Accuracy of recorded body temperature of critically ill patients related to measurement site: a prospective observational study.

Accurate measurement of body temperature is an important indicator of the status of critically ill patients and is therefore essential. While axillary temperature is not considered accurate, it is still the conventional method of measurement in Asian intensive care units. There is uncertainty about the accuracy of thermometers for the critically ill. We compared the accuracy and precision of bladder, axillary and tympanic temperature measurements in critically ill patients. A total of 73 critically ill patients admitted to the intensive care unit of a teaching hospital were prospectively enrolled. Every four hours, we measured body temperature at three sites (bladder, axillary and tympanic). If the patient had received an indwelling pulmonary artery catheter, blood temperature was also recorded and this was compared with bladder, axillary and tympanic temperature readings. For all patients, axillary and tympanic temperature readings were compared with bladder temperature readings. Accuracy and precision were analysed using Bland-Altman analysis. When blood temperature data was available, the mean difference between blood and bladder temperature readings was small (0.02±0.21°C). Compared with bladder temperature, mean difference for axillary temperature was -0.33±0.55°C and for tympanic temperature it was -0.51±1.02°C. For critically ill patients, recorded axillary temperature was closer to bladder temperature than tympanic temperature.

Drosophila AD3 mutation of synaptotagmin impairs calcium-dependent self-oligomerization activity.

Genetic analysis of a Drosophila synaptotagmin (Syt) I mutant (AD3) has revealed that Tyr-334 within the C2B domain is essential for efficient Ca(2+)-dependent neurotransmitter release. However, little is known as to why a missense mutation (Tyr-334-Asn) disrupts the function of the C2B domain at the molecular level. Here, we present evidence that a Tyr-312 to Asn substitution in mouse Syt II, which corresponds to the Drosophila AD3 mutation, completely impairs Ca(2+)-dependent self-oligomerization activity mediated by the C2B domain but allows partial interaction with wild-type proteins in a Ca(2+)-dependent manner. This observation is consistent with the fact that the AD3 allele is homozygous lethal but complements another mutant phenotype. We also showed that the Ca(2+)-dependent C2B self-oligomerization is inhibited by inositol 1,3,4, 5-tetrakisphosphate, a potent inhibitor of neurotransmitter release. All of these findings strongly support the idea that self-oligomerization of Syt I or II is essential for neurotransmitter release in vivo.

Synaptotagmin IV is present at the Golgi and distal parts of neurites.

Synaptotagmin IV (SytIV) is an immediate early gene induced by membrane depolarization in PC12 cells and in rat brain. However, little is known about the function of SytIV or the functional relationship between SytIV and SytI, because SytIV has yet to be localized. Here we show that SytIV was localized at the Golgi and distal part of neurites in nerve growth factor-differentiated PC12 cells and cultured hippocampal neurons by immunocytochemistry using an isoform-specific antibody (anti-SytIV). These SytIV signals were not colocalized well with SytI signals. Upon membrane depolarization, SytIV signals were increased at both the Golgi and distal part of neurites within several hours in both types of cells. We further show that the increase of SytIV protein levels results from protein kinase A-dependent gene up-regulation. In hippocampal neurons, SytIV was developmentally regulated. These results suggest that SytIV may play a role at the Golgi and tips of neurites during development and synaptic plasticity.

Role of synaptotagmin, a Ca2+ and inositol polyphosphate binding protein, in neurotransmitter release and neurite outgrowth.

Synaptotagmin I (or II), a possible Ca(2+)-sensor of synaptic vesicles, has two functionally distinct C2 domains: the C2A domain binds Ca2+ and the C2B domain binds inositol high polyphosphates (IP4, IP5, and IP6). Ca(2+)-regulated exocytosis of secretory vesicles is proposed to be activated by Ca2+ binding to the C2A domain and inhibited by inositol polyphosphate binding to the C2B domain. Synaptotagmins now constitute a large family and are thought to be involved in both regulated and constitutive vesicular trafficking. They are classified from their distribution as neuronal (synaptotagmin I-V, X, and XI) and the ubiquitous type (synaptotagmin VI-IX). Among them, synaptotagmins III, V, VI and X are deficient in IP4 binding activity due to the amino acid substitutions in the C-terminal region of the C2B domain, suggesting that these isoforms can work for vesicular trafficking even in the presence of inositol high polyphosphates. Synaptotagmin I is also known to be present in neuronal growth cone vesicles. Antibody against the C2A domain (anti-C2A) that inhibits Ca(2+)-regulated exocytosis also blocked neurite outgrowth of the chick dorsal root ganglion (DRG) neuron, suggesting that Ca(2+)-dependent synaptotagmin activation is also crucial for neurite outgrowth.

Functional involvement of synaptotagmin I/II C2A domain in neurite outgrowth of chick dorsal root ganglion neuron.

Synaptotagmin I or II (Syt I/II) is involved in Ca2+-regulated exocytosis of secretory vesicles, probably serving as a Ca2+-sensor via its C2A domain. Synaptotagmin is also known to be expressed in neuronal growth cone vesicles, but its functional involvement in neurite outgrowth remains largely unknown. In this study, we examined the function of Syt I/II in neurite outgrowth in cultured chick dorsal root ganglion neurons using an anti-synaptotagmin I and II C2A domain (anti-STI/II-C2A) antibody that inhibits Ca2+-regulated exocytosis. Immunoblots confirmed the high specificity of the anti-STI/II-C2A antibody and showed the expression of synaptotagmin I or II in chick dorsal root ganglion neurons. Immunocytochemistry revealed that synaptotagmin I or II is enriched at the growth cone region of chick dorsal root ganglion neurons, in both lamellipodia and filopodia. Whole or Fab-fragment of the anti-STI/II-C2A antibody loaded into dorsal root ganglion neurons by trituration significantly inhibited neurite outgrowth, whereas preimmune immunoglobulin G had no effect. These results showed that the C2A domain of synaptotagmin I or II plays a crucial role in neurite outgrowth.

[The new criteria for skin prick test of atopic early infants--diagnosis for hypersensitivity of egg white].

To investigate the new criteria for skin prick test (SPT) of seventy-four atopic infants (2-5 months of age at the first visit, Mean 3.8 months, M:F = 54:20) to diagnose for hypersensitivity to egg white. It was classified into three groups by reaction type of SPT in the first visit. Group A were the infants who seemed only late (6 hours) or delayed (48 hours) reaction (n = 26). Group B were seemed immediate (15 minutes) and late or delayed reactions (n = 26), Group C were seemed only immediate reaction (n = 23). Atopic infants and controlled infants without no symptom but have any atopic disease a relative in the third degree, agreed to undergo SPT in the first visit, the prior were undergo 9-12 months of age, too. Serum total IgE (RIST), serum specific IgE antibody of egg white (EWRAST) and peripheral eosinophil counts in the blood (Eo. counts) were determined at the same time of SPT in atopic infants. The best criterion for SPT was the longest diameter of a erythema were greater than 3 mm at late and/or delayed reaction (Sensitivity: 100%, Specificity: 60%) in group A. Two third of infants in group. A were seemed immediate reaction and EWRAST levels were increased to larger than gread two at 9-12 months (p < 0.001). RIST levels and Eo. counts at the first visit were increased compared with the normal levels in the all groups, the prior and EWRAST levels in group B were higher than group A or C (p < 0.05, p < 0.05). RIST and EWRAST levels in group A at 9-12 months were higher than the first visit (p < 0.05, p < 0.01). In conclusion, SPT in atopic early infants were seemed several reactions at the first visit, but all reactions were useful for diagnose for hypersensitivity to egg white.

Mutation of the pleckstrin homology domain of Bruton's tyrosine kinase in immunodeficiency impaired inositol 1,3,4,5-tetrakisphosphate binding capacity.

Bruton's tyrosine kinase (Btk), a cytoplasmic protein-tyrosine kinase, plays a pivotal role in B cell activation and development. Mutations in the pleckstrin homology (PH) domain of the Btk gene cause human X-linked agammaglobulinemia (XLA) and murine X-linked immunodeficiency (Xid). In this paper, we report that the PH domain of Btk functions as an inositol 1,3,4,5-tetrakisphosphate (IP4), inositol 1,3,4,5,6-pentakisphosphate, and inositol 1,2,3,4,5,6-hexakisphosphate (IP6) binding domain (Kd of approximately 40 nM for IP4), and that all of the XLA (Phe replaced by Ser at position 25 (F25S), R28H, T33P, V64F, and V113D) and Xid mutations (R28C) found in the PH domain result in a dramatic reduction of IP4 binding activity. Furthermore, the rare alternative splicing variant, with 33 amino acids deleted in the PH domain, corresponding to exon 3 of the Btk gene, also impaired IP4 binding capacity. In contrast, a gain-of-function mutant called Btk*, which carries a E41K mutation in the PH domain, binds IP6 with two times higher affinity than the wild type. Our data suggest that B cell differentiation is closely correlated with the IP4 binding capacity of the PH domain of Btk.

[A simplified method of HLA-DR typing with the PCR method using the genomic DNA extracted from blood-absorbed filter].

The enzymatic amplification of specific DNA sequences by the polymerase chain reaction (PCR) has provided a new approach to genetic typing of HLA-class II region specificities. However, we found the existing PCR protocols to be adequate and/or time consuming, especially when large numbers of samples need to be processed. We have devised a simplified method which alleviates these problems. Genomic DNA prepared from peripheral blood nucleated cells was facilitated using the punched-out filter paper absorbed with the peripheral blood. Second exon HLA-DRB segments of the genomic DNA were amplified by the polymerase chain reaction (PCR) (30 cycles: denaturing at 94 C for 1 min; annealing at 55 for 1 min, extension at 72 C for 2 min) using HLA-DRB primer DRB 5'-1 (ACCGGTCGTTCITGTCCCCICAGCA) and DRB 3'-1 (CTCGCCICTGCACIGTIAAGC) designed in our laboratory. The presence of specific alleles in a PCR-amplified sample was determined by dot-blot hybridization with 32 P-labeled sequence-specific-oligonucleotides (SSOs). Quantitation of the producer is being evaluated using an automated scanner. These newly designed primers allowed amplification of all the known expressed alleles of DRB 1, B 3, B 4 and B 5 loci. This type of reproducible and precise assay is very useful in the HLA-class II typing of large number of patients.

[A comparison of two delivery methods on the bronchodilator effects of beta 2-stimulant].

The airway response to a bronchodilator, salbutamol delivered by a new handy ultrasonic nebulizer and the compressed air nebulizer was studied in a blind manner. Patients by the ultrasonic nebulizer had better percent increase in FVC and FEV1 significantly and in other parameters showed a tendency to being better than those by the compressed air nebulizer. These results suggest that this ultrasonic nebulizer may be more useful than the compressed air nebulizer as inhalation therapy for bronchial asthma. This ultrasonic nebulizer has several advantages over the compressed air nebulizer. This nebulizer is compact size, light weight and has dry batteries as power source, making it more convenient for travel and use at workplace.

Regulation of lrp gene expression by H-NS and Lrp proteins in Escherichia coli: dominant negative mutations in lrp.

Lrp (leucine-responsive regulatory protein) is a global transcription factor of Escherichia coli and regulates, negatively or positively, many genes including lysU, which encodes lysyl-tRNA synthetase. Dominant negative mutations that derepress lysU expression were isolated in this study. These mutations affected a predicted DNA-binding domain of Lrp and mutants were defective DNA-binding domain of Lrp and mutants were defective both in activation of ilvIH expression and in repression of lysU expression. Consistent with the previous notion that lrp is autoregulated, lrp expression was derepressed by these mutations and repressed by multi-copy plasmids carrying lrp+. Moreover, we found by gene fusion and Northern blot hybridization that the "histone-like" protein, H-NS, bound specifically to a promoter segment of lrp in vitro, and the level of lrp expression increased in the hns null mutant. These results indicated that the lrp gene is not only feedback regulated by Lrp but is also controlled by H-NS protein.

Pharmacokinetics and pharmacodynamics of the tulobuterol patch, HN-078, in childhood asthma.

BACKGROUND: Recently, a novel percutaneous adhesive tulobuterol preparation, HN-078, has been developed and tests using healthy adult subjects have indicated it to be effective for controlling exacerbations early in the morning if applied at bedtime. In children, percutaneous application is very important to eliminate side effects, including abdominal pain and appetite loss. OBJECTIVE: We report the pharmacokinetics and pharmacodynamics of tulobuterol patch, HN-078, in the treatment of childhood asthma. METHODS: Single applications of HN-078 were applied transdermally in six children with asthma who had been admitted to a hospital. Subjects weighing less than 30 kg received 1 mg of tulobuterol while subjects weighing 30 kg or above received 2 mg on the chest for 24 hours. Serum tulobuterol levels and peak expiratory flow rate were determined before and after each application. RESULTS: Cmax of tulobuterol was determined to be 1.33 +/- 0.21 ng/mL, Tmax was 14.0 +/- 2.0 hours, and AUCO-t was 27.1 +/- 4.2 ng.hr/mL. These pharmacokinetic parameters per body surface area of children were nearly equivalent to those of adults obtained in other studies. Peak expiratory flow rate values obtained after application of HN-078 significantly increased in comparison to those obtained before application. No significant changes were observed in pulse rate or blood pressure, and no side effects were found with regard to the subjective symptoms and skin conditions. CONCLUSIONS: These results suggest that the patch formulation of tulobuterol, HN-078, will be very useful for the treatment of pediatric asthma. It is especially significant that no side effects were observed.