The re-incarnation of an old question--clomid effect on oocyte and embryo?
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Abortion, Spontaneous↗
Biomedical subjects
P Berkowitz
Publications and source records attributed to P Berkowitz.
After slow start, project to channel drug company funds to universities builds steam.
Drug companies appear to have been listening when researchers began complaining about their lack of participation in the MRC/PMAC Health Program, for the 30-month-old project appears to have taken on new life. It is designed to increase collaboration between university and pharmaceutical industry researchers by directing more of the industry's growing investment in Canada through the MRC's peer-review process. By mid-May, program commitments stood at $60 million.
Canada↗
The correlation of ankle oscillometric blood pressures and segmental pulse volumes to Doppler systolic pressures in arterial occlusive disease.
PURPOSE: This study was designed to evaluate the accuracy and failure rates of automatically collected oscillometric ankle systolic pressures (Psys) and pulse volumes (Pvol) using a new algorithm as compared with Psys obtained by standard manual Doppler-and-cuff technique. METHODS: One hundred ten consecutive patients at a vascular laboratory had brachial and ankle Psys measured with the two methods. Pvol at or near the mean arterial pressure was also obtained automatically by the oscillometric device. RESULTS: Both methods showed a 6.6% failure rate when measuring Psys at the ankle. Oscillometric Psys measurement was possible when Doppler Psys failed as a result of nonoccluding arteries. No difference was found between the two methods in occluding limbs with ankle-brachial indexes of 1.30 or more. Sequential brachial Psys values had a mean difference (Doppler-oscillometric) or 2 +/- 10.9 mm Hg and a correlation coefficient (r) of 0.92. Measurements at the ankle had a mean difference of -8.4 +/- 16.8 mm Hg and r = 0.90. These differences were not statistically significant. Mean arterial pressure Pvol recorded at the ankle also correlated with ankle Doppler Psys (r = 0.71) and showed a 1.9% failure rate. CONCLUSION: Both automatic oscillometric plethysmographic Psys and Pvol at the ankle are shown to correlate well with Doppler-and-cuff Psys in patients with vascular disease. Oscillometric measurements can replace Doppler measurements in most clinical situations.
Adult↗
A metal ion-binding site in the kringle region of bovine prothrombin fragment 1.
45Ca(II) binding studies (equilibrium dialysis) on the kringle domain of bovine prothrombin fragment 1 were conducted using a mixture of peptides (residues 43-156 and 46-156) resulting from limited alpha-chymotryptic hydrolysis of fragment 1. Analysis of the Scatchard plot of these data indicates a single, low affinity Ca(II)-binding site to be present. Similar results were obtained from studies on the decarboxylated fragment 1 derivative, 10-gamma-MGlu-fragment 1. Acetylation of bovine fragment 1 in the absence of Ca(II) or Mg(II) ions results in the loss of the metal ion-promoted quenching of the intrinsic Trp fluorescence of the protein and the Ca(II)-mediated binding to phosphatidylserine/phosphatidylcholine (PS/PC) vesicles. The acetylation of the NH2 alpha-group of Ala-1 has been shown (Welsch, D. J., and Nelsestuen, G. L. (1988) Biochemistry 27, 4946-4952) to abolish the PS/PC binding property of fragment 1. The present study demonstrates that acetylation of a second site possibly Ser-79 or Thr-81 using the conditions described in the preceding paper results in loss of both the fluorescence transition and the Ca(II)-mediated PS/PC binding of the resulting protein derivative. Removal of the O-acetyl group at the Ser-79/Thr-81 site is accomplished by aminolysis with 0.2 M hydroxylamine, pH 10, 50 degrees C; the fluorescence transition is partially restored. PS/PC binding is partially restored if the NH2 alpha-group of Ala-1 is trinitrophenylated but is not restored if the NH2 alpha-group of Ala-1 is acetylated. We conclude that the Ser-79/Thr-81 site may represent a portion of the metal ion-binding site within the kringle domain of fragment 1. Occupancy of this site by a Ca(II) ion appears to be important in the binding of the protein to PS/PC vesicles.
Animals↗
Modifications of bovine prothrombin fragment 1 in the presence and absence of Ca(II) ions. Loss of positive cooperativity in Ca(II) ion binding for the modified proteins.
Chemical modification of bovine prothrombin fragment 1 according to the procedure of D. J. Welsch and G. L. Nelsestuen (1988) [Biochemistry 27, 4946-4952 and ealier papers] provided a series of fragment 1 derivatives in which various nitrogen-containing side chains were N-acetylated and/or N-2,4,6-trinitrophenylated. In addition the des-[Ala-1,Asn-2]- and des-[Ala-1,Asn-2,Lys-3]-fragment 1 derivatives were prepared by limited enzymatic hydrolysis of fragment 1 using cathepsin C and plasmin, respectively. Quantitative studies on the Ca(II) binding of these proteins have been accomplished using 45Ca(II) equilibrium dialysis. Binding of these fragment 1 derivatives to phosphatidylserine/phosphatidylcholine (PS/PC) vesicles (25:75) in the presence of Ca(II) ions has been studied using the light-scattering technique. Acylation of the 5 lysine residues of fragment 1 by the action of acetic anhydride (500-fold molar excess) in the presence of 75 mM Ca(II), pH 8.0, results in loss of positive cooperativity in Ca(II) binding (Scatchard plot) and an increase in the number of Ca(II) ions bound. The Ca(II)-dependent PS/PC binding of the acylated protein is reduced. Removal of 2 and 3 residues from the amino terminus likewise leads to loss of positive cooperativity in Ca(II) binding and reduced binding affinity to PS/PC vesicles. The important role of the amino-terminal 1-10 sequence is discussed. We conclude that positive cooperativity in Ca(II) binding is not a prerequisite for the Ca(II)-dependent binding of bovine prothrombin fragment 1 to PS/PC vesicles.
Animals↗
Glomerular capsular herniation in focal glomerular sclerosis.
Renal biopsy from a patient with focal segmental sclerosing glomerulopathy (FSSG) and nephrotic syndrome showed localized rupture of Bowman's capsule with herniation of a glomerular loop into the adjacent periglomerular connective tissue. Another glomerulus contained similar changes. Glomerular capsular herniation may be frequent, but unrecognized, in FSSG and would explain many of the morphological features seen in both primary and secondary forms.
Adult↗
Determination of strontium binding to macromolecules.
An equilibrium dialysis technique for determining the binding of strontium to macromolecules is described. The major difficulty to be overcome is that 90Sr has a decay product, 90Y, which is also a beta-emitter. The described protocol is used to determine the Sr binding isotherm to bovine prothrombin fragment 1. The binding is found to be cooperative, somewhat weaker than Ca binding, and to involve approximately nine strontium sites. The stoichiometric equilibrium constants are determined by nonlinear regression. The procedure should be of great utility for many macromolecules that show strontium affinity.
Animals↗
Chemical modification of bovine prothrombin fragment 1 in the presence of Tb3+ ions. Sequence studies on 3-gamma-MGlu-fragment.
Chemical modification of the gamma-carboxyglutamyl (Gla) residues of bovine prothrombin fragment 1 using the formaldehyde-morpholine method in the presence of 100 Kappm Tb3+ ions at pH 5.0 provided a modified protein containing 3 gamma-methyleneglutamyl residues (gamma-MGlu) and 7 Gla residues (bovine 3-gamma-MGlu-fragment 1). The modified protein bound the same number of Ca2+ ions as the native protein (six to seven), exhibited 28Mg2+-binding properties identical to native fragment 1 (five Mg2+ ions bound), exhibited the metal ion-promoted quenching of the intrinsic fluorescence in a manner similar to the native protein, but did not bind to phosphatidylserine (PS)/phosphatidylcholine (PC) vesicles in the presence of Ca2+ ions. Modification of the bovine protein using [14C]formaldehyde-morpholine provided a 14C-labeled 3-gamma-MGlu-fragment 1 suitable for sequence analysis. Edman sequencing of the peptides released by a tryptic digest of the reduced and carboxymethylated bovine [14C]3-gamma-MGlu-fragment 1 indicated that Gla residues at positions 7, 8, and 33 had been converted to [14C]gamma-methyleneglutamyl residues. In addition Lys97 was found to contain a 14C label. Similar analysis of the human [14C]3-gamma-MGlu-fragment 1 indicated that Gla residues at positions 7 and 32 were major modification sites and that Gla residues at positions 6 and 14 were partially modified. Lysine 96 was also modified in the human protein. The incorporation of a 14C label at Lys97 in bovine 3-gamma-MGlu-fragment 1 protein is not responsible for the loss of Ca2+-promoted binding to PS/PC vesicles. We suggest that Gla residues 7, 8, and 33 are elements of the first Ca2+-binding site; occupancy of this site establishes the Ca2+-specific conformation which is essential for the Ca2+-promoted interaction of the bovine protein with PS/PC vesicles. These studies also suggest that the loss of Gla residues at positions 7 and 32 prevents the formation of the initial Ca2+-binding site in the human protein.
Amino Acid Sequence↗
Relative affinity of Ca(II) and Mg(II) ions for human and bovine prothrombin and fragment 1.
Equilibrium dialysis results are presented for Ca(II) and Mg(II) ion binding to human and bovine prothrombin and fragment 1. Ca(II) ions bind cooperatively, Mg(II) does not.
Animals↗
Mg(II) binding by bovine prothrombin fragment 1 via equilibrium dialysis and the relative roles of Mg(II) and Ca(II) in blood coagulation.
The first direct equilibrium dialysis titration of the blood coagulation protein bovine prothrombin fragment 1 with Mg(II) is presented. Fragment 1 has fewer thermodynamic binding sites for Mg(II) than Ca(II), less overall binding affinity, and significantly less cooperativity. Several nonlinear curve fitting models were tested for describing the binding of fragment 1 with Mg(II), Ca(II), and mixed metal binding data. The Mg(II) data is represented by essentially five equivalent, noninteracting sites; for Ca(II), a model with three tight, cooperative sites and four "loose", equal affinity, noninteracting sites provides the best model. Based on the reported equilibrium dialysis data and in conjunction with other experimental data, a model for the binding of Ca(II) and Mg(II) to bovine prothrombin fragment 1 is proposed. The key difference between the binding of these divalent ions is that Ca(II) apparently causes a specific conformational change reflected by the cooperativity observed in the Ca(II) titration. The binding of Ca(II) ions to the three tight, cooperative sites establishes a conformation that is essential for phospholipid X Ca(II) X protein binding. The filling of the loose sites with Ca(II) ions leads to charge reduction and subsequent phospholipid X Ca(II) X protein complex interaction. Binding of Mg(II) to bovine prothrombin fragment 1 does not yield a complex with the necessary phospholipid-binding conformation. However, Mg(II) is apparently capable of stabilizing the Ca(II) conformation as is observed in the mixed metal ion binding data and the synergism in thrombin formation.
Animals↗
Determination of magnesium binding to macromolecules.
An equilibrium dialysis technique for examining magnesium binding to macromolecules is described. The technique is used to determine the binding constants of magnesium to human prothrombin. This procedure should be of great utility for many biochemical systems which exhibit magnesium affinity.
Dialysis↗
Chemical modification of bovine prothrombin fragment 1 in the presence of Tb3+ ions.
The formaldehyde-morpholine method for the conversion of gamma-carboxyglutamyl (Gla) residues to gamma-methyleneglutamyl (gamma-MGlu) residues has been applied to the modification of bovine prothrombin fragment 1. In the absence of Tb3+ ions or at Tb3+ ion concentrations of 2 Km app and 25 Km app the action of 10,000-fold molar excess of formaldehyde and morpholine, pH 5.0, converts the 10 Gla residues of the protein into 10 gamma-MGlu residues. Modification of the protein using the same conditions but increasing the Tb3+ concentration to 100 Km app provided a homogeneous protein containing 3 gamma-MGlu and 7 Gla residues, bovine 3 gamma-MGlu-fragment 1. The modified protein binds the same number of Ca2+ ions (6-7) as bovine fragment 1. However, the positive cooperatively associated with Ca2+ binding is abolished and the overall affinity for Ca2+ ions is reduced. Fluorescence titrations of 3 gamma-MGlu-fragment 1 using either Ca2+ or Mg2+ ions indicate that the modified protein retains a fluorescence quenching behavior similar to that of the native protein. The modified protein does not bind to phosphatidylserine/phosphatidylcholine vesicles in the presence of Ca2+ ions. Thus the metal ion-induced fluorescence transition exhibited by the bovine protein appears to be a necessary but not sufficient condition for phospholipid binding.
Amino Acids↗
The effect of divalent metal ions on the electrophoretic mobility of bovine prothrombin and bovine prothrombin fragment 1.
Examination of metal ion-dependent effects on the electrophoretic mobility of bovine prothrombin and fragment 1 provides a useful and sensitive method for investigation of conformational processes in these proteins. Utilization of this method reveals a conformational change in bovine prothrombin and fragment 1 which occurs at low metal ion concentrations. Equilibrium dialysis studies indicate that the metal ion-induced shape change occurs concomitant with binding of a single calcium ion/molecule of prothrombin or fragment 1. Mixed metal electrophoretic mobility studies with Mg2+ and Ca2+ have demonstrated the "synergistic" effect for fragment 1 observed by others. Mixed metal equilibrium dialysis has provided experimental support for this observation and allows us to conclude that two tight Ca2+ sites are not affected by low Mg2+ concentrations and that the third Ca2+ site is also a tight site for Mg2+. Thus, at low Mg2+ concentrations and upon the addition of Ca2+, there are effectively three tight sites; consequently more Ca2+ will bind to the protein at lower total Ca2+ ion concentrations.
Animals↗
Pentoxifylline treatment of moderate to severe chronic occlusive arterial disease.
A pilot study of the effects of pentoxifylline in 19 patients with moderate to severe chronic occlusive arterial disease (COAD) is described. The severity of disease was assessed by the degree of limitation in the walking distance on the flat surface (less than 100 m), the absence of peripheral pulses on palpation, the diminished Doppler tibial/brachial pressure (the ischemic index) at rest, and by contrast arteriography, when available. After a 2-week washout phase, all subjects received pentoxifylline (1200 mg/day) in an open-label manner for a total of 12 weeks. Twelve of the nineteen patients showed a definite increase in exercise tolerance by the end of the study, with a concomitant reduction in ischemic symptoms. All except 3 patients felt they had derived benefit from the medication. In contrast to the clear improvements in walking distance and symptoms, only small effects on noninvasive vascular laboratory measurements were noted. Platelet aggregation, induced by ADP, epinephrine, and collagen, gradually decreased over the study period. Pentoxifylline appears to be useful in the medical management of patients with moderate to severe chronic occlusive arterial disease; future controlled trials in such patients are now justified.
Aged↗
Prostaglandin formation in the isolated human ductus arteriosus, aorta, pulmonary and umbilical arteries.
The prostaglandins comprise a large family of substances that includes primary prostaglandins, prostacyclin and thromboxane, all of which exhibit some vascular activity. The activity of each prostaglandin may be species - and organ - dependent, and the type of prostaglandin produced in a tissue is often dependent on the presence of terminal enzyme systems in that tissue. The prostaglandin endoperoxide PGH2 serves as a common intermediate for the enzymatic production of prostaglandins, thromboxanes and prostacyclin. We have obtained information on the biosynthesis of these compounds by the human ductus arteriosus, aorta, pulmonary and umbilical arteries in vitro. Vascular tissue samples were obtained from two fetuses of 16 to 18 weeks of gestation, two newborns of 26 and 35 weeks of gestation and in nine term infants. The vascular tissue samples were incubated with [1-14C]-arachidonic acid and/or [1-14C]-prostaglandin endoperoxide (PGH2). The study demonstrates the formation of prostaglandins and prostacyclins from all the vascular tissues and the formation of thromboxanes from the umbilical artery. The study implies that the above vessels contain "prostaglandin synthetase" enzymes as early as 16 weeks of gestation.
6-Ketoprostaglandin F1 alpha↗
Intraoperative anticoagulation during arterial reconstructive procedures.
The technique of thromboelastography was used to monitor the intraoperative administration of heparin during vascular surgical procedures. Interpatient response to 70 units per kilogram of heparin varied widely. There was no need, in any patient, for the use of protamine to reverse the effect of heparin. This, coupled with the absence of hemorrhagic complications, indicates that thromboelastography is a useful method for monitoring the administration of heparin during vascular surgical procedures.
Aged↗
Pseudophakic bullous keratopathy. A clinical-pathologic analysis.
Of 226 patients who had an intraocular lens implanted over a three year period, 6.2% developed bullous keratopathy compared with 2.2% of 90 patients who had a standard cataract extraction performed by the same surgeon. An additional 4.0% of the lens implant patients developed peripheral or intermittent corneal edema. The factors related to the higher incidence of this complication in lens implant patients included corneal touch during implantation, prolonged postoperative iritis and, in cases of mild or peripheral corneal edema without bullous keratopathy, recurrent lens dislocation. The most consistent pathologic findings in 10 corneas that required keratoplasty were epithelial edema and loss of corneal endothelium. Means to prevent this complication are discussed.
Aged↗
Synthesis and antitumor activity of 1,2-dihydro-1-(2-deosy-beta-D-erythro-pentofuranosyl)-2-oxo-5-methylpyrazine 4-oxide, a structural analogue of thymidine.
1,2-Dihydro-1-(2-deoxy-beta-D-erythro-pentofuranosyl)-2-oxo-5-methylpyrazine 4-oxide was synthesized by condensation of the silylated pyrazine base with the blocked chloro sugar, followed by removal of the protecting groups. The compound inhibited the growth of leukemia L1210 cells in vitro by 50% at 2 x 10-7 M. At 400 mg/kg/day x 6 it increased the life-span of leukemia L1210 bearing mice by approximately 55%, without apparent toxicity to the host.
Animals↗