Tetsuaki Hara

Cell-free Fetal DNA in Maternal Circulation after Amniocentesis

After amniocentesis, 5–20% of patients have evidence of fetal-maternal hemorrhage as indicated by increases in maternal serum α-fetoprotein (1)(2)(3)(4)(5) or by the Betke–Kleihauer test (6)(7)(8). The Betke–Kleihauer test can differentiate fetal from maternal erythrocytes by the relative resistance of hemoglobin F-containing cells to acid elution, and it is the most popular method of diagnosing and assessing the severity of fetal-maternal hemorrhage (9). The reliability of this test has been questioned, however, because numerous sources of error are associated with it (10). These sources of error possibly contribute to the wide variation in the reported incidence of fetal-maternal hemorrhage; a more accurate method of assessing fetal-maternal hemorrhage is therefore required in the clinical setting of rhesus D-negative pregnant women. The discovery of cell-free fetal DNA in maternal serum and plasma has opened a new avenue for noninvasive prenatal diagnosis and has provided a useful marker of complicated pregnancies (11)(12)(13)(14)(15)(16). The analysis of fetal DNA in maternal serum or plasma has afforded diagnoses of fetal rhesus D status (12) and single-gene disorders (13), as well as the determination of fetal gender (14)(15). Cell-free DNA may be liberated directly from the fetal-placental interface into the maternal circulation (16), or it may be transferred into the maternal …

Blue rubber bleb nevus syndrome: Report of a patient with hemangiomas of the vaginal portion of the cervix appearing during pregnancy

Blue rubber bleb nevus syndrome (BRBNS) is a rare disorder showing venous malformations in the skin and gastrointestinal tract, and other internal organs. We encountered a patient with BRBNS in whom hemangiomas of the uterine cervix appeared during pregnancy. This was apparently the first reported occurrence. To avoid unexpected bleeding from hemangiomas, patients with BRBNS should be examined repeatedly for hemangiomas of the birth canal, and special care should be taken in deciding the mode of delivery.

Detection of chromosomal abnormalities in uterine leiomyoma using conventional cytogenetic method and interphase fluorescence in situ hybridization

Seventy-nine uterine leiomyomas were examined using a conventional cytogenetic method and fluorescence in situ hybridization (FISH) for detection of chromosomal abnormalities of chromosome 12. Nine (17.6%) of 51 tumor samples examined showed chromosomal abnormalities by conventional cytogenetic analysis. Rearrangements of chromosome 12 were detected in two tumors. Other tumors showed abnormalities affecting chromosomes 1, 4, 6, 7, 10, 13, 14, and 22. For FISH, the whole-chromosome painting probe and the D12Z3 probe specific for the centromeric region were used to detect structural and numerical abnormalities of chromosome 12. Of forty-one tumor samples, six showed structural aberrations and four showed numerical aberrations of chromosome 12 by FISH analysis. Of the tumors with structural aberrations identified by FISH, two had normal karyotypes, two showed structural rearrangements of chromosome 12 cytogenetically, and two could not be analyzed because of an insufficient number of metaphases. There were no correlations between the cytogenetic data and clinical parameters. The results indicate that chromosomal abnormalities are important in the biology of at least some types of uterine leiomyomas, and that FISH is a useful complement to conventional cytogenetic analysis in the study of solid tumors.

Chromosomal FISH analysis of unfertilized human oocytes and polar bodies

AbstractThe incidence of chromosomal aneuploidy was studied in 208 unfertilized metaphase II human oocytes from an in vitro fertilization program by fluorescence in situ hybridization using probes for chromosomes 18, 21, and X. Chromosome spreads were prepared by a gradual fixation-air-drying method. We obtained analyzable results from 183 oocytes and 93 polar bodies; 167 oocytes (91%) were normal, 11 (6%) were diploid, and 5 (3%) were aneuploid. Of the five aneuploid oocytes, four involved chromosome 21, and one involved the X chromosome. In this study, oocyte aneuploidy caused by both nondisjunction of bivalent chromosomes and predivision of univalent chromosomes was observed. The aneuploidy rate (9.8%) in the oocytes from women aged ≧35 years was significantly higher than that (0.7%) in those aged 23 to 34 years (P= 0.0017).

A tilting embryo culture system increases the number of high-grade human blastocysts with high implantation competence

Human embryos normally experience mechanical stimuli during development in vivo. To apply appropriate stimuli to embryos, this study group developed a tilting embryo culture system (TECS) and investigated whether it could improve the grade of fresh human embryos compared with a control static culture system. A total of 450 retrieved oocytes from 32 IVF or intracytoplasmic sperm injection cycles of 32 women were cultured for 5-6 days. Oocytes were divided randomly into TECS and control groups and then were inseminated in vitro. All embryos were evaluated at days 3 and 5 using standard grading criteria for embryo quality. The rates of fertilization per mature oocyte and high-grade cleavage-stage embryo formation in the TECS group were similar to those in the control group. The rates of blastocyst formation and of blastocysts graded 3BB or higher at day 5 were significantly higher in the TECS group than those in the control group: 45.3% (67/148) versus 32.1% (51/159) (P=0.018) and 29.1% (43/148) versus 17.6% (28/159) (P=0.018), respectively. The TECS group produced more high-grade blastocysts than the control group. Embryo movement or mechanical stimulation during embryo culture may be beneficial for human embryonic development.

Detection of chromosomal abnormalities of chromosome 12 in uterine leiomyoma using fluorescence in situ hybridization

SummaryFifty uterine leiomyomas were examined using conventional cytogenetic method and fluorescence in situ hybridization (FISH) for detection of chromosomal, abnormalities of chromosome 12. Of the 50 tumors, nine were examined using FISH on the non-cultured samples. Two (4.0%) of 50 tumor samples examined showed chromosomal abnormalities of chromosome 12 by the conventional cytogenetic analysis. For FISH, the whole-chromosome painting probe and D12Z3 probe specific for the centromeric region were used. Of the 50 cultured samples, 10 showed structural aberrations and four showed numerical aberrations of chromosome 12 by FISH analysis. Of the nine non-cultured samples, four showed structural abnormalities of chromosome 12, all of which also showed structural abnormalities of chromosome 12 on the cultured samples. These results indicate that chromosomal abnormalities of chromosome 12 are important in the biology of at least some types of uterine leiomyoma, and that FISH is a useful complement to the conventional cytogenetic analysis in the study of solid tumors.

Pregnancy rate, multiple pregnancy rate, and embryo quality: Clues for single blastocyst transfer from double blastocyst transfer in an unselected population

ObjectiveMinimizing multiple pregnancy is a priority in assisted reproduction. As implantation rates are critical to success and reduce multiple pregnancy, we investigated whether blastocyst grade determined implantation rate following double blastocyst transfer in unselected cases.Materials and MethodsWe studied 69 three-cleavage stage embryo transfers and 64 two-blastocyst transfers. Two blastocysts, or one when two blastocysts were not available, were transfered after evaluating the grade of blastocysts. The difference in pregnancy and implantation rates to patient age, the number of retrieved oocytes and grade of blastocysts were analyzed.ResultsBlastocyst and grade 3AA rates per fertilized egg were 50.3% and 26.0%, respectively. Following two-blastocyst transfer, pregnancy rate per transfer, implantation rate per embryo, and multiple pregnancy rate per pregnancy were 39.1%, 26.5%, and 24.0%, respectively. Two-blastocyst transfer achieved implantation more often than three-cleavagestage embryo transfer, but did not reduce multiple pregnancy. Pregnancy, implantation, and multiple pregnancy rates did not reflect maternal age. Higher pregnancy and implantation rates per transfer were attained for with six or more oocytes retrieved or transfer of two-blastocyst graded 3AA or higher especially when two or more blastocysts graded 3AA or higher are available, but the latter showed a high multiple pregnancy rate (38.5%).ConclusionsSingle embryo transfer could be carried out when two or more blastocysts of grade 3AA or higher have been developed.

Clinical Application of Elective Single Embryo Transfer at the Blastocyst Stage

ABSTRACT Single embryo transfer is advocated as a strategy to reduce the frequency of multiple births after in vitro fertilization. Transfer of a single, blastocyst stage embryo increases the likelihood of pregnancy and delivery as compared with transfer of a single, cleavage-stage embryo in women under 36 years of age undergoing their first or second trial of in vitro fertilization. The reason for the higher success rate with blactocysts might be mainly related to an embryo selection process. In the near future, this approach might be appropriate for women older than 36 years. A potential drawback to the use of blastocysts is failure to transfer any embryos. Additional concerns are the possibilities of an increased risk of monozygotic twins and an altered sex ratio of births.