Single 6-formylcytidine was introduced into a oligonucleotide duplex (23 mers) as a substitute for thymidine in the Myb binding sequence of 3'-TTGAC-5'. The modified duplex showed Tm of 67 degrees C, which was six degrees lower than the Tm of the native duplex. Binding affinity of the 23-mers to the Myb protein was estimated by electrophoretic mobility shift assays, and the binding was almost completely abolished.

Binding Sites↗

PubMedDate not supplied

DNA sequence recognition by NF kappa B p50 homodimer: strict and obscure recognition sites in the binding sequence.

5-Formyl- and 5-(formylmethyl)-2'-deoxyuridines are introduced into a kappa B site instead of thymidine(s) in order to understand target sequence specificity of NF kappa B. It was found that one thymidine in the kappa B site is particularly important for the sequence specific recognition by NF kappa B.

Base Sequence↗

PubMedDate not supplied

Development of a stopped flow reactor for the rapid, high sensitivity measurement of endotoxin concentration in dialysate.

Continuous measurement of endotoxin concentration in dialysate, using a separated endotoxin-specific limulus reagent, promises rapid measurement without the complex operating procedures of the limulus reagent. To achieve high sensitivity measurements in a short period of time, an improved system featuring stopped-flow operation was developed. To prevent dispersion of the limulus reagent and residence of reacting solution containing the limulus reagent in the system reactor, the circuit in the reactor was changed from a coil configuration to a straight line, and its length was reduced. An endotoxin test solution was supplied at 760 microl/min, into which 40 microl of limulus reagent was pulse-injected. Flow was stopped at the point where the test solution entered the reactor. After the completion of the reaction, the solution was passed through a spectrophotometer and the relationship between reaction time and absorbance was determined. Peak tailing was less than that obtained by the conventional technique, good correlation was obtained from the peak height, and a decrease in sensitivity caused by broadening of the peak was suppressed. The lower detection limit of dialysate was 100 endotoxin units (EU)/L at a reaction time of 20 minutes, and 60 EU/L at 30 minutes. Change from the monitoring system to stop-flow operation made high sensitivity monitoring of endotoxin concentration with a short reaction time possible.

Animals↗

PubMedDate not supplied

Utilization of a partially threaded Kirschner wire in the treatment of femoral neck fractures.

OBJECTIVE: This report introduces a new type of pin for treatment of femoral neck fractures. DESIGN: Retrospective analysis. METHODS: The pin used in this study has a tapered thread in its middle portion to facilitate fixation in the lateral femoral cortex, as well as a simple tip that allows percutaneous pinning without displacement of the reduced position. Between 1978 and 1993, we used our pins on 218 hips in 214 patients, all age sixty years or more. We were able to observe postoperative osteosynthesis in 176 hips (172 patients) during a mean follow-up period of thirty-two months; the incidence of osteosynthesis and local postoperative complications was also charted. RESULTS: Osteosynthesis was obtained in 153 hips (87 percent); for groups based on Garden's classification, results were as follows: twenty-four of twenty-four hips in type 1, fifteen of sixteen in type 2, sixty-three of seventy-one in type 3, and fifty-one of sixty-five in type 4. Some local postoperative complications were observed, including second fractures at the pin insertion point (three hips), pin penetration of the femoral head (twenty-four hips), and pin slippage (three hips). CONCLUSION: This pin is effective in the treatment of femoral neck fractures and has the added benefits of being minimally invasive and low in cost.

Aged↗

PubMedDate not supplied

In vivo effects of apoptosis in asthma examined by a murine model.

BACKGROUND: One of the characteristic features of bronchial asthma is the accumulation of various inflammatory cells, predominantly eosinophils, at the subepithelial region beneath the basement membrane of the airway. Apoptosis is a form of physiological cell death, through which the cellular contents including biologically active substances are kept in the cell membrane and are removed without their harmful effects. So, attempts were made to clarify whether the induction of apoptosis is beneficial in asthma by using a murine model with ovalbumin (OA) as responsible allergen. METHODS: A/J mice, which are genetically predisposed to be hyperresponsive to acetylcholine, were immunized with OA and alum, accompanied by OA inhalation for 2 weeks, during which some of the mice were also treated with either anti-Fas monoclonal antibody or sham control hamster IgG intranasally. Airway responsiveness to acetylcholine was then analyzed by measuring airway resistance with a body plethysmograph box. Apoptosis was assessed by propidium iodide and TUNEL staining. RESULTS: Inhalation of OA increased both airway responsiveness to acetylcholine and the number of cells, mostly eosinophils, infiltrated into the airway. Administration of anti-Fas antibody induced apoptosis in the infiltrated eosinophils and abolished augmentation of airway hyperresponsiveness caused by OA inhalation. CONCLUSION: Induction of apoptosis in proinflammatory cells including eosinophils at the airway may have a beneficial effect on suppressing airway hyperresponsiveness.

Acetylcholine↗

PubMedDate not supplied

Direct measurement of nitric oxide during experimental cardiopulmonary bypass.

We developed a system to measure nitric oxide (NO) concentration during cardiopulmonary bypass in anaesthetized pigs (n = 6). A T-shaped connector, attached to an NO sensor, was mounted in the extracorporeal circuit at two measuring sites: proximal to the membrane oxygenator (venous side) and distal to the arterial line filter (arterial side). After performing a preliminary validation study, we measured plasma NO concentration before and during total cardiopulmonary bypass circulation (non-pulsatile flow 1.5 l/min) and without pulmonary ventilation. After establishing bypass, PaO2 was 318 - 393 mmHg; when PaO2 was decreased to 80 - 100 mmHg, plasma NO concentration in the arterial circuit fell by 39.2 +/- 15.6 nM. There was no observable change in plasma NO concentration at the venous circuit. This new system could be useful in monitoring NO concentration during cardiac surgery with cardiopulmonary bypass, and for understanding the possible pathophysiological roles of hyper-nitric oxaemia in cardiopulmonary bypass-related cardiovascular complications.

Animals↗

PubMedDate not supplied

Development of a glucose sensor with on/off control of enzyme activity without the effects of protein adsorption.

The main factor mitigating against the realization of a hypodermically inserted glucose sensor for an artificial pancreas is the change in response current due to fibroblast adhesion and protein adsorption to the sensor surface. To overcome this problem, we have developed a method whereby the activity of glucose oxidase (GOD) fixed on the membrane of the sensor surface is switched on and off, and measurements are made during a transient state in which the glucose concentration gradient within the GOD membrane is small. Measuring in a transient state while GOD activity is being controlled, a correlation was observed between glucose concentration and response current in a phosphate buffer solution. Calibration curves of response current against glucose concentration in aqueous solutions of human serum albumin and in phosphate buffer solution were then compared using the transient method and a steady state method without control of GOD activity. In addition, glucose concentration was measured in bovine plasma for 480 min, and the time courses of the response currents for the transient and steady state measurements were compared. It was found that in both experiments the response current decreased greatly under steady state measurement as a result of protein adsorption, but during the transient measurement, response current was virtually unchanged. By measuring glucose concentration in the transient state while controlling GOD activity, it is possible to inhibit the effects of protein adsorption.

Adsorption↗