Hideaki Masuzaki

Detection of cell free placental DNA in maternal plasma: direct evidence from three cases of confined placental mosaicism

Fetal cells are consistently found in the maternal circulation, and polymerase chain reaction based studies have led to the identification of cell free fetal DNA (fetal DNA) in maternal blood. Approximately 1.2 nucleated fetal cells/ml of whole blood from women carrying a male fetus were detectable,1 and relative enrichment of fetal DNA was detected in the maternal plasma and serum.2 The amount of fetal DNA in the maternal blood increases with progression of pregnancy, and 3.4–6.2% of the total maternal plasma DNA during pregnancy was of fetal origin.3 Therefore, cell free fetal DNA in pregnant women’s plasma is useful for non-invasive prenatal diagnosis, especially for detection of fetal sex,3,4 RhD blood type,5–7 and gene mutations of paternal origin.8–10 Previous studies indicated that pregnant women with pre-eclampsia,11 placenta previa12 and fetal chromosome abnormalities13 tend to have elevated levels of fetal DNA in their plasma. Since functional or structural abnormalities of the placenta and destruction of the trophoblast may be associated with these diseases,14 it is suggested that cell free fetal DNA is of placental origin. This implies that quantitative analysis of fetal DNA may be valuable to screen for placental dysfunction. Ng et al15 recently reported that placental mRNA is present in the maternal circulation, and suggested that the same might occur for placental DNA,16 However, no direct evidence has been given for placenta derived cell free fetal DNA in the maternal blood, although its clinical use is growing.17nnConfined placental mosaicism, which is defined by the presence of abnormal karyotypes only in the placenta while the fetus itself is usually diploid,18 may occur through a loss of the extra chromosome in a trisomic zygote during an early mitotic cell division in only the …

Immunoexpression of hepatocyte growth factor and c-Met receptor in the eutopic endometrium predicts the activity of ectopic endometrium

OBJECTIVEnTo investigate the mitogenic and angiogenic activity of the eutopic and ectopic endometrium throughout the menstrual cycle and to examine whether the activity of the eutopic endometrium is useful to predict greater activity of the ectopic endometrium.nnnDESIGNnControlled clinicopathologic study using intact tissue.nnnSETTINGnNagasaki University School of Medicine, Nagasaki, Japan.nnnPATIENT(S)nFifteen infertile women with pelvic endometriosis and 10 women without endometriosis undergoing laparoscopy.nnnINTERVENTION(S)nBiopsies from the ectopic endometrium and the corresponding eutopic endometrium were collected. Immunohistochemical staining was performed using respective antibodies, and a computer analyzed modified quantitative-histogram (Q-H) score was used to quantify immunostaining.nnnMAIN OUTCOME MEASURE(S)nThe immunoreactions of hepatocyte growth factor (HGF), its receptor, c-Met, vascular endothelial growth factor (VEGF), proliferating cell nuclear antigen (PCNA), and von Willebrand factor (VWF) in eutopic and ectopic endometrium were examined, and their relation with different revised American Society for Reproductive Medicine (r-ASRM) stages and the morphology of endometriosis was evaluated.nnnRESULT(S)nThe immunoexpressions of HGF and c-Met were significantly higher in the eutopic endometrium of patients with endometriosis than in that of controls. The Q-H scores of HGF, c-Met, VEGF, PCNA, and microvessel density (MVD) were markedly higher in red peritoneal lesions when compared with other lesions. The Q-H scores did not reveal r-ASRM stage-dependent variation in any of these markers. We observed a significant correlation between the immunoexpressions of HGF, c-Met, and PCNA or microvessel counts. When we combined the Q-H scores of the glandular epithelium and stroma, we found that increased activity of the eutopic endometrium as measured by the immunoreaction of HGF, c-Met, VEGF, PCNA, and MVD was similar to highly active red lesions and was significantly higher than that of controls and other lesions.nnnCONCLUSION(S)nImmunoexpression of HGF and c-Met in the eutopic endometrium of patients with pelvic endometrioisis is possibly useful to predict greater activity of the ectopic endometrium.

Microarray comparative genomic hybridization (CGH)-based prenatal diagnosis for chromosome abnormalities using cell-free fetal DNA in amniotic fluid

AbstractCell-free fetal DNA (cffDNA) in the supernatant of amniotic fluid, which is usually discarded, can be used as a sample for prenatal diagnosis. For rapid prenatal diagnosis of frequent chromosome abnormalities, for example trisomies 13, 18, and 21, and monosomy X, using cffDNA, we have developed a targeted microarray-based comparative genomic hybridization (CGH) panel on which BAC clones from chromosomes 13, 18, 21, X, and Y were spotted. Microarray-CGH analysis was performed for a total of 13 fetuses with congenital anomalies using cffDNA from their uncultured amniotic fluid. Microarray CGH with cffDNA led to successful molecular karyotyping for 12 of 13 fetuses within 5 days. Karyotypes of the 12 fetuses (one case of trisomy 13, two of trisomy 18, two of trisomy 21, one of monosomy X, and six of normal karyotype) were later confirmed by conventional chromosome analysis using cultured amniocytes. The one fetus whose molecular-karyotype was indicated as normal by microarray CGH actually had a balanced translocation, 45,XY,der(14;21)(q10;q10). The results indicated that microarray CGH with cffDNA is a useful rapid prenatal diagnostic method at late gestation for chromosome abnormalities with copy-number changes, especially when combined with conventional karyotyping of cultured amniocytes.

Role of DNA methylation and histone H3 lysine 27 methylation in tissue-specific imprinting of mouse Grb10.

ABSTRACT Mouse Grb10 is a tissue-specific imprinted gene with promoter-specific expression. In most tissues, Grb10 is expressed exclusively from the major-type promoter of the maternal allele, whereas in the brain, it is expressed predominantly from the brain type promoter of the paternal allele. Such reciprocally imprinted expression in the brain and other tissues is thought to be regulated by DNA methylation and the Polycomb group (PcG) protein Eed. To investigate how DNA methylation and chromatin remodeling by PcG proteins coordinate tissue-specific imprinting of Grb10, we analyzed epigenetic modifications associated with Grb10 expression in cultured brain cells. Reverse transcriptase PCR analysis revealed that the imprinted paternal expression of Grb10 in the brain implied neuron-specific and developmental stage-specific expression from the paternal brain type promoter, whereas in glial cells and fibroblasts, Grb10 was reciprocally expressed from the maternal major-type promoter. The cell-specific imprinted expression was not directly related to allele-specific DNA methylation in the promoters because the major-type promoter remained biallelically hypomethylated regardless of its activity, whereas gametic DNA methylation in the brain type promoter was maintained during differentiation. Histone modification analysis showed that allelic methylation of histone H3 lysine 4 and H3 lysine 9 were associated with gametic DNA methylation in the brain type promoter, whereas that of H3 lysine 27 regulated by the Eed PcG complex was detected in the paternal major-type promoter, corresponding to its allele-specific silencing. Here, we propose a molecular model that gametic DNA methylation and chromatin remodeling by PcG proteins during cell differentiation cause tissue-specific imprinting in embryonic tissues.

Higher activity by opaque endometriotic lesions than nonopaque lesions

Background.u2002 Higher activity by early endometriosis than advanced endometriosis has been reported. However, the pattern of activity in individual colored endometriotic lesions in pelvic cavity is unknown. We investigated the variation in activity of the different colored morphologic lesions as proposed by the current revised American Society of Reproductive Medicine (ASRM) classification in women with endometriosis.

Modified Reduction Surgery for Adenomyosis

We describe the preliminary clinical results of a modified method of reduction surgery for easy approach and effective removal of lesions in women with adenomyosis. Old classical reduction surgery was performed in 5 women with imaging diagnosis of adenomyosis who were selected retrospectively among 104 patients undergoing conservative surgery. A transverse H incision method in the reduction surgery was applied to 6 of 83 patients wishing to preserve fertility. Benefit in operative procedure, complications, patients’ compliance, and pregnancy outcome were analyzed and compared between 5 women with the classical method and 6 women with the H incision method for adenomyosis. No apparent difference was observed in the operation time, blood loss and volume of excised specimens between these two groups. The major complication of perforation during surgery occurred in 2 patients (40%) by the classical method and in only 1 patient (17%) by the H incision technique. The subjective relief of pain was relatively more evident in the modified than in the classical group. There was no case of pregnancy in the classical group; however, 1 patient conceived spontaneously 4 months after operation by this H incision procedure. Our H incision technique may be considered a useful method for an easy surgical approach and satisfactory removal of adenomyotic lesions and may be better than the old classical method of reduction surgery.

Association of interleukin-6 and estradiol with hepatocyte growth factor in peritoneal fluid of women with endometriosis

Background.u2003 Different cytokines and ovarian steroid hormones have been reported to regulate the growth and maintenance of endometriosis. We determined the relationship between peritoneal fluid concentrations of interleukin‐6, ovarian steroids and hepatocyte growth factor in different revised American Fertility Society (AFS) staging and morphologic appearances of endometriosis.

A strong association between human earwax-type and apocrine colostrum secretion from the mammary gland

Here we provided the first genetic evidence for an association between the degree of apocrine colostrum secretion and human earwax type. Genotyping at the earwax-type locus, rs17822931 within the ABCC11 gene, revealed that 155 of 225 Japanese women were dry-type and 70 wet-type. Frequency of women without colostrum among dry-type women was significantly higher than that among wet-type women (Pxa0<xa00.0002), and the measurable colostrum volume in dry-type women was significantly smaller than in wet-type women (Pxa0=xa00.0341).

Increased level of cell-free placental mRNA in a subgroup of placenta previa that needs hysterectomy.

The purpose of this study was to investigate whether cell‐free placental mRNA levels have the potential to predict a placenta previa resulting in hysterectomy.

Management of prenatal ovarian cysts

OBJECTIVESnThe aim of the present study was to analyze the antenatal and postnatal outcome of fetal ovarian cysts in relation to their ultrasonographic pattern and size.nnnMETHODSnSixteen fetal ovarian cysts were diagnosed in 16 fetuses and followed with serial ultrasonograms in utero and after birth until spontaneous or surgical resolution.nnnRESULTSnEleven fetal ovarian cysts were simple cysts at first prenatal scan but 3 of the 11 became complex cysts at last prenatal scan and required postnatal laparoscopic surgery. Seven of the 11 simple cysts (63%) disappeared on follow-up imaging by ultrasonograms or MRI during pregnancy or within 2 months after birth. The rate of spontaneous resolution of simple cysts was higher than that of complex cysts (40.0%). The mean maximum diameter of the ovarian cysts before delivery that were subsequently excised surgically at postnatal period (50+/-13.4 mm) was not different from that of ovarian cysts that resolved spontaneously (42.8+/-12.8 mm, P=0.2918).nnnCONCLUSIONnIn our study, cyst size did not predict the risk of ovarian loss. The opportunity of laparoscopic exploration versus conservative management needs to be investigated because some complex cysts resolved spontaneously in the postnatal period.