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PubMed2003

Kinetics and thermodynamics of nick sealing by T4 DNA ligase.

T4 DNA ligase is an Mg2+-dependent and ATP-dependent enzyme that seals DNA nicks in three steps: it covalently binds AMP, transadenylates the nick phosphate, and catalyses formation of the phosphodiester bond releasing AMP. In this kinetic study, we further detail the reaction mechanism, showing that the overall ligation reaction is a superimposition of two parallel processes: a 'processive' ligation, in which the enzyme transadenylates and seals the nick without dissociating from dsDNA, and a 'nonprocessive' ligation, in which the enzyme takes part in the abortive adenylation cycle (covalent binding of AMP, transadenylation of the nick, and dissociation). At low concentrations of ATP (<10 microM) and when the DNA nick is sealed with mismatching base pairs (e.g. five adjacent), this superimposition resolves into two kinetic phases, a burst ligation (approximately 0.2 min(-1)) and a subsequent slow ligation (approximately 2x10(-3) min(-1)). The relative rate and extent of each phase depend on the concentrations of ATP and Mg2+. The activation energies of self-adenylation (16.2 kcal.mol(-1)), transadenylation of the nick (0.9 kcal.mol(-1)), and nick-sealing (16.3-18.8 kcal.mol(-1)) were determined for several DNA substrates. The low activation energy of transadenylation implies that the transfer of AMP to the terminal DNA phosphate is a spontaneous reaction, and that the T4 DNA ligase-AMP complex is a high-energy intermediate. To summarize current findings in the DNA ligation field, we delineate a kinetic mechanism of T4 DNA ligase catalysis.

Adenosine Monophosphate↗
PubMed2002

The in vitro and in vivo ultrasonographic appearances of the angio-seal percutaneous closure device.

PURPOSE: To assess the ultrasound appearances of the Angio-Seal device in an animal model and in twenty patients following catheterization of the femoral artery. MATERIALS AND METHODS: Ten patients were scanned within 8 hours of their procedure (early group) and 10 at two to four days (delayed group) using Acuson Aspen and Sequoia scanners. Colour flow images were taken and Doppler spectral analysis was performed proximal proximal to, at and distal to the Angio-Seal device. Early and delayed in vitro images were taken in a water bath in which the Angio-Seal device was deployed across a normal porcine aorta for comparison. RESULT: In the ten early patients colour and Doppler drop out were seen in 100 percent and 90 percent respectively compared with 90 percent and 60 percent in the delayed group. The device was seen in 30 percent and 90 percent in early and delayed groups respectively. In the in vitro study the components of the device were seen and a posterior acustic shadow noted. This shadow was less obvious and the polymer anchor more easily seen in the delayed group. CONCLUSION: The Angio-Seal device produces a consistent artifact when scanned soon after deployment. These appearances could potentially be mistaken for a vascular occlusion by the unwary in the appropriate clinical setting of acute post catheterisation lower limb ischaemia however an awareness of the normal sonographic appearances of this device would make this misinterpretation unlikely.

Absorbable Implants↗
PubMedDate not supplied

Evaluation of a vessel-sealing device for use in laparoscopic ovariectomy in mares.

OBJECTIVE: To evaluate a vessel-sealing instrument (LigaSure) as a method for hemostasis of the ovarian vasculature. STUDY DESIGN: Prospective study. ANIMALS OR SAMPLE POPULATION: Thirteen mares (8 experimental, 5 patients), aged 2 to 20 years and weighing 405 to 500 kg. METHODS: Thirteen mares had standing bilateral laparoscopic ovariectomy using a vessel-sealing device (LigaSure) to provide hemostasis. Eight reproductively normal experimental mares were divided into 2 groups: 1 group was re-examined laparoscopically 72 hours and the other group 10 days after the initial standing laparoscopic ovariectomy. The vessel-sealing device uses high current and low voltage, along with pressure, to reorganize the collagen into a translucent seal to achieve hemostasis of the ovarian vasculature. RESULTS: No major operative or postoperative complications were encountered. Complete hemostasis of the ovarian pedicle was accomplished. One mare had a fever for 24 hours' postoperatively; this responded to a single dose of flunixin meglumine. CONCLUSIONS: The LigaSure appears to be a safe method for hemostasis of the ovarian vasculature. CLINICAL RELEVANCE: Benefits of the LigaSure include no foreign material remaining in the abdomen and minimal to no need for surgical dissection before application. The LigaSure eliminates complications with potential ligature slippage and bleeding during dissection.

Animals↗
PubMed2001

In vitro sealing of punctured fetal membranes: potential treatment for midtrimester premature rupture of membranes.

OBJECTIVE: Midtrimester premature rupture of membranes causes significant perinatal morbidity and death. No effective treatment exists. We investigated (1) whether a needle puncture in the fetal membranes could be sealed in vitro and (2) the optimal composition of the sealant to be used. STUDY DESIGN: Membranes from second trimester pregnancies (16-24 weeks of gestation) were stretched over a modified syringe with a 2.5-cm open diameter. The syringe was filled with 20 mL of second trimester amniotic fluid, and the membrane was punctured with a 20-gauge needle. Sealants were injected into the amniotic fluid. The primary outcome variable was time for leakage of amniotic fluid. Median times for leakage for the formulations were compared by Wilcoxon exact rank sum test. RESULTS: Platelets alone failed to seal the puncture site. All other formulations stopped leakage temporarily. Tisseel (Baxter Corp, Glendale, Calif) and cryoprecipitate/thrombin preparations led to more prolonged sealing of punctured amniotic membranes than platelets (P <.01) and were not significantly different from each other. CONCLUSION: Of the sealants tested in vitro, amniotic membranes are best sealed by a fibrin/thrombin-based sealant.

Amnion↗
PubMed1990

Sealed rupture of abdominal aortic aneurysms.

Sealed rupture of abdominal aortic aneurysms, even if uncommon, deserves particular attention for the possibility of misdiagnosis and for the deleterious effects of such a misdiagnosis. Sixteen patients (mean age 72 years; range 65 to 84 years) with chronic sealed rupture of abdominal aortic aneurysms are reported. Two patients had acute rupture of the aneurysm, and at operation chronic contained rupture was found along with the recent hemorrhage. One patient died after surgery. The remaining patients underwent successful resection with long-term survival and regression of symptoms. Consideration of sealed abdominal aortic aneurysm rupture should be included when examining elderly patients with history of unexplained back pain or femoral neuropathy. Computed tomography is a useful aid in the diagnosis of sealed rupture. Ultrasonography is less accurate; in three patients ultrasonography failed to diagnose the presence of the rupture.

Aged↗
PubMed2003

Sealed bacterial ghosts--novel targeting vehicles for advanced drug delivery of water-soluble substances.

The purpose of the present study was to develop a drug delivery model for water soluble drug substances using the bacterial ghost platform technology. Bacterial ghosts are non-denatured bacterial cell envelopes that are produced by the plasmid encoded gene E mediated lysis. We present a novel method to fill and seal bacterial ghosts for the application as a drug delivery system for fluid, non-anchored substances. E. coli ghosts were filled with the reporter substance calcein and sealed by fusion with membrane vesicles. By flow cytometry and fluorescence microscopy it was shown that bacterial ghosts can be filled with calcein, and that the bacterial ghosts can be sealed by restoring the membranes integrity. The adherence and uptake studies showed that almost all murine macrophages and a lower proportion of human colorectal adenocarcinoma cells took up fluorescence labeled bacterial ghosts. Moreover, these cells also took up effectively sealed E. coli ghosts filled with calcein, which then was released within the cells. Therefore, we propose bacterial ghosts as alternative drug delivery and release vehicles for advanced cell targeting.

Animals↗
PubMed2004

Osmotic properties of the sealed tubular system of toad and rat skeletal muscle.

A method was developed that allows conversion of changes in maximum Ca(2+)-dependent fluorescence of a fixed amount of fluo-3 into volume changes of the fluo-3-containing solution. This method was then applied to investigate by confocal microscopy the osmotic properties of the sealed tubular (t-) system of toad and rat mechanically skinned fibers in which a certain amount of fluo-3 was trapped. When the osmolality of the myoplasmic environment was altered by simple dilution or addition of sucrose within the range 190-638 mosmol kg(-1), the sealed t-system of toad fibers behaved almost like an ideal osmometer, changing its volume inverse proportionally to osmolality. However, increasing the osmolality above 638 to 2,550 mosmol kg(-1) caused hardly any change in t-system volume. In myoplasmic solutions made hypotonic to 128 mosmol kg(-1), a loss of Ca(2+) from the sealed t-system of toad fibers occurred, presumably through either stretch-activated cationic channels or store-operated Ca(2+) channels. In contrast to the behavior of the t-system in toad fibers, the volume of the sealed t-system of rat fibers changed little (by <20%) when the osmolality of the myoplasmic environment changed between 210 and 2,800 mosmol kg(-1). Results were also validated with calcein. Clear differences between rat and toad fibers were also found with respect to the t-system permeability for glycerol. Thus, glycerol equilibrated across the rat t-system within seconds to minutes, but was not equilibrated across the t-system of toad fibers even after 20 min. These results have broad implications for understanding osmotic properties of the t-system and reversible vacuolation in muscle fibers. Furthermore, we observed for the first time in mammalian fibers an orderly lateral shift of the t-system networks whereby t-tubule networks to the left of the Z-line crossover to become t-tubule networks to the right of the Z-line in the adjacent sarcomere (and vice versa). This orderly rearrangement can provide a pathway for longitudinal continuity of the t-system along the fiber axis.

Aniline Compounds↗
PubMed2006

Use of Surgicel for sealing nephrostomy tract after totally tubeless percutaneous nephrolithotomy.

BACKGROUND AND PURPOSE: Fibrin glue and gelatin matrix have been used to seal nephrostomy tracts to reduce bleeding and extravasation after tubeless percutaneous nephrolithotomy (PCNL). In this study, Surgicel (oxidized cellulose) was used to seal the nephrostomy tract after totally tubeless PNL. PATIENTS AND METHODS: Twenty patients with kidney calculi were treated with totally tubeless PNL. According to randomization, at the conclusion of surgery, the nephrostomy tracts were sealed with Surgicel in ten patients and left unsealed in the other ten. Postoperatively, the two groups were compared with respect to hematocrit changes and extravasation as detected by abdominal ultrasonography and wound-dressing inspection. RESULTS: There was statistically significant decrease in the hematocrit in both the study (P = 0.017) and the control (P = 0.003) group. When the two groups were compared with respect to the decrease in hematocrit, no statistically significant difference was seen (P = 0.241). Similarly, extravasation from the nephrostomy tract was not significantly different in the two groups. CONCLUSION: Sealing the nephrostomy tract with Surgicel after totally tubeless PNL did not decrease bleeding or extravasation from the tract.

Adult↗
PubMed2007

Deinococcus radiodurans RNA ligase exemplifies a novel ligase clade with a distinctive N-terminal module that is important for 5'-PO4 nick sealing and ligase adenylylation but dispensable for phosphodiester formation at an adenylylated nick.

Deinococcus radiodurans RNA ligase (DraRnl) is a template-directed ligase that seals nicked duplexes in which the 3'-OH strand is RNA. DraRnl is a 342 amino acid polypeptide composed of a C-terminal adenylyltransferase domain fused to a distinctive 126 amino acid N-terminal module (a putative OB-fold). An alanine scan of the C domain identified 9 amino acids essential for nick ligation, which are located within nucleotidyltransferase motifs I, Ia, III, IIIa, IV and V. Seven mutants were dysfunctional by virtue of defects in ligase adenylylation: T163A, H167A, G168A, K186A, E230A, F281A and E305A. Four of these were also defective in phosphodiester formation at a preadenylylated nick: G168A, E230A, F281A and E305A. Two nick sealing-defective mutants were active in ligase adenylylation and sealing a preadenylylated nick, thereby implicating Ser185 and Lys326 in transfer of AMP from the enzyme to the nick 5'-PO(4). Whereas deletion of the N-terminal domain suppressed overall nick ligation and ligase adenylylation, it did not compromise sealing at a preadenylylated nick. Mutational analysis of 15 residues of the N domain identified Lys26, Gln31 and Arg79 as key constituents. Structure-activity relationships at the essential residues were determined via conservative substitutions. We propose that DraRnl typifies a new clade of polynucleotide ligases. DraRnl homologs are detected in several eukaryal proteomes.

Alanine↗
PubMed2000

Sealing furcation perforations with silver glass ionomer cement: an in vitro evaluation.

Furcation perforations sealed with silver glass ionomer cement (Chelon Silver) were evaluated in vitro compared with amalgam. Access cavities were prepared in 25 extracted human molar teeth. The coronal orifices of the root canals were sealed with amalgam and varnish. Naturally occurring coronal leakage through the intact pulp chamber floor was determined quantitatively for each tooth, using a modified fluid transport model, under pressure of 1.2 Atm. Each tooth was then disconnected from the system, perforated at the furcation, and the perforation sealed with either Chelon Silver (10 teeth) or amalgam (10 teeth); five remaining teeth served as a negative control. After incubation for 24 h at 37 degrees C in 100% humidity, teeth were reconnected to the modified fluid transport system, and coronal leakage under pressure was evaluated at 1, 2, 6, 15, and 24 h. Leakage through each tooth was compared with that of its own intact pulp chamber floor before perforation and the groups compared with each other. No significant difference was found between the mean leakage of the intact pulp chamber floors of the two groups. Chelon Silver had a significantly better sealing ability than amalgam (p < 0.01): leakage rate of 0.007 and 0.017 microliter/min, respectively. It is concluded that Chelon Silver could be an adequate sealer for furcation perforations.

Cermet Cements↗
PubMed2000

The walking bleach procedure: an in vitro study to measure microleakage of five temporary sealing agents.

The purpose of this study was to compare the in vitro sealing capacity of five materials, each used as a temporary sealing agent for the walking bleach technique. All teeth received traditional biomechanical root canal instrumentation, after which the walking bleach agent was placed in the pulp chamber space. The occlusal access was sealed with one of five temporary materials: two hydraulic filling materials, a photoactivated resin composite, a zinc oxide-eugenol cement, and a zinc oxide phosphate cement with/without the placement of a piece of rubber sheet that was placed as a barrier to isolate filling material from the bleaching agent. All teeth were stored in a 1% solution of Alcian blue with thermal cycling stress. After 1 wk, they were sectioned longitudinally, and ranked by graded scores of 0 to 3, according to the degree of the dye penetration. Significantly less dye microleakage was observed in the two hydraulic materials than in the photoactivated resin. Both zinc oxide-eugenol and zinc phosphate cements showed a considerable amount of microleakage. There were no significant differences between the groups with and without a rubber sheet. Our data demonstrate that hydraulic filling materials provide the most favorable cavosurface seal when they are firmly packed into the cavity space to prevent microleakage.

Alcian Blue↗
PubMed1999

Combat trauma airway management: endotracheal intubation versus laryngeal mask airway versus combitube use by Navy SEAL and Reconnaissance combat corpsmen.

BACKGROUND: Airway management takes precedence regardless of what type of life support is taking place. The gold standard for airway control and ventilation in the hands of the experienced paramedic remains unarguably the endotracheal tube. Unfortunately, laryngoscopy and endotracheal intubation require a skilled provider who performs this procedure on a frequent basis. Special Operations corpsmen and medics receive training in the use of the endotracheal tube, but they use it infrequently. The use of alternative airways by Navy SEAL and Reconnaissance combat corpsmen has not been evaluated. Our objective was to compare the ability of Special Operations corpsmen to use the endotracheal tube (ETT), laryngeal mask airway (LMA), and esophageal-tracheal combitube (ETC) under combat conditions. METHODS: This study used a prospective, randomized, crossover design. Twelve Navy SEAL or Reconnaissance combat corpsmen participated in a 2-week Advanced Battlefield Trauma course. During the first week, instruction included the use of ETT, LMA, and ETC, viewing of videotapes for ETC and LMA, and mannequin training. The Special Operations corpsmen were required to reliably insert each airway within 40 seconds. During the second week, participants dealt with a number of active combat trauma scenarios under fire in combat conditions. Each SEAL or Reconnaissance corpsman was asked to control his "casualty's" airway with a randomized device. All participants were evaluated in the use of each of the three airways. RESULTS: Thirty-six airway insertions were evaluated. No failures occurred. All incorrect placements were detected and corrected. Mean time to place the ETT was 36.5 seconds versus 40.0 seconds for the ETC. The LMA insertion time of 22.3 seconds was significantly shorter than the other times (p < 0.05). The mean number of attempts per device was similar with all devices: LMA (1.17), ETC (1.17), and ETT (1.25). CONCLUSION: The Special Operations corpsmen easily learned how to use the ETC and LMA. In this study, they showed the ability to appropriately use the ETT as well as the ETC and LMA. For SEAL corpsmen, the alternative airways should not replace the ETT; however, on occasion an advanced combat casualty care provider may not be able to use the laryngoscope or may be unable to place the ETT. The LMA and ETC are useful alternatives in this situation. If none of these airways are feasible, cricothyrotomy remains an option. Regardless of the airway device, refresher training must take place frequently.

Allied Health Personnel↗
PubMed2003

Fibrin sealing improves stability of corneal prostheses during vitreoretinal procedures.

PURPOSE: The aim of this study was to test the effectiveness of using a fibrin sealant as an aid to stabilize temporarily sutured keratoprostheses. METHODS: Ex vivo the corneas were removed from six porcine eyes, and an Eckardt prosthesis was sutured and additionally fixed with fibrin sealant. The outflow resistance was recorded from eyes with and without fibrin-sealed keratoprostheses at different levels of intraocular pressure. The method of sealing the prosthesis was applied in the clinic and documented in four patients during intravitreal surgery. RESULTS: The flow volume of the irrigating system was measured ex vivo. Control trials were performed to investigate the effects of graduated increases in hydrostatic pressure on the system. Leakage areas were calculated, and the stability of the system was monitored during the surgical procedure. Slit-lamp biomicroscopy, funduscopy, and visual outcome were documented in the patients. Ex vivo the differences in the leakage areas between the fibrin-sealed and the unsealed conditions were statistically significant up to 51.45 mmHg (P = 0.01). In clinical applications, the fibrin sealant stabilized the keratoprosthesis, and no significant leakage or system instabilities occurred. CONCLUSIONS: The results confirm that sealing increases the stability of the keratoprosthesis and may enable greater surgical control.

Adolescent↗
PubMed2005

How long should you wait for a chest radiograph after placing a chest tube on water seal? A prospective study.

OBJECTIVE: The purpose of this study was to determine the optimal time interval for identifying a pneumothorax (PTX) on chest radiograph (CXR) after placing a chest tube on water seal. METHODS: One hundred nineteen chest tubes were placed on water seal according to a prospective, observational study protocol. After water seal, both an early (3.1 +/- 2.1 hours) and a late (17.6 +/- 8.0 hours) CXR was obtained. RESULTS: Thirty-one patients had a PTX on follow-up CXRs. There were 22 early and 9 late PTXs identified. Three patients in the early group had a clinically significant PTX or an increase in the size of PTX on follow-up CXR. None of the patients in the late group had a clinically significant PTX (any worsening of their PTX) or required further intervention. CONCLUSION: A normal chest radiograph obtained 3 hours after placing a chest tube on water seal effectively excludes development of a clinically significant pneumothorax.

Adolescent↗
PubMed2006

Functional and aggregative responses of harbour seals to changes in salmonid abundance.

There is intense debate over the potential impact of seal predation on declining salmon stocks in both the Pacific and Atlantic oceans. However, efforts to model such interactions have been constrained by a lack of data on the functional and numerical responses of these predators. Based upon theory, and data from small-scale terrestrial and freshwater systems, a type 3 functional response is expected to best describe predation by generalist pinnipeds. Similarly, theory also predicts that seal numbers should increase with salmon density in rivers following an aggregative response of predator to prey. We tested these predictions by studying the diet and local density of harbour seals in relation to seasonal variations in the abundance of salmonid in a Scottish river system. As predicted, the abundance of seals in the river was directly related to the abundance of returning salmon, and dietary data supported the type 3 functional response to changes in salmonid abundance. These studies provide empirical support for the use of type 3 response in modelling studies.

Animals↗
PubMed1991

The adaptation and sealing ability of light-cured glass ionomer retrograde root fillings.

The adaptation and sealing ability of a light-cured glass ionomer cement when used as a retrograde root filling was assessed using a confocal optical microscope with and without a fluorescent dye. This material was compared with a conventional glass ionomer cement and amalgam. The root canals of 60 extracted human single-rooted teeth were prepared and filled with gutta-percha and sealer. All the teeth were apicected, retrograde cavities were prepared, and then divided into three groups of 20 teeth each and filled with the test materials. The light-cured glass ionomer cement was well adapted to the retrograde cavity and apicected root surface. Within the retrograde cavity the cement was often well adapted to one wall, but gaps were found on the opposite cavity wall. This was probably caused by the polymerization contraction of the material. In contrast, the amalgam retrograde root fillings were poorly adapted to the cavities, with gaps between the cavity walls and amalgam. This group exhibited the poorest sealing ability as measured by the extent of dye penetration. The conventional glass ionomer cement was often found smeared over the root face, and there were unfilled voids at the base of some retrograde cavities. The results of the dye leakage study were analysed statistically. The sealing ability of the light-cured glass ionomer cement was significantly better than that of amalgam (P less than 0.001). The dye penetration around the light-cured glass ionomer cement and the conventional glass ionomer cement was not significantly different (P greater than 0.05). However, the sealing ability of the conventional glass ionomer cement was significantly better than that of amalgam (P less than 0.05).

Analysis of Variance↗
PubMed1992

The physical properties of a new sealing cement.

This study was performed on a newly developed sealing cement, TS60, to evaluate its physical properties, namely sealing, setting time, compressive strength, solubility and ease of removal. It was compared with Temporary Stopping (GC Dental Industrial Corp., Tokyo, Japan), Eugedain (zinc oxide-eugenol based material, Showa Yakuhin Kako Co. Ltd, Tokyo, Japan) and Cavit-G (calcium sulphate based material, Espe Gmbh, Seefeld, Germany), which are typical commercially available sealing cements. The results of the study revealed that TS60 had excellent sealing properties, good compressive strength, low solubility, and was readily removable by heating and melting.

Dental Cements↗
PubMed2003

Comparative in vivo analysis of the sealing ability of three endodontic sealers in post-prepared root canals.

AIM: To compare the sealing ability of the endodontic sealers AH Plus, Sealer 26 and Endofill in premolar teeth of dogs exposed to the oral cavity after post-preparation. METHODOLOGY: Forty teeth with two canals each underwent root canal cleaning and shaping. Before filling, the canals were randomly distributed into three groups according to the sealer to be used: Group 1 - AH Plus; Group 2 - Sealer 26; and Group 3 - Endofill (Dentsply, Indústria e Comércio Ltda.). Immediately after filling, the gutta percha and sealer were partially removed from the canals, leaving material only in the apical third of the root. The teeth were temporarily sealed with glass ionomer sealer for 72 h to ensure setting. The coronal seal was then removed and the canals were exposed to the oral cavity for 45 days. The animals were euthanased and their mandibles and maxillae were removed. After abundant irrigation with distilled water, the canals were dried and filled with India ink. The teeth were sealed again for 96 h before extraction. The roots of the extracted premolars were separated and stored in labelled test tubes. The roots were cleared and the extent of dye penetration was measured with a 20x stereoscopic magnifying lens. RESULTS: Statistical analysis revealed that there were significant differences between the sealers studied (P < 0.001). Means for the extent of dye penetration for AH Plus, Endofill and Sealer 26 were, respectively, 0.13, 2.27 and 3.08 mm. CONCLUSIONS: After 45 days exposure to the oral cavity, none of the sealers was capable of preventing leakage and coronal dye penetration. There were significant differences between the sealers studied, in terms of mean dye penetration.

Analysis of Variance↗