Effect of the prostacyclin synthetase inhibitor tranylcypromine on uterine blood flow in pregnancy

Prostacyclin is a potent vasodilator in a number of vascular beds including the uterus. However, the role of prostacyclin in maintaining uterine blood flow during pregnancy is not well established. Recent reports have appeared suggesting that tranylcypromine can selectively inhibit prostacyclin synthesis. Thus, the present study was undertaken using an unanesthetized chronically catheterized pregnant sheep preparation to evaluate the effects of direct intra-arterial infusions of tranylcypromine on the uterine vasculature of late-term pregnant ewes. Infusions of 1, 3 and 10 mg/min of tranylcypromine led to dose-related reduction in uterine blood flow (16, 21 and 47 percent, respectively) and increased blood pressure (7, 10 and 23 percent, respectively). However, these alterations were not associated with reductions in the uterine production rates of the prostacyclin metabolite, 6-keto-PGF_1α, as determined by unextracted plasma RIA. In addition, pre-treatment of animals with the α-adrenergic blocking agent, phenoxybenzamine, almost totally abolished uterine and systemic blood pressure responses to tranylcypromine. These data suggest that tranylcypromine either releases or elevates levels of an alpha adrenergic stimulant which constricts the uterine and systemic vasculature and does not alter prostacyclin levels at the dose tested.     

Effect of bisenoic prostaglandins on the uterine vasculature of the nonpregnant sheep

The effects of the bisenoic prostaglandins on the uterine vasculature and uterine contractile activity have been evaluated in an unanesthetized chronically catheterized nonpregnant sheep preparation. Changes in uterine blood flow were monitored with electromagnetic flow probes while uterine contractile activity and tone were determined via an intra-uterine balloon connected to a pressure transducer. Prostaglandins A₂, D₂, E₂, and prostacyclin (PGI₂) were all found to be vasodilators. PGD₂ and PGI₂ were much more potent than PGA₂ and PGE₂ in dilating the uterine vasculature. The prostacyclin breakdown product 6-keto PGF_1α, PGF_2α, thromboxane B₂, and the endoperoxide analogues U44069 and U46619 produced vasoconstriction of the uterine vasculature. Prostaglandins A₂, D₂ and F_2α increased while PGI₂ decreased uterine contractile activity. PGF_2α also increased uterine tone suggesting that a portion of its vasoconstrictor activity may be due to mechanical compression of the uterine vasculature.     

Equilibrium and structural studies of silicon(IV) and aluminium(III) in aqueous solution. 12. A potentiometric and 29Si-NMR study of silicon tropolonates

Complexation in the H⁺-Si(OH)₄-tropolone (HL) system was studied in 0.6 M (Na)Cl medium at 25° C. Speciation and formation constants were determined from potentiometric (glass electrode) and ²⁹Si-NMR data. Experimental data cover the ranges 1.5 ? - log[H⁺] ? 8.4, 0.002 ? B ? 0.012 M, and 0 ? C ? 0.060 M (B and C stand for the total concentration of Si and tropolone, respectively). In acid solutions (-log[H⁺] ? 3) a hexacoordinated cationic complex, SiL₃⁺, is formed with log K(Si(OH)₄ + 3HL + H⁺ XXX SiL₃⁺ + 4H₂O) = 7.08 ± 0.03. Furthermore, the formation of a disilicic acid was established from ²⁹Si-NMR data. The dimerization constant of Si(OH)₄ was found to be 10 exp (1.2 ± 0.1). In model calculations the solubility of quartz and amorphous SiO₂ in the presence of tropolone is demonstrated. Data were analyzed using the least-squares computer program LETAGROPVRID.     

Leukotrienes and myometrial activity of the term pregnant uterus

Biopsies from different segments of the pregnant human uterus were superfused in organ chambers and contractile activity was registered. Leukotriene C₄(LTC₄) caused inhibition of spontaneous but not noradrenaline induced contractile activity in strips from the cervix. This effect occured both in early pregnancy and at term. However, the lower and the upper uterine segment of the term pregnant uterus did not respond to LTC₄. The results represent a documentation of the segmental differentiation in the uterine response to eicosanoids.     

Copper(I) complexes of penicillamine and glutathione

Equilibrium analysis of a model system for the in vivo reactions between penicillamine and Cu(I), the penicillamine-glutathione-Cu(I) system, indicates that in a certain concentration range the use of penicillamine as a drug will not disturb the normal Cu(I) metabolism. The equilibrium data required for this analysis were obtained by emf titrations on the Cu(I)-glutathione (H3A) and the Cu(I)-pencillamine (H2A) systems at 25 degrees C. in 0.5 M NaClO4 medium, using glass and copper amalgam electrodes; the data were analyzed first by various graphical methods and then by a general least squares computer program. The results show that mononuclear Cu(I) species Cu(HA)2 form in both systems with stability constants log beta 122 of 38.8 (glutathione) and 39.18 (penicillamine); in addition, the polynuclear Cu5A43- species forms in the penicillamine system and the mononuclear CuHA- species might form in the glutathione system. The results are discussed in relation to the therapeutic use of penicillamine as well as in relation to the toxic action of copper on living cells.     

The effect of prostaglandin I2 on the contractility of the term pregnant human myometrium

Small myometrial strips were dissected from the upper and lower segments of the term pregnant human uterus. The specimens were superfused in organ chambers and contractile activity was recorded isometrically.In strips from the upper segment, prostacyclin (PGI₂), induced an initial excitatory response followed in the majority of experiments by transient inhibition. In the lower segment the response was generally the same although direct inhibition without initial stimulation occurred in some cases.During the period of inhibition the specimens were refractory to iterated exposure to PGI₂. Furthermore, during this period of PGI₂-induced inhibition the muscle strip was also refractory to PGE₂ but responded to PGF_2α and oxytocin by stimulation.After inhibition of spontaneous contractile activity induced by indomethacin PGI₂ induced an excitatory response.The results do not indicate any critical change in the myometrial responsiveness of the upper uterine segment to PGI₂ during labor. In strips from the lower segment obtained before labor there tended to be a dominance of non-responders and inhibition only as compared to the results during labor. Nevertheless, whether or not PGI₂ under physiological or pharmacological conditions has any significant influence on the contractility of the term pregnant human uterus, still remains obscure.As judged from earlier reports from our laboratory and the present study it is evident that the uterine vessels are considerably more sensitive to the action of PGI₂ than the myometrium.     

Simultaneous measurements of testosterone and three 5a -reduced androgens in the venous effluent of immature rat testes in situ

Simultaneous measurements were made by radioimmunoassay of testosterone, 17β-hydroxy-5 a -androstan-3-one (DHT), 5 a -androstane-3 a, 17 β -diol (3a-diol), and 5 a-androstane-3 β, 17β-diol (3β-diol) in the testicular venous plasma (TVP) and peripheral plasma (PP) of 30, 45, and 55 day old rats. At 30 days of age, the preponderant androgen in both plasmas was 3a-diol but testosterone predominated by day 55. Testosterone levels increased with age in both TVP (6.39, 15.08, and 54.93 ng/ml on days 30, 45, and 55 respectively) and PP (0.13, 0.56, and 1.02 ng/ml on days 30, 45 and 55 respectively) whereas 3a-diol concentrations decreased in TVP (48.07 ng/ml, day 30; 24.85 ng/ml, day 55) though not peripherally (range: 0.41–0.52 ng/ml). DHT was low in both TVP and PP and appeared to rise only slightly although the increase was not statistically significant. Levels of 3β-diol remained low and unchanged. These observations suggest that the total androgen content of the venous effluent from the prepubertal rat test is is quite high and that significant changes in peripheral interconversions of androgens are occurring during sexual maturation.     

Radioimmunoassay of serum and ovarian pregnenolone in immature rats

Antiserum was generated in a rabbit against pregnenolone-16α-car-boxyethyl thioether conjugated to bovine serum albumin. The antibody, used for the assay of pregnenolone in extracts of serum and tissue homogenates, proved sufficiently specific to allow direct assay of extracts without chromatography. Sensitivity, defined as that point on the standard inhibition curve equal to the lower value 2 SD from the mean radioactivity bound to antibody in the buffer control (zero hormone) tubes, was 0.02 ng. The accuracy was confirmed by the recovery of known amounts of pregnenolone added to serum and aqueous buffer medium. The inter- and intra-assay coefficients of variation were respectively 10.6% and 9.1%. Specificity was confirmed by checking cross reactivity of various steroids and by finding comparable pregnenolone levels in samples before and after chromatography. The administration of aminoglutethimide to rats resulted in the reduction of ovarian and serum pregnenolone levels. Administration of isoxazole, on the other hand, caused an increase in both serum and tissue pregnenolone levels. These findings are in agreement with accepted views of the action of these two metabolic blocking agents.     

Deep hypothermia and circulatory arrest: effect on cerebrospinal fluid electrolytes in newborn lambs.

Cerebrospinal fluid (CSF) Na, K, and acid-base changes were studied in 13 new-born lambs anesthetized with α-chloralose (60 mg/kg) or diethylether during 90 min of normothermic (37 °C) or hypothermic (20 °C) circulatory arrest. CSF K concentration increased linearly from 3.1 to 23.2 meq/liter during 90 min of normothermic circulatory arrest. During hypothermic circulatory arrest, animals anesthetized with α-chloralose exhibited an exponential increase in CSF K concentration from 3.1 to 13.6 meq/liter and animals anesthetized with diethylether had an exponential increase in CSF K concentration from 3.3 to 12.7 meq/liter. The rate of increase in CSF K concentration in hypothermic and normothermic animals between 60 and 90 min of circulatory arrest was the same. CSF Na concentration decreased slightly in both hypothermic and normothermic animals, with a greater decrease in the normothermic group.Although CSF pH and bicarbonate were significantly decreased during normothermia as well as hypothermia, both CSF pH and bicarbonate showed greater decreases during normothermia. Mean pH values after 90 min of circulatory arrest were 6.34, 6.87, and 6.77, respectively, in the normothermic, α-chloralose-hypothermic, and diethylether-hypothermic groups; corresponding values for bicarbonate were 7.7, 13.8, and 12.2 meq/liter.CSF pCO₂ increased linearly from 40.2 to 190.0 Torr during 90 min of normothermic circulatory arrest, from 28.6 to 92 Torr in the ether-hypothermic group, and from 28 to 81 Torr in the α-chloralose-hypothermic group.     

Prostaglandins, not cyclic GMP, inhibit oxytocinase isoenzymes from human amniotic fluid in vitro

Two isoenzymes of oxytocinase (EC 3.4.11.3) activity were fractionated from human amniotic fluid samples between the 14th and 22nd weeks of gestation by Ultrogel acrylamide-agarose gel filtration and partially characterized. The isoenzymes were competitively inhibited by PGE₁, PGE₂ and PGF_2α more at pH 6.2 than at pH 6.8, whereas cyclic GMP (cGMP) and its 8-bromo derivative had no effect at either pH. The implications of these findings are discussed and it is suggested that since the activity of amniotic fluid oxytocinases is very low or minimal at or near term, inhibition of these by prostaglandins may not have physiological significance in the initiation of human parturition.     

Effectiveness of prostagland in F2α in restoration of HMG-HCG induced ovulation in indomethacin-treated rhesus monkeys

Six mature female rhesus monkeys were treated with HMG-HCG in control cycles at doses adjusted to induce ovulation while avoiding superovulation. Occurrence of ovulation was determined by observation of fresh ovulation points at laparotomy 48 to 120 hours following HCG. In subsequent cycles animals were treated with indomethacin (treatment days 4 through 10) together with the established dose of HMG-HCG. In 8 cycles indomethacin 5 mg/kg was given i.m. once daily; in 9 cycles 10 mg/kg i.m. was administered in 2 divided doses. Following this, PGF₂α (3 mg t.i.d. s.c.) was administered for 3 days together with indomethacin 10 mg/kg and HMG-HCG, beginning on the day prior to HCG. Determinations of progesterone were performed by RIA on treatment days 4, 7, 10, and 11. Eleven of the 13 control cycles were ovulatory. With indomethacin 5 mg/kg/day, 5 of 8 cycles were ovulatory but ovulation was delayed in 2 instances. Of 9 cycles using indomethacin 10 mg/kg/day only 1 was ovulatory. When PGF₂α was administered in subsequent cycles along with indomethacin (10 mg/kg) and HMG-HCG, ovulation occurred in 13 of 19 cycles. These data suggest that local ovarian PGF₂α may be essential in the mechanics of follicle rupture in gonadotropin-treated rhesus monkeys.     

Chlamydia trachomatis,Chlamydial Heat Shock Protein 60 and Anti-Chlamydial Antibodies in Women with Epithelial Ovarian Tumors

OBJECTIVE: Chlamydia trachomatis (C. trachomatis) infection has been suggested to promote epithelial ovarian cancer (EOC) development. This study sought to explore the presence of C. trachomatis DNA and chlamydial heat shock protein 60 (chsp60) in ovarian tissue, as well as anti-chlamydial IgG antibodies in plasma, in relation to subtypes of EOC. METHODS: This cross-sectional cohort consisted of 69 women who underwent surgery due to suspected ovarian pathology. Ovarian tissue and corresponding blood samples were collected at the time of diagnosis. In ovarian tumor tissue, p53, p16, Ki67 and chsp60 were analyzed immunohistochemically, and PCR was used to detect C. trachomatis DNA. Plasma C. trachomatis IgG and cHSP60 IgG were analyzed with a commercial MIF-test and ELISA, respectively. RESULTS: Eight out of 69 women had C. trachomatis DNA in their ovarian tissue, all were invasive ovarian cancer cases (16.7% of invasive EOC). The prevalence of the chsp60 protein, C. trachomatis IgG and cHSP60 IgG in HGSC, compared to other ovarian tumors, was 56.0% vs. 37.2% P = .13, 15.4% vs. 9.3% P = .46 and 63.6% vs. 45.5% P = .33 respectively. None of the markers of C. trachomatis infection were associated with p53, p16 or Ki67. CONCLUSIONS: C. trachomatis was detected in invasive ovarian cancer, supporting a possible role in carcinogenesis of EOC. However, there were no statistically significant associations of chsp60 in ovarian tissue, or plasma anti-chlamydial IgG antibodies, with any of the subtypes of ovarian tumors.     

Alterations in uterine and placental prostaglandin F and E with gestational age in the rat

Experiments were designed to determine the chronological alterations in placental and uterine prostaglandin F and E (PGF and PGE) during pregnancy in the rat. Pregnant rats (sperm in the vagina = day 0) were sacrified at days 15, 18,19, 20, 21 and delivery (day 21 ) and placental and uterine tissues assayed (RIA) for PGF and PGE immediately (“ ”) or after 1 hour incubation (“ ”). Uterine content of PGF and PGE (ng PG/mg DNA) was increased significantly by day 19 and further increases were seen through delivery. Incubation of uterine tissue resulted in enhanced net production of PGF and PGE (p <.05) per mg DNA (as judged by tissue content and release into the incubation medium) by day 18 of pregnancy vs. day 15. Net production peaked around the time of delivery thus paralleling the alterations in tissue content .By contrast, no differences with gestational age were found in placental content of PGF and PGE , the concentrations throughout late gestation remaining in the range of uterine PGs at day 15. However, production of PGs per mg placental DNA increased markedly during incubation with significant enhancement detected by day 19 vs. 15, achieving levels even greater than the uterus .The and findings for the uterus are consistent with the hypothesis that increases in uterine PGs levels at the end of pregnancy may play an important role in parturition. The experiences with placental tissue suggest that the potential for PG production per placental cell may also increase in late gestation and thereby contribute to the augmented intrauterine availability of PGs at that time.     

Expression Analysis of Platinum Sensitive and Resistant Epithelial Ovarian Cancer Patient Samples Reveals New Candidates for Targeted Therapies

Ovarian cancer has the highest mortality rate of all gynecologic malignancies. Identification of new biomarkers is highly needed due to its late diagnosis and high recurrence rate. The objective of this study was to identify mechanisms of therapy resistance and potential biomarkers by analyzing mRNA and protein expression from samples derived from patients with platinum-sensitive and -resistant ovarian cancer (total cohort n?=?53). The data revealed new candidates for targeted therapies, such as GREB1 and ROR2. We showed that the development of platinum resistance correlated with upregulation of ROR2, whereas GREB1 was downregulated. Moreover, we demonstrated that high levels of ROR2 in platinum-resistant samples were associated with upregulation of Wnt5a, STAT3 and NF-kB levels, suggesting that a crosstalk between the non-canonical Wnt5a-ROR2 and STAT3/NF-kB signaling pathways. Upregulation of ROR2, Wnt5a, STAT3 and NF-kB was further detected in a platinum-resistant cell-line model. The results of the present study provided insight into molecular mechanisms associated with platinum resistance that could be further investigated to improve treatment strategies in this clinically challenging gynecological cancer.     

The copper-enzyme dopamine beta-monooxygenase: studies on the ability of several metals to inhibit the enzyme activity and to replace the copper

The ability of several metals to inhibit dopamine beta-monooxygenase was measured and compared with their ability to compete with the binding of 64Cu to the water-soluble form of the bovine adrenal enzyme at pH 6.0. In the presence of an optimal concentration of copper (0.5 microM in the present assay system), an inhibition was observed upon addition of Hg(II), Zn(II), or Ni(II). Only a small fraction of the inhibition with these metals may be due to uncoupling of electron transport from hydroxylation. Preincubation of these metals with the Cu-depleted apoenzyme before addition of copper, revealed a stronger inhibition than if copper was added before the other metals. Hg(II), Zn(II), and Ni(II) also compete with the binding of 64Cu(II) to the protein. Hg(II) was the most effective and Ni(II) the least effective of these metals, both with respect to inhibition of the enzyme activity and to prevent the binding of 64Cu(II). Competition experiments on the binding of Zn(II) and 64Cu in the presence and absence of ascorbate, indicated i) a similar affinity of Cu(I) and Cu(II) to the native enzyme, and ii) a more rapid binding of Cu(I) than Cu(II) to the Cu-depleted and Zn-containing enzyme. Al(III), Fe(II), Mg(II), Mn(II), Co(II), Cd(II), and Pb(II) neither inhibited the enzyme activity nor competed with the binding of 64Cu(II) to the protein (Fe(II) was not tested for binding). Of those metals cited above only Cu(II)/Cu(I) was able to reactivate the apoenzyme.     

Changes in inducibility of ornithine decarboxylase activity in differentiating human neuroblastoma cells

Human SH-SY5Y neuroblastoma cells could be induced to differentiate morphologically and biochemically in the presence of 12-O-tetradecanoylphorbol-13-acetate (TPA), retinoic acid (RA), or a combination of these two substances. The phenotypical changes induced by these substances differed, but one effect of both was an inhibition of the cell growth. Addition of TPA or RA to non-treated cells had no effect on the activation of ornithine decarboxylase (ODC, EC 4.1.1.17.), while a change to fresh medium stimulated the ODC to maximum activity after 4-6 h. The activity was not altered by the presence of RA in the fresh medium, but TPA partially inhibited the medium-stimulated ODC activity. Cells treated for 4 or 8 days with TPA or a combination of TPA and RA had a low ODC activity which could not be induced by fresh medium. However, RA-treated (and thus growth-inhibited) cells still responded to a change of medium by exhibiting an ODC activity of the same magnitude and duration as in medium-stimulated control cells. The results seem to suggest that the growth inhibition induced by TPA and RA, respectively, is mediated by different mechanisms.     

Localization of calcium in an annelid visceral muscle by pyroantimonate deposition and by X-ray microprobe analysis

The loci of calcium distribution in Nereis pharangeal visceral muscle have been examined by cytochemical precipitation using potassium pyroantimonate. In Na-, Ca- and Mg-free media, pyroantimonate incubation was used to pinpoint loci of intracellularly bound calcium. This method also revealed heavy deposition on the inner face of the plasma membrane, in the sarcoplasmic reticulum and nucleus. X-ray microprobe analysis of the precipitate confirmed the presence of calcium and antimony peaks. It is concluded that the plasma membrane may constitute a major calcium pool for the activation of contraction in this muscle.     

Changes in L-ornithine decarboxylase activity during the cell cycle

The activity of L-ornithine decarboxylase (L-ornithine carboxy-lyase; EC 4.1.1.17), the enzyme that catalyzes the initial and rate-limiting step in polyamine biosynthesis, has been studied in Chinese hamster ovary fibroblasts synchronized by selective detachment of mitotic cells. At various times after plating the distribution of cells among the G1, S and G2+M phases of the cell cycle was calculated from DNA distributions obtained by high-speed flow cytometric analysis. At these same times determination of the cellular L-ornithine decarboxylase activity showed that polyamine (putrescine) synthesis was initiated in mid-G1, that the rate of synthesis was maximal prior to DNA synthesis, and that it decreased during the S phase. A second increase in enzyme activity occurred before mitosis.     

A multinuclear nmr relaxation study of the interaction of divalent metal ions with l-aspartic acid

Carbon-13 spin-lattice relaxation times, T₁, have been measured for aqueous solutions of L-aspartic acid, L-alanine, O-phospho-L-serine, and 2-mercapto-L-succinic acid in the presence of the paramagnetic metal ions, Cu²⁺ and Mn²⁺, and Mg²⁺ as a diamagnetic control, at ambient temperature and neutral pH. Nitrogen-15, oxygen-17 and proton relaxation times were also obtained for L-aspartic acid and phosphorus-31 relaxation times for O-phospho-L-serine under similar conditions. The structures of these complexes in solution were determined from the various metal ion-nuclei distances calculated from the paramagaetically-induced relaxation. These results indicate that the Cu²⁺ interaction with L-aspartic acid is through α-amino and β-carboxyl groups while Mn²⁺ coordinates most strongly through α-and β-carboxyl groups, with the possibility of a weak interaction through the amino group.An examination of the coordination of these divalent metal ions to an analog of L-aspartic acid in which the β-carboxyl group is replaced by a phosphate group (O-phospho-L-serine) indicated that Cu²⁺ coordination is now probably through the α-amino and phosphate groups, while this analog is a monodentate ligand for Mn²⁺ coordinating through the phosphate group. Removal of the β-carboxyl group (L-alanine) also results in Cu²⁺ coordination through the α-carboxyl and α-amino groups, and the same ligand interactions are observed with Mn²⁺. Replacement of the α-amino group of L-aspartic acid with an - SH group (2-mercapto-L-succinate) is sufficient to eliminate any specific coordination with either Cu²⁺ or Mn²⁺.     

Triplet state properties of the methylmercury(II)-tyrosine complex

CH₃Hg(II)OH forms complexes at pH 8 with tyrosine and with tyrosine ethyl ester (TEE) that are detected by ultraviolet difference absorption spectra. With K_f defined by CH₃HgOH + HB

CH₃HgB + H₂O, we find log K_f = 3.61 (tyrosine) and 3.36 (TEE). A heavy-atom effect is observed in frozen glasses of the complexes; this indicates a close interaction between Hg and the chromophore. No UV difference spectrum or heavy-atom effect is observed with N-acetyl tyrosine ethyl ester, indicating that complexing at the phenol O does not occur, and suggesting that binding occurs at the amine N. Zero field optically detected magnetic resonance (ODMR) measurements of the CH₃Hg(II)-tyrosine triplet state give (D, E) = (0.129, 0.047) or (0.134, 0.041) cm^?1 depending upon assignment of transitions. D of tyrosine is relatively unaffected, but E is reduced by CH₃Hg(II) complexing. Low-temperature kinetic measurements show that the shortest lived sublevel of the complex is T_z, where z lies along the phenol long axis in tyrosine. A dominant 11.6-msec component in the 77 K decay of the phosphorescence is consistent with the individual sublevel lifetimes obtained by ODMR.