Molecular cloning of a human oxytocin receptor.
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Amino Acid Sequence↗
Biomedical subjects
C Azuma
Publications and source records attributed to C Azuma.
Expression and localization of oxytocin receptor gene in human uterine endometrium in relation to the menstrual cycle.
The expression of oxytocin receptor mRNA in the human uterus was investigated by polymerase chain reaction after mRNA reverse transcription; in addition, the mRNA was localized by means of in situ hybridization histochemistry. Strong positive signals were observed in myometrial cells at parturition. In nonpregnant human uterine endometrium, oxytocin receptor mRNA was expressed in glandular epithelial cells, but was not observed in stromal cells. The signal in the epithelial cells was stronger at the time of ovulation than during any other phase. The sequential changes in oxytocin receptor mRNA expression in nonpregnant endometrium suggest some alternative biological function of oxytocin in endometrium during the menstrual cycle.
Base Sequence↗
Histogenetic analysis of ovarian germ cell tumors by DNA fingerprinting.
The histogenesis of ovarian germ cell tumors (11 mature teratomas, three malignant transformations of mature teratomas, two immature teratomas, and four dysgerminomas) was investigated genetically using minisatellite DNA probes 33.15 and 33.6 for person-specific restriction fragment length polymorphism (DNA fingerprint) analysis. The DNA fingerprints of six ovarian teratomas were identical with those of mononuclear cells from each host, while some polymorphic bands observed in the host mononuclear cells were lost in the DNA fingerprints of the other five cases. The cases of malignant transformation of mature teratoma and immature teratoma showed that some polymorphic bands of DNA fingerprints from the host mononuclear cells were absent in the tumor tissues. In four cases with dysgerminomas, the DNA fingerprints of tumors were completely identical with those of the respective host mononuclear cells. The present results suggest that mature cystic teratomas of the ovary arise from germ cells arrested at various stages of meiosis, while immature teratomas are derived from postmeiotic germ cells. Malignant transformation may occur exclusively in the mature teratomas arising from postmeiotic germ cells. Dysgerminomas develop from premeiotic oogonia (primordial germ cells). Thus, DNA fingerprints are a useful and sensitive tool for identifying the pathogenesis of germ cell tumours.
Adolescent↗
Malignant cell-specific gelatinase activity in human endometrial carcinoma.
BACKGROUND: The protease activity leading to degradation of the extracellular matrix was compared between human endometrial cancer and normal uterine endometrium. METHODS: Conditioned medium from tumor cells and normal endometrial cells was subjected to electrophoresis on sodium dodecyl sulfate (SDS)-polyacrylamide gel containing gelatin as a substrate. After electrophoresis, the gel was stained with Coomassie blue, and then the enzyme activity, expressed as the zone of dye clearing, was analyzed by densitometry. RESULTS: Densitometric analysis showed that all the endometrial cancers expressed a very high molecular weight enzyme activity (Mr 220,000), which was not detected in medium from normal endometrial cells. The analysis also showed that in endometrial cancer the activity of a Mr 92,000 enzyme was always superior to that of a Mr 64,000 enzyme, which was in contrast to the situation for normal endometrium. CONCLUSIONS: These results indicate that the expression of Mr 220,000 enzyme activity and the higher activity of the Mr 92,000 enzyme than the Mr 64,000 enzyme are involved in the malignant phenotype of native endometrial cancer.
Adenocarcinoma↗
Triplet pregnancy involving complete hydatidiform mole and two fetuses: genetic analysis by deoxyribonucleic acid fingerprint.
A case of a triplet pregnancy involving a dizygous twin pregnancy and a complete hydatidiform mole after therapy with human menopausal gonadotropin and human chorionic gonadotropin is reported. Two female fetuses, two placentas in one mass with two amnions and two chorions, and a tumor mass with a grapelike appearance were spontaneously delivered at 19 weeks of gestation. The deoxyribonucleic acid fingerprints of the two placentas and tumor tissue were compared with those of the parents. The fingerprints of the placentas showed patterns different from each other; however, all their polymorphic fragments could be traced back to either the father or mother. All polymorphic fragments of the tumor tissue were inherited only from the father (androgenesis). These results indicated that this triplet pregnancy involved a dizygous twin pregnancy and a complete hydatidiform mole.
Adult↗
Estimation by an electrophysiological method of the expression of oxytocin receptor mRNA in human myometrium during pregnancy.
In order to evaluate the changes in uterine oxytocin receptor-specific mRNA during pregnancy, receptor expression in Xenopus oocytes are examined electrophysiologically following microinjection of mRNA from human uterus. In voltage-clamped oocytes injected with term myometrial mRNA, oxytocin elicited an inward current response. The amplitude of the oxytocin-induced current increased with increasing dose of oxytocin, but no current was elicited following stimulation with vasopressin. The oxytocin-induced current was completely eliminated as a result of pretreatment with a specific oxytocin antagonist. 21 of 27 oocytes injected with term myometrial mRNA showed a large amplitude (77.0 +/- 16.1 nA) reaction to oxytocin. In comparison, only 3 of 13 oocytes injected with early gestational myometrial mRNA exhibited a small amplitude (4.6 +/- 1.4 nA) reaction to oxytocin. No oxytocin response was observed in oocytes injected with non-pregnant myometrial mRNA. These results indicate that the striking increment in oxytocin sensitivity in term uterus depends on the increase in mRNA encoding oxytocin receptors.
Animals↗
The fetus as an allograft: immunobiologic role of human trophoblasts for fetal survival.
OBJECTIVE: The hypothesis of this study is that trophoblasts contribute to the survival of the mammalian fetal allograft. STUDY DESIGN: Immunologic and molecular biologic studies were performed with human trophoblasts or trophoblastic tumor cells to investigate the expression of class I human leukocyte antigens, the susceptibility to natural killer and lymphokine-activated cells, the ontogeny of Fc gamma-receptors, and the production of immunosuppressive factors. RESULTS: Heterogeneous expression of class I human leukocyte antigens on trophoblasts was regulated at transcriptional level. Trophoblasts showed low susceptibility to natural killer and lymphokine-activated cells. Trophoblasts expressed natural killer cell-type Fc gamma-receptor III, which mediates phagocytosis. The trophoblast-derived immunosuppressive factor was very similar to transforming growth factor-beta. CONCLUSION: The trophoblasts, which ultimately form the fetoplacental interface, constitute a major immune barrier for the survival of the allogenic conceptus.
Female↗
Functional expression of human myometrial endothelin receptors in Xenopus laevis oocytes.
We demonstrate the existence of functional endothelin receptors in human uterine myometrium using the Xenopus oocyte expression system. Fifty nanograms of poly(A)+RNA from myometrium was injected into Xenopus laevis oocytes and incubated for 70-80 h. The membrane potential of the oocyte was clamped at -60 mV and membrane current was measured during and after endothelin stimulation. Endothelin-1 elicited a large inward membrane current in the oocytes injected with poly(A)+RNA; endothelin-2 elicited a small current; while endothelin-3 did not induce any membrane current. These results indicate the existence of messenger RNA encoding functional endothelin-1 receptors in human uterine myometrium.
Animals↗
The biological effects of macrophage-colony-stimulating factor induced by progestin on growth and differentiation of endometrial adenocarcinoma cells.
Progestins have biological effects of regression and differentiation on human endometrial adenocarcinoma. We investigated the effects of progestin on the induction of macrophage-colony-stimulating factor (M-CSF) and its receptor messenger RNAs in the human endometrial adenocarcinoma cell line Ishikawa which has receptors for both estrogen and progesterone. Poly(A)+RNA extracted from Ishikawa cells cultured with or without synthetic progestin R5020 was subjected to Northern blot hybridization using M-CSF and c-fms cDNA probes. The expression of M-CSF mRNA in Ishikawa cells increased about 2.3 times following treatment with R5020 at 10(-7) M. Induction of M-CSF mRNA by R5020 was antagonized by anti-progestin RU486 in a dose-dependent manner. However, c-fms mRNA, coding the M-CSF receptor, was expressed constitutively in Ishikawa cells and its expression was not affected by hormonal treatment. We further examined the biological effects of M-CSF on endometrial cancer cells. Colony formation of Ishikawa cells in soft agar, which represents anchorage-independent cell growth, was inhibited by M-CSF treatment. On the other hand, accumulation of glycogen granules in cytoplasm detected by periodicacid-Schiff staining was observed in Ishikawa cells treated with M-CSF. These results indicate that M-CSF, whose gene expression was enhanced by progestin, suppressed growth and induced differentiation of endometrial adenocarcinoma cells. These effects of M-CSF on endometrial cancer cells are similar to those of progestins, so the effects of progestins on these cells are, at least in part, probably mediated by M-CSF in an autocrine or paracrine manner.
Adenocarcinoma↗
Application of gene amplification by polymerase chain reaction to genetic analysis of molar mitochondrial DNA: the detection of anuclear empty ovum as the cause of complete mole.
To investigate the pathogenesis of complete hydatidiform mole (complete mole), we employed a newly developed gene amplification method by polymerase chain reaction (PCR) for the restriction fragment length polymorphism (RFLPs) analysis of extranuclear DNA (mitochondrial DNA) of complete mole. Whole cellular DNA was extracted from six molar tissues and from peripheral blood mononuclear cells of parents. Two hyperpolymorphic regions of mitochondrial DNA, a 1.5-kb-long fragment and a 1.9-kb-long fragment, were selectively amplified from the extracted DNA by the PCR method. The amplification products amounted to over 10 micrograms after 30 cycles of PCR. The PCR products were digested with endonucleases (HaeIII, HinfI, AluI, and TaqI) and then electrophoresed on agarose gel. The electrophoretic pattern of digested DNA showed that the RFLPs of molar mitochondrial DNA coincided with those of the patient, indicating that the mitochondrial DNA of complete mole was inherited from the ovum. As it has been identified that the intranuclear DNA of complete mole is transmitted only from the spermatozoa, our results verify that complete mole results from the fertilization of an anuclear "empty" ovum with normal sperm at the molecular level.
Base Sequence↗
Histogenetic analysis of ovarian teratomas by DNA fingerprinting.
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DNA Fingerprinting↗
The gene expressions of macrophage colony-stimulating factor (MCSF) and MCSF receptor in the human myometrium during pregnancy: regulation by sex steroid hormones.
We investigated the biological effect of sex-steroid hormones, secreted from the corpus luteum and placenta, on the induction of mRNA encoding macrophage colony-stimulating factor (MCSF) and c-fms proto-oncogene (MCSF receptor) in the human uterine myometrium. Poly(A)+RNA was extracted from the myometrium of pregnant and non-pregnant uterine myometrium and then Northern blot analysis was performed on poly(A)+RNA. The myometrium of non-pregnant women expressed neither mRNA of macrophage colony-stimulating factor (MCSF) nor any transcript related to the c-fms proto-oncogene. On the other hand the myometrium of pregnant women expressed MCSF mRNA (4.7 kb) and two kinds of transcript related to the c-fms proto-oncogene (3.9 and 1.3 kb). The mRNAs of both MCSF and c-fms proto-oncogene were induced in the uterine myometrium of non-pregnant women under pseudopregnant therapy of mestranol and norethindrone. These results indicate that sex steroid hormone secreted from the corpus luteum of pregnancy and/or placenta may be deeply involved in the hypertrophic change of uterus during pregnancy by inducing MCSF and MCSF receptor (c-fms proto-oncogene protein product) in the myometrium.
Corpus Luteum↗
Perinatal infection of human T-lymphotropic virus type I, the etiologic virus of adult T-cell leukemia/lymphoma. DNA amplification of specific human T-lymphotropic virus type I sequences.
A gene amplification technique, polymerase chain reaction, was used to detect human T-lymphotropic virus type I (HTLV-I), the etiologic agent of adult T-cell leukemia/lymphoma, in mononuclear cells in peripheral blood and breast milk of ten HTLV-I carrier gravida. The DNA in umbilical cord blood mononuclear cells of the neonates born to the HTLV-I carrier gravida were also amplified and examined for the possibility of HTLV-I infection via placenta during pregnancy. The HTLV-I sequences were detected both in the peripheral blood and milk of all ten carrier gravida by Southern blot analysis of amplified DNA. However, HTLV-I proviral DNA could not be detected in the cord blood of the carriers' neonates, indicating that transplacental infection of HTLV-I should be rare and that postpartum infection via breast milk is a likely major perinatal transmission route.
Antibodies, Neoplasm↗
Studies on the pathogenesis of choriocarcinoma by analysis of restriction fragment length polymorphisms.
The association of complete hydatidiform mole with choriocarcinoma has long been recognized, but it is unknown whether the pathogenesis of the two are identical. We investigated the pathogenesis of these trophoblastic tumors by analyzing restriction fragment length polymorphisms using a minisatellite DNA probe to choriocarcinoma, the complete mole, and normal trophoblasts as well as the parental cells. The polymorphic fragments of the complete mole were all transmitted from the paternal DNA, but some polymorphic fragments of the paternal DNA were not recognized in the complete mole. This confirms at a molecular level the androgenetic origin of the complete mole. In some cases of choriocarcinoma, the pattern of inheritance of restriction fragment length polymorphisms was the same as that in the complete mole, whereas in others all the polymorphic fragments in tumor tissues were identical to those in the host DNA. These results suggest that the pathogenesis of choriocarcinoma varies, being completely different from that of the complete hydatidiform mole in some cases.
Choriocarcinoma↗
Differential diagnosis between complete mole and hydropic abortus by deoxyribonucleic acid fingerprints.
We used a new method of deoxyribonucleic acid fingerprint analysis to obtain the differential diagnosis between complete mole and hydropic abortus. This method with a deoxyribonucleic acid minisatellite probe requires only a small amount of tissue sample and peripheral blood, and presents individual specific restriction fragment length polymorphisms (deoxyribonucleic acid "fingerprints") by simultaneous detection of many hypervariable regions (minisatellite regions) widely dispersed in the human genome. Southern blot hybridization showed that in cases of complete mole, all polymorphic fragments were exclusively inherited from the father. Some of the polymorphic bands of paternal deoxyribonucleic acid were not observed in molar deoxyribonucleic acid. However, in the hydropic abortus, the polymorphic fragments could be traced back to its parent. These results indicate that deoxyribonucleic acid fingerprints could distinguish the abnormal fertilization of complete mole (androgenesis) from the normal fertilization of hydropic abortus by identifying the difference in genetic variations between complete mole and hydropic abortus at the deoxyribonucleic acid level.
Blotting, Southern↗
Expression of the corticotropin-releasing hormone (CRH) gene in human placenta and amniotic membrane.
Immunoreactive corticotropin-releasing hormone (IR-CRH) in maternal plasma increases progressively during pregnancy and decreases rapidly after delivery, suggesting that IR-CRH is produced in the placenta. We studied the expression of the CRH gene in developing human chorionic tissue, the amniotic membrane, the uterine myometrium and a fresh surgical specimen of hydatidiform mole by Northern blot analysis. Our results were as follows: (1) CRH mRNA was demonstrated in the placenta in the third trimester and at term, but under detectable level in the first and second trimesters. (2) CRH mRNA expression was observed in the amniotic membrane, but its expression in the myometrium in normal pregnancy was under detectable level at term. (3) CRH mRNA was also under detectable level in trophoblasts of a hydatidiform mole. These results suggest that the sources of the increased level of IR-CRH in human plasma and amniotic fluid during pregnancy are the placenta and amniotic membrane, and that gene expression of placental CRH increases during pregnancy.
Amnion↗
Gene expression of macrophage colony-stimulating factor and its receptor in human placenta and decidua.
Macrophage colony-stimulating factor (M-CSF) induces proliferation of monocyte/macrophage progenitor cells and can also activate some functions of mature cells including fetally derived placental cells. To study the role of M-CSF in the pregnant female reproductive tract, the expression of M-CSF mRNA and its receptor, c-fms proto-oncogene, in human placenta and decidua was identified. M-CSF and c-fms mRNAs, 4.7Kb and 3.9Kb respectively, were detected by Northern blotting in the early stage placenta and subsequently increased during pregnancy. These mRNAs were not detected in the nonpregnant endometrium but were strongly induced in maternal decidua with the same mRNA size as in the placenta. Northern blot hybridization on the endometrium of a pseudopregnant uterus revealed that the expression of endometrial M-CSF and c-fms mRNAs is regulated by synergistic action of female sex steroid hormones. These findings indicate that, in an autocrine and/or paracrine manner, M-CSF is deeply involved in the local proliferation and differentiation of cells at the materno-fetal interface, and support the placental immunotrophism hypothesis.
Blotting, Northern↗
Site-specific mutagenesis of human chorionic gonadotrophin (hCG)-beta subunit: influence of mutation on hCG production.
The heterodimeric glycoprotein hormones, human chorionic gonadotrophin (hCG), LH, TSH and FSH, consist of two non-covalently linked subunits, the alpha and beta subunits. The beta subunit is specific for each hormone and is responsible for the biological specificity, but the beta subunits of different hormones show some degree of structural homology. The CAGY (cysteine-alanine-glycine-tyrosine) region is one of the amino acid sequences that is homologous in different beta subunits and is highly conserved between species. In the present study, site-specific in-vitro mutagenesis was used to change three individual nucleotides in the centre of the CAGY region of the hCG-beta subunit, and the effects of these mutations on hCG production was determined by in-vitro transcription and then translation in Xenopus laevis oocytes. The results indicate that the CAGY region, particularly the glycine residue at position 36 in the beta subunit, is essential for the production of hCG. This finding is consistent with previous studies showing that this region is necessary for the biological activity of human TSH.
Amino Acid Sequence↗