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PubMed1997

Expression of Bcl-2 and PCNA in duct cells after pancreatic duct ligation in rats.

Obstruction of the pancreatic duct induces acinar cell deletion followed by duct proliferation and interstitial fibrosis. Apoptosis has been reported to be involved in the induction of acinar cell deletion after pancreatic duct ligation (PDL) in rats, however, the mechanism of pancreatic duct cell proliferation is still unknown. We hypothesized that Bcl-2 (antiapoptosis protein) and PCNA (cell cycle-related protein) could be involved in the mechanism of pancreatic duct cell proliferation after PDL. In PDL, rats, acinar cells decreased in number and disappeared completely after duct ligation and duct-lining cells increased in number and formed duct-tubular complexes. Immunohistochemical study showed that PCNA expression appeared in the ductules and centroacinar cells from early stages after duct ligation and that Bcl-2 expression in duct cells, which was faint in normal pancreas, increased significantly when acinar cells were diminishing. Western blotting demonstrated that Bcl-2 was detected as a single band at 26 kDa, and the intensity of Bcl-2 in PDL rats was approximately ninefold stronger than in normal pancreas. Expression of Bcl-2 and PCNA after pancreatic duct ligation may be related to the prevention of apoptosis and cell proliferation of pancreatic duct cells in rats.

Animals↗
PubMed1994

T-cassette ligation: a method for direct sequencing and cloning of PCR-amplified DNA fragments.

We describe a method to ligate a PCR-amplified DNA fragment with T-protruding cassettes, which have multiple sites for endonuclease, promoter sequences of T3 and T7, and annealing sites for the universal M13 forward and reverse primers. This method, which we named T-cassette ligation, substantially facilitated direct sequencing and subcloning of PCR products. Two T-cassettes with a protruding T at their 3' ends were obtained by annealing adaptor oligomers to XbaI- or XhoI-digested pBluescript. A DNA fragment amplified by the first PCR was ligated separately to one of the two T-cassettes. The second PCR was performed using a primer complementary to one of the two T-cassettes and one of the two primers used for the first PCR to amplify the ligation product in low abundance. The resultant two DNA fragments had the T3 and T7 promoter, and an annealing site for the universal M13 forward and reverse primer, respectively. These DNA fragments were applicable to direct sequencing. For the purpose of subcloning, the third PCR was carried out with two primers each complementary to each of two T-cassettes and two PCR-amplified DNAs by the second PCR, as templates. The PCR product of the third PCR had multiple sites of the pBluescript polycloning site at both ends, facilitating its subcloning into pBluescript.

Base Sequence↗
PubMed1998

Mutation detection by ligation to complete n-mer DNA arrays.

A new approach to comparative nucleic acid sequence analysis is described that uses the ligation of DNA targets to high-density arrays containing complete sets of covalently attached oligonucleotides of length eight and nine. The combination of enzymatic or chemical ligation with a directed comparative analysis avoids many of the intrinsic difficulties associated with hybridization-based de novo sequence reconstruction methods described previously. Double-stranded DNA targets were fragmented and labeled to produce quasirandom populations of 5' termini suitable for ligation and detection on the arrays. Kilobase-size DNA targets were used to demonstrate that complete n-mer arrays can correctly verify known sequences and can determine the presence of sequence differences relative to a reference. By use of 9-mer arrays, sequences of 1.2-kb targets were verified with >99.9% accuracy. Mutations in target sequences were detected by directly comparing the intensity pattern obtained for an unknown with that obtained for a known reference sequence. For targets of moderate length (1.2 kb), 100% of the mutations in the queried sequences were detected with 9-mer arrays. For higher complexity targets (2.5 and 16.6 kb), a relatively high percentage of mutations (90% and 66%, respectively) were correctly identified with a low false-positive rate of <0.03 percent. The methods described provide a general approach to analyzing nucleic acid samples on the basis of the interpretation of sequence-specific patterns of hybridization and ligation on complete n-mer oligonucleotide arrays.

Base Sequence↗
PubMed1994

Ligation of leukocyte function-associated (LFA) molecule-1 provides an accessory signal for T-cell activation with pokeweed mitogen.

The ligation between leucocyte-Function-Associated Molecule 1 (LFA-1) and Intercellular Adhesion Molecules (ICAM) is thought to be an important component in the stimulatory interaction between antigen-presenting cells (APC) and T cells. Similar to antigenic stimulation, T-cell stimulation with pokeweed mitogen (PWM) is highly dependent on monocytes as accessory cells. This is partly a consequence of the requirement for mitogen presentation by the monocytes. The study described here addressed the question of whether LFA-1 ligation by accessory cells influences the activation of T cells with PWM. To avoid multiple costimulatory interactions between T cells and monocytes, experiments were performed with purified T cells, which were stimulated with PWM bound on autologous red blood cells (PWM-RBC). Binding on the RBC substituted partly for PWM presentation by the monocytes. Anti-LFA-1 MoAbs were presented in the immobilized form in order to mimick LFA-1 ligation by cell-bound ICAM. Three out of three different MoAbs against the beta-chain of the LFA-1 molecule (CD18) and two out of three MoAbs against the alpha-chain (CD11a) had an enhancing effect on T-cell proliferation. Proliferation was increased further by simultaneous addition of interleukin (IL)-1 beta and IL-6. Ligation of the LFA-1 molecule was found to enhance IL-2 production and tumour necrosis factor-alpha (TNF-alpha) production. The results suggest that interaction of LFA-1 with ICAM on the monocytes contributes to the accessory signal activity of monocytes in T-cell activation with PWM.

Adult↗
PubMed1988

Effect of histone H1, poly(ethyleneglycol) and DNA concentration on intermolecular and intramolecular ligation by T4 DNA ligase.

The efficiency of ligation of linear DNA and the relative amounts of intramolecular versus intermolecular ligation may be triggered by a number of additive agents. The results show that it is possible to mimic the effect of poly(ethyleneglycol) 6000 by simply increasing DNA concentration about 15-fold: both the rate and the extent of the reaction are greatly enhanced, and intermolecular ligation is largely favored. However, in this case the stimulation by salts, which occurs in poly(ethyleneglycol) solutions, is not observed; we suggest that salts enhance the hydrophobic interactions between ligase and DNA that take place in the presence of poly(ethyleneglycol). We also show that histone H1, which is involved in the formation of chromatin fibers, is able to stimulate intermolecular ligation by T4 ligase. This effect is more specific than a simple neutralisation of the phosphate groups of the DNA by positive charges of the histone; it still occurs at 125 mM NaCl and in the presence of the four core histones. The implications of the finding concerning the mode of action of histone H1 on DNA are discussed.

DNA Ligases↗
PubMed2007

Ligation of left renal vein for large spontaneous splenorenal shunt to prevent portal flow steal in adult living donor liver transplantation.

Persistance of a large spontaneous splenorenal shunt (SRS) may result in graft failure in adult living donor liver transplantation (LDLT) because it reduces the effective portal perfusion to the partial liver graft by diversion of hepatotrophic portal flow into this hepatofugal pathway. We performed a prospective study to evaluate the efficacy of ligation of left renal vein (LRV) to prevent portal flow steal and the safety of this procedure to the renal function in adult LDLT patients with SRS. Between October 2001 and January 2005, 44 cirrhotic patients with large SRS underwent LDLT with ligation of LRV. Each patient received pre- and postoperative computed tomography and Doppler USG to assess the changes of collaterals and portal flow, as well as serial renal and liver function tests. Portal flow after ligation of LRV was statistically and significantly increased when compared with pre-operative value (P = 0.001). Whereas four patients (9.1%) demonstrated sustained, elevated serum creatinine levels after operation, the renal function tests returned to normal in 40 patients. All patients recovered with satisfactory regeneration of the partial liver graft and there was no procedure-related permanent renal dysfunction. In conclusion, ligation of LRV to prevent a 'portal steal phenomenon' seems to be a safe and effective graft salvage procedure for large spontaneous SRS (>10-mm diameter) in adult LDLT.

Adult↗
PubMed1998

Electrophysiological characterization of spinal neuronal response properties in anaesthetized rats after ligation of spinal nerves L5-L6.

1. Despite a number of models of nerve injury, few studies have examined how peripheral nerve injury influences spinal somatosensory processing. 2. Ligation of two (L5-L6) of the three spinal nerves that form the sciatic nerve produces a partial denervation of the hindlimb. Following ligation, rats exhibited withdrawal responses to normally innocuous punctate mechanical and cooling stimuli (acetone) applied to the lesioned hindpaw. Such mechanical and cooling allodynia was not observed in sham-operated rats. 3. A significantly greater proportion of spinal neurones of ligated rats exhibited spontaneous activity at post-operative (PO) days 7-10 (P = 0.03) and 14-17 (P = 0.0001), compared with sham controls. The frequency of the spontaneous activity was significantly higher than that of the sham controls (P = 0.03 and P = 0.02 for days 7-10 and days 14-17, respectively). 4. At the earlier PO period, significantly (P = 0.02) more neurones of spinal nerve-ligated (SNL) rats responded to brush compared with the sham controls; at the later PO period the proportion of neurones of SNL rats responsive to prod was significantly (P = 0.007) reduced compared with the sham controls. The magnitude of the evoked neuronal response of SNL rats at PO days 7-10 was comparable to that of the sham controls. The magnitudes of brush- and prod-evoked neuronal responses of SNL rats were significantly smaller (P = 0.05 and P = 0.002, respectively) than the sham controls at PO days 14-17. In addition, neuronal responses of SNL rats to mechanical punctate stimuli and the C fibre-evoked neuronal responses were significantly reduced at the later PO period, compared with sham controls. Abeta-fibre-induced wind-up was not observed under any conditions. 5. These complex changes in neuronal responses are both time and modality dependent. The plasticity of some of the neuronal and behavioural responses following nerve injury was difficult to reconcile. We suggest that an interplay between pathological peripheral and central mechanisms may account for some of the changes that could contribute to allodynia and hyperalgesia.

Acetone↗
PubMed1983

Effects of prostaglandin E2, propranolol and nitroglycerine with halothane, pethidine or pentobarbitone anaesthesia on arrhythmias and other responses to ligation of a coronary artery in rats.

The effects of various cardiovascular drugs (prostaglandin E2 (PGE2), propranolol and nitroglycerine) and anaesthetic regimens (halothane, pethidine and pentobarbitone), upon the outcome of coronary artery ligation in acutely prepared rats were determined. Effects upon arrhythmias, blood pressure, heart rate, mortality, ECG and the size of the occluded zone were determined for each drug in the presence of each anaesthetic. PGE2 and nitroglycerine had no statistically significant effects on the outcome of ligation whatever the anaesthetic. Propranolol had limited antiarrhythmic actions. The anaesthetic used had major effects upon the outcome of ligation, regardless of the cardiovascular drugs administered. Pentobarbitone anaesthesia resulted in the highest mortality, and most arrhythmias. Pethidine-N2O anaesthesia was associated with fewer arrhythmias. Halothane-N2O anaesthesia markedly decreased the incidence and severity of arrhythmias, independent of the cardiovascular drug. It was concluded that the anaesthetic used can have a major influence on ligation-induced arrhythmias in acutely prepared anaesthetized rats.

Anesthetics↗
PubMed2004

Effect of tubal ligation on ovarian reserve and the ovarian stromal blood supply.

AIM: Our aim was to evaluate possible adverse effects of tubal ligation on ovarian reserve and ovarian stromal vascular support. METHODS: Ninety-one women who underwent tubal ligation, using a modified Pomeroy technique, were studied. The main outcome measurements were serum follicle stimulating hormone, luteinising hormone and oestradiol levels, ovarian volume, number of antral follicles and Doppler study of ovarian stromal artery on the third day of the menstrual cycle immediately before, 1 month and 12 months after the surgical intervention. RESULTS: Statistically significant elevation of follicle stimulating hormone levels was observed at 1 and 12 months after tubal ligation (P < 0.05). CONCLUSION: Doppler flow studies of the ovarian stromal arteries showed that there were no significant differences before and after tubal ligation.

Adult↗
PubMedDate not supplied

Selective vessel ligation in the pelvis: an invaluable tool in certain surgical procedures.

While developing the technique of abdominal radical trachelectomy for conservative cervical cancer management, the vascular supply of the uterus was thoroughly examined. The question of how many vessels the uterus requires to ensure its viability arose. Following an abdominal radical trachelectomy for stage IB cervical carcinoma, blood supply of the body of the uterus is successfully maintained by only the two infundibulopelvic vessels (n= 34). Pregnancy has resulted following this technique (n= 2). Selective ligation of the pelvic vasculature has been utilized in the abdominal radical trachelectomy procedure. The objectives of this study were to investigate the vasculature of the infundibulopelvic and broad ligaments, to assess the contribution of the ovarian and uterine vessels to overall uterine perfusion, and to consider the clinical applications of selective pelvic vessel ligation. Ten fresh dissections of the infundibulopelvic vessels, broad ligaments of benign total abdominal hysterectomy, and bilateral salpingo-oophorectomy specimens were performed. Perfusion index (PI) and oxygen saturation (O(2)Sat) measurements using a modified probe were taken at specified intervals at the uterine cornu during ten routine benign abdominal hysterectomies to assess the contribution of the ovarian and uterine vessels to overall uterine perfusion and the concepts studied were utilized in certain gynecological procedures. The ovarian/infundibulopelvic vessels course medially through the broad ligament toward the uterine cornu and consistently give off a branch to the ovary on its lateral border. In addition, further vessels were noted to run laterally from the uterine cornu along the ovarian ligament to the medial aspect of the ovary. PI and O(2)Sat measurements imply that the uterine and ovarian vessels contribute almost equally to uterine perfusion. Clinical application by selective ligation of the pelvic vasculature has been utilized in certain gynecological procedures often prone to torrential life-threatening uterine hemorrhage. Selective temporary ligation of the uterine and ovarian vessels has proven useful in the surgical management of chemoresistant gestational trophoblastic disease, in the Strassman procedure, fertility-sparing surgery in ruptured cornual ectopic pregnancies, and unrelenting postpartum hemorrhage. Of the six supplying vessels (ovarian, uterine, and vaginal) to the uterus only two (ovarian or uterine or a combination thereof) are required for uterine viability.

Female↗
PubMed1994

Synthesis of proteins by native chemical ligation.

A simple technique has been devised that allows the direct synthesis of native backbone proteins of moderate size. Chemoselective reaction of two unprotected peptide segments gives an initial thioester-linked species. Spontaneous rearrangement of this transient intermediate yields a full-length product with a native peptide bond at the ligation site. The utility of native chemical ligation was demonstrated by the one-step preparation of a cytokine containing multiple disulfides. The polypeptide ligation product was folded and oxidized to form the native disulfide-containing protein molecule. Native chemical ligation is an important step toward the general application of chemistry to proteins.

Amino Acid Sequence↗
PubMed2006

Gingipains of Porphyromonas gingivalis modulate leukocyte adhesion molecule expression induced in human endothelial cells by ligation of CD99.

Porphyromonas gingivalis has been implicated as a key etiologic agent in the pathogenesis of destructive chronic periodontitis. Among virulence factors of this organism are cysteine proteinases, or gingipains, that have the capacity to modulate host inflammatory defenses. Intercellular adhesion molecule expression by vascular endothelium represents a crucial process for leukocyte transendothelial migration into inflamed tissue. Ligation of CD99 on endothelial cells was shown to induce expression of endothelial leukocyte adhesion molecule 1, vascular cell adhesion molecule 1, intercellular adhesion molecule 1, and major histocompatibility complex class II molecules and to increase adhesion of leukocytes. CD99 ligation was also found to induce nuclear translocation of NF-kappaB. These results indicate that endothelial cell activation by CD99 ligation may lead to the up-regulation of adhesion molecule expression via NF-kappaB activation. However, pretreatment of endothelial cells with gingipains caused a dose-dependent reduction of adhesion molecule expression and leukocyte adhesion induced by ligation of CD99 on endothelial cells. The data provide evidence that the gingipains can reduce the functional expression of CD99 on endothelial cells, leading indirectly to the disruption of adhesion molecule expression and of leukocyte recruitment to inflammatory foci.

12E7 Antigen↗
PubMed2000

Recombinogenic flap ligation pathway for intrinsic repair of topoisomerase IB-induced double-strand breaks.

Topoisomerase IB catalyzes recombinogenic DNA strand transfer reactions in vitro and in vivo. Here we characterize a new pathway of topoisomerase-mediated DNA ligation in vitro (flap ligation) in which vaccinia virus topoisomerase bound to a blunt-end DNA joins the covalently held strand to a 5' resected end of a duplex DNA containing a 3' tail. The joining reaction occurs with high efficiency when the sequence of the 3' tail is complementary to that of the scissile strand immediately 5' of the cleavage site. A 6-nucleotide segment of complementarity suffices for efficient flap ligation. Invasion of the flap into the duplex apparently occurs while topoisomerase remains bound to DNA, thereby implying a conformational flexibility of the topoisomerase clamp around the DNA target site. The 3' flap acceptor DNA mimics a processed end in the double-strand-break-repair recombination pathway. Our findings suggest that topoisomerase-induced breaks may be rectified by flap ligation, with ensuing genomic deletions or translocations.

Base Pairing↗
PubMed2005

Signal regulatory protein alpha ligation induces macrophage nitric oxide production through JAK/STAT- and phosphatidylinositol 3-kinase/Rac1/NAPDH oxidase/H2O2-dependent pathways.

Signal regulatory protein alpha (SIRPalpha) is a glycoprotein receptor that recruits and signals via the tyrosine phosphatases SHP-1 and SHP-2. In macrophages SIRPalpha can negatively regulate the phagocytosis of host cells and the production of tumor necrosis factor alpha. Here we provide evidence that SIRPalpha can also stimulate macrophage activities, in particular the production of nitric oxide (NO) and reactive oxygen species. Ligation of SIRPalpha by antibodies or soluble CD47 triggers inducible nitric oxide synthase expression and production of NO. This was not caused by blocking negative-regulatory SIRPalpha-CD47 interactions. SIRPalpha-induced NO production was prevented by inhibition of the tyrosine kinase JAK2. JAK2 was found to associate with SIRPalpha in macrophages, particularly after SIRPalpha ligation, and SIRPalpha stimulation resulted in JAK2 and STAT1 tyrosine phosphorylation. Furthermore, SIRPalpha-induced NO production required the generation of hydrogen peroxide (H(2)O(2)) by a NADPH oxidase (NOX) and the phosphatidylinositol 3-kinase (PI3-K)-dependent activation of Rac1, an intrinsic NOX component. Finally, SIRPalpha ligation promoted SHP-1 and SHP-2 recruitment, which was both JAK2 and PI3-K dependent. These findings demonstrate that SIRPalpha ligation induces macrophage NO production through the cooperative action of JAK/STAT and PI3-K/Rac1/NOX/H(2)O(2) signaling pathways. Therefore, we propose that SIRPalpha is able to function as an activating receptor.

Animals↗
PubMed1976

Mortality and cerebral metabolism after bilateral carotid artery ligation in normotensive and spontaneously hypertensive rats.

Mortality and cerebral glycolytic metabolism were studied after bilateral ligation of the common carotid artery in normotensive Wistar rats (NTR), and spontaneously hypertensive rats (SHR) derived from Wistar strain. In the first 24 hours after occlusion of carotid arteries, 72 per cent of 108 SHR died, whereas it was fatal in only 16 per cent of 43 NTR. In SHR, cerebral lactate and cerebral lactate/pyruvate ratio (L/P ratio) increased by 12.4 and 12.1 times the control, respectively at five to six hours after ligation, and remained raised even in rats surviving for two to three days thereafter. Changes in cerebral lactate and L/P ratio were minimal in NTR. Cerebral ATP decreased markedly at five to six hours after ligation in SHR studied. These results indicate that bilateral carotid artery ligation causes severe brain damage in SHR but not in NTR, suggesting hypertension per se to be operative for the development of cerebral ischaemia.

Acid-Base Imbalance↗
PubMed2000

p53-dependent acinar cell apoptosis triggers epithelial proliferation in duct-ligated murine pancreas.

The mechanisms linking acinar cell apoptosis and ductal epithelial proliferation remain unknown. To determine the relationship between these events, pancreatic duct ligation (PDL) was performed on p53(+/+) and p53(-/-) mice. In mice bearing a wild-type p53 allele, PDL resulted in upregulation of p53 protein in both acinar cells and proliferating duct-like epithelium. In contrast, upregulation of Bcl-2 occurred only in duct-like epithelium. Both p21(WAF1/CIP1) and Bax were also upregulated in duct-ligated lobes. After PDL in p53(+/+) mice, acinar cells underwent widespread apoptosis, while duct-like epithelium underwent proliferative expansion. In the absence of p53, upregulation of p53 target genes and acinar cell apoptosis did not occur. The absence of acinar cell apoptosis in p53(-/-) mice also eliminated the proliferative response to duct ligation. These data demonstrate that PDL-induced acinar cell apoptosis is a p53-dependent event and suggest a direct link between acinar cell apoptosis and proliferation of duct-like epithelium in duct-ligated pancreas.

Animals↗
PubMed1982

Angiotensin does contribute to drinking induced by caval ligation in rat.

In small (0.5 mg/kg) subcutaneous doses, the angiotensin-converting enzyme inhibitor, captopril, greatly enhanced drinking in response to caval ligation in the rat. Drinking was not secondary to urinary water loss since the rats developed a substantial positive fluid balance. High (50 mg/kg) subcutaneous doses of captopril reduced drinking to a level below that following caval ligation alone. This effect could be mimicked by giving repeated intracerebroventricular injections of captopril (total amount 110 micrograms) to rats treated with the lower subcutaneous dose of captopril. With this combination, therefore, not only did the lower dose enhancement disappear, the basal caval ligation drinking response was also reduced with a total dose of captopril of less than 2% of the higher subcutaneous dose alone. These results show that, when conversion of angiotensin I to angiotensin II is prevented in the brain as well as systemically, drinking in response to caval ligation is reduced although not entirely prevented. The original report that such drinking is multifactorial, depending on angiotensin as well as nonangiotensin mechanisms, is confirmed.

Angiotensin I↗
PubMed2006

Does pharmaconutrition with L-arginine and/or alpha-tocopherol improve the gut barrier in bile duct ligated rats?

BACKGROUND/AIM: Nitric oxide supplementation and antioxidant therapy modulate gut barrier function, but the relationships between enhanced nitric oxide production, antioxidant administration, and biliary obstruction remain unclear. We evaluated the role of nitric oxide and alpha-tocopherol supplementation in bile duct ligated rats. METHODS: Fifty male Wistar albino rats underwent sham operation (group I; control animals) or bile duct ligation (groups II, III, IV, and V). The ligation groups received the following regimens: standard pellet diet (group II), pellet diet plus intramuscularly administered alpha-tocopherol (group III), and L-arginine-enriched pellet diet without (group IV) or with (group V) alpha-tocopherol. Nitric oxide, malondialdehyde, and alpha-tocopherol concentrations were assessed at the end of 3 weeks. Liver and intestinal samples were scored histologically. Mesenteric lymph node and liver cultures were assessed for bacterial translocation. RESULTS: The liver malondialdehyde concentration was highest in group III. The nitric oxide content in the liver was higher in groups III and V, as were the blood alpha-tocopherol levels. Bacterial translocation was evident following bile duct ligation, but did not differ among the treatment groups. Intestinal histology revealed that group III had the lowest villus height, that group V had the least villus count, and that group II had the highest mucous cell count. The fibrosis scores were higher in groups IV and V. CONCLUSIONS: An obvious effect of alpha-tocopherol (with or without L-arginine) on the gut barrier could not be demonstrated. Moreover, the L-arginine-enriched diet promoted fibrosis in the liver. Thus, while biliary duct obstruction triggers bacterial translocation, nitric oxide and/or alpha-tocopherol supplementation did not seem to improve the gut barrier in our model.

Administration, Oral↗