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Biomedical subjects

R D Miller

Publications and source records attributed to R D Miller.

At least 55 records · Page 3Linked to original sources
PubMed1998

Marsupial light chains: complexity and conservation of lambda in the opossum Monodelphis domestica.

The Ig lambda chains in the South American opossum, Monodelphis domestica, were analyzed at the expressed cDNA and genomic organization level, the first described for a nonplacental mammal. The V lambda segment repertoire in the opossum was found to be comprised of at least three diverse V lambda families. Each of these families appears to be related to distinct V lambda families present in placental mammals, suggesting the divergence of these genes before the separation of metatherians and eutherians more than 100 million years ago. Based on framework and constant region sequences from full-length cDNAs and intron sequences from genomic clones, it appears that there are multiple functional J lambda-C lambda pairs in the opossum locus. The opossum J lambda-C lambda sequences are phylogenetically clustered, suggesting that these gene duplications are more recent and species specific. Sequence analysis of a large set of functional, expressed V lambda-J lambda recombinations is consistent with an unbiased, highly diverse lambda light chain repertoire in the adult opossum. Overall, the complexity of the Ig lambda locus appears to be greater than that found in the Ig heavy chain locus in the opossum, and light chains are therefore likely to contribute significantly to Ig diversity in this species.

Amino Acid Sequence↗
PubMed1998

V(H) repertoire of a marsupial (Monodelphis domestica).

When contrasted with information available for placental mammals, very little is known about the development of immunocompetence in marsupials. Marsupials, however, provide interesting immunology problems, since most appear to be born at a stage of development much less mature than that of placental mammals. To further understand the marsupial immune system, the Ig repertoire of the short-tailed opossum, Monodelphis domestica, was characterized. The majority of the V(H) clones were isolated in an unbiased manner by screening a spleen cDNA phage library, using C region probes, or anchored PCR, using C region-specific primers paired with vector specific primers. Analysis of 54 unique V(H) sequences from this marsupial revealed the presence of two V(H) families in the expressed Ig repertoire. The larger family, which contributed the majority of the clones identified, appears to be derived from 10 to 12 germline V(H) segments. The second family of clones is derived from a single germline V(H). Both V(H) families are related to the group III sequences described in other vertebrates. Unusual codon bias differences between the two families may result in very different patterns of somatic mutation within the opossum Ig repertoire.

Amino Acid Sequence↗
PubMed1998

Inhibition of Chlamydia pneumoniae replication in HEp-2 cells by interferon-gamma: role of tryptophan catabolism.

Interferon-gamma (IFN-gamma) induces tryptophan catabolism in HEp-2 cells, possibly via stimulation of host cell indoleamine-2,3-dioxygenase activity, in a dose-dependent (12.5-1600 U/mL) fashion after 24 h, resulting in a 99% conversion to its metabolites at 1600 U/mL. Replication of Chlamydia pneumoniae isolates A-03 and BAL-16 was inhibited in HEp-2 cells following treatment with 50 and 100 U/mL IFN-gamma, respectively; however, addition of excess L-tryptophan (200 microg/mL) to monolayers infected with C. pneumoniae resulted in unrestricted growth of both isolates up to 1600 U/mL IFN-gamma. C. pneumoniae could be recovered from IFN-gamma-treated monolayers, indicating the potential for this bacterium to undergo an altered life cycle, in vitro, analogous to that described in detail for Chlamydia trachomatis. The ability of C. pneumoniae to persist in host tissue despite an immunologic response would be an important attribute in order to cause or exacerbate chronic infections.

Animals↗
PubMed1998

Do the pharmacokinetics of vecuronium change during prolonged administration in critically ill patients?

Neuromuscular blocking drugs may be administered over several days to patients in the intensive care unit (ICU), but their pharmacokinetics have been studied at only one point in time, or assumed to be constant throughout the period of administration. We sought to determine if, in individual patients, the pharmacokinetics of vecuronium changed over the course of its administration in the ICU. In six critically ill patients, we measured plasma vecuronium concentrations during two periods: first, during initial administration of vecuronium and second, after its administration continuously for 3-6 days. A pharmacokinetic model was fitted to these plasma concentration data, and its parameters permitted to vary between the periods to determine if they had altered. Individual clearance values during the study ranged from 1.4 to 4.4 ml kg-1 min-1. During prolonged administration, vecuronium clearance increased in three and decreased in two patients. This change ranged from a 61% decrease to a 58% increase, and was not linked to any clinical factor. The steady-state volume of distribution (range 368-1765 ml kg-1; median 494 ml kg-1) did not change in any patient during the study. The change in clearance of vecuronium during its prolonged administration in critically ill patients suggests that future studies of neuromuscular blocking drugs in the ICU should take account of their changing pharmacokinetics over the course of administration.

Adult↗
PubMed1998

Characterization of a strain of Chlamydia pneumoniae isolated from a coronary atheroma by analysis of the omp1 gene and biological activity in human endothelial cells.

Chlamydia pneumoniae is a respiratory pathogen that has been associated with chronic inflammatory diseases such as asthma and atherosclerosis. Recent isolation of C. pneumoniae from human carotid and coronary atheromas provides additional support for a role of this organism in atherogenesis. We characterized the coronary strain C. pneumoniae A-03 by sequence analysis of the major outer membrane protein gene (omp1). In addition, the in vitro activities of A-03 and three respiratory strains of C. pneumoniae (BAL-16, TW-183, and T-2634) were examined in infected human umbilical vein endothelial cells (HUVEC) by analysis of the production of interleukin-8 (IL-8), monocyte chemotactic protein 1 (MCP-1), and soluble intercellular cell adhesion molecule 1 (sICAM-1). Sequence analysis of omp1 of C. pneumoniae A-03, compared to prototype strains TW-183 and AR-39, revealed five nucleotide changes resulting in nonsynonymous codons. Of interest was a nonconservative amino acid substitution (Ser to Pro) in position 61 of variable segment 1. In vitro, the extent of MCP-1, IL-8, and sICAM-1 production was dependent on the C. pneumoniae strain examined at low multiplicities of infection following 24 h of incubation. Strain A-03 displayed the lowest stimulatory activity in infected HUVEC, while T-2634 induced the highest levels of MCP-1, IL-8, and sICAM-1 among all strains examined. Heat-inactivated C. pneumoniae failed to stimulate production of these proteins by all strains tested. In contrast, only partial inhibition was observed by UV-inactivated organisms. Results from this study demonstrate that unlike prototype respiratory strains of C. pneumoniae, the coronary strain A-03 displays divergence in the omp1 gene. In addition, the stimulation of chemokines and adhesion molecules involved in the recruitment of leukocytes to sites of inflammation by C. pneumoniae may be important in the pathogenesis of diseases associated with this organism, including atherosclerosis.

Amino Acid Sequence↗
PubMed1998

Signal transduction during Legionella pneumophila entry into human monocytes.

Legionella pneumophila causes Legionnaires' disease by replication in alveolar macrophages and monocytes. The bacteria are internalized most efficiently by opsonin-dependent, CR3-mediated phagocytosis. This investigation focused on determining the role of actin polymerization and phosphorylation signals in this uptake mechanism. Uptake inhibition assays and confocal microscopic analysis indicated that entry of L. pneumophila activated tyrosine kinase (TK) and protein kinase C (PKC) and induced actin polymerization at the site of bacterial entry. Upon L. pneumophila entry, six major cellular proteins (75, 71, 59, 56, 53, and 52 kDa) were TK phosphorylated in soluble fractions of monocytes, and three of these proteins (52, 53, and 56 kDa) were consistently found in insoluble (i.e., cytoskeletal) fractions of monocytes as well. Tyrosine phosphorylation was suppressed when cells were pretreated with the kinase inhibitor genistein, tyrphostin, or staurosporine. A similar tyrosine-phosphorylated protein pattern was observed with CR3-mediated entry of avirulent L. pneumophila, Escherichia coli, or zymosan into monocytes. This study has shown that PKC and TK signals which activate actin polymerization during the process of phagocytosis are induced upon L. pneumophila entry. In addition, CR3 receptor-mediated phagocytosis into monocytes may involve tyrosine phosphorylation of similar proteins, regardless of the particle being phagocytosed. Therefore, the tyrosine-induced phosphorylation observed during opsonized L. pneumophila entry is not a virulence-associated event.

Actins↗
PubMed1998

Overexpression of protein kinase C-delta increases tight junction permeability in LLC-PK1 epithelia.

The Ca2+-independent delta-isoform of protein kinase C (PKC-delta) was overexpressed in LLC-PK1 epithelia and placed under control of a tetracycline-responsive expression system. In the absence of tetracycline, the exogenous PKC-delta is expressed. Western immunoblots show that the overexpressed PKC-delta is found in the cytosolic, membrane-associated, and Triton-insoluble fractions. Overexpression of PKC-delta produced subconfluent and confluent epithelial morphologies similar to that observed on exposure of wild-type cells to the phorbol ester 12-O-tetradecanoylphorbol-13-acetate. Transepithelial electrical resistance (RT) in cell sheets overexpressing PKC-delta was only 20% of that in cell sheets incubated in the presence of tetracycline, in which the amount of PKC-delta and RT were similar to those in LLC-PK1 parental cell sheets. Overexpression of PKC-delta also elicited a significant increase in transepithelial flux of D-[14C]mannitol and a radiolabeled 2 x 10(6)-molecular-weight dextran, suggesting with the RT decrease that overexpression increased paracellular, tight junctional permeability. Electron microscopy showed that PKC-delta overexpression results in a multilayered cell sheet, the tight junctions of which are almost uniformly permeable to ruthenium red. Freeze-fracture electron microscopy indicates that overexpression of PKC-delta results in a more disorganized arrangement of tight junctional strands. As with LLC-PK1 cell sheets treated with 12-O-tetradecanoylphorbol-13-acetate, the reduced RT, increased D-mannitol flux, and tight junctional leakiness to ruthenium red that are seen with PKC-delta overexpression suggest the involvement of PKC-delta in regulation of tight junctional permeability.

Animals↗
PubMed1998

Advance directives for psychiatric treatment: a view from the trenches.

Medical advance directives for end-of-life decisions at times that patients cannot express their preferences have become a major trend in recent state and federal legislation. The author argues that an extension of directives to cover future psychiatric treatment are fraught with both practical and ethical dilemmas. Although the potential for increasing patient autonomy is certainly positive, the potential for implementing covert ideological agendas has been present from the beginning, and the significant differences between medical end-of-life decisions and ongoing treatment of psychiatric patients must weigh against psychiatric advance directives unless major changes are made in existing procedures.

Advance Directives↗
PubMed1997

A family of fibrinogen-related proteins that precipitates parasite-derived molecules is produced by an invertebrate after infection.

After infection with the digenetic trematode Echinostoma paraensei, hemolymph of the snail Biomphalaria glabrata contains lectins comprised of 65-kDa subunits that precipitate polypeptides secreted by E. paraensei intramolluscan larvae. Comparable activity is lacking in hemolymph of uninfected snails. Three different cDNAs with sequence similarities to peptides derived from the 65-kDa lectins were obtained and unexpectedly found to encode fibrinogen-related proteins (FREPs). These FREPs also contained regions with sequence similarity to Ig superfamily members. B. glabrata has at least five FREP genes, three of which are expressed at increased levels after infection. Elucidation of components of the defense system of B. glabrata is relevant because this snail is an intermediate host for Schistosoma mansoni, the most widely distributed causative agent of human schistosomiasis. These results are novel in suggesting a role for invertebrate FREPs in recognition of parasite-derived molecules and also provide a model for investigating the diversity of molecules functioning in nonself-recognition in an invertebrate.

Amino Acid Sequence↗
PubMed1997

Duration of rocuronium-induced neuromuscular block during liver transplantation: a predictor of primary allograft function.

The prolongation of vecuronium-induced neuromuscular block has been reported as a predictor of hepatic allograft dysfunction. This study investigates the duration of action of rocuronium, which also relies on hepatic clearance, to examine whether it also is prolonged with allograft dysfunction. Fifty-seven patients undergoing orthotopic liver transplant were given rocuronium (0.6 mg/kg) prior to allograft placement and the recovery of contraction of the orbicularis oculi muscle to a 2-Hz train-of-four stimulus was recorded. Fifteen minutes after reperfusion of the allograft, rocuronium (0.6 mg/kg) was administered and the time to recovery of muscle contraction to a train-of-four stimulus (train-of-four time) was again recorded. The patients were divided into two groups according to posttransplant liver function. Group I consisted of 50 patients with immediate normal liver function. Group II contained 7 patients with primary dysfunctional livers. Primary dysfunction was determined by peak serum aspartate aminotransferase and alanine aminotransferase levels > 2000 U/L, and prothrombin time > 16 s. The train-of-four time in Group II was prolonged compared with Group I (P < 0.05). Immediate graft function testing using the recovery time from rocuronium of > 150 min has a positive predictive value of 100% and a negative predictive value of 96%. The sensitivity and specificity is 71% and 100%, respectively. Receiver operating characteristic analysis supports this conclusion.

Adolescent↗
PubMed1997

Pharmacokinetics of rocuronium during the three stages of liver transplantation.

BACKGROUND: Little is known about the influence of liver transplantation on the pharmacokinetics of most anesthetic drugs. The authors determined the pharmacokinetics of rocuronium during liver transplantation and examined whether variability in pharmacokinetics could explain variability in recovery of neuromuscular function. METHODS: Twenty patients undergoing liver transplantation were given rocuronium, 600 microg/kg, after induction of anesthesia and again after perfusion of the transplanted liver. Plasma was sampled to determine rocuronium concentrations. Pharmacokinetic models were fit to rocuronium concentrations versus time data using a mixed-effects population approach. Various models permitted changes in clearance (Cl) or central compartment volume to account for changes in hepatic function and circulatory status during the paleohepatic, anhepatic, and neohepatic periods. Time to initial recovery of four twitches of the orbicularis oculi was determined. RESULTS: During the paleohepatic and anhepatic periods, the typical value of Cl was 2.47 ml x kg(-1) x min(-1) and was not influenced by the magnitude of preexisting liver disease (as evidenced by prothrombin time, bilirubin, serum albumin, alanine transaminase [ALT], and aspartate transaminase [AST]). During the neohepatic period, the typical value of Cl varied as a function of the duration of warm ischemia of the hepatic allograft and was 2.72 ml x kg(-1) x min(-1) for a patient with an average 60-min period of warm ischemia; time to neuromuscular recovery varied as a function of Cl. CONCLUSIONS: Despite prolonged hypothermic ischemia, the newly transplanted liver eliminates rocuronium as well as the diseased native liver (and comparably with historical control values). However, some patients had decreased rocuronium Cl during the neohepatic period, apparently a result of prolonged graft warm ischemia. The authors' finding of preservation of hepatic drug elimination in the hepatic allograft is consistent with limited data for other drugs evaluated during anesthesia.

Adult↗
PubMed1997

Use of heteroduplex analysis to classify legionellae on the basis of 5S rRNA gene sequences.

Seventeen different species of Legionella, 12 serogroups of Legionella pneumophila, and 2 Legionella-like amoebal pathogens (LLAP1 and Sarcobium lyticum) were examined by heteroduplex analysis of PCR products of the 5S rRNA gene. Eight different banding patterns were identified, indicating that heteroduplex analysis of this gene can be used to classify these bacteria according to base substitutions between species. This classification may have future applications in clinical and epidemiological studies.

Bacterial Typing Techniques↗