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K M Murphy

Publications and source records attributed to K M Murphy.

At least 19 recordsLinked to original sources
PubMed2002

Immediate-early gene response to methamphetamine, haloperidol, and quinolinic acid is not impaired in Huntington's disease transgenic mice.

Striatal neurons in symptomatic Huntington's disease (HD) transgenic mice are resistant to a variety of toxic insults, including quinolinic acid (QA), kainic acid and 3-nitropropionic acid. The basis for this resistance is currently unknown. To investigate the possibility that the immediate-early gene (IEG) response is defective in symptomatic HD mice leading to a lack of response to these compounds, we examined the expression of c-Fos and Krox 24 after administration of the indirect dopamine agonist methamphetamine, the dopamine D(2) receptor antagonist haloperidol and the neurotoxin QA in 5- and 10-week-old R6/2 transgenic HD and wild-type mice. Unlike wild-type and pre-symptomatic R6/2 transgenic HD mice, 10-week-old symptomatic HD mice were resistant to methamphetamine-induced gliosis and QA lesion. There was, however, no difference in the number or distribution of c-Fos-immunoreactive nuclei 2 hr after single injections of methamphetamine or haloperidol among 5- and 10-week-old wild-type mice and 5- and 10-week-old R6/2 HD mice. Similarly, despite their resistance to QA-induced lesioning and lower basal levels of krox-24 mRNA, the symptomatic R6/2 mice had equivalent increases in the amount of c-fos and krox-24 mRNA compared to wild-type and pre-symptomatic R6/2 HD mice as determined by in situ hybridization and densitometry 2 hr after QA administration. These data demonstrate that the c-Fos and Krox 24 IEG response to dopamine agonists, dopamine antagonists and neurotoxic insult is functional in symptomatic R6/2 HD mice. Resistance to toxic insult in R6/2 mice may be conferred by interactions of mutant huntingtin with proteins or transcriptional processes further along the toxic cascade.

Animals↗
PubMed2001

Friend of GATA-1 represses GATA-3-dependent activity in CD4+ T cells.

The development of naive CD4+ T cells into a T helper (Th) 2 subset capable of producing interleukin (IL)-4, IL-5, and IL-13 involves a signal transducer and activator of transcription (Stat)6-dependent induction of GATA-3 expression, followed by Stat6-independent GATA-3 autoactivation. The friend of GATA (FOG)-1 protein regulates GATA transcription factor activity in several stages of hematopoietic development including erythrocyte and megakaryocyte differentiation, but whether FOG-1 regulates GATA-3 in T cells is uncertain. We show that FOG-1 can repress GATA-3-dependent activation of the IL-5 promoter in T cells. Also, FOG-1 overexpression during primary activation of naive T cells inhibited Th2 development in CD4+ T cells. FOG-1 fully repressed GATA-3-dependent Th2 development and GATA-3 autoactivation, but not Stat6-dependent induction of GATA-3. FOG-1 overexpression repressed development of Th2 cells from naive T cells, but did not reverse the phenotype of fully committed Th2 cells. Thus, FOG-1 may be one factor capable of regulating the Th2 development.

Animals↗
PubMed2001

MRP1 gene expression level regulates the death and differentiation response of neuroblastoma cells.

We have previously reported a strong correlation between poor prognosis in childhood neuroblastoma (NB) patients and high-level expression of the transmembrane efflux pump, Multidrug Resistance-associated Protein (MRP1), in NB tumour tissue. In this study, we inhibited the endogenous expression of MRP1 in 2 different NB tumour cell lines by stably transfecting an MRP1 antisense expression vector (MRP-AS). Compared with control cells, MRP-AS transfectant cells demonstrated a higher proportion of dead and morphologically apoptotic cells, spontaneous neuritogenesis, and, increased synaptophysin and neurofilament expression. Bcl-2 protein expression was markedly reduced in MRP-AS cells compared to controls. Conversely, we found that the same NB tumour cell line overexpressing the full-length MRP1 cDNA in sense orientation (MRP-S) demonstrated resistance to the neuritogenic effect of the differentiating agent, all-trans-retinoic acid. Taken together, the results suggest that the level of MRP1 expression in NB tumour cells may influence the capacity of NB cells for spontaneous regression in vivo through cell differentiation and death.

Apoptosis↗
PubMed2001

Unexpected characteristics of the IFN-gamma reporters in nontransformed T cells.

Analysis of the IFN-gamma promoter has primarily been conducted by transient expression of reporter constructs in transformed cells. However, the activity of cis elements may differ when expressed transiently compared with their activity within native chromatin. Furthermore, the transcription factors and signaling mechanisms in transformed cells may differ from those in normal T cells. To analyze IFN-gamma promoter regulation in normal T cells, we developed a novel retroviral bottom-strand reporter system to allow the chromatin integration of promoter regions in primary developing T cells. As controls, both the IL-2 and IL-4 promoters were inducible in this system, with the IL-4 reporter having Th2-specific activity. Strikingly, the IFN-gamma promoter exhibited constitutive activity in both Th1 and Th2 subsets, in contrast to the behavior of the endogenous IFN-gamma gene, which is inducible only in Th1 cells. In mapping this activity, we found that the AP-1/GM-CSF site in the distal promoter element is the most critical element for the constitutive activity. Transgenic reporter lines for the IFN-gamma promoter confirmed the constitutive behavior of the isolated IFN-gamma promoter. This constitutive activity was resistant to inhibition by cyclosporin A and was independent of Stat4 and p38 mitogen-activated protein kinase. These results suggest that IFN-gamma promoter regulation may require cis elements residing either downstream or >3.4 kb upstream of the transcriptional start site, involving repression of constitutive activity.

Animals↗
PubMed2001

ACVR1B (ALK4, activin receptor type 1B) gene mutations in pancreatic carcinoma.

DPC4 is known to mediate signals initiated by type beta transforming growth factor (TGFbeta) as well as by other TGFbeta superfamily ligands such as activin and BMP (bone morphogenic proteins), but mutational surveys of such non-TGFbeta receptors have been negative to date. Here we describe the gene structure and novel somatic mutations of the activin type I receptor, ACVR1B, in pancreatic cancer. ACVR1B has not been described previously as a mutated tumor-suppressor gene.

Activin Receptors, Type I↗
PubMed2001

An instructive component in T helper cell type 2 (Th2) development mediated by GATA-3.

Although interleukin (IL)-12 and IL-4 polarize naive CD4(+) T cells toward T helper cell type 1 (Th1) or Th2 phenotypes, it is not known whether cytokines instruct the developmental fate in uncommitted progenitors or select for outgrowth of cells that have stochastically committed to a particular fate. To distinguish these instructive and selective models, we used surface affinity matrix technology to isolate committed progenitors based on cytokine secretion phenotype and developed retroviral-based tagging approaches to directly monitor individual progenitor fate decisions at the clonal and population levels. We observe IL-4-dependent redirection of phenotype in cells that have already committed to a non-IL-4-producing fate, inconsistent with predictions of the selective model. Further, retroviral tagging of naive progenitors with the Th2-specific transcription factor GATA-3 provided direct evidence for instructive differentiation, and no evidence for the selective outgrowth of cells committed to either the Th1 or Th2 fate. These data would seem to exclude selection as an exclusive mechanism in Th1/Th2 differentiation, and support an instructive model of cytokine-driven transcriptional programming of cell fate decisions.

Animals↗
PubMed2001

Stability and commitment in T helper cell development.

Specialized effector activities that are required to eliminate various pathogens involve cytokines produced by specialized CD4(+) T cells subsets, dogmatically termed Th1 and Th2 cells. Despite some oversimplifications, this paradigm is useful for organizing the complex pathways that control forward and backward movements along the road of T cell differentiation. Effective immune memory relies, in part, on the maintenance of the T helper phenotype. This review will address basic issues that relate to the maintenance or reversibility of Th1/Th2 states within the CD4(+) T cell lineage.

Animals↗
PubMed2001

IL-18-stimulated GADD45 beta required in cytokine-induced, but not TCR-induced, IFN-gamma production.

Interleukin-12 (IL-12) and IL-18 induce synergistic transcription of interferon gamma (IFN-gamma) that is T cell receptor (TCR)-independent, not inhibited by cyclosporin A and requires new protein synthesis. To characterize this pathway, we screened for genes that are induced in IL-12- and IL-18-treated T helper type 1 cells. GADD45 beta, which activates mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase kinase 4 (MEKK4), was induced by IL-18 and augmented by IL-12. GADD45 beta expression in naïve CD4+ T cells activated p38 MAPK and selectively increased cytokine-induced, but not TCR-induced, IFN-gamma production. Kinase-inactive MEKK4 and inhibition of the p38 MAPK pathway both selectively inhibit cytokine-induced, but not TCR-induced, IFN-gamma production. Thus, the synergy between IL-12 and IL-18 may involve GADD45 beta induction, which can maintain the MEKK4 and p38 MAPK activation that is necessary for cytokine-induced, but not TCR-induced, IFN-gamma production.

Animals↗
PubMed2001

Characterization of the inflammatory infiltrate during IgE-mediated late phase reactions in the skin of normal and atopic dogs.

In canine and human atopic patients, the intracutaneous injection of offending allergens is followed by the development of both immediate and late-phase reactions. The present study was performed to expand on the characterization and dynamics of inflammatory cell subsets during IgE-mediated late-phase reactions in canine skin. Three normal dogs and three Dermatophagoides farinae-allergic dogs were selected for this experiment. All dogs were challenged intradermally with mite allergen, purified anticanine IgE antibodies (positive control) or phosphate-buffered saline (negative control). Skin biopsies were obtained before and 6, 12 and 24 h post-injection. Sections were stained with metachromatic and eosinophil-specific histological stains. Additionally, we used an immunohistochemical method with antibodies specific for canine leukocyte antigens. This study confirmed the occurrence of a late-phase reaction in atopic skin following allergen challenge, and in normal and atopic canine skin after intradermal injection of IgE-specific antibodies. Whereas early emigrating dermal cells were composed chiefly of neutrophil and activated eosinophil granulocytes, there was an influx of alpha beta T-lymphocytes and dermal dendritic cells in later stages of the late-phase reactions. Because IgE-mediated late-phase reactions resemble spontaneous atopic canine skin lesions, both at macroscopic and microscopic levels, we propose the use of similar challenges to study the anti-inflammatory effects of anti-allergic drugs in a pre-clinical setting.

Allergens↗
PubMed2001

Racial and state differences in the designation of advance directives in nursing home residents.

OBJECTIVES: To examine racial and state differences in the use of advance directives and surrogate decision-making in a nursing home population. DESIGN: A retrospective cohort study. SETTING: Nursing homes in the states of California (CA), Massachusetts (MA), New York (NY), and Ohio (OH). PARTICIPANTS: Nursing home residents: 130,308 in CA, 59,691 in MA, 112,080 in NY, and 98,954 in OH. MEASUREMENTS: Minimum Data Set information concerning resident race and whether or not residents have a living will (LW), a do not resuscitate (DNR) order, or a surrogate decision-maker (SDM). RESULTS: The proportion of LWs, DNR orders, and SDMs varied significantly (P < .0001) by racial categories in each state. In general, whites were distinctly different from other racial categories. Whites were significantly more likely to have a LW (odds ratio (OR) = 1.9 (CA), OR = 2.2 (NY), OR = 4.9 (OH)), a DNR order (OR = 2.4 (CA), OR = 2.4 (MA), OR = 3.3 (NY), OR = 3.2 (OH)), and a SDM (OR = 1.1 (CA), OR = 1.2 (NY), OR = 1.6 (OH)) than were nonwhites, after adjusting for potentially confounding factors. Significant state differences (P < .0001) were observed in LWs, DNR orders, and SDMs and were most pronounced in residents of Ohio, who were significantly more likely to have a LW than were residents in other states (OR = 9.3). CONCLUSIONS: Various resident characteristics explain some of the racial differences, although whites are still more likely to have a LW, a DNR order, or an SDM independent of various resident characteristics included in the adjusted analyses. This pattern is observed in all states, although the ORs varied by state. Some of these differences may be due to distinct cultural approaches to end-of-life care and lack of knowledge and understanding of advance directives. The distinctly higher rates of LWs among all racial groups in Ohio than in other states suggest that states can potentially increase the use of advance directives through intervention.

Advance Directives↗
PubMed2001

A review of techniques for the investigation of otitis externa and otitis media.

Otitis externa, inflammation of the externa ear canal, is relatively easy to diagnose based on the history and physical examination findings. The diagnosis of otitis media, inflammation of the middle ear cavity, is more challenging, with the work-up being both costly and, at times, invasive. The pathogenesis of otitis externa has been classified into predisposing, primary, and perpetuating factors. It is critical to the management of ear disease that the clinician recognize and investigate which factors are contributing to each individual patient's ear disease. Failure to identify and address the primary and/or predisposing factors is the most common cause of chronic recurrent otitis externa. Chronic inflammation of the ear canal leads to the development of the perpetuating factors, which may be the major reason for treatment failure, regardless of the primary cause of the ear disease. In this article, the predisposing, primary, and perpetuating factors involved in ear disease will be presented, along with a review of the techniques used in the diagnosis of otitis externa and otitis media.

Animals↗
PubMed2001

Development of cytochrome oxidase blobs in visual cortex of normal and visually deprived cats.

Cytochrome oxidase (CO) blobs are central to our understanding of the columnar organization and parallel pathways in primate and cat visual cortex. In primates, development of blobs and their relationship with other columnar features of the visual cortex begins pre-natally, before visual experience. In kittens, the supragranular layers differentiate post-natally, after eye opening, raising the possibility that visual experience may influence the development of blobs in cat V1. We have examined the development of blobs in unfolded and flattened sections through the visual cortex of normally reared, dark-reared, monocularly deprived and binocularly deprived kittens. Blobs were found in superficial layers of V1 of normally reared kittens as early as 2 weeks of age, although at this age the overall CO staining in V1 was lighter than in V2. By 6 weeks of age the blobs were adult-like. A patchy pattern of CO staining was also found in V2 of young kittens but not in adults. Visual experience was not necessary for expression of the blobs and monocularly deprived kittens had well developed blobs, indicating that strong Y cell drive is not necessary for the development of blobs in cat V1. CO blobs appear in kitten V1 very early in post-natal development and their expression is independent of visual experience, suggesting that they may be an intrinsic feature of V1 organization.

Age Factors↗
PubMed2001

Structure and specificity of GATA proteins in Th2 development.

Development of Th2 subset of CD4+ T cells involves the interleukin-4 (IL-4)- and Stat6-dependent increase in GATA-3 expression during primary activation. Recently we reported that the phenotypic stability and factor independence of Th2 cells involves acquisition of an intracellular pathway that maintains GATA-3 expression. Evidence from retroviral expression studies implied that this pathway involved an autoactivation of GATA-3 expression, since Stat6-deficient T cells induced endogenous GATA-3 when infected with GATA-3-expressing retroviruses. That study left unresolved the issue of whether GATA-3 autoactivation was direct or indirect. Several other Th2-specific transcription factors have been described, including c-Maf and JunB. We therefore examined the ability of these other transcription factors to induce GATA-3 expression and promote Th2 development. Neither c-Maf nor JunB induced Th2 development in Stat6-deficient CD4+ T cells, in contrast to GATA-3. Consistent with this indication of a possible direct autoactivation pathway, we also observed that heterologous GATA family proteins GATA-1, GATA-2, and GATA-4 were also capable of inducing GATA-3 expression in developing Stat6-deficient T cells and promote Th2 development. Mutational analysis revealed evidence for two distinct mechanisms of GATA-3 action. IL-4 induction by GATA-3 required each of the functional domains to be present, whereas repression of gamma interferon could occur even when mutants of GATA-3 lacking the second transactivation domain, TA2, were expressed. The GATA-dependent induction of the GATA-3 but not the other GATA genes in T cells suggests that T-cell-specific cis elements within the GATA-3 locus likely cooperate with a general GATA recognition motif to allow GATA-3-dependent autoactivation.

Animals↗
PubMed2001

Immune system dysfunction and autoimmune disease in mice lacking Emk (Par-1) protein kinase.

Emk is a serine/threonine protein kinase implicated in regulating polarity, cell cycle progression, and microtubule dynamics. To delineate the role of Emk in development and adult tissues, mice lacking Emk were generated by targeted gene disruption. Emk(-/-) mice displayed growth retardation and immune cell dysfunction. Although B- and T-cell development were normal, CD4(+)T cells lacking Emk exhibited a marked upregulation of the memory marker CD44/pgp-1 and produced more gamma interferon and interleukin-4 on stimulation through the T-cell receptor in vitro. In addition, B-cell responses to T-cell-dependent and -independent antigen challenge were altered in vivo. As Emk(-/-) animals aged, they developed splenomegaly, lymphadenopathy, membranoproliferative glomerulonephritis, and lymphocytic infiltrates in the lungs, parotid glands and kidneys. Taken together, these results demonstrate that the Emk protein kinase is essential for maintaining immune system homeostasis and that loss of Emk may contribute to autoimmune disease in mammals.

Animals↗