Effect of molecular weight on brittle‐to‐ductile transition temperature of polyetherimide

The fracture and yield strength of polyetherimide was evaluated over a temperature range of 23 to 140°C for materials with number‐average (M_n) and weight‐average molecular weight (M_w) ranging from 15.6 to 22.8 and 36.6 to 52.3 kg/mol, respectively. The brittle‐to‐ductile transition temperature, where an equal probability exists that an impact will result in a brittle or ductile failure, was determined by evaluating the temperature at which fracture and yield strength are equal. The transition temperature decreased from 155 to 60°C with increasing molecular weight and provided a measure of relative ductility between material samples. As a case study, the practical impact strength of an injection‐molded food service tray was determined at 20°C and correlated with fracture strength as a function of molecular weight. © 2004 Wiley Periodicals, Inc. J Appl Polym Sci 92: 1666–1671, 2004     

Na+,K(+)-ATPase phosphorylation in the choroid plexus: synergistic regulation by serotonin/protein kinase C and isoproterenol/cAMP-PK/PP-1 pathways

BACKGROUND: The ion pump Na+,K(+)-ATPase is responsible for the secretion of cerebrospinal fluid from the choroid plexus. In this tissue, the activity of Na+,K(+)-ATPase is inhibited by serotonin via stimulation of protein kinase C-catalyzed phosphorylation. The choroid plexus is highly enriched in two phosphoproteins which act as regulators of protein phosphatase-1 activity, DARPP-32 and inhibitor-1. Phosphorylation catalyzed by cAMP-dependent protein kinase on a single threonyl residue converts DARPP-32 and inhibitor-1 into potent inhibitors of protein phosphatase-1. Previous work has shown that in the choroid plexus, phosphorylation of DARPP-32 and I-1 is enhanced by isoproterenol and other agents that activate cAMP-PK. We have now examined the possible involvement of the cAMP-PK/protein phosphatase-1 pathway in the regulation of Na+,K(+)-ATPase. MATERIALS AND METHODS: The state of phosphorylation of Na+,K(+)-ATPase was measured by determining the amount of radioactivity incorporated into the ion pump following immunoprecipitation from 32P-prelabeled choroid plexuses incubated with various drugs (see below). Two-dimensional phosphopeptide mapping was employed to identify the protein kinase involved in the phosphorylation of Na+,K(+)-ATPase. RESULTS: The serotonin-mediated increase in Na+,K(+)-ATPase phosphorylation is potentiated by okadaic acid, an inhibitor of protein phosphatases-1 and -2A, as well as by forskolin or the beta-adrenergic agonist, isoproterenol, activators of cAMP-dependent protein kinase. Two-dimensional phosphopeptide maps suggest that this potentiating action occurs at the level of a protein kinase C phosphorylation site. Forskolin and isoproterenol also stimulate the phosphorylation of DARPP-32 and protein phosphatase inhibitor-1, which in their phosphorylated form are potent inhibitors of protein phosphatase-1. CONCLUSIONS: The results presented here support a model in which okadaic acid, forskolin, and isoproterenol achieve their synergistic effects with serotonin through phosphorylation of DARPP-32 and inhibitor-1, inhibition of protein phosphatase-1, and a reduction of dephosphorylation of Na+,K(+)-ATPase at a protein kinase C phosphorylation site.     

Sequential peritoneal lavage and early diagnosis of colon perforation

In order to determine the usefulness of the lavage white blood cell count as a diagnostic indicator of occult colon perforation, we subjected 20 mongrel dogs to sham laparotomy, isolated liver stab wound, or closed colon perforation and then performed sequential peritoneal lavage. The lavage white cell count rose steadily in those animals with colon perforations, and 100% of these animals had a positive lavage at six hours post-injury. After correction for the white cells shed intraperitoneally secondary to hemorrhage, the lavage white count of animals with liver wounds and of those with sham laparotomy failed to rise significantly during the period of observation. Gram stain of the lavage effluent as well as lavage amylase determination were not of diagnostic value. For solid viscus injury, lavage white cell count rises in proportion to organ hemorrhage; in hollow viscus perforation, lavage white cell count rises disproportionately and is higher than one would expect on the basis of the red cell count. We believe sequential peritoneal lavage white cell counts are of value in the diagnosis of occult colon perforation.     

Specific binding of a designed pyrrolidine abasic site analog to multiple DNA glycosylases

In the base excision DNA repair pathway, DNA glycosylases recognize damaged bases in DNA and catalyze their excision through hydrolysis of the N-glycosidic bond. Attempts to understand the structural basis for DNA damage recognition by DNA glycosylases have been hampered by the short-lived association of these enzymes with their DNA substrates. To overcome this problem, we have employed an approach involving the design and synthesis of inhibitors that form stable complexes with DNA glycosylases, which can then be studied biochemically and structurally. We have previously reported that double-stranded DNA containing a pyrrolidine abasic site analog (PYR) forms an extremely stable complex with the DNA glycosylase AlkA and potently inhibits the reaction catalyzed by the enzyme (Sch?rer, O. D., Ortholand, J.-Y., Ganesan, A., Ezaz-Nikpay, K., and Verdine, G. L. (1995) J. Am. Chem. Soc. 117, 6623-6624). Here we investigate the interaction of this inhibitor with a variety of additional DNA glycosylases. With the exception of uracil DNA glycosylase all the glycosylases tested bind specifically to PYR-containing oligonucleotides. By comparing the interaction of DNA glycosylases with PYR and the structurally related tetrahydrofuran abasic site analog, we assess the importance of the positively charged ammonium group of the pyrrolidine in binding to the active site of these enzymes. Such a general inhibitor of DNA glycosyases provides a valuable tool to study stable complexes of these enzymes bound to substrate-like molecules.     

Inhibition of neuronal Na+ channels by antidepressant drugs

Although tricyclic antidepressant (TCA) blockade of cardiac Na+ channels is appreciated, actions on neuronal Na+ channels are less clear. Therefore, the effects of TCAs (amitriptyline, doxepin and desipramine) as well as trazdone and fluoxetine on voltage-gated Na+ current (INa) were examined in bovine adrenal chromaffin cells using the whole-cell patch-clamp method. Amitriptyline produced concentration-dependent depression of peak INa evoked from a holding potential of -80 mV with KD value of 20.2 microM and a Hill coefficient of 1.2. Although 20 microM amitriptyline induced no change in the rate or voltage dependence of INa activation, steady-state inactivation demonstrated a 15-mV hyperpolarizing shift. Similar results were observed for doxepin and desipramine. This shift in steady-state inactivation was associated with a slowed rate of recovery from the inactivated state. Contrasting results were observed with the atypical antidepressants: while 20 microM fluoxetine depressed peak INa by 61% and caused a 7-mV hyperpolarizing shift in steady-state inactivation, 100 microM trazodone decreased peak INa by only 19% and caused only a 3-mV shift. Although the magnitude of fluoxetine effects was similar to those of the TCAs, the onset of fluoxetine effects was substantially slower than for amitriptyline. In voltage-clamp and current-clamp measurements from neonatal rat dorsal root ganglion neurons, 20 microM amitriptyline decreased INa by 52% and depressed action potential dynamics consistent with enhanced Na+ channel inactivation. The effects of the TCAs on INa are similar to local anesthetic behavior and could contribute to certain analgesic actions.     

Evaluation of four methods for rapid identification of Staphylococcus aureus from blood cultures

The identification of Staphylococcus aureus directly from blood cultures is clinically relevant, but it requires a test that is both rapid and reliable. Previously, biochemical, immunological, tube coagulase, and thermostable-endonuclease methods have shown variable sensitivity and specificity. Testing directly from blood culture broth has not been described for the latex kit Staphaurex Plus (Murex Diagnostics Ltd.), and the modified conventional tests have not been used with the newer, continuously monitored blood culture systems. In addition, the commercial RAPIDEC staph kit (bioMerieux Vitek, Inc.) has been used to detect S. aureus directly from the Vital blood culture system (bioMerieux, Marcy l'Etoile, France), but its performance has not been evaluated with other continuously monitored systems. A total of 201 clinical blood cultures (BACTEC 9240 culture system; Johnston Laboratories, Inc.) in which a Gram stain showed gram-positive cocci resembling staphylococci were evaluated prospectively. The Staphaurex Plus kit, the tube coagulase test, the thermostable-endonuclease test, and the RAPIDEC staph kit were compared. The sensitivities were 23, 92, 85, and 98% and the specificities were 99, 100, 93, and 100%, respectively. The RAPIDEC staph kit was the most reliable test, with a diagnostic accuracy comparable to that of the best published results for any of the rapid tests. However, it was the most expensive of the tests and relatively labor-intensive. The tube coagulase test was also sensitive, the simplest to perform, and inexpensive.     

Development of a highly efficient purification process for recombinant adenoviral vectors for oral gene delivery

This study compared catalytic and immunochemical properties of drug metabolizing phase I and II enzyme systems in houbara bustard (Chlamydotis undulata) liver and kidney and rat liver. P450 content in bustard liver (0.34 +/- 0.03 nmol mg-1 protein) was 50% lower than that of rat liver (0.70 +/- 0.02 nmol mg-1 protein). With the exception of aniline hydroxylase activity, monooxygenase activities using aminopyrine, ethoxyresorufin and ethoxycoumarin as substrates were all significantly lower than corresponding rat liver enzymes. As found in mammalian systems the P450 activities in the bird liver were higher than in the kidney. Immunohistochemical analysis of microsomes using antibodies to rat hepatic P450 demonstrated that bustard liver and kidney express P4502C11 homologous protein; no appreciable cross-reactivity was observed in bustards using antibodies to P4502E1, 1A1 or 1A2 isoenzymes. Glutathione content and glutathione S-transferase (GST) activity in bustard liver were comparable with those of rat liver. GST activity in the kidney was 65% lower than the liver. Western blotting of liver and kidney cytosol with human GST isoenzyme-specific antibodies revealed that the expression of alpha-class of antibodies exceeds mu in the bustard. In contrast, the pi-class of GST was not detected in the bustard liver. This data demonstrates that hepatic and renal microsomes from the bustard have multiple forms of phase I and phase II enzymes. The multiplicity and tissue specific expression of xenobiotic metabolizing enzymes in bustards may play a significant role in determining the pharmacokinetics of drugs and susceptibility of the birds to various environmental pollutants and toxic insults.     

Pulsed dye laser lithotripsy for treatment of urolithiasis in two geldings

Transendoscopic pulsed dye laser lithotripsy was effective in the treatment of calcium carbonate urothlithiasis in 2 adult geldings. Perineal urethrotomy provided convenient access for standing transendoscopic lithotripsy and evacuation of a large cystic calculus in one gelding. In the second horse, an obstructive urethral calculus was fragmented and removed by a transurethroscopic approach. Pulsed dye laser lithotriptor is effective in fragmentation of the most common form (calcium carbonate) of uroliths in horses and may be performed in standing horses with reduced surgical invasiveness and trauma to the urinary tract, compared with conventional approaches. The principle disadvantages include cost of the procedure, which is comparable to laparocystotomy, and the time delay required to make arrangements for use of the pulsed dye laser lithotriptor.