慢性化脓性中耳炎与对侧迟发性膜迷路积水

梅尼埃病 (ｍ啨ｎｉ埁ｒｅ’ｓｄｉｓｅａｓｅ,ＭＤ)的治疗目前仍然是耳科临床上的难题之一 ,主要是因为其病因尚未完全弄清。近年来的耳免疫学研究显示免疫病理学因素在ＭＤ的发病中可能起着重要的作用 ,部分患者的发病可能与内耳自身免疫性病变有关。最近 ,我们对 7例 (耳 )慢性化脓性中耳乳突炎(ＣＳＯＭ)伴对侧ＭＤ患者行免疫学检查并与 1 0例(耳 )ＣＳＯＭ不伴ＭＤ患者进行对照 (对照组 ) ,借以探讨ＭＤ发病的免疫学因素。报告如下。1　资料与方法1 .1 　 临床资料ＣＳＯＭ伴ＭＤ组 :7例ＣＳＯＭ伴对侧ＭＤ患者 ,男 2例 ,女 5例 ;平均…     

Effects of agonist and phorbol ester on adrenergic receptors of DDT1 MF-2 cells

The effects of the adrenergic agonist epinephrine (EPI) and of phorbol 12-myristate 13-acetate (PMA) on the regulation of alpha-1 adrenergic receptors (AAR) and beta adrenergic receptors (BAR) were compared in DDT1 MF-2 cells grown in suspension culture. Pretreatment of cells with 10 microM EPI for 30 min at 37 degrees C resulted in homologous desensitization of BAR-coupled adenylate cyclase activity assayed in membranes and induced internalization or sequestration of BAR. Pretreatment of cells with PMA did not alter BAR-coupled adenylate cyclase activity or induce internalization of BAR. EPI pretreatment caused a 50% decrease in the subsequent ability of EPI to stimulate AAR-mediated incorporation of 32P into phosphatidylinositol, whereas PMA pretreatment inhibited incorporation by 95%. Neither EPI nor PMA induced the internalization of AAR. Neither EPI nor PMA altered agonist binding properties of AAR in short-time competition binding assays on intact cells, indicating that pretreatment of cells with these agents does not alter the affinity of AAR for agonist. In control cells, agonists converted AAR from a form exhibiting predominantly high affinity for agonists, detected in short-time assays, to a form, exhibiting low apparent affinity for agonist during the course of equilibrium competition binding assays. PMA pretreatment increased the extent of this subsequent agonist-induced conversion to the low affinity form. These results indicate that PMA can mimic agonist-induced desensitization of AAR, but not BAR, and that the desensitization of AAR-coupled phosphatidylinositol turnover induced by EPI and by PMA is not due to altered receptor affinity for EPI or due to receptor internalization.     

The N terminus of the human alpha1D-adrenergic receptor prevents cell surface expression

We previously reported that truncation of the N-terminal 79 amino acids of alpha(1D)-adrenoceptors (Delta(1-79)alpha(1D)-ARs) greatly increases binding site density. In this study, we determined whether this effect was associated with changes in alpha(1D)-AR subcellular localization. Confocal imaging of green fluorescent protein (GFP)-tagged receptors and sucrose density gradient fractionation suggested that full-length alpha(1D)-ARs were found primarily in intracellular compartments, whereas Delta(1-79)alpha(1D)-ARs were translocated to the plasma membrane. This resulted in a 3- to 4-fold increase in intrinsic activity for stimulation of inositol phosphate formation by norepinephrine. We determined whether this effect was transplantable by creating N-terminal chimeras of alpha(1)-ARs containing the body of one subtype and the N terminus of another (alpha(1A)NT-D, alpha(1B)NT-D, alpha(1D)NT-A, and alpha(1D)NT-B). When expressed in human embryonic kidney 293 cells, radioligand binding revealed that binding densities of alpha(1A)-or alpha(1B)-ARs containing the alpha(1D)-N terminus decreased by 86 to 93%, whereas substitution of alpha(1A)- or alpha(1B)-N termini increased alpha(1D)-AR binding site density by 2- to 3-fold. Confocal microscopy showed that GFP-tagged alpha(1D)NT-B-ARs were found only on the cell surface, whereas GFP-tagged alpha(1B)NT-D-ARs were completely intracellular. Radioligand binding and confocal imaging of GFP-tagged alpha(1D)- and Delta(1-79)alpha(1D)-ARs expressed in rat aortic smooth muscle cells produced similar results, suggesting these effects are generalizable to cell types that endogenously express alpha(1D)-ARs. These findings demonstrate that the N-terminal region of alpha(1D)-ARs contain a transplantable signal that is critical for regulating formation of functional bindings, through regulating cellular localization.     

Nuclear RNA sequences coding for alpha and beta globins in erythroid cells: evidence for multiple intermediate molecules

The poly (A)-containing nuclear RNA from dimethylsulfoxide-induced Friend leukemia cells was fractionated by acrylamide gel electrophoresis in denaturing conditions and analyzed for alpha and beta globin RNA sequences. The results indicate that nuclear RNA contains one species of large-size RNA (0.6 X 10(6) daltons), which is the putative precursor for beta globin mRNA only. In addition, it was shown by electrophoretic analysis that the complex of RNA molecules not resolved by sucrose gradient centrifugation (11S) comprises sequences of decreasing size (0.34, 0.28, and 0.26 X 10(6) daltons), which might be the precursors of alpha and beta globin mRNA.     

Identification of spontaneous feline idiopathic pulmonary fibrosis: morphology and ultrastructural evidence for a type II pneumocyte defect

STUDY OBJECTIVES: Idiopathic pulmonary fibrosis (IPF) is a poorly understood chronic respiratory disease of humans, which has no correlate in other animals. Understanding the role that inflammation, alveolar epithelial cells, and myofibroblasts play in the progression of the disease is controversial, and hampered by the lack of an animal model. We have identified spontaneous IPF in domestic cats and hypothesized that this newly identified disease shares the pathology of human IPF; further, this work provides data suggesting that the disease is related to a defect in type II pneumocyte biology. SETTING AND SUBJECTS: Chronic respiratory disease with pathology consistent with usual interstitial pneumonia (UIP) spontaneously developed in 16 domestic cats. RESULTS: The histopathology of feline IPF consisted of the following: (1) interstitial fibrosis with fibroblast/myofibroblast foci, (2) honeycombing with alveolar epithelial metaplasia and type II pneumocyte hyperplasia, and (3) alveolar interstitial smooth-muscle metaplasia. Interstitial inflammation was not a prominent feature of the disease. alpha-Smooth muscle actin-positive myofibroblasts were prominent in myofibroblast foci, beneath honeycomb and hyperplastic epithelium, and in alveolar septa away from the remodeling. Feline IPF type II pneumocyte ultrastructure is similar to a heritable form of human IPF, with abnormal cytoplasmic lamellar body-like inclusions. CONCLUSIONS: We conclude the following: (1) chronic respiratory disease with clinical and pathology features of UIP/IPF occurs in the domestic cat; (2) as in human IPF, the type II pneumocyte and myofibroblasts are important cellular constituents of feline IPF; and (3) type II cell ultrastructure suggests feline IPF is a defect in the type II pneumocyte.     

Panoramic view of a superfamily of phosphatases through substrate profiling

Large-scale activity profiling of enzyme superfamilies provides information about cellular functions as well as the intrinsic binding capabilities of conserved folds. Herein, the functional space of the ubiquitous haloalkanoate dehalogenase superfamily (HADSF) was revealed by screening a customized substrate library against >200 enzymes from representative prokaryotic species, enabling inferred annotation of ∼35% of the HADSF. An extremely high level of substrate ambiguity was revealed, with the majority of HADSF enzymes using more than five substrates. Substrate profiling allowed assignment of function to previously unannotated enzymes with known structure, uncovered potential new pathways, and identified iso-functional orthologs from evolutionarily distant taxonomic groups. Intriguingly, the HADSF subfamily having the least structural elaboration of the Rossmann fold catalytic domain was the most specific, consistent with the concept that domain insertions drive the evolution of new functions and that the broad specificity observed in HADSF may be a relic of this process.Since the first genomes were sequenced, there has been an exponential increase in the number of protein sequences deposited into databases worldwide. At the time of this writing the UniProtKB/TrEMBL database contains over 32 million protein sequences. Although this increase in sequence data has dramatically enhanced our understanding of the genomic organization of organisms, as the number of protein sequences grows, the proportion of firm functional assignments diminishes. Traditionally, methods of functional annotation involve comparing sequence identity between experimentally characterized proteins and newly sequenced ones, typically via BLAST (1). In cases where significant sequence similarity cannot be ascertained, proteins are annotated as “hypothetical” or “putative.” Moreover, the decrease in sequence identity leads to an increased uncertainty in functional assignment, especially as the phylogenetic distance between organisms grows, limiting iso-functional ortholog discovery.As the number of newly sequenced genomes grows larger, more protein sequences are likely to be misannotated, oftentimes resulting in the propagation of incorrect functional annotation across newly identified sequences. To tackle the problem of unannotated or misannotated proteins, newer methods for computational assignment have been created with varying degrees of success (2). Although these methods outperform historical methods, continued improvement is necessary to ensure accurate annotation of function (2). A greater swath of functional space can be covered by screening substrates in a high-throughput manner on multiple enzymes from a family (3, 4). Family-wide substrate profiling offers a data-rich resource. The use of sparse screening of sequence space and a diversified library permits the determination of substrate specificity profiles to provide a family-wide view of the range of substrates and insight into the structure of the prototypical substrate. Where structures are available, correlation between substrate range and structural determinants of specificity can be achieved. In addition, the approach has utility in genomic annotation (inferred function), iso-functional ortholog assignment, and the assignment of in vitro substrate profiles to orphaned PDB entries (enzymes with structure but no function, or SNFs). Here we report the application of in vitro high-throughput functional screening of metabolites and related compounds at the superfamily level. We use as an example prokaryotic members of the haloalkanoic acid dehalogenase superfamily (HADSF), a diverse superfamily of enzymes (5) that catalyze a wide range of reactions involving the formation of a covalent intermediate with an active-site aspartate. Reactions catalyzed by this superfamily include dehalogenation (6) as well as Mg²⁺-dependent phosphoryltransfer, although the vast majority (∼99%) are phosphotransferases (7). Members of the HADSF share a Rossmannoid fold “core” domain that contains the phosphoryl transfer site (8, 9) and a “cap” domain that provides substrate specificity determinants (10). There are three major types of caps in the HADSF (C0, C1, and C2A/C2B; see SI Appendix, Fig. S1) based on size, position of insert within the Rossmann fold, and overall topology (7). At the time of writing, the HADSF is known to comprise over 120,000 members across the three domains of life with at most 3% associated with an EC identifier (11).In this study, the functional space of the HADSF was sampled by screening a customized substrate library of 167 compounds against over 200 enzymes from numerous prokaryotic species. The study revealed that a large number of family members show a broad substrate range, with the majority of the HADSF enzymes reacting with five or more substrates. Thus, widespread promiscuity is not incompatible with participation in cell metabolism and may be advantageous to the evolution of new enzyme activities. The activity profiling when applied to putative iso-functional orthologs allowed us to infer annotation for ∼35% of the HADSF. Intriguingly, the HADSF subfamily with the least structural elaboration of the core catalytic Rossmann fold was the most specific with respect to substrate range and number, implying that domain insertions drive the evolution of new functions and that the broad specificity observed in the HADSF may be a relic of this process.     

Modified LSA2-L2 treatment in 53 children with E-rosette-positive T- cell leukemia: results and prognostic factors (a Pediatric Oncology Group Study)

In an attempt to improve the poor outlook for children with T-cell leukemia (T-ALL), the Southwest Oncology Group, Pediatric Division, used a modified LSA2-L2 multidrug regimen to treat 53 patients with E- rosette-positive T-ALL. This regimen was chosen because of its demonstrated efficacy in T-cell (mediastinal) non-Hodgkin's lymphoma. Complete remission (CR) rate was 88%. Range of follow-up for those patients remaining in CR is 24-49 mo (median 39 mo). Life table analysis estimates that 40% (SE 8.3%) of all patients who started induction therapy will remain failure-free at 3 yr. For patients achieving CR, 46% (SE 9%) are projected to remain in both marrow and extramedullary CR at 3 yr. Median failure-free duration was 13 mo, but only 1 patient has relapsed beyond 16 mo. Twenty-nine percent of initial relapses were isolated CNS relapses. The following presenting factors did not relate significantly to outcome: hemoglobin, platelet count, uric acid, race, and mediastinal mass. Age greater than 10 yr was a poor prognosis indicator only in the less than 50,000/microliter WBC group. Sex was not a significant factor after adjusting for WBC. WBC was the most important prognostic factor: 19% (SE 8%) of patients with WBC greater than 50,000/microliter are projected to remain failure- free at 3 yr as compared to 67% (SE 11%) of patients with WBC less than 50,000/microliter. Although the overall results are better than those previously reported for pediatric patients with T-ALL, the long-term failure-free rate remains low for patients presenting with greater than 50,000/microliter WBC.     

Changing radiation dose from diagnostic computed tomography examinations in Saskatchewan

Purpose

Follow-up study to observe if provincial mean effective radiation dose for head, chest, and abdomen-pelvis (AP) computed tomographies (CTs) remained stable or changed since the initial 2006 survey.

Methods

Data were collected in July 2008 from Saskatchewan's 13 diagnostic CT scanners of 3358 CT examinations. These data included the number of scan phases and projected dose length product (DLP). Technologists compared projected DLP with 2006 reference data before scanning. Projected DLP was converted to effective dose (ED) for each head, chest, and AP CT. The total dose that the patients received with scans of multiple body parts at the same visit also was determined.

Results

The mean (± SD) provincial ED was 3.4 ± 1.6 mSv for 1023 head scans (2.7 ± 1.6 mSv in 2006), 9.6 ± 4.8 mSv for 588 chest scans (11.3 ± 8.9 mSv in 2006), and 16.1 ± 9.9 mSv for 983 AP scans (15.5 ± 10.0 mSv in 2006). Single-phase multidetector row CT ED decreased by 31% for chest scans (9.5 ± 3.9 mSv vs 13.7 ± 9.7 mSv in 2006) and 17% for AP scans (13.9 ± 6.0 mSv vs 16.8 ± 10.6 mSv in 2006) and increased by 19% for head scans (3.2 ± 1.2 mSv vs 2.7 ± 1.5 mSv in 2006). The total patient dose was highest (33.8 ± 10.1 mSv) for the 20 patients who received head, neck, chest, and AP scans during a single visit. Because of increased utilisation and the increased CT head dose, Saskatchewan per capital radiation dose from CT increased by 21% between 2006 and 2008 (1.14 vs 1.38 mSv/person per year).

Conclusion

Significant dose and variation reduction was seen for single-phase CT chest and AP examinations between 2006 and 2008, whereas CT head dose increased over the same interval. These changes, combined with increased utilisation, resulted in per capita increase in radiation dose from CT between the 2 studies.     

Pulmonary elastase activity in response to Streptococcus pneumoniae and Pseudomonas aeruginosa

Elastase activity generated during lung defense against aerobic bacteria was studied in an animal model. Bronchoalveolar lavage (BAL) fluid from hamsters inoculated with bacteria was assayed for elastase activity at 0, 2, 4, 6, and 8 h after inoculation using a synthetic substrate of elastase, succinyl-trialanine-nitroanilide (SLAPN). Streptococcus pneumoniae type 25 inoculation led to a peak elastase activity of 0.72 +/- 0.27 X 10(-3) units, not significantly different from baseline (0.41 +/- 0.08 X 10(-3) units) or saline control (0.33 +/- 0.18 X 10(-3) units). In contrast, inoculation with Pseudomonas aeruginosa strain PAO-1 (a species known to produce elastase as well as other virulence factors) produced peak elastase activity of 3.0 +/- 1.2 X 10(-3) units in BAL fluid, significantly higher than either pneumococcus type 25 or saline control (p less than 0.025). Inoculation with Pseudomonas aeruginosa strain E-64, an isogenic mutant of PAO-1 that produces a nonfunctional elastase, led to peak levels similar to the PAO-1 strain, suggesting that the presence of bacterial elastase was not the primary factor in BAL fluid elastase activity. Total numbers of granulocytes in BAL fluid from pneumococcus-inoculated animals (144 +/- 31 X 10(6] was significantly higher (p less than 0.05) than from either the PAO-1 (74 +/- 31 X 10(6] or E-64 (99 +/- 27 X 10(6] strains of Pseudomonas, Use of selective enzyme inhibitors of elastase, diisopropyl fluorophosphate and disodium ethylenediaminetetraacetate, implied that the majority of elastase activity in BAL fluid was due to a serine protease, of which granulocyte elastase is the primary source.(ABSTRACT TRUNCATED AT 250 WORDS)