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T M Egan

Publications and source records attributed to T M Egan.

At least 19 recordsLinked to original sources
PubMed2006

Development of the new lung allocation system in the United States.

This article reviews the development of the new U.S. lung allocation system that took effect in spring 2005. In 1998, the Health Resources and Services Administration of the U.S. Department of Health and Human Services published the Organ Procurement and Transplantation Network (OPTN) Final Rule. Under the rule, which became effective in 2000, the OPTN had to demonstrate that existing allocation policies met certain conditions or change the policies to meet a range of criteria, including broader geographic sharing of organs, reducing the use of waiting time as an allocation criterion and creating equitable organ allocation systems using objective medical criteria and medical urgency to allocate donor organs for transplant. This mandate resulted in reviews of all organ allocation policies, and led to the creation of the Lung Allocation Subcommittee of the OPTN Thoracic Organ Transplantation Committee. This paper reviews the deliberations of the Subcommittee in identifying priorities for a new lung allocation system, the analyses undertaken by the OPTN and the Scientific Registry for Transplant Recipients and the evolution of a new lung allocation system that ranks candidates for lungs based on a Lung Allocation Score, incorporating waiting list and posttransplant survival probabilities.

Adolescent↗
PubMed2003

Molecular characterization of the zebrafish P2X receptor subunit gene family.

P2X receptors are non-selective cation channels gated by extracellular ATP and are encoded by a family of seven subunit genes in mammals. These receptors exhibit high permeabilities to calcium and in the mammalian nervous system they have been linked to modulation of neurotransmitter release. Previously, three complementary DNAs (cDNAs) encoding members of the zebrafish gene family have been described. We report here the cloning and characterization of an additional six genes of this family. Sequence analysis of all nine genes suggests that six are orthologs of mammalian genes, two are paralogs of previously described zebrafish subunits, and one remains unclassified. All nine subunits were physically mapped onto the zebrafish genome using radiation hybrid analysis. Of the nine gene products, seven give functional homo-oligomeric receptors when recombinantly expressed in human embryonic kidney cell line 293 cells. In addition, these subunits can form hetero-oligomeric receptors with phenotypes distinct from the parent subunits. Analysis of gene expression patterns was carried out using in situ hybridization, and seven of the nine genes were found to be expressed in embryos at 24 and 48 h post-fertilization. Of the seven that were expressed, six were present in the nervous system and four of these demonstrated considerable overlap in cells present in the sensory nervous system. These results suggest that P2X receptors might play a role in the early development and/or function of the sensory nervous system in vertebrates.

Adenosine Triphosphate↗
PubMed2002

Long term results of lung transplantation for cystic fibrosis.

OBJECTIVES: We reviewed our experience with lung transplant for cystic fibrosis (CF) over a 10-year period to identify factors influencing long-term survival. METHODS: One hundred and twenty-three patients with CF have undergone 131 lung transplant procedures at our institution; 114 have had bilateral sequential lung transplants (DLTX) and nine have had bilateral lower lobe transplants from living donors. Three patients had retransplant for acute graft failure, and five had late retransplant for bronchiolitis obliterans syndrome (BOS). Kaplan-Meier survival was calculated for the entire cohort and for subsets at higher risk of death to determine factors predicting a better outcome. RESULTS: Actuarial survival for the entire group of DLTX CF patients was 81% at 1 year, 59% at 5 years, and 38% at 10 years. Lobar transplant was associated with a poorer survival (37.5% at 1 and 5 years). Among DLTX patients, colonization with Burkholderia cepacia was present in 22 patients and was associated with poorer outcome (1- and 5-year survival 60 and 36% in B. cepacia patients vs. 86 and 64% in non-cepacia patients). DLTX patients younger than age 20 (n=22) had a similar survival to patients age 20 or older (n=90). Being on a ventilator at the time of transplant was not associated with poorer survival (n=8). BOS affects increasing numbers of survivors with time. Five CF patients have been retransplanted due to BOS with one operative death and 1-year survival of 60%. CONCLUSIONS: DLTX has acceptable long term survival in CF adults and children with end stage disease. CF patients colonized with B. cepacia have a worse outcome but transplantation is still warranted.

Adolescent↗
PubMed2001

On the contribution of the first transmembrane domain to whole-cell current through an ATP-gated ionotropic P2X receptor.

Scanning cysteine mutagenesis was used to identify potential pore-forming residues in and around the first transmembrane domains of ionotropic P2X(2) receptor subunits. Twenty-eight unique cysteine-substituted mutants (R28C-Y55C) were individually expressed in HEK293 cells by lipofection. Twenty-three of these were functional as assayed by application of ATP to transfected voltage-clamped cells. Individual mutants varied in their sensitivity to ATP; otherwise, currents through functional mutant receptors resembled those of the homomeric wild-type (WT) receptor. In five (H33C, R34C, I50C, K53C, and S54C) of 23 functional mutants, coapplication of 30 microm ATP and 500 nm Ag(+) irreversibly inhibited inward current evoked by subsequent applications of ATP alone. These inhibitions did not result in a lateral shift in the agonist concentration-response curve and are unlikely to involve a modification of the agonist binding site. Two (K53C and S54C) of the five residues modified by Ag(+) applied in the presence of ATP when the channels were gating were also modified by 1 mm (2-aminoethyl)methanethiosulfonate applied in the absence of ATP when the channels were closed. These data suggest that domains near either end of the first transmembrane domain influence ion conduction through the pore of the P2X(2) receptor.

Adenosine Triphosphate↗
PubMed2001

The first transmembrane domain of the P2X receptor subunit participates in the agonist-induced gating of the channel.

Based on pharmacological properties, the P2X receptor family can be subdivided into those homo-oligomers that are sensitive to the ATP analog alphabeta-methylene ATP(alphabetameATP) (P2X(1) and P2X(3)) and those that are not (P2X(2), P2X(4), P2X(5), P2X(6), and P2X(7)). We exploited this dichotomy through the construction of chimeric receptors and site-directed mutagenesis in order to identify domains responsible for these differences in the abilities of extracellular agonists to gate P2X receptors. Replacement of the extracellular domain of the alphabetameATP-sensitive rat P2X(1) subunit with that of the alphabetameATP-insensitive rat P2X(2) subunit resulted in a receptor that was still alphabetameATP-sensitive, suggesting a non-extracellular domain was responsible for the differential gating of P2X receptors by various agonists. Replacement of the first transmembrane domain of the rat P2X(2) subunit with one from an alphabetameATP-sensitive subunit (either rat P2X(1) or P2X(3) subunit) converted the resulting chimera to alphabetameATP sensitivity. This conversion did not occur when the first transmembrane domain came from a non-alphabetameATP-sensitive subunit. Site-directed mutagenesis indicated that the C-terminal portion of the first transmembrane domain was important in determining the agonist selectivity of channel gating for these chimeras. These results suggest that the first transmembrane domain plays an important role in the agonist operation of the P2X receptor.

Adenosine Triphosphate↗
PubMed2001

Polar residues of the second transmembrane domain influence cation permeability of the ATP-gated P2X(2) receptor.

P2X receptors are simple polypeptide channels that mediate fast purinergic depolarizations in both nerve and muscle. Although the depolarization results mainly from the influx of Na(+), these channels also conduct a significant Ca(2+) current that is large enough to evoke transmitter release from presynaptic neurons. We sought to determine the molecular basis of this Ca(2+) conductance by a mutational analysis of recombinant P2X(2) receptors. Wild type and 31 mutant P2X(2) receptors were expressed in HEK-293 cells and studied under voltage-clamp. We found that the relative Ca(2+) permeability measured from the reversal potentials of ATP-gated currents was unaffected by neutralizing fixed charge (Asp(315), Asp(349)) near the mouths of the channel pore. By contrast, mutations that changed the character or side chain volume of three polar residues (Thr(336), Thr(339), Ser(340)) within the pore led to significant changes in P(Ca)/P(Cs). The largest changes occurred when Thr(339) and Ser(340) were replaced with tyrosine; these mutations almost completely abolished Ca(2+) permeability, reduced P(Li)/P(Cs) by about one-half, and shifted the relative permeability sequence of Cs(+), Rb(+), K(+), and Na(+) to their relative mobility in water. Our results suggest that the permeability sequence of the P2X(2) receptor arises in part from interactions of permeating cations with the polar side chains of three amino acids located in a short stretch of the second transmembrane domain.

Adenosine Triphosphate↗
PubMed2001

Lung retrieval from non-heart beating cadavers with the use of a rat lung transplant model.

BACKGROUND: Lungs retrieved from cadavers after death and circulatory arrest may alleviate the critical shortage of lungs for transplant. We report a rat lung transplantation model that allows serial measurement of arterial blood gases after left single lung transplantation from non-heart beating donors. METHODS: Twelve Sprague-Dawley rats underwent left lung transplantation with a vascular cuff technique. Donor rats were anesthetized with intraperitoneal injection of pentobarbital, heparinized, intubated via tracheotomy, and then killed with pentobarbital. Lungs were retrieved immediately or after 2 hours of oxygen ventilation after death (tidal volume 1 mL/100 g, rate 40/min FIO2 = 1.0, positive end-expiratory pressure 5 cm H2O). Recipient rats were anesthetized, intubated, and ventilated. The carotid artery and jugular vein were cannulated for arterial blood gases and infusion of Ringer's lactate (4 mL/h). Anesthesia was maintained with halothane 0.2%, and recipient arterial blood gases were measured at 4 and 6 hours after lung transplantation after snaring the right pulmonary artery for 5 minutes. Animals were put to death 6 hours after lung transplantation, and portions of transplanted lungs were frozen in liquid nitrogen and assayed for wet/dry ratio, myeloperoxidase as a measure of neutrophil infiltration, and conjugated dienes as a measure of free radical-mediated lipid peroxidation. RESULTS: Arterial PO2 and wet/dry ratio were not significantly different in recipients of non-heart beating donor lungs retrieved immediately after death or after 2 hours of oxygen ventilation. Significant neutrophil infiltration was observed in recipients of non-heart beating donor lungs retrieved 2 hours after death from oxygen-ventilated donors. CONCLUSIONS: Strategies to ameliorate reperfusion injury may allow for successful lung transplantation from non-heart beating donors.

Animals↗
PubMed2001

Maintenance of cAMP in non-heart-beating donor lungs reduces ischemia-reperfusion injury.

Studies suggest that pulmonary vascular ischemia-reperfusion injury (IRI) can be attenuated by increasing intracellular cAMP concentrations. The purpose of this study was to determine the effect of IRI on capillary permeability, assessed by capillary filtration coeficient (Kfc), in lungs retrieved from non-heart-beating donors (NHBDs) and reperfused with the addition of the beta(2)-adrenergic receptor agonist isoproterenol (iso), and rolipram (roli), a phosphodiesterase (type IV) inhibitor. Using an in situ isolated perfused lung model, lungs were retrieved from NHBD rats at varying intervals after death and either ventilated with O(2) or not ventilated. The lungs were reperfused with Earle's solution with or without a combination of iso (10 microM) and roli (2 microM). Kfc, lung viability, and pulmonary hemodynamics were measured. Lung tissue levels of adenine nucleotides and cAMP were measured by HPLC. Combined iso and roli (iso/roli) reperfusion decreased Kfc significantly (p < 0.05) compared with non-iso/roli-reperfused groups after 2 h of postmortem ischemia. Total adenine nucleotide (TAN) levels correlated with Kfc in non-iso/roli-reperfused (r = 0.89) and iso/roli-reperfused (r = 0.97) lungs. cAMP levels correlated with Kfc (r = 0.93) in iso/roli-reperfused lungs. Pharmacologic augmentation of tissue TAN and cAMP levels might ameliorate the increased capillary permeability observed in lungs retrieved from NHBDs.

Adenine Nucleotides↗
PubMed2000

Molecular cloning and functional characterization of the zebrafish ATP-gated ionotropic receptor P2X(3) subunit.

We describe a P2X subunit cloned from the zebrafish (Danio rerio) that is an orthologue of the mammalian P2X(3) subunit. Like the mammalian P2X(3), this receptor desensitizes rapidly in the presence of agonist. However, it differs in that alphabeta-meATP is a much less potent agonist than ATP and the antagonist TNP-ATP is not active at low nanomolar concentrations. Similar to the rat P2X(3) subunit, the zebrafish subunit forms hetero-oligomeric assemblies with the rat P2X(2) that possesses a phenotype distinct from either parent. This novel clone will provide an important basis for future experiments investigating the structure/function relationships of P2X subunit domains.

Amino Acid Sequence↗
PubMed2000

Factors associated with adherence to treatment regimens after lung transplantation.

CONTEXT: The number of patients currently awaiting lung transplantation far exceeds the supply of available organs. Adherence to postoperative treatment regimens is essential for optimal posttransplant success. OBJECTIVE: The present study was designed to examine the demographic and psychological factors associated with compliance in patients who have had lung transplants. DESIGN: Eighteen women and 13 men participated in this study an average of 24 months after transplantation, completing a demographic form, a self-report compliance measure, a social support questionnaire, and the Multidimensional Health Locus of Control Scale. A significant other or family member and the posttransplant nurse coordinator also rated each subject's compliance with the posttransplant regimen. RESULTS: Although patients rated themselves as being compliant with aspects of their self-care, on more subtle measures of compliance, their self-reported compliance was not as impressive. Patients who had had their transplants more recently appeared to be more compliant. Patients with cystic fibrosis used their spirometer more often than patients with other lung diseases. Family support was significantly correlated with self-reported compliance. CONCLUSIONS: This study suggests that how patients are asked about adherence to treatment regimens influences how compliant they appear. The data also indicate that the longer after transplant, the less compliant the patient, and suggests the need for patient reeducation at some point after transplant. Longitudinal studies are needed to assess the degree to which compliance affects the number of rejection and febrile episodes as well as patient mortality after lung transplant.

Adolescent↗
PubMed1999

Identification of a domain involved in ATP-gated ionotropic receptor subunit assembly.

P2X receptors are ATP-gated ion channels found in a variety of tissues and cell types. Seven different subunits (P2X(1)-P2X(7)) have been molecularly cloned and are known to form homomeric, and in some cases heteromeric, channel complexes. However, the molecular determinants leading to the assembly of subunits into P2X receptors are unknown. To address this question we utilized a co-immunoprecipitation assay in which epitope-tagged deletion mutants and chimeric constructs were examined for their ability to co-associate with full-length P2X subunits. Deletion mutants of the P2X(2) receptor subunit were expressed individually and together with P2X(2) or P2X(3) receptor subunits in HEK 293 cells. Deletion of the amino terminus up to the first transmembrane domain (amino acid 28) and beyond (to amino acid 51) did not prevent subunit assembly. Analysis of the carboxyl terminus demonstrated that mutants missing the portion of the protein downstream of the second transmembrane domain could also still co-assemble. However, a mutant terminating 25 amino acids before the second transmembrane domain could not assemble with other subunits or itself, implicating the missing region of the protein in assembly. This finding was supported and extended by data utilizing a chimera strategy that indicated TMD2 is a critical determinant of P2X subunit assembly.

Adenosine Triphosphate↗
PubMed1999

Hetero-oligomeric assembly of P2X receptor subunits. Specificities exist with regard to possible partners.

P2X receptors are a distinct family of ligand-gated ion channels activated by extracellular ATP. Each of the seven identified subunit proteins (P2X1 through P2X7) has been reported to form functional homo-oligomeric channels when expressed in heterologous systems. Functional studies of native receptors, together with patterns of subunit gene expression, suggest that hetero-oligomeric assembly among members of this family may also occur. This prediction is supported by reports describing hetero-oligomeric assembly for three different recombinant subunit combinations. In this report, we systematically examined the ability of all members of the P2X receptor family to interact using a co-immunoprecipitation assay. The seven P2X receptor subunits were differentially epitope-tagged and expressed in various combinations in human embryonic kidney 293 cells. It was found that six of the seven subunits formed homo-oligomeric complexes, the exception being P2X6. When co-assembly between pairs of subunits was examined, all were able to form hetero-oligomeric assemblies with the exception of P2X7. Whereas P2X1, P2X2, P2X5, and P2X6 were able to assemble with most subunits, P2X3 and P2X4 presented a more restricted pattern of co-association. These results suggest that hetero-oligomeric assembly might underlie functional discrepancies observed between P2X responses seen in the native and recombinant settings, while providing for an increased diversity of signaling by ATP.

Adenosine Triphosphate↗
PubMed1999

Inadequacy of computed tomography in assessing patients with esophageal carcinoma after induction chemoradiotherapy.

BACKGROUND: Induction chemoradiotherapy followed by surgery may improve survival of patients with esophageal carcinoma. Computed tomography (CT) has been used to evaluate the tumor response after completing induction chemoradiotherapy. The authors examined the ability of CT to evaluate the pathologic tumor response to induction therapy and to stage the tumor correctly. METHODS: Preinduction and postinduction chemoradiotherapy CT scans were reviewed retrospectively for 50 patients enrolled in a protocol of induction chemoradiotherapy followed by surgery. All studies were performed on third-generation or fourth-generation scanners. Radiographic response was determined using Eastern Cooperative Oncology Group solid tumor response criteria for bidimensional measurable disease. This was compared with the pathologic tumor response. CT-tumor (T) classification using the modified Tio scale was compared with the pathologic T classification. RESULTS: CT-T classification did not correlate with the pathologic stage (P = 0.09) or the pathologic tumor response (P = 0.22). The postinduction chemoradiotherapy CT accurately staged the T classification in 42% of patients but overstaged 36% of patients and understaged 20% of patients. CT had a sensitivity of 65%, a specificity of 33%, a positive predictive value of 58%, and a negative predictive value of 41% in evaluating the pathologic tumor response. CONCLUSIONS: CT is a poor diagnostic study tool for determining the pathologic tumor response or the pathologic disease stage after induction chemoradiotherapy in patients with esophageal carcinoma.

Adult↗
PubMed1999

Reduced ischemia-reperfusion injury with rolipram in rat cadaver lung donors: effect of cyclic adenosine monophosphate.

BACKGROUND: The perfusion of rat lungs retrieved from cadavers with a solution containing isoproterenol has been shown to ameliorate the ischemia-reperfusion injury seen in lungs retrieved after death, and this protective effect parallels increases in tissue cyclic adenosine monophosphate levels. In this study, we investigated the effect of rolipram, a phosphodiesterase inhibitor, on capillary permeability and lung cyclic adenosine monophosphate levels in lungs retrieved from circulation-arrested rats. METHODS: Using an isolated perfused lung circuit, we retrieved lungs from circulation-arrested donor rats either ventilated with 100% oxygen or not ventilated for varying postmortem times. The lungs were reperfused with or without rolipram (2 micromol/L). The capillary filtration coefficient and wet to dry weight ratio, indicators of pulmonary vascular integrity, were determined, and tissue levels of adenine nucleotides and cyclic adenosine monophosphate were measured by high-performance liquid chromatography. RESULTS: The capillary filtration coefficient was significantly reduced in nonventilated cadaver lungs reperfused with rolipram 120 minutes after death (p<0.05). Oxygen ventilation or reperfusion with rolipram had a similar effect on the capillary filtration coefficient. Cyclic adenosine monophosphate levels were significantly higher in rolipram-reperfused lungs retrieved 120 minutes after death in both oxygen-ventilated (p<0.01) and nonventilated (p<0.01) lungs. CONCLUSIONS: In lungs from nonventilated, circulation-arrested donors, reperfusion with rolipram reduces the ischemia-reperfusion injury that may be due to intracellular cyclic adenosine monophosphate. Alteration of perfusate may have an impact on capillary leak caused by antecedent ischemia. Thus, rolipram may be a useful adjunct in the preservation of donor lungs retrieved after death.

Adenine Nucleotides↗
PubMed1999

Properties of the novel ATP-gated ionotropic receptor composed of the P2X(1) and P2X(5) isoforms.

We recently reported that a novel hetero-oligomeric P2X receptor is formed from the P2X(1) and P2X(5) isoforms when coexpressed in human embryonic kidney 293 cells (). A more complete description of the pharmacology of this novel receptor is presented here. A brief application of ATP to a voltage-clamped cell transiently expressing P2X(1/5) receptors resulted in a biphasic current that rapidly reached a peak and then decayed to a sustained plateau. Washout of ATP was accompanied by generation and fade of a pronounced tail of inward current. EC(50) values were determined from concentration-response curves for a range of agonists. The rank order of agonist potency was ATP >/= 2 methylthio ATP > adenosine 5'-O-(3-thiotriphosphate) > alpha,beta-methylene ATP > ADP > CTP. alpha,beta-methylene ADP, UTP, GTP, and AMP were ineffective. Only ATP and 2 methylthio ATP were full agonists. IC(50) values were determined from concentration-response curves for three commonly used purinergic antagonists. Suramin and pyridoxal phosphate-6-azophenyl-2', 4'-disulfonic acid were equipotent at P2X(1) and P2X(1/5) receptors; however, the P2X(1/5) receptor was much less sensitive to TNP-ATP than was the P2X(1) receptor. The amplitude of peak ATP-gated current was relatively insensitive to changes in [Ca(2+)](O) (1-30 mM). Finally, plateau currents were potentiated by low concentrations of pyridoxal phosphate-6-azophenyl-2', 4'-disulfonic acid and by raising [Ca(2+)](O). These results provide additional information on the pharmacological profile of the recombinant P2X(1/5) receptor channel and provide a basis to further evaluate ATP-induced currents in native tissues.

Adenosine Triphosphate↗
PubMed1998

N-Linked glycosylation is essential for the functional expression of the recombinant P2X2 receptor.

P2X receptors are integral membrane proteins that belong to the growing family of transmitter-gated ion channels. The extracellular domain of these receptors contains several consensus sequences for N-linked glycosylation that may contribute to the functional expression of the channel. We have previously reported the extracellular orientation of asparagine residues 182, 239, and 298 of the P2X2 receptor subunit by showing that the protein is glycosylated at each site [Torres, G. E., et al. (1998) FEBS Lett. 425, 19-23 (1)]. In this study, we focused on the consequences of removing N-linked glycosylation from the P2X2 receptor by using two different approaches, tunicamycin treatment or site-directed mutagenesis. HEK-293 cells stably transfected with the P2X2 receptor subunit showed little or no response to ATP after tunicamycin treatment. In addition, loss of function was observed with the elimination of all three N-linked glycosylation sites from P2X2. Cell surface labeling with biotin or indirect immunofluorescence revealed that the expression of the nonglycosylated receptors produced by either tunicamycin or site-directed mutagenesis is greatly reduced at the cell surface, indicating that the nonglycosylated P2X2 receptors are retained inside the cell. These data provide the first direct evidence for a critical role of N-linked glycosylation in the cell surface expression of a P2X receptor subunit.

Animals↗