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PubMed1995

Site-directed mutagenesis of Azotobacter vinelandii ferredoxin I: cysteine ligation of the [4Fe-4S] cluster with protein rearrangement is preferred over serine ligation.

The [4Fe-4S] cluster of Azotobacter vinelandii ferredoxin I receives three of its four ligands from a Cys-Xaa-Xaa-Cys-Xaa-Xaa-Cys sequence at positions 39-45 while the fourth ligand, Cys20, is provided by a distal portion of the sequence. Previously we reported that the site-directed mutation of Cys20 to Ala (C20A protein) resulted in the formation of a new [4Fe-4S] cluster that obtained its fourth ligand from Cys24, a free cysteine in the native structure. That ligand exchange required significant protein rearrangement. Here we report the conversion of Cys20 to Ser (C20S protein), which gives the protein the opportunity either to retain the native structure and use the Ser20 O gamma as a ligand or to rearrange and use Cys24. X-ray crystallography demonstrates that the cluster does not use the Ser20 O gamma as a ligand; rather it rearranges to use Cys24. In the C20S protein the [4Fe-4S] cluster has altered stability and redox properties relative to either C20A or the native protein.

Amino Acid Sequence↗
PubMed1984

Polymer-stimulated ligation: enhanced ligation of oligo- and polynucleotides by T4 RNA ligase in polymer solutions.

The effects of macromolecular crowding were tested on several reactions catalyzed by T4 RNA ligase. The rate of cyclization of oligoriboadenylates was stimulated up to 10-fold by relatively high concentrations of several polymers (polyethylene glycol (PEG) 8000 or 20,000; bovine plasma albumin; Ficoll 70). In addition, higher concentrations of PEG 8000 or PEG 20,000 allowed the novel formation of large linear products from the oligoriboadenylates. Also stimulated by high concentrations of PEG 8000 were the rate at which T4 RNA ligase joined p(dT)10 to oligoriboadenylates and the rate at which the enzyme activated p(dT)n by transfer of an adenylyl moiety from ATP to the oligonucleotides. These results with T4 RNA ligase are compared to earlier studies on the effects of crowding on DNA ligases.

Chemical Phenomena↗
PubMed1998

Non-enzymatic, template-directed ligation of 2'-5' oligoribonucleotides. Joining of a template and a ligator strand.

Decauridylate containing exclusively a 2'-5' phospho-diester bond ([2'-5']U10) served as a template for the synthesis of oligoadenylates [oligo(A)s] from the 5'-phosphorimidazolide of 2'-5' diadenylate (ImpA-2'p5'A). Joining of [2'-5']U10and ImpA2'p5'A also took place in substantial amounts to yield long-chain oligoribonucleotides in the template-directed reaction. An unusual CD spectrum ascribed to helix formation between [2'-5']U10and [2'-5'](pA)2was observed under the same conditions as that of the template-directed reaction. The 3'-5' linked decauridylate ([3'-5']U10) also promoted the template-directed synthesis of oligo(A)s from ImpA2'p5'A, but more slowly compared with [2'-5']U10. The results indicate that short-chain RNA oligomers with a 2'-5' phosphodiester bond could lead to longer oligoribonucleotides by template-directed chain elongation.

Adenine Nucleotides↗
PubMed1975

Tubal ligation and pregnancy: mechanism of recanalization after tubal ligation.

The mechanism of recanalization after resection of a segment of the fallopian tube by the Pomeroy sterilization method is discussed. At the level of resection, the epithelial lining of the fallopian tube tends to regenerate, covering the split ends and planes of cleavage of the resected surfaces and forming slitlike spaces and blind pouches lined by tall columnar cells. Scarring and subsequent retraction of both severed ends of the tubes tend to result in approximation; in some cases, the resected ends and the epithelial lining bridge the gap between the lumina, re-establishing patency of the tube. Surgical procedures that prevent approximation of the resected ends of the fallopian tubes or methods that seal the lumina should produce failure rates lower than those obtained with the classic Pomeroy sterilization method.

Adult↗
PubMed1994

[Ischemia-reperfusion experimental model in the rabbit: ligation of the circumflex coronary artery versus ligation of the anterior descending coronary artery].

INTRODUCTION AND OBJECTIVES: The myocardial damage during ischemia and reperfusion in the rabbit was studied, in order to compare two different models: the occlusion of the circumflex coronary artery (Cx) vs the occlusion of the left anterior descending coronary artery (LAD). METHODS: Each group consisted of 10 New Zealand rabbits; after 50 minutes of occlusion the artery was reopened for 20 minutes and then a biopsy from the area at risk and another from the perfused area were obtained. The specimens were used for chemiluminiscence and electron microscopy. Electrocardiograms were taken throughout the experience. Transverse sections of both ventricles were used for light microscopy. RESULTS: In both groups elevations of the ST segment, more important in the Cx group (p < 0.001) were observed. In the LAD group, the QRS complexes were enlarged in three animals, in 4 rabbits there were reperfusion arrhythmias, but the experimental mortality was zero. In the other group, 50% of the animals died during the experiment. In the LAD group the chemiluminiscence of the area at risk was greater than that of the perfused area (p < 0.001). No differences could be demonstrated between the samples of the Cx group, probably because of the extensive necrotic areas unable to generate photoemission. Ultrastructural and microscopic characteristic lesions of the ischemia-reperfusion phenomena were found in both groups, particularly severe in the Cx group with extensive areas of necrotic tissue. CONCLUSIONS: This experience shows that the myocardial necrosis, the mortality and the malignant arrhythmias were quantitatively different according to which coronary artery was occluded. This fact may be taken into account when an animal model for ischemia-reperfusion is selected.

Animals↗
PubMed1995

Jugular ligation does not increase intracranial pressure but does increase bihemispheric cerebral blood flow and metabolism.

OBJECTIVES: To answer the following questions: a) Does jugular venous ligation (simulating venovenous extracorporeal life support) alter proximal jugular venous pressure, intracranial pressure, hemispheric cerebral blood flow, or cerebral metabolism? b) Does release of ligation reverse these effects? and c) What are the comparative effects of venous ligation alone vs. venous ligation in combination with arterial ligation? DESIGN: Prospective, randomized, laboratory investigation. SETTING: Multidisciplinary laboratory setting. SUBJECTS: Sixteen swine, weighing 8.1 to 12.1 kg, 3 to 4 wks of age. INTERVENTIONS: Sixteen swine were randomly assigned to two groups, utilizing a random sequence of vessel ligation. Nine swine underwent occlusion of the right internal and external jugular veins alone (venovenous ligation) followed by release of the occlusion and then occlusion of the right common carotid artery and the right internal and external jugular veins together (venoarterial ligation). The remaining seven swine underwent venoarterial ligation, followed by release of the occlusion and then venovenous ligation. In the experimental group in which venovenous ligation was performed first, the 5, and 30-min release periods after ligation were taken to represent the effects of draining the right jugular vein during venovenous extracorporeal life support. MEASUREMENTS AND MAIN RESULTS: Data were obtained at baseline, 5, and 30 mins after each ligation/release period. Intracranial pressure, right and left internal jugular pressures/flow rates, and cerebral sinus lactate concentrations were measured. Cerebral blood flow was determined using 133Xe clearance methodology, and the cerebral metabolic rate was calculated. There were no significant differences between the ipsilateral internal jugular pressure or extracorporeal life support at 5 or 30 mins after venovenous or venoarterial ligation compared with baseline values or compared with the release of the ligation at 5 or 30 mins. There was a significant increase in right-side (44.7 +/- 2.0 vs. 38.8 +/- 2.4 mL/kg/min; p < .05) and left-side (42.9 +/- 2.3 vs. 38.7 +/- 1.9 mL/kg/min; p < .05) cerebral blood flow 5 mins after venovenous ligation when compared with baseline values. Similarly, after venoarterial ligation, there was a significant increase in right-side (44.6 +/- 2.2 vs. 38.8 +/- 2.4 mL/kg/min; p < .05) and left-side (43.9 +/- 1.5 vs. 38.7 +/- 1.9 mL/kg/min; p < .05) and cerebral blood flow. Cerebral oxygen consumption was significantly increased after venovenous (2.7 +/- 0.2 to 3.2 +/- 0.2 mL/kg/min; p < .05) and venoarterial (2.7 +/- 0.2 to 3.1 +/- 0.2 mL/kg/min; p < .05) ligation at 5 mins after ligation. This increase persisted at the 30-min period and after release of ligation. CONCLUSIONS: Ligation of the right jugular veins alone (venovenous ligation) or jugular veins and right carotid artery (venoarterial ligation) does not increase jugular venous pressures or intracranial pressure. However, this procedure does increase cerebral blood flow and cerebral oxygen consumption. These findings demonstrate that there is adequate decompression of the venous system by the cerebrovascular system and retrograde decompression during extracorporeal life support appears unwarranted.

Animals↗
PubMed2004

Hemorrhoidal elastic band ligation with flexible videoendoscopes: a prospective, randomized comparison with the conventional technique that uses rigid proctoscopes.

BACKGROUND: Elastic band ligation by means of a rigid proctoscope is the treatment of choice for patients with symptoms caused by internal hemorrhoids of grade 2 to 3. In contrast to the flexible videoendoscope, the rigid proctoscope has limited maneuverability, has a narrower field of view, and does not allow adequate documentation. Therefore, a randomized trial was conducted to compare the safety and the efficacy of conventional elastic band ligation with videoendoscopic elastic band ligation. METHODS: A total of 100 consecutive patients (mean age 47 [12] years) with chronically bleeding grade 2 or 3 internal hemorrhoids were randomized to elastic band ligation or videoendoscopic elastic band ligation. For videoendoscopic elastic band ligation, a reusable multiband ligator was attached to the end of a therapeutic upper videoendoscope. From one to 3 bands were placed per session in both groups. Re-treatment was performed every 2 to 3 weeks in both groups until cessation of bleeding and eradication of the hemorrhoids (at least grade 2) were achieved. Thereafter, the patients were followed to assess complications and efficacy. Recurrent bleeding was considered a treatment failure. RESULTS: To achieve the desired therapeutic aims, a significantly lower number of treatment sessions was required in the videoendoscopic elastic band ligation group (1.8 [0.8] vs. 2.4 [0.9]; p < 0.01) and the total number of bands applied was significantly less (2.8 [1.1] vs. 3.7 [1.4]; p < 0.01). Pain was noted after ligation by 25% of patients in the elastic band ligation group compared with 27% of those who had videoendoscopic elastic band ligation. However, analgesic medications were required in only 7% after elastic band ligation and 9% after videoendoscopic elastic band ligation (NS). Post-ligation bleeding that had to be treated endoscopically occurred in 3.5% of the patients of the elastic band ligation group and 3.2% of those in the videoendoscopic elastic band ligation group (NS). Blood transfusion was not required. At a median follow-up of 12 months, there was no recurrence of bleeding in 40 patients (80%) in the conventional elastic band ligation group vs. 43 (86%) in the videoendoscopic elastic band ligation (NS). CONCLUSIONS: The long-term efficacy and safety of conventional elastic band ligation and videoendoscopic elastic band ligation are highly comparable. However, when videoendoscopic elastic band ligation is performed, significantly fewer treatment sessions are required.

Endoscopes, Gastrointestinal↗
PubMed1997

Chronic cerebral hypoperfusion by permanent internal carotid ligation produces learning impairment without brain damage in rats.

To investigate the influence of cerebral hypoperfusion on learning behaviours, we developed a novel rat cerebral hypoperfusion model, in which the bilateral internal carotid arteries were permanently ligated to reduce the cerebral blood flow, and examined its behavioural and histopathological consequences in comparison to those occurring after bilateral common carotid ligation. In the Morris water maze task, rats with common carotid ligation exhibited a learning deficit, whereas rats with internal carotid ligation exhibited normal learning. Both models exhibited significant learning impairments in the eight-arm radial maze task, although the impairment was less severe in internal carotid-ligated rats than in common carotid-ligated rats. The cerebral blood flow of rats with common carotid ligation was reduced significantly both two and 10 days after ligation, and was still below normal three months after ligation. A milder, but significant reduction in the cerebral blood flow was observed in internal carotid-ligated rats. Shrinkage of the optic nerves and a circadian activity rhythm desynchronized to the light/dark cycle were exhibited by the rats with common carotid ligation, whereas these parameters remained unaffected in the rats with internal carotid ligation, suggesting that permanent ligation of common carotid arteries but not internal carotid arteries impairs visual functions. The main pathological changes observed in the brain following common carotid ligation were rarefaction and gliosis of the white matter and neuronal loss in the hippocampal CA1 region. On the other hand, the rats with internal carotid ligation had no significant brain damage. Chronic treatment with idebenone (1.5 and 15 mg/kg/day), a cerebral energy metabolism enhancer, over a three-month period, commencing five days after ligation, ameliorated the impairment of water maze learning in rats with common carotid ligation. The treatment also significantly improved the learning impairment in the radial maze task of internal carotid-ligated rats. Idebenone had no effect on the histopathological changes that followed cerebral hypoperfusion. It is concluded that cerebral hypoperfusion induced by permanent internal carotid ligation impairs the working memory without causing pathological damage to the brain tissues and the visual system, and the learning impairment can be ameliorated by a cerebral energy metabolism enhancer. These findings have the clinical implication that a reduction in blood flow may be an important factor that causes or exacerbates cognitive decline in dementias.

Animals↗
PubMed1997

Effect of high and intermediate ligation on survival and recurrence rates following curative resection of colorectal cancer.

PURPOSE: How wide excision of the regional mesenteric lymphatic drainage influences survival and recurrence rates following curative resection of colorectal cancers needs to be more clearly defined. METHODS: A series of 2,409 consecutive patients undergoing curative resections with detailed descriptions of the operative procedure and the lymphatic drainage in the surgical specimens provided a unique database to provide meaningful comparisons between high and intermediate level ligation. RESULTS: High ligation made a statistically significant difference in the death rate from recurrent cancer in patients with Dukes B, AC, and C1 cancers. Based on cancer-related deaths, the probability of five-year survival rate increased with high ligation from 73.9 to 84 percent in patients with Dukes B colon cancers and from 49.0 to 58.6 percent in patients with Dukes C1 colon cancers. In patients with Dukes AC cancers, high ligation increased the five-year survival rate from 64.9 to 80.4 percent. In patients with Dukes C cancers with involved middle level lymph nodes, the five-year survival rate increased from 20.5 to 33 percent and the death rate from recurrent cancer fell from 77 to 59 percent with high ligation. In patients with Dukes AC cancers with four or less involved nodes, the five-year survival rate was increased by high ligation from 50 to 78.6 percent in the colon and from 40 to 71.4 percent in the rectum. When more than four lymph nodes were involved, the survival rate was unaffected by the level of ligation. Although high ligation reduced distant recurrences, its greatest effect was observed in the incidence of local and suture line recurrence. The five-year local recurrence rate in patients with Dukes B who were managed by high ligation was 11.4 percent compared with 18.7 percent with intermediate ligation. In patients with Dukes C cancer, the local recurrence rate was 20.8 percent five years following high ligation compared with 30.7 percent for intermediate ligation. In patients with Dukes B cancer who were undergoing curative resections, the incidence of suture line recurrence was 3.9 percent following high ligation compared with 5.5 percent following intermediate ligation. In patients with Dukes C cancer, the incidence of suture line recurrence was 6.9 percent with high ligation and 11.4 percent with intermediate ligation. CONCLUSION: In certain stages of colorectal cancer, the more extensive resection of mesenteric lymphatic drainage associated with high ligation appears to increase the survival rate and reduce the recurrence rate following curative resections.

Colorectal Neoplasms↗
PubMed2000

Defining the role of extended saphenofemoral junction ligation: a prospective comparative study.

OBJECTIVE: This study explores the added effect of extended saphenofemoral junction (SFJ) ligation when the greater saphenous vein (GSV) has been eliminated from participating in thigh reflux by means of endovenous obliteration. GSV obliteration, unlike surgical stripping, can be done with or without SFJ ligation to isolate and study SFJ ligation's specific contribution to treatment results. METHODS: Sixty limbs treated with SFJ ligation and 120 limbs treated without high ligation were selected from an ongoing, multicenter, endovenous obliteration trial on the basis of their having primary varicose veins, GSV reflux, and early treatment dates. RESULTS: Five (8%) high-ligation limbs and seven (6%) limbs without high ligation with patent veins at 6 weeks or less were excluded as unsuccessful obliterations. Treatment significantly reduced symptoms and CEAP clinical class in both groups (P =.0001). Recurrent reflux developed in one (2%) of 49 high-ligation limbs and eight (8%) of 97 limbs without high ligation by 6 months (P =.273). New instances of reflux did not appear thereafter in 57 limbs followed to 12 months. Recurrent varicose veins occurred in three high-ligation limbs and four limbs without high ligation by 6 months and in one additional high-ligation limb and two additional limbs without high ligation by 12 months. Actuarial recurrence curves were not statistically different with or without SFJ ligation (P >.156), predicting greater than 90% freedom from recurrent reflux and varicosities at 1 year for both groups. CONCLUSION: These early results suggest that extended SFJ ligation may add little to effective GSV obliteration, but our findings are not sufficiently robust to warrant abandonment of SFJ ligation as currently practiced in the management of primary varicose veins associated with GSV vein reflux.

Adult↗
PubMed1996

Behavioral and electrophysiological assessment of hyperalgesia and changes in dorsal horn responses following partial sciatic nerve ligation in rats.

Behavioral and electrophysiological methods were used to investigate the hyperalgesia and allodynia, and functional changes in lumbar spinal dorsal horn neurons, in a model of neuropathic pain (Selzer et al. 1990) involving ligation of one-third to one-half of one sciatic nerve in rats. One and 5 weeks following ligation, there was a significant reduction in hind limb withdrawal latency to noxious radiant heat on the operated side and, to a lesser degree, on the unoperated side. By 16 weeks, heat withdrawal latencies were reduced about equally (approximately 40%) on both sides. Withdrawal threshold to mechanical pressure was markedly reduced within 1 week on the operated side, and decreased in a time-dependent manner on the unoperated side. Heat withdrawal latency and von Frey withdrawal thresholds were not significantly affected in sham-operated rats. The same rats were tested in a paradigm measuring the isometric force of hind limb withdrawals elicited by graded noxious contact heat stimuli (38-52 degrees C, 5 sec). Withdrawal force increased monotonically with stimulus temperature starting at a threshold of approximately 40 degrees C. Stimulus-response functions were not significantly different between a sham-operated group and groups tested 5 (acute) and 16 weeks (chronic) after partial sciatic nerve ligation. Following behavioral testing, the animals were deeply anesthetized with pentobarbital sodium to allow electrophysiological recording of responses of single lumbar dorsal horn wide-dynamic range-type neurons to mechanical and noxious thermal stimulation of the hind paw. Recordings were made from 6 sham-operated rats (26 neurons ipsilateral and 31 contralateral to the operated leg), from 7 rats receiving partial sciatic nerve ligation 5 weeks previously (29 ipsilateral and 29 contralateral to ligation), and from 7 rats receiving partial sciatic ligation 16 weeks previously (18 ipsilateral, 29 contralateral to ligation). In several ligated rats we were unable to find heat-responsive neurons with cutaneous receptive fields on the hind paw ipsilateral to the ligation. For the neurons that were sensitive to heat, responses increased monotonically from a threshold of 40-42 degrees C. Neuronal stimulus-response functions for heat were not significantly different between ipsi- and contralateral (to operated) sides in the sham, 5-week or 16-week post-ligation groups, or between sham and 5- or 16-week post-ligation groups. Mechanical receptive field areas were not significantly different between ipsi- and contralateral sides in the sham and 5-week post-ligation groups, or between sham and 5-week post-ligation groups. However, receptive field areas were significantly larger in the 16-week post-ligation group (both ipsi- and contralateral to ligation) compared to sham and 5-week post-ligation groups. The results suggest that allodynia may be associated with a chronic enhancement of neuronal mechanosensitivity, but that the thermal hyperalgesia is not associated with enhanced neuronal responsiveness or force of withdrawal.

Animals↗