Characterization of a Chinese hamster ovary cell line with acquired resistance to the bisdioxopiperazine dexrazoxane (ICRF-187) catalytic inhibitor of topoisomerase II

A Chinese hamster ovary (CHO) cell line highly resistant to the non-cleavable complex-forming topoisomerase II inhibitor dexrazoxane (ICRF-187, Zinecard) was selected. The resistant cell line (DZR) was 1500-fold resistant (IC50 = 2800 vs 1.8 microM) to continuous dexrazoxane exposure. DZR cells were also cross-resistant (8- to 500-fold) to other bisdioxopiperazines (ICRF-193, ICRF-154, and ICRF-186), and somewhat cross-resistant (4- to 14-fold) to anthracyclines (daunorubicin, doxorubicin, epirubicin, and idarubicin) and etoposide (8.5-fold), but not to the other non-cleavable complex-forming topoisomerase II inhibitors suramin and merbarone. The cytotoxicity of dexrazoxane to both cell lines was unchanged in the presence of the membrane-active agent verapamil. DZR cells were 9-fold resistant to dexrazoxane-mediated inhibition of topoisomerase II DNA decatenation activity compared with CHO cells (IC50 = 400 vs 45 microM), but were only 1.4-fold (IC50 = 110 vs 83 microM) resistant to etoposide. DZR cells contained one-half the level of topoisomerase II protein compared with parental CHO cells. However, the specific activity for decatenation using nuclear extract topoisomerase II was unchanged. Etoposide (100 microM)-induced topoisomerase II-DNA complexes in DZR cells and isolated nuclei were similarly one-half the level found in CHO cells and in isolated nuclei. However, the ability of 500 microM dexrazoxane to inhibit etoposide (100 microM)-induced topoisomerase II-DNA covalent complexes was reduced 4- to 6-fold in both DZR cells and nuclei compared with CHO cells and nuclei. In contrast, there was no differential ability of aclarubicin or merbarone to inhibit etoposide-induced topoisomerase II-DNA complexes in CHO compared with DZR cells and isolated nuclei. It was concluded that the DZR cell line acquired its resistance to dexrazoxane mainly through an alteration in the topoisomerase II target.     

Nonsurgical and surgical management of infants and children with obstructive sleep apnea syndrome

Obstructive sleep apnea (OSA) syndrome in infants and children is one of the more common problems treated by the otorhinolaryngologist-head and neck surgeon. The authors discuss diagnostic studies, nonsurgical and surgical therapies, and perioperative care, including indications for inpatient stay following surgery. Most children are successfully treated for obstructive breathing by adenotonsillectomy. An awareness of the wide range of treatment modalities, with attention to careful postoperative care, should allow for successful management of nearly all children with OSA.     

Antigen presentation and T cell development in H2-M-deficient mice

HLA-DM (DM) facilitates peptide loading of major histocompatibility complex class II molecules in human cell lines. Mice lacking functional H2-M, the mouse equivalent of DM, have normal amounts of class II molecules at the cell surface, but most of these are associated with invariant chain-derived CLIP peptides. These mice contain large numbers of CD4+ T cells, which is indicative of positive selection in the thymus. Their CD4+ cells were unresponsive to self H2-M-deficient antigen-presenting cells (APCs) but were hyperreactive to wild-type APCs. H2-M-deficient APCs failed to elicit proliferative responses from wild-type T cells.     

Gestational diabetes: antepartum characteristics that predict postpartum glucose intolerance and type 2 diabetes in Latino women

We examined antepartum clinical characteristics along with measures of glucose tolerance, insulin sensitivity, pancreatic beta-cell function, and body composition in Latino women with gestational diabetes mellitus (GDM) for their ability to predict type 2 diabetes or impaired glucose tolerance (IGT) within 6 months after delivery. A total of 122 islet cell antibody-negative women underwent oral and intravenous glucose tolerance tests (OGTT; IVGTT), hyperinsulinemic-euglycemic clamps, and measurement of body fat between 29 and 36 weeks' gestation and returned between 1 and 6 months postpartum for a 75-g OGTT. Logistic regression analysis was used to examine the relationship between antepartum variables and glucose tolerance status postpartum. At postpartum testing, 40% of the cohort had normal glucose tolerance, 50% had IGT, and 10% had diabetes by American Diabetes Association criteria. Independent antepartum predictors of postpartum diabetes were the 30-min incremental insulin:glucose ratio during a 75-g OGTT (P = 0.0002) and the total area under the diagnostic 100-g glucose tolerance curve (P = 0.003). Independent predictors of postpartum IGT were a low first-phase IVGTT insulin response (P = 0.0001), a diagnosis of GDM before 22 weeks' gestation (P = 0.003), and weight gain between prepregnancy and the postpartum examination (P = 0.03). All subjects had low insulin sensitivity during late pregnancy, but neither glucose clamp nor minimal model measures of insulin sensitivity in the 3rd trimester were associated with the risk of IGT or diabetes within 6 months' postpartum. These results highlight the importance of pancreatic beta-cell dysfunction, detectable under conditions of marked insulin resistance in late pregnancy, to predict abnormalities of glucose tolerance soon after delivery in pregnancies complicated by GDM. Moreover, the association of postpartum IGT with weight gain and an early gestational age at diagnosis of GDM suggests a role for chronic insulin resistance in mediating hyperglycemia outside the 3rd trimester in women with such a beta-cell defect.     

Application of a new method for analysis of exhaled gas in critically ill patients

OBJECTIVE: Application of a new method for analysis of exhaled gas in critically ill patients. DESIGN: Open study. SETTING: Surgical intensive care unit of an university hospital. PATIENTS: Thirty-seven consecutive, critically ill, mechanically ventilated patients. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: Chemical analysis of the patient's exhaled gas was based upon substance adsorption and concentration onto activated charcoal, microwave desorption and gas chromatographic separation. Patients with acute respiratory distress syndrome (ARDS) exhaled less isoprene than those without ARDS [9.8 (8.2-21.6) vs 21.8 (13.9-41.4) nmol/m2 per min [median (95% confidence interval)], p = 0.04]. In patients who developed pulmonary infection, pentane elimination increased from 0.4 (0.0-5.4) to 2.7 (0.6-6.1, p = 0.05) nmol/m2 per min and isoprene elimination decreased from 5.2 (0-33) to 5.0 (0-17, p = 0.05) nmol/m2 per min, resulting in a significant increase in pentane/isoprene ratio from 0.1 (0-0.3) to 0.4 (0-15, p = 0.007) when compared to patients without pulmonary infection. CONCLUSIONS: The new method allows quantitative analysis of human gas samples with low substance concentrations and is well suited for clinical studies which involve the investigation of metabolic processes in the lung and the body.     

Early evolutionary origin of the neurotrophin receptor family

Neurotrophins and their Trk receptors play a crucial role in the development and maintenance of the vertebrate nervous system, but to date no component of this signalling system has been found in invertebrates. We describe a molluscan Trk receptor, designated Ltrk, from the snail Lymnaea stagnalis. The full-length sequence of Ltrk reveals most of the characteristics typical of Trk receptors, including highly conserved transmembrane and intracellular tyrosine kinase domains, and a typical extracellular domain of leucine-rich motifs flanked by cysteine clusters. In addition, Ltrk has a unique N-terminal extension and lacks immunoglobulin-like domains. Ltrk is expressed during development in a stage-specific manner, and also in the adult, where its expression is confined to the central nervous system and its associated endocrine tissues. Ltrk has the highest sequence identity with the TrkC mammalian receptor and, when exogenously expressed in fibroblasts or COS cells, binds human NT-3, but not NGF or BDNF, with an affinity of 2.5 nM. These findings support an early evolutionary origin of the Trk family as neuronal receptor tyrosine kinases and suggest that Trk signalling mechanisms may be highly conserved between vertebrates and invertebrates.     

Localization of glutathione S-transferases alpha and pi in human embryonic tissues at 8 weeks gestational age

Glutathione S-transferases (GST) are a family of enzymes involved in the detoxification of xenobiotics. In humans, GST are divided into four different classes, alpha, mu, pi and theta, with partly overlapping substrate specificity and a tissue-specific expression pattern. We studied the cellular distribution of GST alpha and pi in a variety of human embryonic organs obtained from an extra-uterine monozygotic twin pregnancy at 8 weeks' gestational age. Tissues were fixed in 4% paraformaldehyde and embedded in paraffin. Three 4 microm thick sections were used, one for routine haematein and eosin staining, the others for immunohistochemical determination using monoclonal and polyclonal antibodies against GST alpha and pi, respectively. Both GST alpha and pi were present in hepatocytes, gastrointestinal epithelium, adrenal gland medulla, and tela chorioidea in the telencephalon. GST pi, but not alpha, was found in the epithelium of pancreatic and pulmonary glands, trachea, nephrons and urinary collecting ducts, as well as in the pia mater of the telencephalon and in developing nerve tissue in the gastrointestinal muscularis mucosae. In summary, we have demonstrated that immunoreactive protein for both GST alpha and pi is expressed in the human embryo at 8 weeks' gestational age. The early expression of GST alpha and pi in the epithelia of the urinary and digestive tracts and the respiratory system supports the importance of GST in the detoxification of potentially toxic or carcinogenic compounds. Our results suggest that the embryo itself is capable of detoxifying noxious compounds that are generated intracellularly or that cross the trophoblastic tissue.     

Computation of a location shift between two subsequent intra vascular ultrasound registrations by cross-correlation analysis of the lumen area functions

For longitudinal studies on atherosclerosis and restenosis after angioplasty by intravascular ultrasound (IVUS), it is essential that repeated studies are performed at exactly the same location along an arterial section. In human femoral arteries, lumen and plaque area functions of two subsequent IVUS pullback maneuvers were compared by cross-correlation analysis. In cross-correlation analysis of two functions with equal abscissa values, the data sets are repetitively correlated after incremental shifts of the two functions along the abscissa. This results in multiple correlation coefficients with a maximum at the relative position where the two functions show the closest match. In group A (12 patients), both pullbacks were performed after angioplasty and in group B (17 patients) one pullback was performed before angioplasty and the second immediately after angioplasty. In group A, cross-correlation showed a shift between lumen area functions of 5 mm in one patient and no shift in the other patients. Maximum correlation coefficients in group A ranged from 0.644 to 0.978. Four patients from group B showed shifts from 2.5 to 35 mm. Maximum coefficients were significantly smaller than in group A: 0.259-0.864 (p < 0.01). Plaque area functions in group B showed higher correlations (0.468-0.862, p = 0.034) and only two shifts. Cross-correlation of lumen and plaque area functions may be used to compute location shifts between two subsequent IVUS registrations and to correct such shifts.