As the prevalence of pregnancies with advanced maternal age increases, the risk of fetal chromoso... more As the prevalence of pregnancies with advanced maternal age increases, the risk of fetal chromosomal abnormalities is on the rise. Therefore, prenatal genetic screening and diagnosis have become essential elements in contemporary obstetrical care. Trophoblast retrieval and isolation from the cervix (TRIC) is a non-invasive procedure that can be utilized for prenatal genetic diagnosis. The method involves the isolation of fetal cells (extravillous trophoblasts) by transcervical sampling; along with its non-invasiveness, TRIC exhibits many other advantages such as its usefulness in early pregnancy at 5 weeks of gestation, and no interference by various fetal and maternal factors. Moreover, the trophoblast yields from TRIC can provide valuable information about obstetrical complications related to abnormal placentation even before clinical symptoms arise. The standardization of this clinical tool is still under investigation, and the upcoming advancements in TRIC are expected to meet t...
Trophoblasts retrieval and isolation from the cervix (TRIC) is a non-invasive method which enable... more Trophoblasts retrieval and isolation from the cervix (TRIC) is a non-invasive method which enables analysis of fetal genetic information from the extravillous trophoblast cells (EVTs). The aim of this study was to compare the efficacy of the HLA-G antibodies—G233 and 4H84—in isolating EVT cells and provide an optimized protocol of TRIC. We analyzed EVTs from 23 pregnant women in between 5 to 20 weeks of gestation who underwent invasive prenatal testing. Two HLA-G antibodies—G233 and 4H84—were used in a subgroup of 11 and 12 samples for immunomagnetic isolation. Cells with β-hCG expression were counted to compare the rate of isolated trophoblast cells. The rate of β-hCG positive cells was significantly different between the G233 and the 4H84 by immunefluorescence microscopy (p < 0.001). The percentage of β-hCG expressing cells in G233 and 4H84 groups were 62.4 ± 8.24% and 82.6 ± 7.1%, respectively (p < 0.001). The average fetal cell positive rate was 14.1 ± 3.78 in the G233 gro...
Preterm birth (PTB) is a global health issue and one of the most challenging problems affecting 1... more Preterm birth (PTB) is a global health issue and one of the most challenging problems affecting 12.9 million births worldwide. PTB is a multi-etiological disease and remains incompletely understood. The major cause of PTB is infection or inflammation and disruption of the vaginal microbiome, which affects the maternal immunologic response leading to PTB. The vaginal microbiome composition changes by a shift in the community are typically dominated by Lactobacillus during pregnancy. There are complex interactions between the maternal microbiome in pregnancy and the development of PTB, therefore, researchers have struggled to connect the maternal microbiome with the dysregulation of the maternal immune response in cases of PTB. The host microbiome affects alterations of the microorganisms with external stimuli such as disease, nutrition, immunity, and behavior. In this review, we discuss the complex association between the maternal microbiome and the risk of PTB and also focus on rece...
Objective: In an attempt to further maximize the potential of genetic analysis from fetal cells i... more Objective: In an attempt to further maximize the potential of genetic analysis from fetal cells isolation, fetal nucleated red blood cell (FNRBC) recovery with direct anti-gamma hemoglobin staining after density gradient and depletion was compared with three different whole blood magnetic separations (1-step and 2-step ferrofluid, 2-step Dynal beads). Methods: In model systems such as quantitatively defined spikes of fetal into adult blood, as well as blood samples after surgical termination procedures, fetal cell yield and purity through the results of fluorescence in situ hybridization (FISH), quantitative real time polymerase chain reaction (PCR), and fluorescence-activated cell sorting (FACS) were calculated. Results: The yield of total number of cells with a XY signal after FISH was the highest on direct anti-gamma hemoglobin staining. After normalizing the results of each experiment to the corresponding result from anti-gamma hemoglobin staining (1), ratio is 0.42 in 1-step fe...
The use of nanoscale materials (NMs) could cause problems such as cytotoxicity, genomic aberratio... more The use of nanoscale materials (NMs) could cause problems such as cytotoxicity, genomic aberration, but the impacts of NM exposure during pregnancy remain uncharacterized in the context of clinical applications.
Objective: To investigate whether fetal microchimeric cells were detected in ovarian tissues with... more Objective: To investigate whether fetal microchimeric cells were detected in ovarian tissues with pelvic endometriosis. Methods: Ovarian tissues with endometriosis were obtained from five women who had at least one live-born son and who underwent enucleation of endometriotic cyst or oophorectomy after a diagnosis of endometriotic cyst. Control tissues were obtained from five women with endometriosis who had no pregnant history. Tissue sections were analyzed with fluorescence in situ hybridization for the presence of fetal cells, defined by X and Y chromosome. Results: Fluorescence in situ hybridization using paraffin-embedded ovarian specimens was performed successfully. Male cells were found in ovarian tissues from all five patients. No male cells were found in ovarian tissues from all five controls. Conclusion: Fetal microchimeric cells, possibly from feto-maternal cell trafficking were detected in ovarian tissues with endometriosis were obtained from women who had prior male preg...
To estimate the combined screening performance of first and early second trimester prenatal serum... more To estimate the combined screening performance of first and early second trimester prenatal serum markers for Down syndrome, in screening for the development of preeclampsia, and analyze the correlation among marker levels, week of onset, and severity of the disease. A retrospective cohort study was carried out on 32 women with preeclampsia and 3044 controls. Serum samples from these pregnancies were assayed for pregnancy-associated plasma protein-A (PAPP-A), alpha-fetoprotein (AFP), unconjugated estriol (uE3), human chorionic gonadotrophin (hCG), and inhibin-A. A likelihood ratio and the odds of being affected given a positive result (OAPR) of various combinations of markers were calculated and receiver operating characteristic (ROC) curves analysis was performed. In the pregnancies that subsequently developed preeclampsia, first trimester PAPP-A concentration was significantly lower and concentrations of early second trimester inhibin-A and hCG significantly elevated. Levels of early second trimester uE3 and AFP were not significantly altered. We also found that inhibin-A correlates with both onset of the disease and the severity. Down syndrome biochemical markers levels are altered in those patients who subsequently developed preeclampsia and may be a useful screening test for preeclampsia. Inhibin-A is the most predictive marker and correlates with the severity of subsequent preeclampsia and inversely with the week of occurrence of preeclampsia.
Recurrent pregnancy loss (RPL) is defined as at least three pregnancy losses in series prior to t... more Recurrent pregnancy loss (RPL) is defined as at least three pregnancy losses in series prior to the 20-28 weeks of pregnancy. There are several etiological factors associated with immunology, anatomy, endocrinology, genetic, infection, chromosomal abnormalities, and environmental factors contributing to the condition. The aim of this study was to identify RPL associated factors in human blood using proteomics. Since it is difficult to obtain tissues or follicular fluids, we used blood samples from normal and RPL patients to conduct a comparative proteomic study. Three RPL blood samples and one cocktailed blood sample from 3 normal women were used. We performed 2-DE and selected spots were analyzed with MALDI-TOF/MS. In the three RPL blood samples, 2-DE analysis revealed 549, 563 and 533 spots to be differentially expressed, respectively. Through a comparative analysis between the control and RPL, 21 spots were shown to be differentially expressed. Of these, 5 proteins were confirmed by Western blot analysis. One of these proteins, ITI-H4 (inter-α trypsin inhibitor-heavy chain 4), was weakly expressed at a molecular weight of 120 kDa, but was highly expressed at a modified molecular weight of 36 kDa in RPL patients. These findings suggest that ITI-H4 expression may be used as a biomarker, which could facilitate the development of novel diagnostic and therapeutic tools.
This study examined whether epistasis between single nucleotide polymorphisms (SNPs) within propr... more This study examined whether epistasis between single nucleotide polymorphisms (SNPs) within proprotein convertase subtilisin/kexin type 1 (PCSK1) and dopamine β-hydroxylase (DBH) genes is associated with premature ovarian failure (POF). One hundred twenty women with POF and 222 female controls were recruited for this study. To genotype SNPs within PCSK1 and DBH, we used a GoldenGate assay with VeraCode technology, which uses an allele-specific primer extension method. Two SNPs (rs155979 and rs3762986) within PCSK1 and one SNP (rs1611114) within DBH, which were located in the 5&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39; flanking region, were involved in synergistic interactions. The C allele in the rs155979 SNP showed an increased risk of POF in a dominant model when AA genotype in the rs1611114 SNP was present (odds ratio, 3.60; 95% CI, 1.82-7.14; P = 0.00024), whereas the G allele in the rs1611114 SNP showed a reduced risk of POF in a dominant model when at least one C allele at the rs155979 SNP was present (odds ratio, 0.24; 95% CI, 0.11-0.51; P = 0.00018) or one G allele at the rs3762986 SNP was present (odds ratio, 0.33; 95% CI, 0.19-0.60; P = 0.00023). Epistases between SNPs within PCSK1 and DBH genes are significantly associated with susceptibility or resistance to POF.
To compare gene expression profiles of placentas from preeclamptic and normal pregnancies. We per... more To compare gene expression profiles of placentas from preeclamptic and normal pregnancies. We performed microarray experiments to analyze genome-wide expression profiling for 10 placentas from pregnant women with preeclampsia and 10 placentas from women who experienced noncomplicated pregnancies (CON), and to identify dysregulated signaling pathways as well as genes in preeclampsia. RT-PCR, real-time RT-PCR and/or immunofluorescence analyses were performed to validate the data obtained from microarray experiments. Unsupervised hierarchical clustering showed heterogeneity of preeclampsia at the molecular levels, whereas expression profiles of preeclampsia are distinctly different from those of CON. A list of genes which are differentially expressed between preeclampsia and CON included well known preeclampsia markers, such as Flt-1, leptin, HTRA1 and SIGLEC6. Gene Set Enrichment Analysis, a pathway-oriented analysis method for expression profiles, provided evidence that a number of biological activities including pathways that regulate actin cytoskeleton, TGFβ signaling, oxidative phosphorylation, and proteasome activity were aberrantly either up-regulated or down-regulated in preeclampsia. RT-PCR and real-time-RT-PCR for genes contributing these biological pathways (gene sets) enriched in either CON or preeclampsia reinforced that these biological processes were systemically dysregulated in preeclampsia. Genome-wide expression profiles of preeclampsia showed heterogeneous characteristics of preeclampsia at the molecular levels. Dysregulation of genes and biological pathways could contribute to abnormal behavior of preeclmapsia. Our results will help further understand underlying mechanisms by which preeclampsia affects placental physiology.
As the prevalence of pregnancies with advanced maternal age increases, the risk of fetal chromoso... more As the prevalence of pregnancies with advanced maternal age increases, the risk of fetal chromosomal abnormalities is on the rise. Therefore, prenatal genetic screening and diagnosis have become essential elements in contemporary obstetrical care. Trophoblast retrieval and isolation from the cervix (TRIC) is a non-invasive procedure that can be utilized for prenatal genetic diagnosis. The method involves the isolation of fetal cells (extravillous trophoblasts) by transcervical sampling; along with its non-invasiveness, TRIC exhibits many other advantages such as its usefulness in early pregnancy at 5 weeks of gestation, and no interference by various fetal and maternal factors. Moreover, the trophoblast yields from TRIC can provide valuable information about obstetrical complications related to abnormal placentation even before clinical symptoms arise. The standardization of this clinical tool is still under investigation, and the upcoming advancements in TRIC are expected to meet t...
Trophoblasts retrieval and isolation from the cervix (TRIC) is a non-invasive method which enable... more Trophoblasts retrieval and isolation from the cervix (TRIC) is a non-invasive method which enables analysis of fetal genetic information from the extravillous trophoblast cells (EVTs). The aim of this study was to compare the efficacy of the HLA-G antibodies—G233 and 4H84—in isolating EVT cells and provide an optimized protocol of TRIC. We analyzed EVTs from 23 pregnant women in between 5 to 20 weeks of gestation who underwent invasive prenatal testing. Two HLA-G antibodies—G233 and 4H84—were used in a subgroup of 11 and 12 samples for immunomagnetic isolation. Cells with β-hCG expression were counted to compare the rate of isolated trophoblast cells. The rate of β-hCG positive cells was significantly different between the G233 and the 4H84 by immunefluorescence microscopy (p < 0.001). The percentage of β-hCG expressing cells in G233 and 4H84 groups were 62.4 ± 8.24% and 82.6 ± 7.1%, respectively (p < 0.001). The average fetal cell positive rate was 14.1 ± 3.78 in the G233 gro...
Preterm birth (PTB) is a global health issue and one of the most challenging problems affecting 1... more Preterm birth (PTB) is a global health issue and one of the most challenging problems affecting 12.9 million births worldwide. PTB is a multi-etiological disease and remains incompletely understood. The major cause of PTB is infection or inflammation and disruption of the vaginal microbiome, which affects the maternal immunologic response leading to PTB. The vaginal microbiome composition changes by a shift in the community are typically dominated by Lactobacillus during pregnancy. There are complex interactions between the maternal microbiome in pregnancy and the development of PTB, therefore, researchers have struggled to connect the maternal microbiome with the dysregulation of the maternal immune response in cases of PTB. The host microbiome affects alterations of the microorganisms with external stimuli such as disease, nutrition, immunity, and behavior. In this review, we discuss the complex association between the maternal microbiome and the risk of PTB and also focus on rece...
Objective: In an attempt to further maximize the potential of genetic analysis from fetal cells i... more Objective: In an attempt to further maximize the potential of genetic analysis from fetal cells isolation, fetal nucleated red blood cell (FNRBC) recovery with direct anti-gamma hemoglobin staining after density gradient and depletion was compared with three different whole blood magnetic separations (1-step and 2-step ferrofluid, 2-step Dynal beads). Methods: In model systems such as quantitatively defined spikes of fetal into adult blood, as well as blood samples after surgical termination procedures, fetal cell yield and purity through the results of fluorescence in situ hybridization (FISH), quantitative real time polymerase chain reaction (PCR), and fluorescence-activated cell sorting (FACS) were calculated. Results: The yield of total number of cells with a XY signal after FISH was the highest on direct anti-gamma hemoglobin staining. After normalizing the results of each experiment to the corresponding result from anti-gamma hemoglobin staining (1), ratio is 0.42 in 1-step fe...
The use of nanoscale materials (NMs) could cause problems such as cytotoxicity, genomic aberratio... more The use of nanoscale materials (NMs) could cause problems such as cytotoxicity, genomic aberration, but the impacts of NM exposure during pregnancy remain uncharacterized in the context of clinical applications.
Objective: To investigate whether fetal microchimeric cells were detected in ovarian tissues with... more Objective: To investigate whether fetal microchimeric cells were detected in ovarian tissues with pelvic endometriosis. Methods: Ovarian tissues with endometriosis were obtained from five women who had at least one live-born son and who underwent enucleation of endometriotic cyst or oophorectomy after a diagnosis of endometriotic cyst. Control tissues were obtained from five women with endometriosis who had no pregnant history. Tissue sections were analyzed with fluorescence in situ hybridization for the presence of fetal cells, defined by X and Y chromosome. Results: Fluorescence in situ hybridization using paraffin-embedded ovarian specimens was performed successfully. Male cells were found in ovarian tissues from all five patients. No male cells were found in ovarian tissues from all five controls. Conclusion: Fetal microchimeric cells, possibly from feto-maternal cell trafficking were detected in ovarian tissues with endometriosis were obtained from women who had prior male preg...
To estimate the combined screening performance of first and early second trimester prenatal serum... more To estimate the combined screening performance of first and early second trimester prenatal serum markers for Down syndrome, in screening for the development of preeclampsia, and analyze the correlation among marker levels, week of onset, and severity of the disease. A retrospective cohort study was carried out on 32 women with preeclampsia and 3044 controls. Serum samples from these pregnancies were assayed for pregnancy-associated plasma protein-A (PAPP-A), alpha-fetoprotein (AFP), unconjugated estriol (uE3), human chorionic gonadotrophin (hCG), and inhibin-A. A likelihood ratio and the odds of being affected given a positive result (OAPR) of various combinations of markers were calculated and receiver operating characteristic (ROC) curves analysis was performed. In the pregnancies that subsequently developed preeclampsia, first trimester PAPP-A concentration was significantly lower and concentrations of early second trimester inhibin-A and hCG significantly elevated. Levels of early second trimester uE3 and AFP were not significantly altered. We also found that inhibin-A correlates with both onset of the disease and the severity. Down syndrome biochemical markers levels are altered in those patients who subsequently developed preeclampsia and may be a useful screening test for preeclampsia. Inhibin-A is the most predictive marker and correlates with the severity of subsequent preeclampsia and inversely with the week of occurrence of preeclampsia.
Recurrent pregnancy loss (RPL) is defined as at least three pregnancy losses in series prior to t... more Recurrent pregnancy loss (RPL) is defined as at least three pregnancy losses in series prior to the 20-28 weeks of pregnancy. There are several etiological factors associated with immunology, anatomy, endocrinology, genetic, infection, chromosomal abnormalities, and environmental factors contributing to the condition. The aim of this study was to identify RPL associated factors in human blood using proteomics. Since it is difficult to obtain tissues or follicular fluids, we used blood samples from normal and RPL patients to conduct a comparative proteomic study. Three RPL blood samples and one cocktailed blood sample from 3 normal women were used. We performed 2-DE and selected spots were analyzed with MALDI-TOF/MS. In the three RPL blood samples, 2-DE analysis revealed 549, 563 and 533 spots to be differentially expressed, respectively. Through a comparative analysis between the control and RPL, 21 spots were shown to be differentially expressed. Of these, 5 proteins were confirmed by Western blot analysis. One of these proteins, ITI-H4 (inter-α trypsin inhibitor-heavy chain 4), was weakly expressed at a molecular weight of 120 kDa, but was highly expressed at a modified molecular weight of 36 kDa in RPL patients. These findings suggest that ITI-H4 expression may be used as a biomarker, which could facilitate the development of novel diagnostic and therapeutic tools.
This study examined whether epistasis between single nucleotide polymorphisms (SNPs) within propr... more This study examined whether epistasis between single nucleotide polymorphisms (SNPs) within proprotein convertase subtilisin/kexin type 1 (PCSK1) and dopamine β-hydroxylase (DBH) genes is associated with premature ovarian failure (POF). One hundred twenty women with POF and 222 female controls were recruited for this study. To genotype SNPs within PCSK1 and DBH, we used a GoldenGate assay with VeraCode technology, which uses an allele-specific primer extension method. Two SNPs (rs155979 and rs3762986) within PCSK1 and one SNP (rs1611114) within DBH, which were located in the 5&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;#39; flanking region, were involved in synergistic interactions. The C allele in the rs155979 SNP showed an increased risk of POF in a dominant model when AA genotype in the rs1611114 SNP was present (odds ratio, 3.60; 95% CI, 1.82-7.14; P = 0.00024), whereas the G allele in the rs1611114 SNP showed a reduced risk of POF in a dominant model when at least one C allele at the rs155979 SNP was present (odds ratio, 0.24; 95% CI, 0.11-0.51; P = 0.00018) or one G allele at the rs3762986 SNP was present (odds ratio, 0.33; 95% CI, 0.19-0.60; P = 0.00023). Epistases between SNPs within PCSK1 and DBH genes are significantly associated with susceptibility or resistance to POF.
To compare gene expression profiles of placentas from preeclamptic and normal pregnancies. We per... more To compare gene expression profiles of placentas from preeclamptic and normal pregnancies. We performed microarray experiments to analyze genome-wide expression profiling for 10 placentas from pregnant women with preeclampsia and 10 placentas from women who experienced noncomplicated pregnancies (CON), and to identify dysregulated signaling pathways as well as genes in preeclampsia. RT-PCR, real-time RT-PCR and/or immunofluorescence analyses were performed to validate the data obtained from microarray experiments. Unsupervised hierarchical clustering showed heterogeneity of preeclampsia at the molecular levels, whereas expression profiles of preeclampsia are distinctly different from those of CON. A list of genes which are differentially expressed between preeclampsia and CON included well known preeclampsia markers, such as Flt-1, leptin, HTRA1 and SIGLEC6. Gene Set Enrichment Analysis, a pathway-oriented analysis method for expression profiles, provided evidence that a number of biological activities including pathways that regulate actin cytoskeleton, TGFβ signaling, oxidative phosphorylation, and proteasome activity were aberrantly either up-regulated or down-regulated in preeclampsia. RT-PCR and real-time-RT-PCR for genes contributing these biological pathways (gene sets) enriched in either CON or preeclampsia reinforced that these biological processes were systemically dysregulated in preeclampsia. Genome-wide expression profiles of preeclampsia showed heterogeneous characteristics of preeclampsia at the molecular levels. Dysregulation of genes and biological pathways could contribute to abnormal behavior of preeclmapsia. Our results will help further understand underlying mechanisms by which preeclampsia affects placental physiology.
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