Biomedical subjects

J H White

Publications and source records attributed to J H White.

At least 37 records · Page 2Linked to original sources

PubMed2000

The GABAB receptor interacts directly with the related transcription factors CREB2 and ATFx.

gamma-Aminobutyric acid type B (GABA(B)) receptors mediate the metabotropic actions of the inhibitory neurotransmitter GABA. These seven-transmembrane receptors are known to signal primarily through activation of G proteins to modulate the action of ion channels or second messengers. The functional GABA(B) receptor is made up of a heterodimer consisting of two subunits, GABA(B)-R1 and GABA(B)-R2, which interact via coiled-coil domains in their C-terminal tails. By using a yeast two-hybrid approach, we have identified direct interactions between the C-terminal tails of GABA(B)-R1 and GABA(B)-R2 with two related transcription factors, CREB2 (ATF4) and ATFx. In primary neuronal cultures as well in recombinant Chinese hamster ovary cells expressing GABA(B) receptors, CREB2 is localized within the cytoplasm as well as the nucleus. Activation of the GABA(B) receptor by the specific agonist baclofen leads to a marked translocation and accumulation of CREB2 from the cytoplasm into the nucleus. We demonstrate that receptor stimulation results in activation of transcription from a CREB2 responsive reporter gene. Such a signaling mechanism is unique among Family C G protein-coupled receptors and, in the case of the GABA(B) receptor and CREB2, may play a role in long-term changes in the nervous system.

Amino Acid Sequence↗

PubMed2000

Cellular and sub-cellular localisation of GABA(B1) and GABA(B2) receptor proteins in the rat cerebellum.

Following the recent discovery that GABA(B) receptors expressed in cell lines are only functional when both GABA(B1) and GABA(B2) are expressed, the present study reports on the development of polyclonal antisera specific for carboxyl-terminal portions of the two related GABA(B) receptor components respectively. Western blotting indicated the specificity of affinity-purified antibodies for native or recombinant expressed GABA(BR1) and GABA(BR2), with no cross-reactivity, both antisera detecting the heterodimer in rat cerebellar membranes. Immunohistochemistry revealed a distinct distribution of both receptor proteins in rat cerebellum. GABA(B1) immunoreactivity was primarily located in the granule cell layer and Purkinje cells, with discrete immuno-positive cell bodies being present in the molecular layer. GABA(B2) staining revealed intense immunoreactivity in the molecular layer, with weaker staining in the granule cell layer. Purkinje cell bodies were less intensely immuno-positive for GABA(B2). Co-localisation of both receptor proteins was observed using double immunofluorescence techniques, consistent with the notion that both proteins are required for the formation of functional GABA(B) receptors in vivo. Immunofluorescence also indicated that GABA(B) receptors did not co-localise with glial fibrillary acid protein, confirming a neuronal localisation for GABA(B) receptors. Electron microscopic analysis of the molecular layer revealed that the distribution of immunolabelling for both GABA(B1) and GABA(B2) was mainly located on the membrane of Purkinje cell dendrites and spines and in parallel fibre terminals.

Animals↗

PubMed2000

GABA(B) receptor heterodimer-component localisation in human brain.

In recombinant cell lines, functional GABA(B) receptors are only formed by the heterodimerisation between two related G-protein coupled receptor proteins GABA(B)R1 (GBR1) and GABA(B)R2 (GBR2), whilst the individual GBR1 or GBR2 do not produce fully functional receptors. To determine whether the heterodimerisation occurs in vivo, novel polyclonal antibodies targeting the C termini of GBR1 and GBR2, were raised in different species, characterised, and used to determine the relative localisation of the reported heterodimer components in human brain tissue, using immunohistochemistry. The use of different species for the raising of the antisera allowed double immunofluorescent labelling of the receptors as an indication of GBR1/GBR2 receptor co-localisation in human brain. The presence of both proteins is reported in cerebellum, hippocampus, cortex, thalamus and basal ganglia. Regions of the brainstem including pons and medulla, also express GBR1 and GBR2 protein. The double immunofluorescence demonstrated that GBR1 and GBR2 are co-localised in the human cerebellar cortex. Together these results suggest the widespread distribution of GABA(B) receptors in human brain, and that GABA(B) receptors GBR1 and GBR2 can exist in the same cell, and therefore may function as a heterodimer in the human brain.

Aged↗

PubMed2000

Methylation patterns of human parathyroid hormone (PTH)/PTH-related peptide receptor gene promoters are established several weeks prior to onset of their function.

Expression of the human parathyroid hormone (PTH)/PTH-related peptide receptor (PTHR) gene is controlled by three promoters, P1-P3. P1 functions specifically in kidney, whereas P2 is ubiquitously active. P3 is also widely active, although more so in kidney than other tissues. However, only P2 functions at midgestation. We examined the role of methylation in controlling PTHR promoter activity. Function of all promoters was inhibited by CpG methylation in vitro. Significantly, P1 is selectively hypomethylated in adult kidney in vivo, strongly suggesting that demethylation is required for renal P1 function. Moreover, this pattern is established by 11. 75 weeks of fetal age, several weeks prior to the onset P1 activity. P3 is unmethylated at midgestation, although it is inactive at this stage of development, and thus exhibits characteristics of both tissue-specific and ubiquitously active promoters. These results show that adult methylation patterns of P1 and P3 are established several weeks prior to their induction, indicating that their function requires factors expressed late in development.

Adult↗

PubMed2000

Functional coupling of mammalian receptors to the yeast mating pathway using novel yeast/mammalian G protein alpha-subunit chimeras.

The expression of mammalian G protein coupled receptors (GPCRs) in S. cerevisiae provides a powerful assay system for functional analysis, ligand identification and pharmaceutical screening. However, relatively few receptors have been coupled to the pheromone response pathway via the yeast G(alpha), Gpa1p, or chimeric yeast/mammalian G(alpha) subunits containing long C-terminal regions of mammalian G(alpha) proteins. We tested an extended range of seven such chimeras for G(alpha) sub-types of three major classes (G(alphai/o), G(alphas) and G(alphaq)), against eight human GPCRs (SST(2), SST(5), 5-HT(1A), 5-HT(1Dalpha), ML(1B), P2Y(1) and P2Y(2)). Although the G(alphai/o) chimeras increased the range of receptors that coupled efficiently, the G(alphas) and G(alphaq) chimeras were inactive when expressed using the GPA1 promoter. We describe 10 novel Gpa1p chimeras, designated 'transplants', in which the C-terminal five amino acids of Gpa1p were exchanged with mammalian residues. Coupling efficiency and ligand sensitivity improved significantly using the transplants. For the P2Y purinergic receptors, coupling could only be detected with the transplants; this is the first report of G(q) specificity coupling in yeast. Thus, the transplants offer major advantages over previously described approaches, in terms of both the range of receptors coupled and the efficiency of coupling.

GTP-Binding Proteins↗

PubMed2000

Symptom development in bulimia nervosa: a comparison of women with and without a history of anorexia nervosa.

Identifying women at risk for developing bulimia nervosa (BN) is an important area of study for psychiatric-mental health nurses. Of those who develop BN, a large proportion of women have been diagnosed earlier with anorexia nervosa (AN). It has been posited that women who are treated for AN are often only partially recovered. Although their weights become clinically acceptable, they often continue to restrict food; this typically leads to a binge eating and purging cycle and the development of BN. A history of AN is viewed as a possible risk factor for the development of BN. This study compared women seeking treatment for BN with a prior history of AN, and those without this history on selected variables. A profile emerged and is described supporting a distinct subgroup with prior AN. Findings from this study have implications for early detection of and extended follow-up care after treatment for AN.

Age of Onset↗

PubMed2000

Developing a CNS role to meet the mental health needs of the underserved.

The purpose of this article is to describe a clinical nurse specialist program that prepares psychiatric-mental health nurses for positions in today's mental health care system. Recently, there has been a national increase in psychiatric-mental health nurse practitioner programs and a decrease in psychiatric-mental health-clinical nurse specialist programs. We faced the dilemma that many psychiatric-mental health nurse faculties face in this climate of mental health care delivery changes: how to best prepare psychiatric-mental health nurses for advanced practice. Because of our long history of preparing clinical nurse specialists, we believed the role was a viable one. The clinical nurse specialist role also fit with our university's mission and our beliefs regarding the importance of providing care for underserved populations. The role of the community mental health-clinical nurse specialist is described, and the economic, demographic, and mental health system changes that influenced the development of the community mental health-clinical nurse specialist role are explored. The curriculum and the impact of the program are described and evaluated. Finally, implications regarding the need and the viability of this new role in both psychiatric-mental health nursing and in other specialties as we care for clients in the community are presented.

Community Health Nursing↗

PubMed2000

Comparison of vaginal shapes in Afro-American, caucasian and hispanic women as seen with vinyl polysiloxane casting.

Full vinyl polysiloxane casts of the vagina were obtained from 23 Afro-American, 39 Caucasian and 15 Hispanic women in lying, sitting and standing positions. A new shape, the pumpkin seed, was found in 40% of Afro-American women, but not in Caucasians or Hispanics. Analyses of cast and introital measurements revealed: (1) posterior cast length is significantly longer, anterior cast length is significantly shorter and cast width is significantly larger in Hispanics than in the other two groups and (2) the Caucasian introitus is significantly greater than that of the Afro-American subject.

Black People↗

PubMed2000

Eating disorders in elementary and middle school children: risk factors, early detection, and prevention.

Eating disorders (EDs) are significant problems that are typically diagnosed during adolescence. However, the risk factors for and early symptoms of EDs often develop in the elementary and middle school years. Dieting, body dissatisfaction, obesity, parental attitudes, and the influence of the media are some of the significant identifiable risk factors. Prevention programs need to be developed that focus on education, consultation, and consciousness-raising. Early detection involves screening, assessment, and referral for appropriate treatment. School nurses are skilled, educated, and positioned to develop programs for the prevention and early detection of EDs.

Adolescent↗

PubMed2000

Dissection of differentially regulated (G+C)-rich promoters of the human parathyroid hormone (PTH)/PTH-related peptide receptor gene.

The PTH/PTH-related peptide (PTHrP) receptor (PTHR) is required for normal skeletal development, and a wide array of physiological responses mediated by PTH and PTHrP. We have previously identified three promoters, P1-P3, which control human PTHR gene transcription. P2 and P3 are (G+C)-rich, function in a number of tissues, lie within the same CpG island, and display many hallmarks of housekeeping promoters. However, they are differentially regulated during development as P2, but not P3, functions in fetal tissues. Here, we have used both stably and transiently transfected human osteoblast-like cells to delineate regions of P2 and P3 required for promoter activity. Deletion analyses performed in stably transfected cells indicated that sequences extending from -91 to -12 relative to the transcription start site were required for function of the P2 promoter. No negative regulatory elements were detected in P2. In contrast, deletion of an A-rich region of P3 extending from -147 to -115 was required for optimal basal activity, suggesting that this sequence acts as a repressor of P3. Strikingly, however, whereas the A-rich region also functioned as a negative element when inserted upstream of the (G+C)-rich P2 promoter, it enhanced expression from the thymidine kinase promoter, suggesting that its function depends on other transcription factors bound to promoter sequences. Fine deletion of P3 sequences proximal to -115 implicated Spl motifs and downstream initiation sites in P3 function. These studies indicate that function of P2 and P3 is controlled by ubiquitously expressed transcription factors and raise the possibility that P3 activity is repressed during fetal development.

Base Sequence↗

PubMed2000

Recent studies on the biological action of parathyroid hormone (PTH)-related peptide (PTHrP) and PTH/PTHrP receptor in cartilage and bone.

Mice with a targeted deletion of parathyroid hormone (PTH)-related peptide (PTHrP) develop a form of dyschondroplasia resulting from diminished proliferation and premature maturation of chondrocytes. Abnormal, heterogeneous populations of chondrocytes at different stages of differentiation were seen in the hypertrophic zone of the mutant growth plate. Although the homozygous null animals die within several hours of birth, mice heterozygous for PTHrP gene deletion reach adulthood, at which time they show evidence of osteopenia. Therefore, PTHrP appears to modulate cell proliferation and differentiation in both the pre and post natal period. PTH/PTHrP receptor expression in the mouse is controlled by two promoters. We recently found that, while the downstream promoter controls PTH/PTHrP receptor gene expression in bone and cartilage, it is differentially regulated in the two tissues. 1alpha,25-dihydroxyvitamin D3 downregulated the activity of the downstream promoter in osteoblasts, but not in chondrocytes, both in vivo and in vitro. Most of the biological activity of PTHrP is thought to be mediated by binding of its amino terminus to the PTH/PTHrP receptor. However, recent evidence suggests that amino acids 87-107, outside of the amino terminal binding domain, act as a nucleolar targeting signal. Chondrocytic cell line, CFK2, transfected with wild-type PTHrP cDNA showed PTHrP in the nucleoli as well as in the secretory pathway. Therefore, PTHrP appears to act as a bifunctional modulator of both chondrocyte proliferation and differentiation, through signal transduction linked to the PTH/PTHrP receptor and by its direct action in the nucleolus.

Animals↗

PubMed2000

Developmental and tissue-specific regulation of parathyroid hormone (PTH)/PTH-related peptide receptor gene expression.

Parathyoid hormone (PTH) and PTH-related peptide (PTHrP) signal through a common PTH/PTHrP receptor (PTHR1). PTH is secreted by the parathyroids and functions in an endocrine manner to regulate extracellular fluid calcium concentrations, largely by interacting with receptors expressed in kidney and bone. PTHrP, on the other hand, is widely expressed and acts by autocrine or paracrine signaling to modulate a range of physiological and developmental responses. PTH1R1 signaling is required both in the adult and during development, and ablation of PTHR1 gene expression results in an embryonic lethal phenotype in both mice and humans. Consequently, the PTHR1 is widely expressed, and transcription of its gene is tissue specific and developmentally regulated. This review focuses on our current knowledge of the mechanisms controlling expression of the PTHR1 gene in humans and in rodents, both during development and in the adult. We also compare the conserved and distinct aspects of PTHR1 gene regulation in the two systems and their bearing on signaling by PTH and PTHrP.

Animals↗

PubMed1999

Formation and biochemical characterization of tube/pelle death domain complexes: critical regulators of postreceptor signaling by the Drosophila toll receptor.

In Drosophila, the Toll receptor signaling pathway is required for embryonic dorso-ventral patterning and at later developmental stages for innate immune responses. It is thought that dimerization of the receptor by binding of the ligand spätzle causes the formation of a postreceptor activation complex at the cytoplasmic surface of the membrane. Two components of this complex are the adaptor tube and protein kinase pelle. These proteins both have "death domains", protein interaction motifs found in a number of signaling pathways, particularly those involved in apoptotic cell death. It is thought that pelle is bound by tube during formation of the activation complexes, and that this interaction is mediated by the death domains. In this paper, we show using the yeast two-hybrid system that the wild-type tube and pelle death domains bind together. Mutant tube proteins which do not support signaling in the embryo are also unable to bind pelle in the 2-hybrid assay. We have purified proteins corresponding to the death domains of tube and pelle and show that these form corresponding heterodimeric complexes in vitro. Partial proteolysis reveals a smaller core consisting of the minimal death domain sequences. We have studied the tube/pelle interaction with the techniques of surface plasmon resonance, analytical ultracentrifugation and isothermal titration calorimetry. These measurements produce a value of K(d) for the complex of about 0.5 microM.

Amino Acid Sequence↗

PubMed1999

Effect of salt-dependent stiffness on the conformation of a stressed DNA loop containing initially coplanar bends.

Closed DNA loops containing one or more bent regions are important structures that occur in the regulation of gene expression. We analyze the response of structures of this type to a change in applied rotation (change in linking deficiency, delta Lk). Our results apply to a closed loop formed from an elastic rod that is intrinsically bent in Nb discrete, 20 degrees steps up to a maximum of 240 degrees, the bent regions being initially coplanar with the plane of the relaxed DNA loop. We determine the effect of changing the intrinsic elastic resistance of the DNA loop to bending and torsional deformations. This relative resistance is expressed by Poisson's ratio v, which depends upon the ratio of bending stiffness to torsional rigidity. Poisson's ratio is primarily a function of salt type and concentration. We find that the tertiary structure of DNA loops changes with delta Lk, but that the geometric response can be either of two quite different types, depending upon the precise (Nb, v) pair. For combinations of Nb and v that are above a critical curve (the Fickel curve), the response to increasing delta Lk is nonmonotonic (NMT region): the distance between the loop closure point and its diametric opposite first decreases, then increases, as delta Lk increases. For combinations of Nb and v that are below the Fickel curve (NMT region), the corresponding diameter never increases, but always decreases with increasing delta Lk. In addition to these results, we define and implement a new measure of tertiary structure in closed DNA: the absolute writhe, AWr.

Animals↗

PubMed1999

The development and clinical testing of an outpatient program for women with bulimia nervosa.

This article describes the development of a cost-effective treatment program for women with Bulimia Nervosa (BN). A comprehensive review of the literature was undertaken to determine the most effective programs. The Time Limited Cognitive Behavioral Program (TLCBP) was developed and clinically tested on two groups of women to determine its cost-effectiveness. The program consisted of a 12-week intervention phase followed by a 14-week support/reinforcement phase. The clinical findings were that 9 of the 10 clients were abstinent following the 12-week intervention. Other symptoms improved following the 14-week phase of support/reinforcement. Utilizing an Advanced Practice Psychiatric-Mental Health Registered Nurse (APRN) significantly reduced the cost of the treatment, as did the use of a group model approach. APRN's have bio-behavioral knowledge as well as research skills. They are in a unique position to conduct outcome research with women with BN, as well as with clients who have other disorders with both biological and behavioral symptoms.

Ambulatory Care↗

PubMed1999

Vitamin D3 differentially regulates parathyroid hormone/parathyroid hormone-related peptide receptor expression in bone and cartilage.

Transcription of the mouse parathyroid hormone (PTH)/PTH-related peptide (PTHrP) receptor (PTHR) gene is controlled by promoters P1 and P2. We performed transcript-specific in situ hybridization and found that P2 is the predominant promoter controlling PTHR gene expression in bone and cartilage. Treatment with 1alpha, 25-dihydroxyvitamin D3 (D3) in vivo specifically downregulated P2-specific transcripts in osteoblasts, but not in chondrocytes, under conditions where it enhanced bone resorption. Treatment of the osteoblastic cell line MC3T3-E1 with D3 in vitro reduced expression of both P2-specific transcripts and PTHR protein. This effect was not blocked by cycloheximide, indicating that D3 inhibits PTHR expression by downregulating transcription of the P2 promoter. A similar inhibitory effect of D3 was not observed in the chondrocytic cell line CFK2. Gene-transfer experiments showed that P2, but not P1, is active in both MC3T3-E1 and CFK2 cells, and that D3 specifically inhibited P2 promoter activity in MC3T3-E1, but not in CFK2 cells. Inhibition of P2 activity by D3 required promoter sequences lying more that 1.6 kb upstream of the P2 transcription start site. Thus, the P2 promoter controls PTHR gene expression in both osteoblasts and chondrocytes. D3 downregulates PTHR gene transcription in a cell-specific manner by inhibiting P2 promoter activity in osteoblasts, but not in chondrocytes.

Animals↗

PubMed1999

Mitogen-activated protein kinase inhibits 1,25-dihydroxyvitamin D3-dependent signal transduction by phosphorylating human retinoid X receptor alpha.

Human retinoid X receptor alpha (hRXR alpha) is a member of the nuclear receptor family of transcriptional regulators. It regulates transcription through its association with several heterodimeric partners, including the vitamin D3 receptor (VDR). Signaling through the VDR is essential for normal calcium homeostasis and has been shown to inhibit the proliferation of cancer cells derived from a number of tissues. Here we show that phosphorylation of hRXR alpha in ras-transformed human keratinocytes through the activated Ras-Raf-mitogen-activated protein kinase (Ras-Raf-MAP kinase) pathway results in attenuated transactivation by the VDR and resistance to the growth inhibitory action of 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] and RXR-specific agonist LG1069 (4-[1-(5,6,7, 8-tetrahydro-3,5,5,8,8-pentamethyl-2-naphthalenyl) ethenyl]-benzoic acid). Phosphorylation of hRXR alpha occurs at serine 260, a consensus MAP kinase site. Inhibition of MAP kinase activity or point mutagenesis of serine 260 of hRXR alpha reverses the observed resistance to 1,25(OH)2D3 and LG1069. Thus, hRXR alpha is a downstream target of MAP kinase, and its phosphorylation may play an important role in malignant transformation.

Animals↗

PubMed1999

The independent and combined effects of RAR-, RXR-, and VDR-selective ligands on the growth of squamous cell carcinoma in vitro.

OBJECTIVE: The purpose of our study was to analyze the independent and combined effects of RAR-, RXR-, and VDR-selective ligands on the growth of squamous cell carcinoma to develop new, more effective, and less toxic chemopreventive therapy. METHOD: The effects of 13-cis retinoic acid (13-cis RA), LG1069 (a highly selective RXR ligand), and vitamin D3 (D3) were analyzed in vitro in the SCC-25 human squamous cell carcinoma cell line. SCC-25 cells were grown in culture medium containing vehicle or hormone at the indicated concentrations. Growth of surviving cells was then assessed using a hemocytometer. The presence of functional vitamin D3 receptor (VDR) was confirmed using gene-transfer experiments with a promoter-lac Z reporter plasmid. RESULTS: D3 and 13-cis RA have equipotent antiproliferative effects on SCC-25 cells. Furthermore, equimolar LG1069 completely blocks the growth-suppressive effects of D3 but has no effect on the action of 13-cis RA. CONCLUSION: D3 and its analogs, administered alone or in combination with 13-cis RA, may provide more effective and less toxic chemopreventive therapy for the prevention of second primary carcinomas of the head and neck.

Antineoplastic Agents↗