Koichi Shirakawa

Two cell lines established from mixed Müllerian tumors of the uterus morphologic, immunocytochemical, and cytogenetic analyses

To clarify the cellular origin and characteristics of malignant mixed müllerian tumor (MMMT), the authors investigated two cell lines (designated as FU‐MMT‐1 and FU‐MMT‐2) established from two patients with heterologous MMMT of the uterus. Both cell lines propagated continuously for 83 and 55 serial passages over 1.5 years, respectively. Morphologically, FU‐MMT‐2 was a mixture of carcinoma cells and sarcoma cells with predominance of carcinoma cells; FU‐MMT‐1 only had a sarcomatous element with distinct rhabdomyoblastic differentiation. Immunocytochemically, the sarcoma cells of each cell line expressed, not only myogenic and mesenchymal antigens (desmin, myoglobin, and vimentin), but also epithelial antigens, including epithelial membrane antigen and keratin. The carcinoma cells in FU‐MMT‐2 were positive for the epithelial antigens and vimentin and negative for desmin and myoglobin. Both lines had abnormal karyotypes; the modal chromosome numbers of FU‐MMT‐1 and FU‐MMT‐2 were 47 and 80, respectively. In addition, FU‐MMT‐1 had trisomy 8, and FU‐MMT‐2 had complex structural abnormalities. When transplanted into nude mice, FU‐MMT‐1 reproduced and maintained the characteristics of the original tumor. These cell lines and xenografts appear to provide a useful system for studying the biologic behavior, cytogenetic features, and histogenesis of MMMT. In conclusion, the presence of epithelial antigens in the sarcomatous and carcinomatous elements seemed to support the hypothesis that both elements are derived from a common stem cell.

Effects of putative dopamine D3 receptor agonists, 7-OH-DPAT, and quinpirole, on yawning, stereotypy, and body temperature in rats

7-OH-DPAT ((+/-)-2-(dipropylamino)-7-hydroxy-1,2,3,4-tetrahydronaphthalene) was recently identified as a dopamine receptor agonist having a > 100-, 1,000- and > 10,000-fold higher affinity for dopamine D3 than for D2, D4 and D1 receptors, respectively. Quinpirole (LY 171555) has also been reported to have a 113-fold greater affinity for dopamine D3 receptors than for D2 receptors. Therefore, we investigated the effects of these putative dopamine D3 receptor agonists on yawning, stereotypy and rectal temperature in rats (N = 424). 7-OH-DPAT and quinpirole administered subcutaneously (SC) at respective low doses of 10-250 micrograms/kg and 25-500 micrograms/kg elicited yawning behavior. The yawning induced by these agents was blocked by spiperone (0.5 mg/kg, SC) and scopolamine (0.5 mg/kg, SC) but was increased by intraperitoneal (IP) administration of pindolol (20 mg/kg). The yawning was also potentiated after treatment with reserpine. 7-OH-DPAT and quinpirole at respective high doses of 0.25 mg/kg (SC) and 0.5 mg/kg (SC) evoked slight stereotypy such as sniffing and licking, and this effect was enhanced by a selective dopamine D1 receptor agonist, SK&F 38393 (1-phenyl-2,3,4,5,-tetrahydro-(1H)-3-benzazepine-7,8-diol). 7-OH-DPAT (0.5 mg/kg, SC) and quinpirole (0.5 mg/kg, SC) decreased, but SK&F 38393 (10 mg/kg, SC) increased body temperature. However, the hyperthermia induced by SK&F 38393 was interestingly enhanced by 7-OH-DPAT and quinpirole.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of nitric oxide on vasorelaxation in human umbilical artery

OBJECTIVE Endothelium-derived relaxing factor (or nitric oxide) is thought to play an important role in control of blood flow in umbilical blood vessels at midgestation compared with term. Previous studies suggest that histamine releases endothelium-derived relaxing factor from umbilical arteries. In this study we intended to clarify the mechanism by which histamine releases endothelium-derived relaxing factor and causes vasorelaxation in human umbilical artery at the midstage (18 to 22 weeks) of gestation. STUDY DESIGN By means of very thin muscle strips that allow rapid diffusional access of applied drugs (in a few seconds), contractile properties of human umbilical artery were examined. Isometric tensions were measured in response to potassium chloride (39 mmol/L) or caffeine and inhibitory effects of histamine, A23187, glyceryl trinitrate, and 8-bromo-cyclic guanosine monophosphate on these contractions were also examined. RESULTS Histamine (0.01 to 0.1 mumol/L) did not inhibit 39 mmol/L K(+)-induced contractions of tissues taken at the terminal (38 to 41 weeks) stage of gestation. However, at midgestation histamine (0.01 to 0.1 mumol/L), A23187 (10 mumol/L), and 8-bromo-cyclic guanosine monophosphate (membrane-permeable analog of cyclic guanosine monophosphate, 0.1 mmol/L) inhibited 39 mmol/L K(+)-induced contractions. The inhibitory effects of histamine were antagonized by mepyramine (an H1 antagonist), L-NG-nitro arginine, methylene blue, and Ca++ depletion of the extracellular space but not by cimetidine (an H2 antagonist). Caffeine produced contractions both in the presence and absence of extracellular Ca++ possibly because of the release of Ca++ from intracellular storage sites. Glyceryl trinitrate and 8-bromo-cyclic guanosine monophosphate reduced the caffeine-induced contractions in Ca(++)-free solution. In addition, 10 mumol/L cyclic guanosine monophosphate did not attenuate the Ca++ sensitivity for contractile elements. CONCLUSION These results suggest that (1) histamine coupled to the histamine H1 receptor increases intracellular Ca++ concentration to stimulate nitric oxide synthase in human umbilical endothelial cells, (2) nitric oxide from endothelial cells activates guanylate cyclase to produce cyclic guanosine monophosphate in the umbilical smooth muscle cells, and (3) cyclic guanosine monophosphate relaxes the umbilical tissues, perhaps as a result of the activation of a Ca++ extrusion system.

Occurrence of yawning and decrease of prolactin levels via stimulation of dopamine D2-receptors after administration of SND 919 in rats.

SummarySND 919 ((S)-2-amino-4,5,6,7-tetrahydro-6-propylamino-benzothiazole) is expected to have a potent and selective dopamine D2-receptor agonistic activity. From this information, the present study was performed to investigate effects of SND 919 on yawning behavior and prolactin secretion in rats. Subcutaneous injections of SND 919 (25–500 gm/kg, s. c.) elicited yawning responses. Its dose-response curve was bell-shaped with maximal effects at a dose of 100 μg/kg. Yawning behavior was also evoked by the putative dopamine autoreceptor agonists, talipexole (6-allyl2-amino-5,6,7,8-tetrahydro-4H-thiazolo [4,5-d]azepine) (BHT 920) (5–100 μg/kg, s. c.) and (+)-3-PPP ((+)-3-(3-hydroxyphenyl)-N-n-propylpiperidine) (5−15 mg/kg, s. c.). The yawning induced by SND 919 (100 μg/kg, s. c.) as well as talipexole (25 μg/kg, s. c.) was inhibited by pretreatment with dopamine D2-receptor antagonists such as spiperone (0.5 mg/kg, i.p.) and YM-09151-2 (cis-N-(1-benzyl-2methylpyrrolidin-3-yl)-5-chloro-2-methoxy-4-methylaminobenzamide) (0.1 mg/kg, i. p.), or the muscarinic receptor antagonist, scopolamine (0.5 mg/kg, i.p.). However, the yawning was not affected by the dopamine D1-receptor antagonist, SCH 23390 (R(+)-8-chloro-2,3,4,5-tetrahydro-3methyl-5-phenyl-1 H-3-benzazepine-7-ol) (0.5 mg/kg, i. p.). Stereotypy such as licking and biting was not observed following the administration of SND 919, talipexole and (+)-3-PPP. Administration of SND 919, talipexole or (+)-3-PPP in respective yawn-inducing doses caused a reduction in both the basal prolactin levels and the a-methyl-p-tyrosine-induced hyperprolactinemia. The results suggest that SND 919 as well as talipexole exerts selective agonistic activities for specific dopamine D2-receptors, which are not related to the occurrence of stereotypy and have a high affinity for dopamine receptor agonists quite similar to that of the pituitary lactotroph dopamine D2-receptors.

Study on Nonspecific Immunity in Pregnant Women: II. Effect of Hormones on Chemiluminescence Response of Peripheral Blood Phagocytes

ABSTRACT: To analyze the mechanisms of increased nonspecific immunity in pregnant women, the effect of various hormones on the phagocytic activity was estimated by a luminol‐dependent chemiluminescence (CL) response during phagocytosing opsonized zymosan. The CL response of whole blood supplemented with exogenous human chorionic gonadotropin (hCG) increased significantly in all the male and female subjects and pregnant women. An approximate two‐ to fourfold increase was observed in comparison with the unsupplemented control in each subject at concentrations ranging from 1 to 1,000 IU/ml after 48 h of incubation (P < 0.05). Progesterone slightly stimulated the CL response in female subjects only, but had no effect on male and pregnant women. Estradiol (E2) did not stimulate the CL response in any subject.

Potentiation by serotonergic inhibition of yawning induced by dopamine receptor agonists in rats

Low doses of the dopamine D2-receptor agonist, B-HT 920 (25 micrograms/kg, SC), and the dopamine D1/D2-receptor agonists, apomorphine (50 micrograms/kg, SC) and piribedil (1 mg/kg, SC), evoked yawning. However, the dopamine D1-receptor agonist, SK&F 38393 (2 mg/kg, SC), failed to induce yawning. The yawning responses induced by B-HT 920, apomorphine or piribedil were markedly increased without eliciting stereotypy by pretreatment with reserpine (5 mg/kg, IP, 24 hr). These yawning responses were also enhanced by p-chlorophenylalanine (PCPA) (300 mg/kg, IP, 72 hr), but not by alpha-methyl-p-tyrosine (300 mg/kg, IP, 6 hr). The yawning induced by B-HT 920 given alone and in combination with reserpine or PCPA was inhibited by spiperone (0.5 mg/kg, IP) or scopolamine (0.5 mg/kg, IP), but not by SCH 23390 (0.5 mg/kg, IP). The present results suggest that yawning is evoked by stimulation of dopamine D2-receptors having a high affinity and consequent muscarinic activation, and that the yawning induced by dopamine receptor agonists is potentiated by decreases in serotonergic neuron activity.

The effect of nitric oxide on uterine and umbilical artery flow velocity waveform in pre-eclampsia

We compared the flow velocity waveforms of uterine and umbilical arteries in normotensive and pre-eclamptic patients at mid-gestation. In a randomised controlled trial we tested the effects of isosorbide dinitrate (ISDN, nitric oxide donor) patch therapy on the flow velocity waveform of pre-eclamptic patients at mid-gestation. The resistance indices (RI) of human uterine and umbilical arteries were higher in pre-eclamptic patients compared to the normotensive patients. ISDN patch therapy significantly reduced the increased RI values of the umbilical artery in pre-eclamptic patients without any change in systemic blood pressures, but the RI values of the uterine artery were not significantly attenuated. The change of the umbilical artery might be due to the improvement of end-diastolic flow velocity. These results suggest that the feto-placental circulation in pre-eclampsia, perhaps due to the disturbance of the endothelium-dependent vaso-relaxation system, and that ISDN therapy may improve the impaired endothelium dependent nitric oxide system.

Potentiation of yawning responses to the dopamine receptor agonists B-HT 920 and SND 919 by pindolol in the rat

Subcutaneous injection of B-HT 920, a dopamine D2-receptor agonist, in doses ranging from 5 to 100μg/kg, induced yawning behavior in rats. Yawning was also elicited by low doses (25–500 μg/kg sc) of SND 919, a newly synthesized dopamine receptor agonist. The yawning evoked by B-HT 920 or SND 919 was increased by the β-adrenoceptor antagonist pindolol (20mg/kg ip) which alone did not induce yawning. Stereotyped behavior did not appear after B-HT 920 or SND 919 given alone or in combination with pindolol. The results suggest that SND 919 as well as B-HT 920 has stimulatory activity at dopamine D2-receptors, and that pindolol may exert its enhancement of the yawning response to dopamine receptor agonists via blockade of β-adrenoceptors.

Some mechanical properties of skinned fibres of pregnant human myometrium

The properties of contractile elements and intracellular Ca2+ storage sites of pregnant human myometrium were studied by recording the mechanical responses in skinned (saponin-treated and membrane-permeable) fibres. Calmodulin increased the amplitude of contractions induced by Ca2+ and the Ca2+ sensitivity for contractile elements in small myometrium strips, but PGF2 alpha, PGE2, oxytocin, or cyclic AMP failed to produce similar effects. After accumulation of Ca2+ in intracellular Ca2+ storage sites, 10 mumol/l PGF2 alpha, 10 mumol/l PGE2, 30 mmol/l caffeine, and 20 mumol/l InsP3 (inositol-trisphosphate) produced contractions by releasing Ca2+ from storage sites. However, 20 nmol/l oxytocin had no effects under the same conditions. The InsP3 sensitive Ca2+ store was much larger than those of PGs or caffeine. These results suggest that pregnant human myometrium contracts with low Ca2+ by a calmodulin sensitive system. The data also indicate that direct application of PGF2 alpha, or PGE2 into the cells discharges Ca2+ from Ca2+ storage sites and that oxytocin extricates Ca2+ via a pathway involving InsP3 by activation of phosphoinositide turnover. We suggest that these agents induce added contractile responses due to a Ca2+ release mechanism from store sites in addition to the influx of Ca2+ from the extracellular space.

Setting standards for the levels of endotoxin in the embryo culture media of human in vitro fertilization and embryo transfer

OBJECTIVE To clarify the optimum set standards for the levels of endotoxin in a culture medium for human IVF-ET programs. DESIGN Retrospective and randomized study. SETTING One hundred patients underwent 163 cycles of IVF-ET at Fukuoka University Hospital. INTERVENTIONS Measurements for endotoxin were performed using the Limulus Amoebocyte Lysate test. The negative group was administered media with < 1 pg/mL of endotoxin and the positive group was given media with > or = 1 pg/mL of endotoxin. MAIN OUTCOME MEASURES Human embryo development, clinical outcomes, and pregnancy outcomes were evaluated in each group and also the levels of endotoxin. RESULTS The rates of clinical pregnancy (26.1%) and the live birth rates (20.7%) in the negative media were significantly higher than those in the positive media (9.9% and 5.6%, respectively). Rates of embryo development were decreased and a gestational sac and fetal heart beat were not detected in media with > 2 pg/mL of endotoxin. CONCLUSION The set standard for the level of endotoxin in a medium should be < 1 pg/mL to obtain the best outcome. The allowable levels of endotoxin in a medium should be < or = 2 pg/mL in human IVF.