Purification of synthetic DNA.
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Base Sequence↗
Biomedical subjects
T Brown
Publications and source records attributed to T Brown.
Relationship of cross-brain oxygen content difference, cerebral blood flow, and metabolic rate to neurologic outcome after near-drowning.
We evaluated the relationship of global cerebral blood flow, cross-brain oxygen content difference, cerebral metabolic rate for oxygen, intracranial pressure, and cerebral perfusion pressure to functional neurologic outcome in 12 comatose children on 2 consecutive days after near-drowning. Five children survived with functional neurologic outcome; five died and two survived with severe neurologic damage. Children who survived with functional neurologic outcome had a significantly higher cross-brain oxygen content difference (7.89 +/- 2.62 vs 3.91 +/- 1.59 ml/dl; p = 0.028) at 24 hours and a higher cerebral metabolic rate for oxygen 48 hours after admission (3.19 +/- 2.86 vs 0.96 +/- 0.45 ml/100 gm per minute; p = 0.030) compared with those who died or survived in a damaged state. There were no significant differences in global cerebral blood flow, intracranial pressure, and cerebral perfusion pressure between groups at either 24 or 48 hours. Our preliminary data suggest that a higher cross-brain content difference value is an important early variable associated with functional neurologic recovery after near-drowning. However, a single cross-brain oxygen content difference value must be interpreted with caution because considerable variability may occur among patient groups.
Adolescent↗
Use of comparison radiographs in the diagnosis of traumatic injuries of the elbow.
STUDY OBJECTIVE: To determine whether comparison radiographic views of the uninjured elbow result in increased diagnostic accuracy in elbow trauma. DESIGN: Physicians were provided with a short clinical summary and asked to interpret radiographs of the injured elbow or of both the injured and the uninjured elbow in a randomized fashion. SETTING: The radiology department in a university hospital. METHODS: Fifty sets of radiographs from 25 children with elbow injuries were reviewed by two residents, two emergency physicians, and one pediatric radiologist using a standard classification of injuries. For each child, there were two sets of radiographs: one of the injured elbow and one of both the injured and the uninjured elbow. Descriptive statistics were used to report the results. kappa statistics were used to determine interobserver and intraobserver agreement. Missed diagnoses were divided into those that were clinically relevant and those that were not. RESULTS: The overall percentage of correct diagnoses (one versus two elbow radiographs) were as follows: residents, 69% versus 70%; emergency physicians, 62% versus 67%; and pediatric radiologist, 74% versus 72% (P greater than .05). kappa scores for interobserver variability and intrarater agreement were in the moderate range (.383 to .805; kappa, .08). Clinically relevant diagnoses were missed by trainees and emergency physicians regardless of whether radiographs of just the injured elbow or both the injured and the uninjured elbow were interpreted. Incorrect radiograph interpretations were due to false-positives in 17 of 23 cases. CONCLUSION: Comparison radiographs of the uninjured elbow did not improve diagnostic accuracy in elbow trauma in the pediatric emergency department.
Adolescent↗
An absorptiometry method for the determination of arterial blood concentration of injected iodinated contrast agent.
A single photon absorptiometry method to measure the arterial concentration of injected iodinated contrast agent was developed. A prototype absorptiometry unit was built which consists of either a square or circular cross section acrylic (polymethylmethacrylate) cuvette connected to an arterial catheter at one end and a paristaltic pump at the other via PE60 surgical tubing. At opposing ends of the length of the cuvette were a 0.4 GBq 125I source and a scintillation crystal/photomultiplier tube assembly. This assembly was connected to a single-channel analyser (SCA)/scaler unit to count the transmitted photons through the cuvette. The scaler was interfaced to an IBM PC and counts accumulated in preset time intervals were transmitted to the computer via a serial interface. Experiments were performed to calibrate the unit for measurement of blood concentration of contrast agent (Isovue 300) and to determine the dispersion characteristics of the unit. Deconvolution was used to correct the measured concentration waveform for the dispersion introduced by passage through the lead-in tubing and the cuvette. The precision of concentration measurements was determined to be between 5 and 10% using computer simulations and theoretical calculations. The method was used successfully in a number of patient and animal studies to measure the contrast concentration in blood following intravenous injection of contrast agent.
Absorptiometry, Photon↗
A phase I clinical and pharmacokinetic study of the topoisomerase I inhibitor topotecan (SK&F 104864) given as an intravenous bolus every 21 days.
Topotecan (SK&F 104864) is a novel antitumor agent whose mechanism of action is inhibition of the DNA unwinding protein topoisomerase I. An analog of camptothecin, topotecan was designed to be more water soluble in an effort to decrease the severe and sporadic toxicities experienced during phase I/II trials of the parent compound. In this phase I clinical and pharmacological trial, topotecan was given as a bolus intravenous (i.v.) infusion over 30 min every 21 days. A total of 42 patients entered the study, receiving doses ranging from 2.5 to 22.5 mg/m2. The maximum tolerated dose (MTD) of topotecan given in this schedule was 22.5 mg/m2. Myelosuppression, primarily neutropenia, was dose-limiting. The extent of prior therapy did not predict for more severe neutropenia. Non-hematologic toxicities were mild and included low-grade to moderate fever, nausea, vomiting, alopecia, diarrhea and skin rashes. There were no objective partial or complete responses, although there was a suggestion of antitumor activity in three patients. Topotecan undergoes pH-dependent hydrolysis of the lactone ring; only the closed, lactone form is active. The lactone form predominated during infusion, with hydrolysis occurring rapidly following the end of infusion. There were linear relationships between dose administered and peak plasma lactone concentrations as well as AUC lactone to AUC total. The lactone was rapidly cleared from plasma with a total body clearance of 25.7 (+/- 6.7) l/h/m2. The plasma lactone concentration declined rapidly with a harmonic mean terminal half-life of 3.4 (+/- 1.1)h. Lactone hydrolysis and renal excretion were the major routes of elimination.(ABSTRACT TRUNCATED AT 250 WORDS)
Adult↗
The fifth cause of splenomegaly?--Parvovirus B19.
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Acute Disease↗
The tissue distribution of tumor necrosis factor biosynthesis during endotoxemia.
Tumor necrosis factor (TNF) is a protein hormone implicated in the development of septic shock and other pathologic states. However, complexities inherent in detecting TNF synthesis by individual tissues have left the precise origins of this protein undefined. In addition, the possibility that localized TNF production may contribute to the pathogenesis of organ-specific diseases such as type I diabetes has not been explored in vivo. We have developed a transgenic mouse line bearing a reporter gene construct in which the TNF coding sequence and introns are replaced by a chloramphenicol acetyltransferase (CAT) coding sequence. In normal transgenic animals, CAT activity is expressed only in the thymus. When endotoxin is administered to the animals, CAT activity is also evident in kidney, heart, islets of Langerhans, spleen, lung, fallopian tubes, and uterus, but not in other organs. The biosynthesis of CAT in vivo correlated with tissue capacity to secrete TNF in vitro. Thus, TNF was secreted by all the tissues that expressed CAT, including lung, spleen, thymus, uterus/fallopian tubes, pancreatic islets, renal glomeruli, and cultured cardiac cells after exposure to endotoxin.
Animals↗
Visual compatibility of tacrolimus with commonly used drugs during simulated Y-site injection.
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Chemical Precipitation↗
N.m.r. determination of the solution conformation and dynamics of the A.G mismatch in the d(CGCAAATTGGCG)2 dodecamer.
A.G base-paired mismatches that occur during replication are among the most difficult to detect by repair enzymes. Such purine.purine mispairs can exist in two conformations, one of which is stabilized by protons [Gao & Patel (1988) J. Am. Chem. Soc. 110, 5178-5182]. We have undertaken a 1H-n.m.r. and 31P-n.m.r. study of the mismatched dodecamer d(CGCAAATTGGCG)2 as a function of both temperature and pH to determine the conformational features of the A.G mismatch. At pH greater than 7 the mispaired bases are each in the anti conformation and are stacked in the B-like helix. As the pH is decreased, a second conformation becomes populated (apparent pKa approx. 5.9) with concomitant changes in the chemical shifts of protons of the mispaired bases and their nearest neighbours. Data from two-dimensional nuclear-Overhauser-enhancement spectroscopy show unequivocally that, at low pH, the dominant conformation is one in which the mismatched G residues are in the syn conformation and are hydrogen-bonded to the A residues that remain in the anti conformation. Residues not adjacent to the A.G sites are almost unaffected by the transition or the mispairing, suggesting considerable local flexibility of the unconstrained duplexes. Despite the bulging of the mispaired bases, the conformation of the A(anti).G(anti) duplex is very similar to the native dodecamer, whereas the AH+(anti).G(syn) duplex shows a greater variation in the backbone conformation at the mismatched site. According to the chemical shifts, the duplex retains twofold symmetry in solution. The equilibrium between the syn and anti conformations of G9/G21 is strongly dependent on pH, but only weakly dependent on temperature (delta H approx. 16 kJ.mol-1). The first-order rate constant for the transition is approx. 9 s-1 at 283 K and approx. 60 s-1 at 298 K, with an activation enthalpy of approx. 100 kJ.mol-1. The stabilization of the A(anti).G(syn) conformation by protons is consistent with models invoking N1 protonation of adenine. Using the derived glycosidic torsion angles we have used restrained molecular dynamics to build models of the neutral and protonated d(CGCAAATTGGCG)2 oligomers. The results confirm that the A(anti).G(anti) and AH+(anti).G(syn) conformations are favoured at high pH and low pH respectively, in accord with n.m.r. and single-crystal X-ray data.
Base Sequence↗
The synthesis of oligonucleotides that contain 2,4-dinitrophenyl reporter groups.
The synthesis of non-nucleoside-based phosphoramidites that bear the 2,4-dinitrophenyl group is reported. These labelled phosphoramidites, which have been used in solid-phase oligonucleotide synthesis to attach single and multiple dinitrophenyl groups to the 5'-end of oligonucleotides, are entirely compatible with the normal oligonucleotide synthesis cycle. Multiple labelling can be performed readily in high yield and the resulting oligonucleotides can be purified readily by reversed-phase h.p.l.c. The labelled oligonucleotides have been detected using monoclonal and polyclonal anti-dinitrophenyl antibodies.
2,4-Dinitrophenol↗
Interaction of berenil with the EcoRI dodecamer d(CGCGAATTCGCG)2 in solution studied by NMR.
The conformation of the EcoRI dodecamer d(CGCGAATTCGCG)2 has been examined in solution by 1H and 31P NMR. Spin-spin coupling constants and nuclear Overhauser (NOE) enhancement spectroscopy show that all deoxyriboses lie in the south domain, with a small admixture of the north conformation (0-20%). The time dependence of the nuclear Overhauser enhancements also reveals a relatively uniform conformation at the glycosidic bonds (average angle, chi = -114 degrees). The average helical twist is 36.5 degrees (9.8 base pairs per turn). Tilt angles are small (in the range 0 to -10 degrees), and roll angles are poorly determined. Unlike single-crystal X-ray studies of the same sequence, there is no evidence for asymmetry in the structure. Both the NOE intensities and 31P relaxation data imply conformational anomalies at the C3-G4/C9-G10 and the A5-A6/T7-T8 steps. Berenil binds in 1:1 stoichiometry to the dodecamer with high affinity (Kd = 1 microM at 298 K) and causes substantial changes in chemical shifts of the sugar protons of nucleotides Ado 5-Cyt 9 and of the H2 resonances of the two Ado residues. No significant asymmetry appears to be induced in the DNA conformation on binding, and there is no evidence for intercalation, although the binding site is not centrosymmetric. NOEs are observed between the aromatic protons of berenil and the H1' of both Thy 7 and Thy 8, as well as to Ado 5 and Ado 6 H2. These results firmly establish that berenil binds via the minor groove and closely approaches the nucleotides Ado 6, Thy 7, and Thy 8. On the basis of quantitative NOE spectroscopy and measurements of spin-spin coupling constants, changes in the conformations of the nucleotides are found to be small. Using the observed NOEs between the ligand and the DNA together with the derived glycosidic torsion angles, we have built models that satisfy all of the available solution data. The berenil molecule binds at the 5'-AAT (identical to 5'-ATT on the complementary strand) site such that (i) favorable hydrogen bonds are formed between the charged amidinium groups and the N3 atoms of Ado 6 and Ado 18 and (ii) the ligand is closely isohelical with the floor of the minor groove.
Base Sequence↗
Prenatal diagnosis for dystrophia myotonica using the polymerase chain reaction.
The polymerase chain reaction has been used to detect an abundant class of short repeat DNA families of the form (dC-dA)n.(dG-dT)n, known as microsatellites. These units are found throughout the human genome and have been characterized for several loci including APOC2 on chromosome 19q12-q13.2. The locus APOC2 is linked to the gene for dystrophia myotonica and a microsatellite within this locus was used to derive polymorphisms in a family to predict the inheritance of the disease. Chorionic villus sampling (CVS) was performed at 15 1/2 weeks' gestation. Following DNA extraction from the CVS material and parental blood samples, microsatellite analysis was carried out by the polymerase chain reaction.
Adult↗
Chemical and biological studies on a series of lipid-soluble (trans-(R,R)- and -(S,S)-1,2-diaminocyclohexane)platinum(II) complexes incorporated in liposomes.
cis-Bis(neodecanoato)(trans-(R,R)-1,2-diaminocyclohexane)platinum( II) [L-NDDP] is a liposome incorporated lipophilic cisplatin analogue that has shown promising antitumor activity against tumors resistant to cisplatin and liver metastases in mice. L-NDDP is currently under clinical evaluation. However, NDDP is an isomeric mixture of different species having various isomeric neodecanoic moities as liganded leaving groups. A series of new highly lipid-soluble cis-bis(neodecanoato)(trans-(R,R)- and -(S,S)-1,2-diaminocyclohexane)platinum(II) [Pt] complexes, using single isomers of neodecanoic acid, were synthesized and characterized by analytical and spectroscopic techniques (infrared and 195Pt NMR). Multilamellar vesicles (MLVs) composed of dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG) at a molar ratio of 7:3 were used as carriers of the Pt complexes. The efficiency of incorporation of the liposomal-platinum (L-Pt) preparations was greater than 95% and stability in normal saline at 4 degrees C was greater than 95% at day 14 in each case. The iv LD50 values of all L-Pt preparations tested were in the range of 62.3 to 104 mg/kg. The % T/C obtained after a single ip injection of the optimal dose of L-Pt preparations against L1210 leukemia was in the range of 150 to 253 (160 for cisplatin). When a multiple ip injection schedule was used (on days 1, 5, and 9) the L-Pt preparations of R,R complexes (1, 7, and 9) were more active than cisplatin at the optimal dose (% T/C = 257 for each vs 220 for cisplatin). The L-Pt preparations of R,R complexes were also markedly active against L1210 leukemia resistant to cisplatin (% T/C 355, 231, and 185 respectively vs 112 for cisplatin). These studies show that the single isomers of NDDP are comparable to the original isomeric mixture in terms of toxicity and biological activity.
Animals↗
Relative incidence of intracranial mass lesions and severe torso injury after accidental injury: implications for triage and management.
In a previous study of head injury patients we found that old age, low Glasgow Coma Scale (GCS) score, pupillary inequality, and falls were significant predictors of intracranial mass lesions (IMLs). Injury to motor vehicle occupants was less likely to result in IML. The present study defines predictors of severe torso injury (STI) in 646 patients admitted to a trauma unit and compares these with predictors of IML obtained in the previous study. Tachycardia and low blood pressure were associated with an increased incidence of STI (p = 0.003, p = 0.0000). The incidence of STI in falls differed from that of IML (13.2% vs. 47.7%, p less than 0.001). There was a greater incidence of STI than IML in MVAs (33.6% vs. 14.8%, p less than 0.001). Patients 70 years of age or older had a higher incidence of IMLs than STIs (p less than 0.001). Patients less than 30 years old had a significantly greater incidence of STIs than IMLs (p less than 0.001). These data suggest that in MVA victims who are less than 30 years old, are hypotensive, and tachycardic, the diagnosis and emergent treatment of severe torso injury should take precedence over measures designed to detect and treat intracranial mass lesions. The converse is true for older patients injured in falls.
Abdominal Injuries↗
A papular eruption and acute abdominal pain associated with coxsackie A-14 virus.
A 14-year-old girl who was admitted to hospital with acute abdominal pain developed an erythematous papular eruption which persisted for 4 weeks. Examination of a stool specimen yielded evidence of a coxsackie A-14 virus.
Abdomen, Acute↗
The treatment of facial disharmony and malocclusion by jaw surgery. Case reports.
Examples are given of the Australian Cranio-Facial Unit's methods of dealing with facial disharmony and malocclusion by orthognathic surgery. Members of the Unit specialize in different aspects of craniofacial structure and function, and their teamwork is described. Treatment planning and assessment are illustrated by traditional cephalometric and other analytical procedures, and by pattern profile analysis. An appraisal is made of the Unit's increasing use of computer technology to facilitate these analyses, and to integrate various sources of information about facial disharmony.
Adolescent↗
New aniline blue dye medium for rapid identification and isolation of Candida albicans.
Organic dyes have long been used in diagnostic microbiology to differentiate species by color reactions. We studied the ability of a new noninhibitory medium, YM agar containing 0.01% aniline blue WS dye, Colour Index 42780 (YMAB), to identify Candida albicans among 1,554 yeast specimens obtained from seven clinical laboratories. Appropriate American Type Culture Collection and other characterized strains served as controls. A total of 487 of the clinical strains were identified as C. albicans. The remainder were other Candida species and non-Candida yeasts. Clinical isolates and controls were grown on Sabouraud agar for 18 h at 30 degrees C and then transferred to YMAB. Plates were incubated for 12 to 18 h at 30 degrees C, and colonies were observed for yellow-green fluorescence under long-wave UV light (A365). All control strains of C. albicans and Candida stellatoidea fluoresced, as did 480 of the 490 isolates designated as C. albicans (which included 3 strains of C. stellatoidea). Cells of C. albicans grown on YMAB produced germ tubes in serum. Only five of the other 1,062 non-C. albicans yeasts fluoresced. The sensitivity and specificity were 98.0 and 99.5%, respectively, with a predictive value of 99.1%. A fluorescent metabolite was found in cell wall particulate fractions of C. albicans sonic extracts grown on YMAB but not in non-C. albicans yeasts. This metabolite showed the same spectral curve as those of metabolites from whole cells in a recording spectrofluorometer when it was excited at 400 nm and scanned from 420 to 550 nm. Thus, growth on YMAB generates the production of a fluorescent moiety that can be used to specifically identify C. albicans within 12 to 18 h.
Aniline Compounds↗
A CAT reporter construct allows ultrasensitive estimation of TNF synthesis, and suggests that the TNF gene has been silenced in non-macrophage cell lines.
We have prepared a construct (designated CATTNF) in which the mouse TNF (cachectin) coding sequence is replaced by a sequence encoding chloramphenicol acetyltransferase (CAT), with preservation of the TNF promoter and 3'-untranslated sequences known to be important in the regulation of gene expression. When activated by LPS, permanently transfected RAW 264.7 (mouse macrophage) cells synthesize large quantities of CAT. Unlike TNF itself, CAT is nonsecreted and quite stable in the macrophage cytoplasm. Fewer than 1,000 LPS-induced macrophages can easily be detected by CAT assay. Cells maintain the ability to respond to LPS in vivo; as such, when injected intravenously, they accurately report conditions required for the production of TNF in diverse tissues. These cells may thus be used for the detection of cachectin/TNF synthesis in mice under conditions in which endogenously produced cachectin/TNF would be undetectable. Studies of the expression of CATTNF in nonmacrophage cell lines have revealed that the modified TNF gene is constitutively expressed in L-929 cells, but that its expression is tightly suppressed in HeLa cells and in NIH 3T3 cells. This finding would suggest that certain non-macrophage cells are potentially capable of utilizing the TNF promoter and translating the TNF mRNA; however, the endogenous gene has been developmentally silenced.
Animals↗