Vitamin K-dependent and vitamin K-independent hypocoagulant effects of dietary fish oil in rats.

In rats, dietary fish oil causes a plasma triglyceride-lowering as well as hypocoagulant effect. The latter is apparent from reduced levels of vitamin K-dependent coagulation factors and a decreased thrombin-forming potential of the coagulating plasma. Here, we describe that intervention with low levels of n-3 polyunsaturated fatty acids (n-3 PUFAs, about 2.5% of digestible energy, en%) resulted in no more than a small reduction in coagulation factors, when supplied as part of a high-fat diet relatively rich in vitamin K. Plasma triglycerides also remained unchanged. On the other hand, when feeding rats with low- or high-fat diets restricted in vitamin K, intervention with 3 en% of n-3 PUFAs acids (fish oil) caused only a lowering in triglycerides in combination with high fat. The fish caused a reduction in coagulation potential and levels vitamin K-dependent coagulation factors (prothrombin and factor VII) that was most prominent with the low-fat diet. Fish oil, in combination with low fat but not with high fat, reduced the vitamin K levels in the liver of the animals. In addition, regardless of the fat content, the vitamin K-independent coagulation factor V was decreased in the fish oil groups. Taken together, these results indicate that, in the rat, the hypocoagulant effect of a low dose of n-3 PUFAs is most apparent at low intakes of both vitamin K and fat, is not linked to the triglyceride plasma level, but involves modulation of both vitamin K-dependent and -independent coagulation factors.

Determination of glufosinate ammonium and its metabolite (AE F064619 and AE F061517) residues in water by gas chromatography with tandem mass spectrometry after ion exchange cleanup and derivatization.

An analytical method for the determination of glufosinate ammonium and its principal metabolites, AE F064619 and AE F061517, in water of two different hardnesses (5 and 30 DH, French hardness) has been developed and validated. Samples were spiked at different levels (0. 05 and 0.5 microgram/L) and were purified by column chromatography on ion-exchange resins. After derivatization with glacial acetic acid and trimethylarthoacetate mixture, the derivatives were quantified by using capillary gas chromatography with an ion-trap tandem mass spectrometric detector. Analytical conditions for MS/MS detection were optimized, and the quantification was carried out on the areas of the most representative ions. The limit of quantification was validated at 0.05 microgram/L for each compound. The mean recovery value and the relative standard deviation (n = 20) were 92.0% and 17. 8% for glufosinate ammonium, 90.2% and 15.8% for AE F064619, and 89. 7% and 12.7% for AE F061517.

Determination of glyphosate and aminomethylphosphonic acid residues in water by gas chromatography with tandem mass spectrometry after exchange ion resin purification and derivatization. Application on vegetable matrixes

An analytical method for the determination of glyphosate and its principal metabolite, aminomethylphosphonic acid (AMPA), in water of different hardnesses (5, 20, and 30 degrees DH, french hardness) has been developed. Samples were fortified at different levels (0.05, 0.1, 1, and 5 microg/L) and were purified by column chromatography on ion-exchange resins. After derivatization with TFAA/HFB mixture, the derivatives were quantified by using capillary gas chromatography with an ion-trap tandem mass spectrometric detector. Analytical conditions for MS/MS detection were optimized, and the quantification was carried out on the sum of areas of the three most representative ions: m/z 283, 223, and 181 for AMPA and m/z 440, 321, and 261 for glyphosate. The limit of quantification was demonstrated to be at 0.05 microg/L for each compound. The mean recovery value and the relative standard deviation (n = 65) were 93 and 12% for AMPA and 95 and 13% for glyphosate.

Gastrointestinal injuries during gynaecological laparoscopy.

A retrospective case review study was carried out on gastrointestinal injuries which occur during gynaecological laparoscopy. Fifty-six patients with 62 gastrointestinal injuries were identified. One-third of the complications (32.2%) occurred during the installation phase for laparoscopy. Four of the six complications attributed to electrosurgery were secondary to the use of monopolar coagulation. Diagnosis of these gastrointestinal injuries was made during surgery in only 20 patients (35.7%). The mean time before diagnosis was 4.0 +/- 5.4 (range 0-23) days. Treatment of these complications was performed by laparoscopic surgery in 16.1% of cases. Prevention relies on the surgeon's experience, strict observance of the safety rules, perfect familiarity with the physical properties of the instruments used, systematic use of bowel preparation for patients presenting a risk of bowel complications, systematic supervision of the route taken by the trocars, meticulous inspection on completion of surgery of all areas where bowel adhesiolysis has been used and, in case of any doubt, tests for leakage involving the rectosigmoid. For patients with a risk of bowel complications, the creation of a pneumoperitoneum and performing a mini laparoscopy in the left hypochondrium can be the judicious option.

Operative management of deep endometriosis infiltrating the uterosacral ligaments.

STUDY OBJECTIVE: To describe and assess the efficacy of laparoscopic surgical treatment for patients with pain and deep endometriosis located on the uterosacral ligaments. DESIGN: Retrospective analysis (Canadian Task Force classification II-2). SETTING: University-affiliated hospital. PATIENTS: One hundred ten consecutive women with deep endometriosis infiltrating uterosacral ligaments. INTERVENTION: Operative laparoscopic management of endometriosis. MEASUREMENTS AND MAIN RESULTS: Improvement was reported in 82.3% (70/85) of patients with severe dysmenorrhea and was considered satisfactory in 82.8% (58/70). Improvement also occurred in 88.2% (75/85) of women with deep dyspareunia, and was considered satisfactory in 88.0% (66/75). CONCLUSION: Provided the surgeon is highly skilled in laparoscopy, operative laparoscopy is efficient for the treatment of painful symptoms related to deep endometriosis infiltrating uterosacral ligaments. (J Am Assoc Gynecol Laparosc 6(1):31-37, 1999)

Expression of biological activity of draculin, the anticoagulant factor from vampire bat saliva, is strictly dependent on the appropriate glycosylation of the native molecule.

Draculin, a glycoprotein isolated from vampire bat (Desmodus rotundus) saliva, is a natural anticoagulant which inhibits activated coagulation factors IX (IXa) and X (Xa). The observation that under captivity conditions, the anticoagulant activity present in vampire bat saliva is dependent upon the salivation protocol, led us to investigate the possible relationship between the expression of biological activity of native draculin and the post-translational glycosylation of the protein backbone. Daily salivation of vampire bats yields a saliva that progressively decreases in anticoagulant activity, without any significant change in overall protein content, or in the amount of protein specifically recognized by a polyclonal anti-draculin antibody. Anticoagulant activity of the saliva is restored after a 4-day period of rest. Besides the marked difference in anticoagulant activity, purified native draculin, obtained from high- and low-activity saliva, shows significant differences in: (a) composition of the carbohydrate moiety, and (b) Glycosylation pattern. Furthermore, controlled chemical deglycosylation of native draculin, under conditions that do not affect the polypeptide backbone, progressively leads to complete loss of the biological activity. Our present results implicate that correct glycosylation of draculin is a seminal event for the expression of the biological activity of this glycoprotein.

Peritoneal endometriosis and "endometriotic" nodules of the rectovaginal septum are two different entities.

OBJECTIVE: To compare histologically and stereologically the endometriotic nodule of the rectovaginal septum to peritoneal endometriosis. DESIGN: Morphometric investigation, cytokeratin and vimentin content, and steroid receptor evaluation were performed on endometriotic tissue from the peritoneum (n = 52) and rectovaginal nodules (n = 68). SETTING: An academic teaching hospital. PATIENTS: Biopsies were taken from 120 patients undergoing a laparoscopy for infertility and/ or pelvic pain (52 from typical black peritoneal endometriotic implants and 68 from endometriotic nodule of the rectovaginal septum). None of the patients were treated. RESULTS: Mitotic activity was found to be significantly different in peritoneal and rectovaginal endometriosis. The evaluation suggested that the stroma is not mandatory for the invasion of glandular epithelium in the rectovaginal nodule, which is, like a adenomyoma, a circumscribed nodular aggregate of smooth muscle and glandular elements. Cytokeratin and vimentin content as well as the estrogen receptor (ER) and P receptor (PR) content were significantly lower in both types of lesion when compared with eutopic endometrium. But vimentin immunoreactivity in epithelium, as well as the ER and PR content, were significantly lower in nodules when compared with black peritoneal lesions. CONCLUSION: It is suggested that the rectovaginal endometriotic nodule is a different disease from peritoneal endometriosis and must be called rectovaginal adenomyosis or rectovaginal adenomyoma. Its histopathogenesis probably is not related to the implantation of regurgitated endometrial cells but to the metaplasia of Müllerian rests.

Purification and partial characterization of draculin, the anticoagulant factor present in the saliva of vampire bats (Desmodus rotundus).

From the saliva of the vampire bat Desmodus rotundus, we isolated an unknown anticoagulant protein which we have named draculin. Its molecular mass as determined by non-reduced SDS-PAGE is about 83 kDa. The reduced polypeptide shows a slower migration. HPLC in a molecular sieve matrix yields a single, symmetrical peak corresponding to 88.5 kDa. Isoelectric focusing shows an acidic protein with pI = 4.1-4.2. Aminoacid analysis is compatible with a single chain polypeptide of about 80 kDa. Cyanogen bromide cleavage yields a single 16-aminoacid peptide, corresponding to the amino-terminus of the native molecule. Draculin inhibits the activated form of coagulation factors IX and X. It does not act on thrombin, trypsin, chymotrypsin and does not express fibrinolytic activity. The inhibition is immediate and not readily reversible, with a stoichiometry of about two molecules of draculin per molecule of factor IXa or Xa. Surprisingly, the inhibitory activity against either factor is not affected by the presence of the other. Draculin binds quantitatively to either immobilised factor Xa or factor IXa. Our preliminary interpretation is that there are two forms of draculin that hardly differ in structure. Both bind to factor Xa and to factor IXa but one form inhibits factor Xa and the other inhibits factor IXa. When added to plasma, draculin increases the lag phase as well as the height of the peak of thrombin generation.

Absence of transplacental passage of pentosan polysulfate during mid trimester of pregnancy.

Eight women who were going to have an abortion between the 18th and 23 week of gestation for chromosomal abnormalities or haemoglobinopathies received intravenously 50 mg of pentosan polysulfate (PSP). Maternal results of haemostasis prior and after the injection of the drug were compared. Fetal coagulation parameters were tested on samples obtained by direct puncture of the umbilical cord under ultrasound guidance, 30 min after injection. Results were compared to those of normal fetuses at the same stage of gestation, obtained in the same conditions. In mothers' plasma, 30 min after injection, APTT was prolonged, factor Xa generation was markedly impaired, and factor V level was deeply decreased. By contrast, no modifications of these parameters were observed in fetal plasma, 30 min after the injection of PSP to their related mothers when compared to control fetuses. Thus the absence of biological modifications induced by PSP injection could demonstrate that this drug does not cross through the placenta.

Comparison between the effect of pentosan polysulphate heparin and antithrombin III injections in antithrombin III deficient patients.

Pentosan polysulphate is an heparin analogue which acts via an antithrombin III (AT III) independent pathway. We compared the effect of this drug to that of heparin and AT III infusions in AT III deficient patients. Four patients with AT III congenital deficiency received on three different occasions: (i) an infusion of human AT III concentrate (20 U/kg or 40 U/kg), (ii) an intramuscular injection of pentosan polysulphate (2 mg/kg), (iii) a subcutaneous calcium heparin injection (100 U/kg). AT III infusion inhibits the excessive thrombin generation (46% of inhibition) observed in the plasma of AT III deficient patients during at least 12 hours, but does not modify the factor Xa formation. On the contrary, pentosan polysulphate has a marked effect on both thrombin (62% of inhibition) and factor Xa generation (57% of inhibition) still present 8 hours after injection. Heparin injection has the same effect, more prolonged, as pentosan polysulphate on thrombin generation but is not so effective on impairing factor Xa generation (27% of inhibition). The marked effect of pentosan polysulphate on thrombin and factor Xa generation in these patients is due to its AT III independent mechanism of action.

Chromatic adaptation in a mutant of Fremyella diplosiphon incapable of phycoerythrin synthesis.

A mutant of the chromatically adapting cyanobacterium Fremyella diplosiphon, incapable of phycoerythrin synthesis but responding to wavelength modulation of its biliprotein content, was isolated. The biliprotein composition of the mutant and of the wild type were identical after growth in red light, but green light induced, in the mutant, the synthesis of a biliviolin-type chromophore bound to some of the alpha subunits of its phycocyanin. Implications of the results on the regulation and possible pathways of biliprotein biosynthesis are discussed.

[Prevention of thrombosis by Hemoclar. Preliminary results of a test in a patient with antithrombin III deficiency]. / Prévention des thromboses par l'Hémoclar. Résultats préliminaires d'un essai chez une malade atteinte de déficit en antithrombine III.

We report here preliminary results of a study comparing the effect of Hemoclar and antithrombin III (AT III) in a patient with an AT III congenital deficiency. Both drugs inhibit thrombin generation in the patient's plasma. Unlike AT III, Hemoclar also inhibits factor Xa generation. This led us to propose the use of Hemoclar, which acts via an AT III independent pathway, to prevent thrombosis in A III deficient patients.

Tissue factor activity of rabbit peritoneal macrophages: influence of immune stimulation.

Rabbit peritoneal macrophages exhibit a potent tissue factor activity able to trigger the extrinsic pathway of blood coagulation. This activity is elicited by the inflammatory reaction. Local injection of antigen in immunized animals leads to a lowering of the macrophage procoagulant activity. The mechanism of this phenomenon was investigated in cell culture. After a 24 hour culture, sticking macrophages as well as the culture supernatant exhibit a potent tissue factor activity, both closely related to the initial cell concentration. When the cells are collected from immunized rabbits the presence of the antigen in culture medium is able to significantly enhance the procoagulant activity of the supernatant. These data allow to propose a mechanism for the "macrophage disappearance reaction" and some related phenomena.

Physico-chemical and immunological properties of allophycocyanins.

Allophycocyanins were purified from diverse cyanobacteria and one rhodophytan alga (Cyanidium caldarium). The native proteins are trimeric molecules with the structure (alpha beta)3. Representative native allophycocyanins and their alpha and beta subunits were characterized with respect to molecular weight, amino acid composition, isoelectric point, absorption and fluorescence spectra and immunological properties. All of the allophycocyanins studied were strikingly similar with respect to each of these properties. Renatured alpha and beta subunits of allophycocyanin were distinct immunologically from each other, and both cross-reacted with the antiserum to the native protein. Trimeric allophycocyanin was readily reconstituted from the purified alpha and beta subunits. Formation of hybrid allophycocyanins was demonstrated by direct isolation and characterization of the hybrid proteins and by immunological techniques. The results support the view that allophycocyanins are a highly conserved group of proteins.