binding proteins. The proteins are identical in size, antibody directed against ElrA immunoprecipitates the 36-kDa protein, and the two proteins have the same RNA binding specificity in vitro. C12 and activin receptor mRNAs, both of which contain eCPEs, are detected in immunoprecipitated ElrA-mRNP complexes from eggs and embryos. In addition, this in vivo interaction requires the eCPE. Although a number of experiments failed to define a role for ElrA in cytoplasmic polyadenylation, the expression of a dominant negative ElrA protein in embryos results in an exogastrulation phenotype. The possible functions of ElrA in gastrulation are discussed.

Animals↗
PubMed1997

Cyclin D1 antisense RNA destabilizes pRb and retards lung cancer cell growth.

To investigate the role of cyclin D1 in the regulation of lung cancer cell growth, we created five stably transfected cell lines carrying a cyclin D1 antisense construct. The transfected cells exhibited a marked decrease in the rate of cell growth, in contrast to the original lines (A549 and NCI-H441). The expression of several cell cycle-regulating proteins, including cyclin A, the cyclin-dependent kinases (cdk) 2 and cdk4, in addition to cyclin D1 itself, was markedly decreased. The expression of one cdk inhibitor, p21WAF1/CIP1, increased in the A549-derived cell lines. A specific target of cyclin D1 activity, the growth-suppressing product of the retinoblastoma gene, pRb, exhibited decreased expression and a decreased level of phosphorylation in the transfected cells. Decreased expression of pRb due to a significant increase in its turnover rate suggested that the stability of the protein may depend on phosphorylation by cyclin D1-dependent cdk activity. In addition to the impact on pRb stability, decreased expression of cyclin D1 induced susceptibility to cell death after withdrawal of exogenous growth factors in the antisense transfected cell lines, a response that was not observed in the original cancer cell lines. We conclude that abrogation of cyclin D1 overexpression in lung cancer cells disrupts several key pathways that are required for uncontrolled cell growth and induces those that lead to cell death after growth factor deprivation. Therefore, we speculate that use of antisense cyclin D1 expression in appropriate gene vectors could be a useful method for retarding lung cancer cell growth in accessible tumors such as those of the lung epithelium.

CDC2-CDC28 Kinases↗
PubMed1997

Chart review. A need for reappraisal.

Imperfections of chart review as a data collection method are detailed, with particular emphasis on the issue of reliability. The authors identify 125 journal articles published between 1991 and 1992 in which the authors' conclusions were based principally on chart review findings and were either clinical or health care epidemiology studies, or quality assessment studies. Eight percent of epidemiology studies and 56% of quality assessment studies presented data on interrater reliability. Forty-four percent of epidemiology studies and 20% of quality assessment studies did not describe the number of chart reviewers involved. Forty-three percent of epidemiology studies and 28% of quality assessment studies used supplementary data sources. The authors conclude that the validity and utility of studies based on chart review would be enhanced by attention to interrater reliability and the use of supplementary data sources.

Data Collection↗
PubMed1997

MPM-2 antibody-reactive phosphorylations can be created in detergent-extracted cells by kinetochore-bound and soluble kinases.

The MPM-2 antibody labels mitosis-specific and cell cycle-regulated phosphoproteins. The major phosphoproteins of mitotic chromosomes recognized by the MPM-2 antibody are DNA topoisomerase II (topoII) alpha and beta. In immunofluorescence studies of PtK1 cytoskeletons, prepared by detergent lysis in the presence of potent phosphatase inhibitors, the MPM-2 antibody labels phosphoproteins found at kinetochores, chromosome arms, midbody and spindle poles of mitotic cells. In cells extracted without phosphatase inhibitors, labeling of the MPM-2 antibodies at kinetochores is greatly diminished. However, in cytoskeletons this epitope can be regenerated through the action of kinases stably bound at the kinetochore. Various kinase inhibitors were tested in order to characterize the endogenous kinase responsible for these phosphorylations. We found that the MPM-2 epitope will not rephosphorylate in the presence of the broad specificity kinase inhibitors K-252a, staurosporine and 2-aminopurine. Several other inhibitors had no effect on the rephosphorylation indicating that the endogenous MPM-2 kinase at kinetochores is not p34cdc2, casein kinase II, MAP kinase, protein kinase A or protein kinase C. The addition of N-ethylmaleimide inactivated the endogenous kinetochore kinase; this allowed testing of several purified kinases in the kinetochore rephosphorylation assay. Active p34cdc2-cyclin B, casein kinase II and MAP kinase could not generate the MPM-2 phosphoepitope. However, bacterially expressed NIMA from Aspergillus and ultracentrifuged mitotic HeLa cell extract were able to catalyze the rephosphorylation of the MPM-2 epitope at kinetochores. Furthermore, fractionation of mitotic HeLa cell extract showed that kinases that create the MPM-2 epitope at kinetochores and chromosome arms are distinct. Our results suggest that multiple kinases (either soluble or kinetochore-bound), including a homolog of mammalian NIMA, can create the MPM-2 phosphoepitope. The kinetochore-bound kinase that catalyzes the formation of the MPM-2 phosphoepitope may play an important role in key events such as mitotic kinetochore assembly and sister chromatid separation at anaphase.

Adenosine Triphosphate↗
PubMed1997

Different types of acid-base changes at birth, fetal heart rate patterns, and infant outcome at 4 years of age.

BACKGROUND: To study the relationship between different types of acidemia in umbilical artery blood at birth, fetal heart rate (FHR) patterns during labor, and infant long-term outcome. METHODS: Case-control study of 48 infants with pure high base deficit (base deficit (BD) > or = 12 mmol/L and pCO2 < 8.0 kPa), 51 with mixed acidemia (BD > or = 12 mmol/L and pCO2 > or = 8.0 kPa), born at or after term, and controls matched for maternal age, parity, and gestational age at birth. FHR patterns during labor and the results of developmental screening at age 4 were compared between the groups. RESULTS: Late decelerations were associated with pure high base deficit and complicated variable decelerations with mixed acidemia. Both types of acidemia were correlated with Apgar scores below 7 at 1 minute, and mixed acidemia with more admissions to the neonatal intensive care unit. Developmental screening at age 4 years showed no significant differences between infants with mixed acidemia or pure high base deficit and controls. Twelve infants with mixed acidemia and six controls had deficits in language/speech development. CONCLUSIONS: Late decelerations may be an indicator of a metabolic component of acidemia and complicated variable decelerations an indicator of mixed acidemia. The higher rate of admissions to the neonatal intensive care unit in cases with mixed acidemia may suggest that a concomitant hypercapnia (resulting in lower pH) in metabolic acidemia at birth may be of importance for the outcome. A possible relation between acidemia at birth and deficits in speech/language development should be further evaluated.

Acid-Base Equilibrium↗