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Biomedical subjects

L Conroy

Publications and source records attributed to L Conroy.

At least 19 recordsLinked to original sources
PubMed2001

Fatal falls in the US construction industry, 1990 to 1999.

The Occupational Safety and Health Administration's (OSHA's) Integrated Management Information System (IMIS) database allows for the detailed analysis of risk factors surrounding fatal occupational events. This study used IMIS data to (1) perform a risk factor analysis of fatal construction falls, and (2) assess the impact of the February 1995 29 CFR Part 1926 Subpart M OSHA fall protection regulations for construction by calculating trends in fatal fall rates. In addition, IMIS data on fatal construction falls were compared with data from other occupational fatality surveillance systems. For falls in construction, the study identified several demographic factors that may indicate increased risk. A statistically significant downward trend in fatal falls was evident in all construction and within several construction categories during the decade. Although the study failed to show a statistically significant intervention effect from the new OSHA regulations, it may have lacked the power to do so.

Accidental Falls↗
PubMed1999

Vocational outcome following spinal cord injury.

STUDY DESIGN: Non-experimental (ex post facto) survey research design involving the use of a fixed alternative format questionnaire. OBJECTIVES: To investigate variables influencing vocational outcome, to identify barriers to gaining and sustaining employment and to identify the effects of variables on the type of work engaged in following spinal cord injury. The two sets of independent variables considered were, individual and injury-related factors (age at onset of injury, time since injury, extent/level of injury, highest educational qualification achieved pre-injury, and pre-injury occupation) and circumstantial factors (means of transport, access difficulties, perceived workplace discrimination, financial disincentives to work and perceived level of skill). SETTING: The Princess Alexandra Hospital Spinal Injuries Unit, Queensland, Australia. METHODS: Data on the variables and the vocational outcomes of having ever worked or studied post-injury, current employment status and post-injury occupation were obtained from survey responses. Demographical and medical data were gathered from medical records. RESULTS: Forward stepwise logistic regression revealed that having ever worked or studied post-injury was associated with all individual and injury-related factors except pre-injury occupation, and two circumstantial factors, namely means of transport and access difficulties. Current employment was associated with all circumstantial factors as well as age at injury and pre-injury occupation. Standard multiple regression analyses revealed that post-injury occupation was correlated with all individual and injury-related factors as well as means of transport and perceived workplace discrimination. CONCLUSIONS: Tailored rehabilitation programs for individuals with characteristics associated with less successful vocational outcomes may facilitate their employment status after injury.

Adolescent↗
PubMed1998

CPAN, a human nuclease regulated by the caspase-sensitive inhibitor DFF45.

Induction of apoptosis by death receptors such as Fas or tumour necrosis factor (TNF) R1 leads to distinct changes in cell morphology, activation of the caspase protease cascade, and the degradation of nuclear chromatin by activated nucleases. Here, we describe the purification and cDNA cloning of a novel 40 kDa endonuclease from Jurkat cells that is activated by caspases. This protein, designated caspase-activated nuclease (CPAN), is sufficient to degrade naked DNA and to induce apoptotic morphology and DNA fragmentation in naive nuclei. CPAN is highly homologous to a recently described mouse nuclease, CAD [1], and may represent the human homologue. Our data on the human cDNA as well as additional data on the mouse homologue suggest that a 30 amino-acid portion of the recently published mouse sequence [1] is incorrect. We show that the activity of human CPAN is regulated by DFF45 [2], an inhibitor necessary for CPAN expression and stabilization in an inactive state in living cells. Proteolytic cleavage of DFF45 by caspases in vitro leads to dissociation of DFF45 fragments from CPAN and activation of CPAN as an endonuclease. CPAN is a tightly regulated endonuclease with unique characteristics that might represent a distinctive family of endonucleases.

Amino Acid Sequence↗
PubMed1997

Lead exposure in ironworkers.

In adults, lead toxicity is most commonly caused by occupation in a lead industry. Whereas lead toxicity has been described in workers who are involved in bridge rehabilitation, as of this date there has been no systematic evaluation published regarding the conditions responsible for lead toxicity in ironworkers. This is a report of a study designed to identify risk factors for elevated blood-lead levels in ironworkers. One hundred fifty members of a 2,400-member local ironworkers union volunteered to have their blood drawn for lead and zinc protoporphyrin analysis and to complete a questionnaire regarding demographics, health, and occupation. The relationships between these variables and blood-lead level were analyzed using student's t-test, chi-square, and logistic regression. Current work on a lead job, rivet busting as the predominant job task, and cigarette smoking were all found to be significantly associated with elevated blood-lead level. Whereas cigarette smoking and current work with lead have been previously identified as risk factors for toxicity, interventions to prevent lead toxicity in ironworkers should also focus on work practices during rivet busting.

Adult↗
PubMed1997

Mycobacterium tuberculosis DNA Not Detected by Polymerase Chain Reaction In Air Samples From Hospital Rooms of Tuberculosis Patients.

Background: As the incidence of tuberculosis (TB) increased in the United States, the risk of occupational and nosocomial TB also increased. Airborne Mycobacterium tuberculosis (MTB) is difficult to measure directly. Quantifying MTB DNA by polymerase chain reaction in air samples obtained from isolation rooms of patients with TB would provide a measure of the number of airborne organisms produced by a patient and the efficacy of ventilation, and might predict when an individual patient is no longer infectious. Methods and Results: The air was sampled through cellulose ester filters from the isolation room of a patient with newly diagnosed pulmonary TB, from a patient on therapy for 14 days, and from adjacent offices, and an attempt was made to detect MTB DNA; however, MTB DNA was detected only on positive control filters. Conclusions: Mycobacterium tuberculosis was not detected by polymerase chain reaction in air samples from the rooms of two patients with pulmonary TB. This may have been due to the large number of room air changes with resultant rapid clearance of airborne droplet nuclei or to the limited air volume sampled. A sensitive molecular assay of airborne MTB could be used to monitor the efficacy of infection control measures by sampling a sufficient volume of isolation room air and could aid in determining when an individual patient was no longer infectious.

Journal Article↗
PubMed1996

Affinity, specificity, and kinetics of the interaction of the SHC phosphotyrosine binding domain with asparagine-X-X-phosphotyrosine motifs of growth factor receptors.

The phosphotyrosine binding (PTB) domain specifically binds to tyrosine-phosphorylated proteins, but differs in structure and mechanism of action from the SH2 domain family. We quantitated the affinity, specificity, and kinetics of the interaction of the SHC PTB domain with a sequence motif, asparagine-X-X-phosphotyrosine (NXX(pY)), found in several receptor tyrosine kinases and oncogenic proteins. PTB domain-mediated interaction with the NXX(pY) motif of c-ErbB2 was characterized by similar overall affinity but slower kinetics than that reported for SH2 domains. This suggested that unlike SH2 domains, PTB domains may not rapidly exchange among associated proteins. Furthermore, when directly and quantitatively compared, PTB domain binding specificity did not significantly overlap with a panel of seven SH2 domains. Thus, signaling pathways involving PTB and SH2 domain-mediated interactions can be regulated separately. Finally, our data define the minimal SHC PTB domain binding motif as NXX(pY), not NPX(pY) as suggested by other authors, and suggest a high affinity motif, hydrophobic residue-(D/E)-N-X-X-pY-(W/F), found in the Trk and ErbB receptor tyrosine kinase families. We conclude that PTB domains mediate specific protein-protein interactions independent from those mediated by SH2 domains.

Amino Acid Sequence↗
PubMed1996

Positive tuberculin skin test reactions among house staff at a public hospital in the era of resurgent tuberculosis.

BACKGROUND: The number and significance of tuberculin skin test reactions were compared with self-reported baseline values among house staff working in a public hospital. High-risk medical specialties, locations, and infection control practices were examined. METHODS: House staff interviews, tuberculin skin test applications, review of employee health service records, and environmental monitoring of high-risk areas were performed. RESULTS: Among house staff self-reported as having negative tuberculin skin test status, 46.2% (95% CI 27.0% to 65.4%) of internal medicine house staff, compared with 4.8% (95% CI 4.3% to 13.9%) of house staff from other areas (p < 0.005), had positive results on a repeat tuberculin skin testing before graduation. These differences were not entirely explained by the use of surgical masks, year of training, or previous vaccination with bacille Calmette-Guérin. Most skin test reactions (69%) occurred among house staff who had not been vaccinated with bacille Calmette-Guérin. Increased skin reactivity probably represented excess conversions from unprotected exposure. Tuberculosis transmission was facilitated by delays in diagnosis, inadequate isolation facilities, and suboptimal ventilation. House staff did not comply with recommended tuberculosis surveillance because of time constraints, fear, and misunderstandings about tuberculin skin test interpretations in light of previous bacille Calmette-Guérin vaccination. CONCLUSIONS: House staff in high-exposure settings with suboptimal environmental controls are at increased risk for tuberculosis infection. Participation in surveillance programs can be increased by enlisting the participation and advocacy of respected medical colleagues, screening house staff differentially according to exposure and job classifications, and more accurately interpreting subsequent test results from baseline two-step testing.

BCG Vaccine↗
PubMed1995

Functional wild-type and carboxyl-terminal-tagged rat substance P receptors expressed in baculovirus-infected insect Sf9 cells.

The rat substance P (SP) receptor (SPR) was expressed in insect Sf9 cells by infection with recombinant baculovirus. The receptor bound SP with high affinity (KD = 360 pM) and had a rank order of affinity of SP > neurokinin A > neurokinin B. Ligand activation of the receptor resulted in an increase in both inositol lipid hydrolysis and intracellular Ca2+ concentration ([Ca2+]i). However, high-level expression of the receptor, in the absence of ligand, was correlated with increased basal turnover of inositol lipids and an elevated rate of Ca2+ influx. These results demonstrate that the Sf9 cells provide a suitable environment for the high-level expression of a functionally active SPR. Two carboxy-terminal epitope-tagged receptors (SPR-KT3 = SPR-TPPPEPET, COOH; SPR-Glu = SPR-EEEEYMPME, COOH) were also expressed. The affinity of the KT3-tagged receptor for ligand was similar to that of the wild-type receptor (KD = 405 pM), and that of the Glu-tagged receptor was slightly lower (KD = 1,082 pM). The high-affinity SP binding site of all three receptors was sensitive to guanosine 5'-O-(3-thiotriphosphate) pretreatment. The maximal signal-transducing ability of the epitope-tagged receptors was comparable to that of the wild-type receptor ([Ca2+]i rise as a percentage of wild-type: SPR-KT3, 80-100%; SPR-Glu, 88-100%). These data show that heterologous expression in the baculovirus system results in high expression of functional wild-type and tagged receptors.

Animals↗
PubMed1993

Requirement for posttranslational processing of Rac GTP-binding proteins for activation of human neutrophil NADPH oxidase.

Rac1 and Rac2 are closely related, low molecular weight GTP-binding proteins that have both been implicated in regulation of phagocyte NADPH oxidase. This enzyme system is composed of multiple membrane-bound and cytosolic subunits and when activated catalyzes the one-electron reduction of oxygen to superoxide. Superoxide and its highly reactive derivatives are essential for killing microorganisms. Rac proteins undergo posttranslational processing, primarily the addition of an isoprenyl group to a carboxyl-terminal cysteine residue. We directly compared recombinant Rac1 and Rac2 in a human neutrophil cell-free NADPH oxidase system in which cytosol was replaced by purified recombinant cytosolic components (p47-phox and p67-phox). Processed Rac1 and Rac2 were both highly active in this system and supported comparable rates of superoxide production. Under different cell-free conditions, however, in which suboptimal amounts of cytosol were present in the assay mixture, processed Rac2 worked much better than Rac1 at all but the lowest concentrations. This suggests that a factor in the cytosol may suppress the activity of Rac1 but not of Rac2. Unprocessed Rac proteins were only weakly able to support superoxide generation in either system, but preloading of Rac1 or Rac2 with guanosine 5'-O-(3-thio-triphosphate) (GTP gamma S) restored activity. These results indicate that processing is required for nucleotide exchange but not for interaction with oxidase components.

Amino Acid Sequence↗
PubMed1992

Generation of monoclonal antibodies to human lymphocyte cell surface antigens using insect cells expressing recombinant proteins.

We have expressed human CD40 and human B7 in insect cells using the baculovirus expression system and have used these insect cells to immunize mice for the generation of monoclonal antibodies. We demonstrate here that specific monoclonal antibodies to human CD40 and human B7 were obtained using this approach. One significant advantage of this method is that immunizing mice with insect cells did not evoke an immune response to human cells and, therefore, EBV-transformed human B cells could be used to screen for specific antibody production by the hybridoma clones.

Amino Acid Sequence↗
PubMed1992

Molecular cloning and nucleic acid binding properties of the GAP-associated tyrosine phosphoprotein p62.

p62 is a tyrosine phosphoprotein that associates with p21ras GTPase-activating protein (GAP). Purification and cDNA cloning of p62 reveal extensive sequence similarity to a putative hnRNP protein, GRP33. Recombinant human p62 purified from insect Sf9 cells binds to DNA and to mRNA and, like many proteins involved in mRNA processing, recombinant p62 is modified by dimethylation on multiple arginine residues. p62 also binds tightly to p21ras GAP in vitro: this binding depends on phosphorylation of p62 on tyrosine residues and occurs through SH2 regions of GAP. These data suggest that p120-GAP and p62 play a role in some aspect of mRNA processing or utilization and that this role may be regulated by tyrosine phosphorylation, and indirectly, by p21ras.

3T3 Cells↗
PubMed1992

Somatic mutations in the neurofibromatosis 1 gene in human tumors.

The neurofibromatosis 1 (NF1) gene product, neurofibromin, contains a GTPase-activating protein (GAP)-related domain, or NF1 GRD, that is able to down-regulate p21ras by stimulating its intrinsic GTPase. Since p21ras.GTP is a major regulator of growth and differentiation, mutant neurofibromins resulting from somatic mutations in the NF1 gene might interfere with ras signaling pathways and contribute to the development of tumors. We describe an amino acid substitution in the NF1 GRD, altering Lys-1423, that has occurred in three tumor types: colon adenocarcinoma, myelodysplastic syndrome, and anaplastic astrocytoma, and in one family with neurofibromatosis 1. The GAP activity of the mutant NF1 GRD is 200- to 400-fold lower than that of wild type, whereas binding affinity is unaffected. Thus, germline mutations in NF1 that cause neurofibromatosis 1 can also occur in somatic cells and contribute to the development of sporadic tumors, including tumors not associated with neurofibromatosis 1.

Adenocarcinoma↗
PubMed1992

GAP domains responsible for ras p21-dependent inhibition of muscarinic atrial K+ channel currents.

The interaction between the low molecular weight G protein ras p21 and a guanosine triphosphatase activating protein (GAP) uncouples a heterotrimeric G protein (Gk) from muscarinic receptors. Through the use of isolated atrial cell membranes and genetically engineered GAP deletion mutants, the src homology regions (SH2-SH3) at the amino terminus of GAP have been identified as the domains responsible for this effect. Deletion of the domain required to stimulate the guanosine triphosphatase activity of ras p21 relieves the requirement for ras p21 in this system. A model is presented that suggests that ras p21 induces a conformational change in GAP, which allows the SH2-SH3 regions of GAP to function.

Animals↗
PubMed1992

Localization of the rap1GAP catalytic domain and sites of phosphorylation by mutational analysis.

rap1GAP is a GTPase-activating protein that specifically stimulates the GTP hydrolytic rate of p21rap1. We have defined the catalytic domain of rap1GAP by constructing a series of cDNAs coding for mutant proteins progressively deleted at the amino- and carboxy-terminal ends. Analysis of the purified mutant proteins shows that of 663 amino acid residues, only amino acids 75 to 416 are necessary for full GAP activity. Further truncation at the amino terminus resulted in complete loss of catalytic activity, whereas removal of additional carboxy-terminal residues dramatically accelerated the degradation of the protein in vivo. The catalytic domain we have defined excludes the region of rap1GAP which undergoes phosphorylation on serine residues. We have further defined this phosphoacceptor region of rap1GAP by introducing point mutations at specific serine residues and comparing the phosphopeptide maps of the mutant proteins. Two of the sites of phosphorylation by cyclic AMP (cAMP)-dependent kinase were localized to serine residues 490 and 499, and one site of phosphorylation by p34cdc2 was localized to serine 484. In vivo, rap1GAP undergoes phosphorylation at four distinct sites, two of which appear to be identical to the sites phosphorylated by cAMP-dependent kinase in vitro.

Amino Acid Sequence↗
PubMed1991

Isoprenoid modification of rab proteins terminating in CC or CXC motifs.

Mevalonate starvation of hamster fibroblasts resulted in a shift of rab1b from the membrane to the cytosolic fraction, suggesting that rab1b depends upon an isoprenoid modification for its membrane localization. rab1b and rab3a proteins expressed in insect cells incorporated a product of [3H]mevalonate, and gas chromatography analysis of material released by Raney nickel cleavage demonstrated that rab1b and rab3a are modified by geranylgeranyl groups. Additionally, in vitro prenylation analysis demonstrated farnesyl modification of H-ras but geranylgeranyl modification of five rab proteins (1a, 1b, 2, 3a, and 6). Together, these results suggest that the carboxyl-terminal CC/CXC motifs (X = any amino acid) specifically signal for addition of geranylgeranyl, but not farnesyl, groups. A rab1b mutant protein lacking the two carboxyl-terminal cysteine residues was not prenylated in vitro. However, since a mutant H-ras protein that terminates with tandem cysteine residues was also not modified, the CC motif may be essential, but not sufficient, to signal prenylation of rab1b. Finally, rab1b and rab3a proteins were not efficient substrates for either farnesyl- or geranylgeranyltransferase activities that modify CAAX-containing proteins (A = any aliphatic amino acid). Therefore, rab proteins may be modified by a prenyltransferase(s) distinct from the prenyltransferases that modify carboxyl-terminal CAAX proteins.

Amino Acid Sequence↗
PubMed1991

Molecular cloning of a GTPase activating protein specific for the Krev-1 protein p21rap1.

The rap1/Krev-1 gene encodes a ras-related protein that suppresses transformation by ras oncogenes. We have purified an 88 kd GTPase activating protein (GAP), specific for the rap1/Krev-1 gene product, from bovine brain. Based on partial amino acid sequences obtained from this protein, a 3.3 kb cDNA was isolated from a human brain library. Expression of the cDNA in insect Sf9 cells resulted in high level production of an 85-95 kd rap1GAP that specifically stimulated the GTPase activity of p21rap1. The complete deduced amino acid sequence is not homologous to any known protein sequences, including GAPs specific for p21ras. Northern and Western blotting analysis indicate that rap1GAP is not ubiquitously expressed and appears most abundant in fetal tissues and certain tumor cell lines, particularly the Wilms' kidney tumor, SK-NEP-1, and the melanoma, SK-MEL-3, cell lines.

Amino Acid Sequence↗