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Biomedical subjects

D Johnson

Publications and source records attributed to D Johnson.

At least 19 recordsLinked to original sources
PubMed1999

Cigarette smoke impairs endothelium-dependent relaxation in rabbit superficial femoral veins.

BACKGROUND: The use of autogenous vein for arterial reconstruction provides the optimal conduit for limb salvage. Cigarette smoking is a risk factor for vascular disease and may adversely affect graft patency and limb preservation rates of extremities reconstructed with autogenous vein. This study was performed in order to determine the effects of cigarette smoke on venous endothelium-dependent relaxation which is mediated by nitric oxide. MATERIALS AND METHODS: New Zealand white rabbits were exposed to cigarette smoke in a 240-ft3 air-flow chamber for 3 h per day, 5 days per week, for 8 weeks. A control group was treated similarly without infusion of smoke into the chamber. Elevated serum cotinine and carboxyhemoglobin levels comparable to those of chronic smokers were observed in the experimental group. After 8 weeks, the superficial femoral veins were explanted, cut into 3-mm segments, and studied in organ chambers. RESULTS: No difference in contractile response to KCl (80 mM) (control, 0.10 +/- 0.06; smoke, 0.17 +/- 0.04) or norepinephrine (EC50) (control, 0.78 +/- 0.18; smoke, 0.87 +/- 0.11) was seen. A significant decrease in relaxation was noted with all doses of acetylcholine (M) (control, 10(-8) - 50.35 +/- 8.37, 3 x 10(-8) - 71.20 +/- 9.05, 10(-7) - 88.32 +/- 13.72, 3 x 10(-7) - 92. 86 +/- 13.69; and smoke, 10(-8) - 8.25 +/- 1.83*, 3 x 10(-8) - 19.11 +/- 5.11*, 10(-7) - 31.84 +/- 7.90*, 3 x 10(-7) - 39.74 +/- 8.65*; *P < 0.05). Both control and smoke veins relaxed completely with sodium nitroprusside. CONCLUSIONS: Superficial femoral veins, when exposed to cigarette smoke, demonstrated a significant decrease in endothelium-dependent relaxation in response to acetylcholine without smooth muscle injury. This loss of vasomotor tone could be detrimental over time in veins which have been placed as arterial conduits.

Acetylcholine↗
PubMed1999

De novo alu-element insertions in FGFR2 identify a distinct pathological basis for Apert syndrome.

Apert syndrome, one of five craniosynostosis syndromes caused by allelic mutations of fibroblast growth-factor receptor 2 (FGFR2), is characterized by symmetrical bony syndactyly of the hands and feet. We have analyzed 260 unrelated patients, all but 2 of whom have missense mutations in exon 7, which affect a dipeptide in the linker region between the second and third immunoglobulin-like domains. Hence, the molecular mechanism of Apert syndrome is exquisitely specific. FGFR2 mutations in the remaining two patients are distinct in position and nature. Surprisingly, each patient harbors an Alu-element insertion of approximately 360 bp, in one case just upstream of exon 9 and in the other case within exon 9 itself. The insertions are likely to be pathological, because they have arisen de novo; in both cases this occurred on the paternal chromosome. FGFR2 is present in alternatively spliced isoforms characterized by either the IIIb (exon 8) or IIIc (exon 9) domains (keratinocyte growth-factor receptor [KGFR] and bacterially expressed kinase, respectively), which are differentially expressed in mouse limbs on embryonic day 13. Splicing of exon 9 was examined in RNA extracted from fibroblasts and keratinocytes from one patient with an Alu insertion and two patients with Pfeiffer syndrome who had nucleotide substitutions of the exon 9 acceptor splice site. Ectopic expression of KGFR in the fibroblast lines correlated with the severity of limb abnormalities. This provides the first genetic evidence that signaling through KGFR causes syndactyly in Apert syndrome.

Acrocephalosyndactylia↗
PubMed1998

Safety and immunogenicity of HIV recombinant envelope vaccines in HIV-infected infants and children. National Institutes of Health-sponsored Pediatric AIDS Clinical Trials Group (ACTG-218).

Study objectives were to evaluate the safety and immunogenicity of three HIV recombinant glycoproteins in HIV-infected infants and children between 1 month and 18 years of age with asymptomatic (P-1) infection. Using Chiron rgp 120 (SF-2) 15 or 50 microg; MicroGeneSys rgp 160 (IIIB) 40 or 320 microg; Genentech rgp120 (MN) 75 or 300 microg; or adjuvant control (Alum or MF-59), children were randomized to a double-blind, placebo-controlled, dose-escalating study of vaccine administered intramuscularly at entry and 1, 2, 3, 4, and 6 months later. No adverse events were attributed to study vaccines. Between 30% and 56% of volunteers exhibited a lymphoproliferative response as defined in terms of stimulation index (SI) to vaccine antigens; 65% of vaccinees but none of placebo recipients exhibited moderate or strong responses after enzyme immunoassay to HIV specific antigens. CD4 cell counts and quantitative HIV culture did not differ significantly among vaccine and control groups, nor were differences found among groups in HIV disease progression. The rgp160 and gp120 subunit vaccines were safe and immunogenic in this population.

AIDS Vaccines↗
PubMed1998

Characterization of the neuropeptide Y5 receptor in the human hypothalamus: a lack of correlation between Y5 mRNA levels and binding sites.

Neuropeptide Y (NPY) is a 36-amino-acid peptide that appears to play a central role in the control of feeding behavior. Recently, a cDNA encoding a novel NPY receptor subtype (Y5) was cloned from the rat and human hypothalamus, and shown to have a pharmacology consistent with NPY-induced feeding. We have subsequently cloned this cDNA from human hypothalamus and stably expressed it in CHO cells. Consistent with earlier reports, hY5 has a high affinity for NPY, [Leu31, Pro34]NPY, and NPY(3-36), but low affinity for larger C-terminal deletions of NPY and BIBP3226. High levels of hY5 mRNA were found in the human testis, brain, spleen and pancreas, with lower levels in several other tissues. In the human brain, hY5 mRNA levels were typically higher than hY2, but lower in comparison to hY1 receptor mRNA. To quantify the relative amounts of hY1, hY2 and hY5 mRNA in the human hypothalamus, we employed competitive RT-PCR. Interestingly, the relative amount of hY5 mRNA was substantially higher than either hY1 or hY2. However, pharmacological characterization of NPY binding sites in human hypothalamus membranes revealed predominantly the hY2 subtype. These data establish that while hY5 mRNA levels are very high in the human hypothalamus, conventional radioligand binding techniques do not detect hY5-like binding site. Whether hY5-like binding sites exist in the other human tissues that express hY5 mRNA (and what function hY5 has in those tissues) awaits future investigation.

Animals↗
PubMed1998

The major myosin phosphatase in skeletal muscle is a complex between the beta-isoform of protein phosphatase 1 and the MYPT2 gene product.

Myosin is dephosphorylated by distinct forms of protein phosphatase 1 (PP1) in smooth muscle and skeletal muscle that are composed of PP1 complexed to different regulatory subunits. The smooth muscle myosin phosphatase (smPP1M) has been characterised previously and is composed of PP1beta complexed to M110 and M21 subunits that enhance the dephosphorylation of smooth muscle myosin, but not skeletal muscle myosin. In contrast, the regulatory subunit(s) of skeletal muscle myosin phosphatase (skPP1M) greatly enhance(s) the dephosphorylation of skeletal muscle myosin. Here we identify a regulatory subunit of skPP1M as the product of the MYPT2 gene, a protein whose sequence is 61% identical to the M110 subunit of smPP1M. Surprisingly, the M21 subunit of smPP1M appears to be produced from the same gene that encodes MYPT2.

Amino Acid Sequence↗
PubMed1998

Interstitial duplications of chromosome region 15q11q13: clinical and molecular characterization.

Duplications of chromosome region 15q11q13 often occur as a supernumerary chromosome 15. Less frequently they occur as interstitial duplications [dup(15)]. We describe the clinical and molecular characteristics of three patients with de novo dup(15). The patients, two males and one female (ages 3-21 years), had nonspecific findings that included autistic behavior, hypotonia, and variable degrees of mental retardation. The extent, orientation, and parental origin of the duplications were assessed by fluorescent in situ hybridization, microsatellite analyses, and methylation status at D15S63. Two patients had large direct duplications of 15q11q13 [dir dup(15)(q11q13)] that extended through the entire Angelman syndrome/Prader-Willi syndrome (AS/PWS) chromosomal region. Their proximal and distal breaks, at D15S541 or D15S9 and between D15S12 and D15S24, respectively, were comparable to those found in the common AS/PWS deletions. This suggests that duplications and deletions may be the reciprocal product of an unequal recombination event. These two duplications were maternally derived, but the origin of the chromatids involved in the unequal crossing over in meiosis differs. In one patient, the duplication originated from two different maternal chromosomes, while in the other patient it arose from the same maternal chromosome. The third patient had a much smaller duplication that involved only D15S11 and parental origin could not be determined. There was no obvious correlation between phenotype and extent of the duplication in these patients.

Adult↗
PubMed1998

Evidence for complex nuclear inheritance in a pedigree with nonsyndromic deafness due to a homoplasmic mitochondrial mutation.

The relationship between mitochondrial genotype and clinical phenotype is complicated in most instances by the heteroplasmic nature of pathogenic mitochondrial mutations. We have previously shown that maternally inherited hearing loss in a large Arab-Israeli kindred is due to the homoplasmic A1555G mutation in the mitochondrial 12S ribosomal RNA gene [Prezant et al., 1993: Nat Genet 4:289-294]. Family members with this mutation have phenotypes ranging from profound hearing loss to completely normal hearing, and we have shown that there is genetic and biochemical evidence for nuclear gene involvement in this family [Bu et al., 1993: Genet Epidemiol 9:27-44; Guan et al., 1996: Hum Mol Genet 5:963-971]. To identify such a nuclear locus, two candidate genes were excluded through linkage analysis and sequencing, and a genome-wide linkage search in family members who all have the identical homoplasmic mitochondrial mutation, but differ in their hearing status, was performed. In two stages a total of 560 polymorphic genetic markers was genotyped, and the data were analyzed under model-dependent and model-free assumptions. No chromosomal region was identified as a major contributor to the phenotypic expression of the mitochondrial mutation. Thus, in this simplified paradigm of a homoplasmic mitochondrial mutation in a single kindred who all live in the similar environment of a small village, the penetrance of the mitochondrial mutation appears to depend on the interaction of multiple nuclear genes.

Chromosome Mapping↗
PubMed1998

Expression of the v-Src oncoprotein in fibroblasts disrupts normal regulation of the CDK inhibitor p27 and inhibits quiescence.

To determine how an oncogenic tyrosine kinase disturbs cell cycle control we examined expression of cell cycle proteins and growth of fibroblasts reversibly transformed by a temperature sensitive mutant of v-Src (ts LA 29). ts LA 29 Rat-1 cells and normal Rat-1 cells had similar growth rates but the transformed cells traversed the G1 phase of the cell cycle more rapidly and failed to exit cycle efficiently in response to serum starvation and cell confluence. Cyclin D1 and cyclin E levels were not elevated in growing ts LA 29 Rat-1 cells and the abbreviated G1 was further accelerated by overexpression of cyclin E. A fall in cyclin E and cyclin A dependent kinase activities in Rat-1 cells in response to inhibitory growth conditions was abrogated in ts LA 29 Rat-1 cells and correlated with lack of p27 accumulation or cyclin A down regulation, the latter due to sustained cyclin A promoter activity. The expression of p27 mRNA was lower in ts LA 29 Rat-1 cells than Rat-1 cells and was elevated following v-Src inactivation concurrent with an increase in p27 promoter activity and temporary cell cycle exit. The suppression of mRNA or transcription is a novel way an oncoprotein can induce down regulation of p27 and contributes to the G1 shortening and perturbed cell cycle regulation of the v-Src transformed cells.

Animals↗
PubMed1998

Characterization and cloning of the gene encoding the vacuolar membrane protein EXP-2 from Plasmodium falciparum.

As a contribution to the characterization of the parasitophorous vacuolar membrane from Plasmodium falciparum we have begun the identification of vacuolar membrane proteins. Exported protein-2 (EXP-2) is a vacuolar membrane protein exposed into the vacuolar space. To further characterize EXP-2, it was purified, and the 45 N-terminal amino acids were determined by micro-sequencing. Based on this information, partial cDNA and genomic fragments were amplified by PCR and used as probes for the isolation of complete cDNA and genomic DNA clones. The single copy gene is located on chromosome 14, and is transcribed during the ring stage of parasite development. The open reading frame encodes an N-terminal signal sequence which is cleaved from the mature protein. The amino acid composition of EXP-2 is characterized by charged amino acids, with a high abundance of aspartate residues in the C-terminal portion of the protein. In contrast to EXP-1, an integral protein of the vacuolar membrane, EXP-2 lacks a typical hydrophobic transmembrane domain. We suggest that EXP-2 may associate with the vacuolar membrane via an amphipathic helix located in the N-terminal half of the protein.

Amino Acid Sequence↗
PubMed1998

Gene structure and properties of TIGR, an olfactomedin-related glycoprotein cloned from glucocorticoid-induced trabecular meshwork cells.

Expression of the trabecular meshwork inducible glucocorticoid response (TIGR) gene progressively increases from barely detectable levels to greater than 2% of total cellular mRNA over 10 days exposure of trabecular meshwork (TM) cells to dexamethasone. Cycloheximide blocked most of the TIGR mRNA induction, suggesting a requirement for ongoing protein synthesis. The genomic structure of TIGR (approximately 20 kilobases) consists of 3 exons, and a 5-kilobase promoter region that contains 13 predicted hormone response elements, including several glucocorticoid regulatory elements, and other potentially important regulatory motifs. TIGR cDNA encodes an olfactomedin-related glycoprotein of 504 amino acids with motifs for N- and O-linked glycosylation, glycosaminoglycan initiation, hyaluronic acid binding, and leucine zippers. Recombinant TIGR (rTIGR) showed oligomerization and specific binding to TM cells. Anti-rTIGR antibody detected multiple translational/post-translational forms of TIGR produced by the cells (including secreted 66 kDa/55 kDa glycoproteins/proteins in the media and 55 kDa cellular proteins), whereas Northern blot showed a single mRNA species. The findings suggest potential mechanisms by which TIGR could obstruct the aqueous humor fluid flow and participate in the pathogenesis of glaucoma.

Amino Acid Sequence↗
PubMed1998

Aminopyrimidines with high affinity for both serotonin and dopamine receptors.

A series of [4-[2(4-arylpiperazin-1-yl)alkyl]cyclohexyl]pyrimidin-2-ylamine s was prepared and found to have receptor binding affinity for D2 and D3 dopamine (DA) receptors and serotonin 5-HT1A receptors. The structural contributions to D2/D3 and 5-HT1A receptor binding of the aminopyrimidine, cycloalkyl, and phenylpiperazine portions of the molecule were examined. From these studies compounds 14, 39, 42, 43, having potent affinity for both DA D2 and 5-HT1A receptors, were evaluated for intrinsic activity at these receptors, in vitro and in vivo. Compound 14 (PD 158771) had a profile indicative of partial agonist activity at both D2 and 5-HT1A receptors causing partially decreased synthesis of the neurotransmitters DA and 5-HT and their metabolites. This compound has a profile in behavioral tests that is predictive of antipsychotic activity, suggesting that mixed partial agonists such as 14 may have utility as antipsychotic agents with increased efficacy and decreased side effects.

Animals↗
PubMed1998

A new pitheciin primate from the middle Miocene of Argentina.

We report here a new fossil primate from the middle Miocene of Argentina. The material consists of isolated teeth, mandibular fragments, and a talus. The fossils were collected in the Collón Cura formation at Cañadón del Tordillo in Neuquén Province. An age of 15.71 +/- 0.07 Ma has been reported for the Pilcaniyeu Ignimbrite, which lies just below the paleosols in which the fossils were found. This material is thus the youngest occurrence of fossil primates in Argentina (hitherto documented in the Santacrucian and older land mammal ages) but still is older than the middle Miocene platyrrhine primates from La Venta, Colombia, in particular the pitheciins Nuciruptor and Cebupithecia. The material is recognized as a new genus and species of Pitheciinae, Propithecia neuquenensis. The mesiodistally compressed, high-crowned incisors are specialized and similar to species in the tribe Pithecini and to the nonpitheciin Soriacebus (early Miocene, Patagonia). We rule out a phylogenetic relationship to the latter because of differences in molar morphology. Propithecia does, however, fit well into the pattern of pitheciin evolution, being more derived than the middle Miocene pitheciin Nuciruptor but not as much as another middle Miocene taxon, Cebupithecia. As such, this makes Propithecia the oldest taxon that can be confidently placed within this modern New World monkey subfamily. By analogy with the molar structures and diets of extant platyrrhines, Propithecia has a molar structure consistent with a variety of low-fiber diets ranging from fruit and gum to seeds. Its incisors suggest seed-eating in much the same way as extant pitheciins, like Pithecia. The talus resembles that of Callicebus, suggesting arboreal quadrupedal locomotion.

Animals↗
PubMed1998

The nonspecific inflammatory response to injury.

PURPOSE: The role of the nonspecific inflammatory response in causing injury related to surgery has become better understood over the last decade. There are complex interactions between neutrophils, cytokines and nitric oxide metabolites that may cause organ injury following surgery. The purpose of this review is to summarize some of the processes causing injury through these nonspecific pathways. METHODS: A review of the medical and anaesthetic literature related to inflammation, neutrophils and pro-inflammatory cytokines were performed using Medline. Bibliographies of relevant articles were searched and additional articles were then selected and reviewed. RESULTS: Pro-inflammatory cytokines, such as tumour necrosis factor, are released in response to a variety of noxious stimuli (e.g. burns, sepsis, or CABG surgery). These cytokines cause activation of neutrophils with increased upregulation of adhesion complexes on neutrophils and vascular endothelium. Nitric oxide synthase activity is also increased with a resultant increased production of nitric oxide. The increased nitric oxide concentration in the presence of superoxide free radicals secreted by activated neutrophils forms peroxynitrite, a more reactive and toxic molecule. Once this process is initiated, diffuse organ injury can result. Although some information related to specific anaesthetics is available, firm recommendations related to clinical practice cannot be made. CONCLUSIONS: There is a complex interplay of inflammatory mediators that can cause injury. Although specific clinical applications for manipulating these pathways are not yet generally available, this area holds promise to develop new techniques to improve outcomes following surgery.

Cell Adhesion Molecules↗
PubMed1998

The selective use of a single-stage and a two-stage technique for hypospadias correction in 157 consecutive cases with the aim of normal appearance and function.

This paper establishes the aim of normal function and appearance in hypospadias surgery. In order to meet this aim we have used a combination of a single-stage and a two-stage repair, both incorporating foreskin reconstruction. The technique of single-stage repair is illustrated using operative photographs. Our experience with 157 repairs in a 3-year period is described. Thirty-five single-stage repairs were completed with no complications; 122 two-stage repairs were carried out, in which a 5% fistula rate was achieved in primary cases. Salvage reconstruction had a higher incidence of complications.

Aged↗